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Blog Archives
Guide to what's on in London in February 2019
Guide to what's on in London this February 2019, including festivals, Chinese New Year events, winter pop-ups and more.
It's February and winter is in full swing. The weather is pretty freezing, so a host of London's events are taking place inside. Halfway through the month, things get a bit soppy for Valentine's Day. Throughout the month, there's music, beer and winter pop-ups. There will also be celebrations for Chinese New Year in the West End, while half-term means there's plenty of family-friendly events on around town.
For a guide to London's Valentine's Day events for singles and couples, click here.
- 1 – 2 February : London Remixed Festival
A two-day celebration of emerging talents from the worlds of Latin Grooves, Afro beats, Tropical Bass, Vintage-Remix, Desert Remix Balkan Beats, Urban Roots, Acoustic Soundclash and Brass Band Remix and more. Open: Fri 8pm-1am, Sat 8pm-4am. Tickets: £11.19–£21.79. Rich Mix, 35-47 Bethnal Green Road, Shoreditch, E1 6LA. Nearest station: Shoreditch High Street or Old Street. For tickets, visit the London Remixed Festival website. Check out Metro Girl's blog post on the London Remixed Festival.
- 1 February – 7 April : Lost Lagoon
Bompas & Parr have created an immersive, subterranean journey inspired by the world's most famous buccaneer Captain Henry Morgan. Expect whirlpool cocktail station, beach-inspired food and entertainment. Tickets: £20-£30. West 12 shopping centre, Shepherd's Bush, W12 8PP. Nearest station: Shepherd's Bush. For booking, visit the Lost Lagoon website.
- Now until 1 February : French Winter Village @ The Langham
The Langham Hotel are teaming up with Grey Goose to create a winter paradise in their courtyard garden. Expect hot and cold Grey Goose vodka cocktails, festive food from the woodfire, heaters, cosy blankets and more. Open Wed-Sat 4pm–10pm. Minimum spend of £60pp (for bookings between 6-60 people). The Langham Hotel, 1c Portland Place, Regent Street, W1B 1JA. Nearest station: Oxford Circus. For more information, visit The Langham website.
- 2 February – 14 July : Christian Dior: Designer of Dreams @ V&A
Exhibition of Christian Dior designs, from 1947 to the present day. Open daily 10am-5.45pm. Tickets: £20-£24. Victoria & Albert Museum, Cromwell Road, South Kensington, SW7 2RL. Nearest station: South Kensington. For more information, visit the V&A website.
- 3 February : The Vintage Collections
Vintage fashion fair comes to the stunning art deco surrounds of Freemasons Hall, featuring 50 traders, cocktails and afternoon tea. 11am-5:30pm. Entry: £5 (£2 with NUS card). Freemasons Hall, 60 Great Queen Street, Covent Garden, WC2B 5AZ. Nearest station: Holborn or Covent Garden. For more information, visit the Clerkenwell Vintage Fashion Fair website.
- 3 – 9 February : Pisco Sour Week @ Mr Foggs Society of Exploration
A special menu will be on offer all week celebrating Peru's iconic cocktail. Relax with a concoction in the Map Room or the Train Carriage. Open Mon-Sat 4pm-12am. Mr Foggs Society of Exploration, 1A Bedford Street, Covent Garden, WC2E 9HH. Nearest station: Charing Cross or Covent Garden. For more information, visit Mr Foggs website.
- 3 February – 19 March : Festival of New Masculinity
Inaugural festival from the Book of Man, featuring 10 events across two east London venues. Exploring issues facing modern men today. Including live podcasts, special guest panels, photography exhibition, grooming lounges, boys brunches, and a Penis Gallery. Ticket prices vary. Hoxton Bar & Kitchen, 2-4 Hoxton Square, Hoxton, N1 6NU and the Book Club, 100-106 Leonard Street, Shoreditch, EC2A 4RH. Nearest station: Old Street. For more information, visit the Book Of Man website.
- Now until 3 February : London International Mime Festival
Festival at various events across London including performances of physical theatre, dance, circus, puppetry and live art. Workshops also available. Tickets range from £12.50-£30. Venues include the Barbican, Jacksons Lane, Platform Theatre, Sadler's Wells, Shoreditch Town Hall and Southbank Centre. For more information, visit the Mime London website.
- Now until 3 February : Take A View exhibition
Exhibition of the winning and shortlisted images from this year's Landscape Photographer Of The Year are on display on the balcony at Waterloo station. Open during station hours. Free. Mezzanine level, Waterloo Station, SE1 8SW. Nearest station: Waterloo. For more information, visit the Take A View website.
- Now until 3 February : Destinations – the Holiday & Travel Show
Travel show, featuring tourist boards, travel agencies, cultural entertainment, world food and drink, travel celebrities, talks, presentations, workshops, panel sessions and more. Open 10am-5.30pm. Tickets: £11 in advance. Olympia London, Hammersmith Road, Kensington, W14 8UX. Nearest station: Kensington Olympia. For more information and tickets, visit the Destinations – Holiday & Travel Show website.
- 4 February : The WineBarn's Annual Portfolio Tasting
An evening of German wine, featuring over 120 to choose from and learn more about how the wines are made. 6pm-8pm. Tickets: £35pp. Army & Navy Club, 36-39 Pall Mall, St James, SW1Y 5JN. Nearest station: Green Park. For more information, visit the Wine Barn website.
- 6 – 7 February : Dining With Daisy Supper Club @ The Little Blue Door
Hollyoaks actress and foodie Daisy Wood Davis teams up with the flatmates to host a supper club over two nights. The four-course menu features sentimental dishes with names such as The Hangover, Scampi Fry @ The SU, and Lambrini Sundae. 7pm-9.30pm. Tickets: £40. The Little Blue Door, 871-873 Fulham Road, Fulham, SW6 5HP. Nearest station: Parsons Green. For more information and booking, visit DesignMyNight. Read Metro Girl's review of, The Little Blue Door.
- 8 February – 2 June : Swinging Sixties exhibition
New exhibition celebrates the fashion, textiles, homewares and furniture of the Swinging Sixties. Including vintage designs by Mary Quant, Laura Ashley and Terence Conran. Open Tues-Sat 11am–6pm, Thurs until 8pm, Sun 11am–5pm. Tickets: £9.90. Fashion & Textile Museum, 83 Bermondsey Street, SE1 3XF. Nearest station: London Bridge or Tower Hill. For more information, visit the museum website.
- 9 February – 10 March : Orchid Festival
An orchid display is coming to the Princess of Wales Conservatory for four weeks, with this year being inspired by Colombia. General entrance tickets to Kew Gardens includes orchid exhibition: Adults £18.15, Children (4-16yr) £4.95. Kew Gardens, Brentford Gate, Kew, TW9 3AB. Nearest station: Kew Gardens. For more information, visit the Kew Gardens website.
- 10 February : Chinese New Year
Festivities to mark the Year of the Pig take place across Chinatown, Shaftesbury Avenue and Trafalgar Square. The parade from north of Trafalgar Square to Shaftesbury Avenue will start at 10am. From 11am-6pm there will be performances from Chinese artists in Trafalgar Square. Free. Nearest stations: Charing Cross, Piccadilly Circus or Leicester Square. For more information, check out Visit London.
- 11 – 28 February : #LoveSpot installation
Over Valentine's Day and throughout February will be a special light installation in Wembley Park. The #LoveSpot is a huge pulsating love heart that will provide the perfect backdrop for your next Instagram shot. Located outside Sunglasses Hut at London Designer Outlet, Wembley Park Boulevard, Wembley Park, HA9 0QL. Nearest stations: Wembley Central or Wembley Park. For more information, visit the London Designer Outlet website.
- 13 – 17 February : Ealing Music and Film Festival
Five day festival in Ealing featuring musical performances, film screenings and walks. Events take place at various venues, including St Mary's Church in South Ealing, St Mary's Perivale, University Of West London, the Brentham Club, St Barnabas Church and MetFilm School. Tickets range from free to £25. Nearest stations: Ealing Broadway or South Ealing. For booking, visit the Ealing Music & Film website.
- 13 – 24 February : Imagine Children's Festival
Two week children's festival at the Southbank Centre, including art, theatre, books, music, performances and workshops. Including Jacqueline Wilson, Malorie Blackman, Chris Riddell, Kiran Millwood Hargrave, Peppa Pig, Nadine Kaadan, the Singing Mermaid and more. 10.30am-4pm daily. Many activities and events are free, but some go up to £16. Southbank Centre, Belvedere Road, South Bank, SE1 8XX. Nearest station: Waterloo. For more information and booking visit the Southbank Centre website. Read the rest of this entry
Latin grooves, Afro beats, brass band remix and more at the London Remixed Festival 2019
Returning to East London to brighten up gloomy February is the eclectic London Remixed Festival. Over two days, 25 musical acts and DJs will be taking over four stages at Richmix in Shoreditch. Promising all of the fun and sounds of a festival without the mud and cold, the London Remixed Festival will feature Latin Grooves, Afro beats, Tropical Bass, Vintage-Remix, Desert Remix, Balkan Beats, Urban Roots, Acoustic Soundclash and Brass Band Remix.
Among the fabulous names performing are Sam And The Womp with their soundsystem show of Balkan-influenced heavy dance beats. London Remixed Festival's very own new UK Garage Orchestra will be performing UK garage classics with the Blue Lion Band and special guests MCs. Meanwhile, The True Stays will be playing vintage rock 'n' roll in the Folk Ghetto.
On Friday night, the Brass Off stage curated by Continental Drifts features Deadbeat Brass vs Das Brass. On Saturday, revellers will be spoiled for choice with severals stages to choose from, including Tropicarnival curated by Wormfood + Movimientos; Polka Club curated by Continental Drifts + MARSM; and the Folk Ghetto curated by Fire in the Mountain + Woodburner. There will also be the popular Silent Disco, with music from PantherPanther, DJ Hiphoppapotamus, Ecklectic Mick and Madame Electrifie.
- The London Remixed Festival takes place from 1-2 February 2019. Open: Fri 8pm-1am, Sat 8pm-4am. Tickets: £11.19–£21.79. Rich Mix, 35-47 Bethnal Green Road, Shoreditch, E1 6LA. Nearest station: Shoreditch High Street or Old Street. For tickets, visit the London Remixed Festival website.
For a guide to what else is on in London in Febuary, click here.


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Trackless Train
- $275.00
Good news... Best Jump has the area's BEST LOOKING trackless train around! Let us bring the train to your neighborhood, church, school, or special event. Covered seating for up to 18 people means there's plenty of room and plenty of shade. An experienced train conductor drives the train and gives you the complete train party experience! Our train comes fully equipped with whistle and bell, giving you an authentic and magical experience for both kids and parents. It has seatbelts on every seat for saefty and security.
DELIVERY AREA: We bring our train to Mandeville, Covington, Slidell, Hammond, Independence, Tickfaw, Madisonville, Ponchatoula, Folsom, Bush, Lacombe, and Robert, Pearl River, Loranger, Bogalusa, and Franklinton (delivery fees may apply to some locations).

subset: commoncrawl


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Did I mention yet how Pumpkin embarrassed me for the first time in public today? We walked into the locker room at the Y to get ready to swim, and there was a very elderly woman there in the process of getting dressed with help from who I assume was her daughter.
We rounded the bank of lockers to reveal a very naked elderly behind, and Pumpkin very loudly exclaimed, "Look at that big, stinky BUTT!"
I was so mortified I had no idea how to proceed, so I chose to pretend I didn't hear her. I hope THEY didn't, but the chances are good that they did.
Photo is from October 2012. I love how it has captured her sass.


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Traffic sucks, so why not start your morning off with some music? You provide the toast and we'll provide the jams.
Somewhere in the zone between rap and vaporwave lies this cut by DJ Screw of Al B Sure! I have listened to it a few weeks ago and I'm not sure the song ever ended. It's still going. It is winding its way between the folds of my brain. I see birds slowing down midair. I can count each leaf waving in the breeze.


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Saturday, April 23, 2005
Majestic Bath Art Show
We had the Majestic Bath Show last night. Over all it went pretty well. Majestic Bath is a bath fixtures store. They work in conjunction with Hotel Charlotte, a nice little restaurant a few doors down. I didn't sell anything, but I met some really nice fellow artists. lol Most of the art was abstract or impressionistic, I think I was the only one realist there. There was a portrait artist, is that the same genre as realism or is portraiture in a group by itself? hmmm There was a thunderstorm and high wind warning right in the middle of the show, which I am sure kept many would be visitors away. It was a little disconcerting to see my work displayed next to toilets.. very nice expensive toilets...but still:
Everyone seemed to like the strawberries and crystal. I am entering it a juried show (meaning they can kick it out if they don't like it, and you might win a prize if they keep it ;) ) The winners of last year's show were abstracts for the most part...but we have a different judge this year. Keep your fingers crossed for me.
And I am taking a break from the chickadee and blossoms and working on more gemstones for this weekend's gem and mineral show in Charleston. About to go do the rough drawing for that. Dev has to work this evening. I'm so glad he can dial in from home on the weekends!
Posted by emcrorie at 5:47 PM
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Svevind Solutions AB is a subsidiary owned by Svevind AB and its directors, tasked with maintaining and optimizing wind farms. Asset management is performed of wind farms in Svevind's own portfolio and those of third parties.
For more information about our services please see our presentation: Presentation Svevind Solutions.pdf
Contact details:
Per Olofsson, COO
Svevind Solutions AB
+46 90 349 44 25
In a perfect world wind turbines would never need maintenance and produce renewable energy without interruptions. However, in reality, someone needs to take care of the turbines when they stop or when something breaks down. This is where Svevind Solutions comes in - we monitor performance of turbines in real time, meaning that technicians can be dispatched as soon as a problem arises.
A big part of the work that Svevind Solutions do it optimizing wind farms, to maximize production of wind turbines. This is done by analyzing data collected from the turbines to see what causes troubles and what solutions, for e.g. preventing icing, works best. To enable a safe and high performance operation, Svevind Solutions equips its wind farms with additional measurement sensors e.g. cameras, temperature sensors inside and outside WEC components.


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4-6 weeks
I'll do my best to meet these shipping estimates, but can't guarantee them. Actual delivery time will depend on the shipping method you choose.
Buyers are responsible for any customs and import taxes that may apply. I'm not responsible for delays due to customs.
Just contact me within: 7 days of delivery
Ship items back to me within: 14 days of delivery
Request a cancellation within: 24 hours of purchase
But please contact me if you have any problems with your order.
Because of the nature of these items, unless they arrive damaged or defective, I can't accept returns for:
Buyers are responsible for return shipping costs. If the item is not returned in its original condition, the buyer is responsible for any loss in value.
Please contact me if you have any problems with your order.
I can not exchange items that have been shipped Internationally do to the high shipping cost. sorry.
Jul 15, 2018
Beautiful well made. I look forward to using this item!
Jan 16, 2018
Lovely, high quality and quick turnaround. We love our tree skirt.
Dec 23, 2017
AMAZING!! I've been looking for a washable treeskirt that I like for YEARS and saw this and new it would be perfect. Cannot express how much I really love it! Additionally... it's gotten a TON of compliments. woohoo!
Jan 2, 2011
Exactly what I wanted...Its perfect!


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Richmond Family Dentistry
Dentistry, General Physician(10) Average rating of 5 out of 5 stars Richmond Family Dentistry 10446 Ridgefield Pkwy, Richmond, Virginia, 23233, USA (804) 381-6238 Contact This Business
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Services Offeredgeneral Dentistry Same Day Crowns Dental Bridges Dental Fillings Dental Implants Teeth Cleaning Dentures Tooth Extractions Gum Disease Treatment Restorative Dentistry Family Dentistry Veneers Ceramic Implants Ceramic Crowns Clear Braces Teeth Whitening Richmond Family Dentistry of Richmond, Virginia is committed to offering biological dentistry for families who are looking for cosmetic and general dental procedures.They ensure that all treatments are beneficial for each phase of dentistry, and are happy to give patients the opportunity to improve their smile in just days instead of years.
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Richmond Family Dentistry
Joseph Guadagno has given a 5 out of 5 star rating on October 2, 2018
Dr. Hart is not my Family Dentist…but I wish she was! I recently sought out Dr. Hart for removal of amalgam/mercury fillings. In my quest to find the "best" dentist for the work, I found Dr. Hart to be highly recommended by other dentists as well as within my online research. I also realized that not all dentist are created equal, qualified, or up to the challenge of safely removing mercury fillings. Not only is Dr. Hart preeminently qualified and certified by the IAOMT to safely remove mercury fillings, but she combines this with holistic, homeopathic, and cutting edge technologies that result in the absolute best experience and results. In my experience, I also found Dr. Hart to be the most engaging and compassionate dentist. She is a great listener with zeal to provide you the best in dental care. She is a great listener, engaged in your overall supporting health, and forthright in her explanation of required protocols. In the end I had 12 mercury fillings removed by Dr. Hart. Dr. Hart's professionalism, compassion, and unique approach to dentistry extends to her staff as well. Each contributing to an overall great experience on all fronts. I lacked for nothing and am extremely pleased with the results. In fact, I was so impressed with Dr. Hart and her staff that I may have to change dentist and make my wish come true, despite the 1.5 hour drive!
Richmond Family Dentistry
Paula Horne has given a 5 out of 5 star rating on January 5, 2018
Dr. Hart and her staff are incredibly friendly and knowledgeable. They've helped me overcome my fear of the dentist. She's my dentist for life! 🙂
Richmond Family Dentistry
S. McCullough has given a 5 out of 5 star rating on November 7, 2017
Dr. Hart and her team are amazing! Dr. Hart applied a holistic approach by explaining how oral hygiene effects the entire body and vice versa. She found a metal sensitivity that no other dentist had detected. And, when I had a root canal extracted, they supported my decision to heal with natural remedies as opposed to antibiotics and prescription pain killers. Plus, her team was very patient with my children. My son actually doesn't mind going to the dentist now!
Richmond Family Dentistry
Tom Bradford has given a 5 out of 5 star rating on November 30, 2016
I had been looking for a good biological dentist who would be able to remove my old amalgam fillings without adding more mercury to my body. I discovered Richmond Family Dentistry and Dr. Hart. The web site looked good but it was a 160 mile trip from my house so I was not thrilled about the journey up to Richmond. Once I talked on the phone with Sandy, I became confident that this was a good choice. Sandy gave me a lot of helpful information and was the nicest person to work with. As soon as I arrived and was escorted to the dental chair I knew that I had made the correct choice. All of the equipment was first rate and professional. The area was relaxing and very comfortable. They even have TV sets that you can look up and watch as they do their work. It is a very high quality place. Then I met Charleen. She was extremely warm and welcoming. She took time to explain how everything would work. She is a very impressive and talented person. Dr. Hart came and examined me and gave me a lot of useful insights to my situation, even beyond the reason that I came. Dr. Hart and Charleen were very patient with me, taking time to answer all my questions. Both are very kind, considerate and gentle people. Once they started to work, they were extremely professional. They worked exceptionally well together. Dr. Hart has a really great touch with the needles. I never felt them. There was no pain before, during or after the procedures. In about 3 hours of work they had removed and replaced all of my old fillings. While they worked I could see the special attention they took in solving my dental problems. I felt they had real sensitivity to my situation. When they finished, I hated to leave. It was the best dental experience I had in my life, and I feel that I made some new friends – really nice people. They even did a great job providing me with an understandable bill and treatment plan. I could not be more pleased with their professionalism and their kind, gentle manner. Wonderful people and wonderful results. It is a long trip for me to come, but any time I have a serious procedure, I will make ever effort to come back to Dr. Hart, Charleen and Sandy. It was a wonderful experience. Thanks so much.
Richmond Family Dentistry
Patrick Brady has given a 5 out of 5 star rating on November 14, 2016
Wow, wow, wow, Dr. Hart is very knowledgeable, professional, kind and sweet. Her staff is excellent too. Sandy and Charlene have a great positive attitude/energy and always make you feel welcomed. I had a fair amount of work that had to be done. I had 10 teeth with 16 old mercury fillings and one molar that needed a crown. I went to Richmond family dentistry first to get an idea on treatment and cost. I then went to two other well-known dentist for the same consultation and review of my remedies. Without hesitation, I returned to Richmond family dentistry for the above mentioned reasons. I have since had all of my old fillings removed, a new crown and all of this excellent work was done under 30 days. I am a giant fan and will continue to return for all of my dental needs and highly suggest you stop by and see for yourself this special place and special people.
Richmond Family Dentistry
Carol has given a 5 out of 5 star rating on August 5, 2016
Wonderful dental office! All dental work has been excellent and they treat any dental anxiety with compassion. Everyone is professional and friendly.
Richmond Family Dentistry
Jody J. has given a 5 out of 5 star rating on March 11, 2016
I will only go to biologic dentists and my first experience with Dr. Hart was an excellent one. She is thorough, pays attention to details and answers questions with sound, fact-based information. I had a filling and crown preparation done and will be going back for the final crown placement. Dr. Hart is the kind of dentist you travel out of your way to go see.
Richmond Family Dentistry
Ashleigh has given a 5 out of 5 star rating on January 12, 2016
I have searched for a dentist that placed biological needs first and foremost and I have found the perfect dentist. She truly cares for each and every patient and treats me with natural materials. I love coming to this office!
Richmond Family Dentistry
Oressie W. has given a 5 out of 5 star rating on January 12, 2016
They paid close attention to me and all of my questions were answered. Their staff is very friendly and professional. I was very satisfied with my visit. They were helpful and I left feeling encouraged. The office was easy enough to find. I'll definitely be going back. I didn't have a bad experience. I have been going there for 10 years, driving a 50 mile distance because of great service.
Richmond Family Dentistry
Erin C. has given a 5 out of 5 star rating on January 12, 2016
Everyone in the office is really nice and friendly. As Doug said, you don't have to wait long. They will also talk to you like you're a human being and accommodate any requests. I'm normally not afraid of dentists at all, but when I saw that big ole novocaine needle my mouth snapped shut, "Could I? Could I get some gas or something?" Within seconds I had a mask on my face and was chilling out, needles be damned! They didn't judge or hem and haw at all. While I think that maybe the cleaning they gave me could have been a little more thorough the fillings I need were done quickly and professionally. I can't even tell that they're there!


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"Good writing excites me, and makes life worth living" - Harold Pinter
With thousands of recruiting blogs published every day, how do you know what to read? Cutting through the noise and finding real quality can be a tough proposition.
Hopefully the Beamery Blog is one of your first choices, but we do a fair bit of reading at Beamery so we know exactly how many other fantastic recruiting bloggers are out there.
We like to think that we know good writing when we see it, so...
Here are the 20 best recruiting articles of 2015:
Section 1: Social Recruiting
With almost every candidate active on social media, the way we think about social recruiting is only going to become more important. It's easy to get social wrong though, how do you make sure that you add value to candidates and don't just compete for clicks?
1. This social recruitment thing just isn't working
Every company has a battle plan for social recruiting - typically it revolves around using social media to broadcast roles and find candidates.
How social is this? Kasia Borowicz believes that we've lost our way on social media. If we want to boost response rates and increase engagement, we need to make our interactions genuinely valuable for candidates.
This article provides plenty of food for thought - do your candidates enjoy interacting with you on social?
2. The deadly sins of social recruiting
Retweets, likes and clicks make it easy to feel good about your social recruiting strategy.
Tony Restell wants you to question this warm fuzzy feeling though - are your social efforts actually leading to genuine business opportunities? Are they bringing passive candidates to your door?
If they're not, you may well be making one of the 4 deadly sins of social recruiting. It can be hard to get quantify social impact, so this article is great reading for anyone looking for more value from their social recruiting strategy.
Section 2: Technical Recruiting
Hiring great engineers has become the greatest pain point for many top organisations. The key problem is competition. How can you differentiate yourself and attract the best? What do developers really want?
3. This is how Coursera competes against Google and Facebook for the best talent
It's fair to say that great technical recruiting is a science. It's such a competitive market that companies tend to keep their tactics pretty close to the chest.
All this makes John Ciancutti's playbook even more special. He breaks down the exact tactics that Coursera use to compete for the best engineers. From email templates, to interview tactics it's all here.
This comprehensive piece is totally invaluable for anyone charged with bringing in top technical talent.
4. Building a product in the technical recruiting space? Read this first
This article is written for people considering building a product to 'fix' technical recruiting. What it also does though, is provide an excellent synopsis of several problems that recruiters face when trying to hire engineers.
Aline Lerner argues that the it's not finding great candidates that's the issue, it's finding great candidates that want to talk to you! The key problem engagement - how do you make someone interested?
Section 3: Recruitment marketing
Recruitment marketing gives recruiters a new framework to approach and engage with top talent. As many traditional hiring tactics lose efficacy, using marketing tactics can give recruiters a major advantage over competitors.
5. Why job adverts suck and what you can do about it
Most of us our pretty aware that our job adverts are far from perfect. They tend revolve around bland 'copy and pasted' text that does little to inspire an application.
Matt Buckland uses Maslow's 'Hierarchy of Needs' to map out the 5 things that every great job advert should touch upon. Forget perks like "on site masseuses and hot and cold running champagne" and focus on the candidate hierarchy of needs!
If you want to understand candidate motivations and how you can craft job adverts that actually work, this article is a fantastic starting point.
6. Recruitment is marketing. Are you ready for that?
Question: does a candidate need to be actively looking for you to place or hire them?
Greg Savage argues that although roughly 75% of candidates aren't job hunting, they are 'placeable', these 'hard-to-reach' candidates are the ones that you can really get paid on!
These candidates are unlikely to turn directly to an agency, so the key is to engage them consistently and build a 'resevoir' of talent that you can add to your pipeline in the future.
With so much out there on recruitment marketing for in house teams, this is a great resource for agency recruiters.
7. Recruiting content is a game anyone can win
How do you compete in a world where some companies e.g. Google, Facebook don't even think about limiting the amount that they spend on recruiting?
Traditional recruiting is a rigged game. Budget size dictates how many job ads you can buy and how far you can promote your roles. James Ellis believes that to win you have to change the game.
Recruiting content something many of us are guilty of ignoring is the answer. Content lets you control your company messaging and target the candidates that you really want to apply. It's a game you can win, but you need to play!
8. 3 ways that recruiting must copy marketing
Is recruiting in need of a reboot?
Marvin Smith argues that, in the face of widespread digital disruption, recruiters need to adopt new tactics to connect with high value talent.
In an age where bad candidate experience spreads as fast as cute cat memes, evaluating hiring through marketing lens can make a huge difference to your employer brand and help you attract better candidates.
9. Build talent pipelines with permission marketing and lower recruiting costs
What happens to the people that you turn down? In most companies, these candidates gather dust in the ATS, possibly with some kind of 'revisit later' tag attached!
Sean Rehder sees a seriously wasted opportunity here. Recruiters should 'nurture' these candidates with targeted messages.
You have bucketloads of information on every candidate. This lets you segment your data and send relevant emails to every person. For example, sales candidates should only be sent 'sales' based information.
This trick has been part of the marketing armoury for a while now and should help you convert previous applicants back into applicants in the future!
Section 4: Sourcing
In today's market, one of the key problems is that the best talent isn't applying. This makes the sourcing function even more important. Are you engaging and connecting with great candidate and building a hiring pipeline?
10. How to create candidate personas to recruit talent
Do you know who you're trying to hire?
It sounds like a silly question, but Andy Headworth argues that, beyond job title and company, many recruiters work without a clear idea of exactly which candidates they're looking for.
The answer to this is to build candidate personas. Think about what your ideal new hire looks - what is their experience, where might they work, what are their goals, what messages would they respond to?
Answers to questions like these and the others that Andy raises will help you connect with the candidates that you're really looking for.
11. A Recruiter's Guide To Email Marketing: Making Passive Candidates Care Enough To Click
Email is a crucial tool in your fight to win the hearts and minds of passive candidates. The problem is just getting them to 'open' and 'click'.
Robert Carroll argues that email marketing is your best bet and breaks down tactics to help you create emails powerful enough to take candidates from passive to active.
They key? Avoid the two major mistakes that recruiters make when firing off emails. Lack of clarity around the role and why the candidate fits, and a reliance on generic templates.
12. How recruiter spam is ruining recruiting brands
Familiar with the #recruiterspam movement? If not, you should be…
This has nothing to do with the canned meat, and everything to do with recruiter carelessness. Marie Burns and others want to take a stand against spam and make sure that every candidate is treated as a person first and foremost.
If you want to see where spam is most prevalent, and learn more about the movement, then I really recommend checking this one out.
Section 5: Analytics
If you don't measure what you do, how do you know what's working? It's a simple question, but it's still not front of mind for enough organisations. Analytics need to have a place in your recruiting process...
13. Sourcing & Recruiting Candidate Funnel & Output Calculators
Ever wondered exactly how many candidates you have to identify to make a successful hire? Or maybe you'd like to know how efficient your hiring process is...
Well, wonder no longer, Glen Cathey has created awesome calculators to provide insights into your candidate funnel and your team's output.
If you have unanswered questions, I urge you to take a good look at these calculators, the data could be invaluable in managing the expectations of your hiring team.
14. The one recruiting metric we all overlook
Technology has opened up a whole new world for recruiters. Analytics tools provide a veritable candy store of metrics and learnings.
Despite this, Jim Stroud argues that we're ignoring one key metric: retention. Surely it makes sense to judge recruiters on the quality of their hires and their fit with the company. Retention is the best way to measure this, and Jim even provides a few formulas to help you get the data you need.
Section 6: Candidate Experience
While it may be one of our favourite recruiting buzzwords, we can all agree that candidate experience gets more important by the day. In the social age, you treat applicants badly at your peril...
15. Candidates are NOT customers...not yet
The sheer volume of information available on every company lets candidates research job applications the same way as they would any other major purchasing decision.
Gerry Crispin argues that we're fast approaching a stage where the candidate should be seen as a 'partner' in the recruiting process. Companies that don't focus on candidate experience are certain to lose out.
16. What the candidate experience really looks like
Ever wondered what it's actually like to apply to one of your jobs? Thought about what your candidate experience is really like?
Matt Charney takes you on a typical application journey. From Google search, to form filling, this article shows you exactly how unnecessary and possibly ridiculous so many facets of job applications are.
If you haven't applied for one of your own jobs I suggest you do it now!
Section 7: Best of the Rest
We found these articles tricky to slot into a specific category, but they were simply too good to leave out. Take a quick browse through them and I'm sure you'll agree!
17. The 4 trends driving the future of recruitment
A quick Google search will show you that there's a constant stream of articles on the future of recruiting - few hit the nail on the head as cleanly as this one though!
Mervyn Dinnen cuts straight to the heart of recruiting: people. Our connections, the strength of our relationships, and the way that our brand is perceived by candidates all matter more than ever.
Recruiting has always been about relationships and this looks set to continue.
18. What a Recruiter REALLY Does- #iamarecruiter
Does the average candidate actually know what a recruiter does?
Will Thomson argues that the general perception of the industry has been ruined. Lazy recruiters are happy to spam 1000 candidates in the hope of a single response.
To counter this, he puts forward 8 excellent rules that every recruiter should live by to succeed and take pride in their profession.
19. What it really takes to attract top talent
Want to know how to attract the right candidates? It's simple… Think about it from the perspective of those that you're recruiting!
Peter Capelli reminds us that the goal should be to find one person who really fits your needs not thousands to apply. Figure out what candidates are a good fit and then meet their particular needs - this is the way to succeed.
2o. Always be closing almost never works in recruiting
Cult classic Glengarry Glen Ross has solidified the archetype of the aggressive salesman that's 'always closing' and will stop at nothing to win.
When you're 'selling' jobs though, this model is out of place. Bryan Chaney reminds us that it isn't just filling a role that's important in recruiting, it's building the right relationships so you can fill the role with the right person.
This takes time and relies on engagement, messaging and relationship building. If you want to know how to get started, take a look at Bryan's article.
Section 8: Bonus
Just because it's nearly Christmas, here's one more awesome recruiting article. This one breaks down a really interesting new movement: #OpenSourceHR.
21. A vision for open source recruiting
Ever thought about sharing the secrets behind your recruiting success?
It might seem crazy at first, but this is what Lars Schmidt asks you to do. Open source recruiting asks that you share case studies and best practices to help move the industry forward.
The open source model has been central to the development of engineering: could it work for recruiters?
Recruitment Marketing University
If you're looking for more information on how to build and scale an outstanding talent acquisition operation, check out our online Recruitment Marketing University.


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"When a daughter loses a mother, the intervals between grief responses lengthen over time, but her longing never disappears. It always hovers at the edge of her awareness, prepared to surface at any time, in any place, in the least expected ways." Hope Edelman (Motherless Daughters: The Legacy of Loss)
I became a motherless daughter when I was sixteen years old. This was almost 11 years ago… I lost her to cancer. As I have aged and gone through different phases and stages of life the "longing" for her has come like rapid fire emerging out of nowhere. While other times it is subtle and more like a pinched nerve that only hurts when I move a certain way. Being pregnant without my mom has definitely felt like that pinched nerve at certain moments and other moments it's like I'm reliving her death and that gasping-for-air-grief all over again.
During my first trimester when I was hospitalized multiple times due to being dehydrated and so sick, I wondered what her pregnancies were like and if she was sick too. I laid up in my mother-in-law's house for weeks, puking every 20-40 minutes, dry heaving even 12+ hours after I attempted to eat. I laid there everyday so thankful I had this wonderful lady who took care of me like she was my mom, but I also laid there feeling guilty and sad.
As I progressed into the second trimester and started feeling Geneveive kick, I wanted to talk to my mom about those wonderful flutters. I wanted to ask her about stretch marks and how much weight she gained. Now, in my third trimester, I long for her to know her first grand-baby. To have her in the room with me during labor, to call and cry to her when I'm having an anxiety attack because I don't feel prepared, to simply talk to her about this new journey. Some days it's in the back of my mind and others it is a blazing forest fire ripping through me.
I want my mom here and that is selfish because I know heaven is beyond anything I could imagine. I feel guilty because I have been blessed with different motherly figures throughout my adulthood and it's a fine line between letting them love me like their own and feeling like I'm leaving my mom behind. There are so many emotions (some irrational) that come with grief and loss. I feel heartbroken because my daughter will never know this amazing woman who I called my mom. I feel ashamed when I think about all the 'fill-in' grandma's my daughter will have because what if that hurts my mom? I know it's probably irrational to think that way because God gave me these incredible women to talk to and love me–but it does not make the thoughts go away.
There are times when her death lays on my chest like an elephant suffocating me… I wonder if I am allowed to be loved by other motherly figures. It is okay to have these second families that I spend my time, holidays, etc with? Does it make me a bad daughter to not mourn her at all times? I feel it becomes harder to accept love from others–that I should return the love, gifts, favors, advice with another gift or money. I know God did not take my mom as punishment, but I find myself wanting to punish myself for letting other people in.
I know as I grow older, the longing for her will still not fade. There will always be a need for my mom–questions I have, stories I want to share, other babies I want her to meet… I was blessed to have my mother for sixteen years and I am so thankful for our time, but it was not enough.
Side Note: losing my mother so young did create a special bond between my siblings and I. My little brother has blossomed into this amazing young man who is wise beyond his years. My older sister is a role model, a take-charge and keep us in-line beautiful soul who consistently reminds us the value and importance of family. She is so much like my mom and it is wonderful having her around to keep that memory even more vibrant. God works in mysterious ways that we don't always understand, but I know our relationship was a small part of his bigger plan.


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Nobody ever expects to come home and find a raccoon rooting through their kitchen pantry, but it can happen. No matter how unlikely it seems, it's important to know what to do when an animal gets in your home. It's also just as important you know what NOT to do.
If you ever find a wild animal inside your home, these are the things you SHOULD and SHOULD NOT do. Follow these "DO" steps exactly and you'll be able to remove the animal quickly, safely, and humanely. Avoid making the "DO NOT" mistakes and you'll avoid endangering yourself, your family, or your pets. If you find a wild animal in your home…
Stay calm
Whatever you do, don't panic. If you're freaking out, the animal will freak out. You want to keep the animal as calm as possible until you can have it removed. The best way to do that is to keep calm yourself. Move slowly, speak quietly, and take deep, slow breaths.
Call for help right away
This is the most important step. If there's an animal in your home, you need professional assistance right away. Call Varment Guard as soon as you notice the animal. Let us know it's an emergency.
Keep your distance
Give the animal space to move around. Ideally, you should stay out of its line of sight at all times. Try not to get in the animal's way. You can always repair or replace home damage, but you can't undo injuries.
Keep an eye on the animal
While you should give the animal space, don't give it too much space. Make sure you know exactly where the animal is at all times. Never let it out of your sight. Don't stop the animal from hiding, but make sure you know where it's hiding.
Take a picture
Use your phone to take a couple pictures of the animal when you have a chance. Make sure your flash is off and your phone is on silent to avoid scaring it. If you lose sight of the animal, you can show the pictures to animal control when we arrive. You should also take pictures of any damage the animal inflicts on your home. Just don't get so focused on taking pictures that you get too close to the animal!
Separate your pets
Keeping your pet away from the wild animal in your home is very important for everyone involved. Pets are often naturally defensive and react to an unwelcome animal's presence aggressively. Take whatever steps you can to separate your pet from the animal safely. Do NOT attempt to restrain or move the wild animal; focus on isolating your pet instead. Ideally, you should try to keep your pets from seeing the animal at all.
…Do NOT:
Attempt to shoo the animal
Do not try to force the animal to move or go away. Approaching the animal may make it defensive and therefore dangerous. Instead, keep your distance and let the animal go wherever it's going. Your #1 objective is to keep the animal calm until animal control can take it away.
Approach it
If possible, do your best not to approach the wild animal at all. If you absolutely need to move closer to it for some reason, do so very slowly. Do not stay close to the animal for any longer than is absolutely necessary. The more space you can give it, the better. Be especially careful about approaching animals while they're not looking at you. Animals are at their most dangerous when they're startled. Speaking of which…
Attempt to scare it
This is very important. Don't do anything that could scare the wild animal. Don't make sudden movements or loud noises. DO NOT throw anything at the animal, lunge at it, prod it, or otherwise actively attempt to startle it. Leave the animal alone as much as you possibly can, even if it's inflicting damage on your home. Trying to bully an animal never ends well.
Block its path
This rule might be difficult to follow, but it's important. In general, you should let the animal move where it wants to move. If it's headed toward a door, don't slam that door. We're not saying you have to open your pantry doors, but you shouldn't actively impede the animal either. Never stand in the animal's way or attempt to shoo it toward another area of the home. It might be hard to watch a skunk lumber toward your clean carpet, but it beats the alternative.
Leave your home
You might want to evacuate your home immediately, but you should resist that impulse. Instead, stay where you are, contact us immediately, and watch the animal. If you leave your home, you won't see where the animal goes. It could inflict damage you won't notice right away, and it'll be harder to find when we arrive. Stick around until animal control arrives.
To reiterate: the #1 most important thing to do when dealing with a wild animal is CALL THE PROS. The faster you call us, the faster we can get there. The faster we get there, the sooner we can make sure you, your home, and your family are safe. Don't wait! Call Varment Guard the moment you encounter a wild animal in your home. We'll be there as fast as possible!BACK TO BLOG


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For many aspiring models, it's easy to recognize the many obvious modeling scams like all the different variations of the Nigerian (advance fee) scams. But there is a whole other category of legally operating businesses that don't fall into this category but should still be avoided. Why should they be avoided? Well I just don't think they offer good value for your money. This is a good time to clearly state that this is my opinion. No doubt many on this list will get pissed off and suddenly become internet lawyers threatening all kinds of things because they think my opinion is libelous. Let me take a few moments to teach them a short lesson. Let's say I don't like the new Apple iPad because it does not support flash. I can say "I don't recommend the iPad because it's too shiny and the lack of flash means I can't watch Hulu.com". That is an opinion. I can also simply say "The iPad sucks". That too is an opinion. What I can't say is "The iPad has a super secret chip inside that will steal your banking info off your wifi so don't buy it". That is an incorrect statement of fact that is potentially libelous. Now I doubt Apple will give a crap if I said it but they may not like it so much if Time magazine said it. Get it so far?
Ok. On to companies I do not recommend because I think they suck.
- Let's get the obvious ones out of the way. John Robert Powers, John Cassablancas Modeling and Career Centers, and Barbizon. They will all take anyone and everyone with a valid credit card or a check. You will spend thousands on modeling lessons taught by someone whose last job was selling TVs at Bestbuy. If you get lucky, you will get a modeling job handing out flyers at the local mall.
- Model Management, Inc. in Los Angeles (currently shutdown). This was a typical portfolio mill charging thousands for useless pictures under the ruse of an agency. They appear to be shut down and their web site is gone but keep an eye out for them to pop up under another name. You can see how they typically operated here.
- Wilhelmina PA. They have been aggressively advertising on Craigslist trying to get anyone and everyone to go to their open call (aka sales pitch). They have nothing to do with Wilhelmina NY other than paying to license the name. They want at least $2,400 to shoot your portfolio with their own photographer and will try to sell you on all kinds of useless classes. Here's a link for those too stupid to use Google. There are only three legitimate Wilhelmina modeling agencies. They are in NY, LA, and Miami. The others are all affiliates so do your research.
- MTC/ Model Training Center in San Diego. Models don't need training. Any required training should be done by legitimate agencies after the model is signed. Training cannot make models taller and you can't train the ugly out of someone.
- Elite Modeling Academy. Same as above. They also charge $5,200 for courses. No, that was not a typo. More here.
- IMTA, Proscout, MSA. For the vast majority, some who spend over$10,000 to attend these conventions, it's a complete waste of time. They do have legitimate and real agency reps there that are sometimes paid to be there and are required to do a minimum number of call backs. Call backs mean exactly that. They call you back. If they call you back and talk about the weather or tell you you're interesting, that counts. It does not mean those who got call backs got any offers. In some rare cases, it can be beneficial to a very small percentage of undiscovered talent but you probably don't fit into that category. If you are near a modeling market, go to open call instead and save yourself $10,000.
- Glamour Model Talent Inc. and Latte Model & Talent Agency Ltd in Chicago, IL. They are getting shut down by the Attorney General there. Enough said.
- Orange Model Management in Toronto, ON. From the complaints online, this is either a typical portfolio mill or a poorly managed agency, probably a combination of both. They require models to pay upfront for images shot by photographers with whom they have a financial relationship.
- Panache Model Management in Winnipeg, Manitoba. This one is another modeling school. They say it right there on their home page. Save your money. Not everyone should be a model despite it being your "passion". Here's a real life testimonial for why you shouldn't go there. Then look at the bullshit they are trying to sell on their own website.
- Most online casting sites. With the exception of sites recommended to you by legitimate modeling agencies, most online casting sites are worthless. They just scour Craigslist and other modeling sites and report worthless outdated castings, many of which are scams to begin with. The following casting sites are legitimate but are mostly for actors, not models.
- Styles Model Management Inc. They list their phone number in this press release as 416-895-3011. It's the same number used to recruit for hookers in this ad.
- Anyone listed here. ACTRA Toronto "No Cheque list"
Do you have suggestions for additions? Post a comment with relevant links.


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CXCR4 is a G protein–coupled chemokine receptor that has been implicated in the pathogenesis of primary immunodeficiency disorders and cancer. Autosomal dominant gain-of-function truncations of CXCR4 are associated with warts, hypo-gammaglobulinemia, infections, and myelokathexis (WHIM) syndrome, a primary immunodeficiency disorder characterized by neutropenia and recurrent infections. Recent progress has implicated CXCR4-SDF1 (stromal cell-derived factor 1) signaling in regulating neutrophil homeostasis, but the precise role of CXCR4-SDF1 interactions in regulating neutrophil motility in vivo is not known. Here, we use the optical transparency of zebrafish to visualize neutrophil trafficking in vivo in a zebrafish model of WHIM syndrome. We demonstrate that expression of WHIM mutations in zebrafish neutrophils induces neutrophil retention in hematopoietic tissue, impairing neutrophil motility and wound recruitment. The neutrophil retention signal induced by WHIM truncation mutations is SDF1 dependent, because depletion of SDF1 with the use of morpholino oligonucleotides restores neutrophil chemotaxis to wounds. Moreover, localized activation of a genetically encoded, photoactivatable Rac guanosine triphosphatase is sufficient to direct migration of neutrophils that express the WHIM mutation. The findings suggest that this transgenic zebrafish model of WHIM syndrome may provide a valuable tool to screen for agents that modify CXCR4-SDF1 retention signals.
Stromal cell-derived factor 1 (SDF1, CXCL12)–mediated activation1 of the chemokine receptor CXCR4 is important for both normal and pathologic processes, including primordial germ cell migration, HIV pathogenesis, invasive migration of cancer cells, and leukocyte trafficking.2-5 Therefore, there is substantial interest in understanding how CXCR4-SDF1 signaling regulates cell motility and how these mechanisms can be targeted to treat human disease. CXCR4 signaling is attenuated by receptor internalization, which is regulated by phosphorylation events and binding of regulatory proteins to the cytoplasmic tail.6 The functional importance of CXCR4 internalization is highlighted by the dominantly inherited primary immunodeficiency, warts, hypogammaglobulinemia, infections, and myelokathexis (WHIM) syndrome, in which truncations of CXCR4 lead to altered signaling and gain of function.7-9
WHIM syndrome is characterized by warts, hypogammaglobulinemia, infections, and myelokathexis, a severe chronic neutropenia.10,11 Substantial evidence supports the importance of CXCR4 signaling in regulating neutrophil homeostasis and release from the bone marrow (BM).5 It has been postulated that the neutropenia in patients with WHIM syndrome results from both neutrophil retention in the BM and enhanced neutrophil apoptosis of retained neutrophils.11 Direct evidence to support this hypothesis has been provided by a mouse model of WHIM syndrome induced by the ectopic expression of WHIM truncation mutations of CXCR4 in hematopoietic stem cells that show impaired neutrophil release into the blood and increased rates of apoptosis in the BM.12 Previous reports indicate that neutrophils from patients with WHIM show increased signaling13 and chemotaxis8,9 in response to SDF1. However, some reports have suggested that the C-terminus of CXCR4 can both positively and negatively regulate cell motility8,9,14 and, alternatively, may be involved in modulating the precise targeting of cells in vivo.15
Despite the importance of CXCR4-SDF1 signaling, few animal models of WHIM syndrome are amenable to imaging or screening for drugs that modulate CXCR4-SDF1 function in vivo. Modeling WHIM syndrome is particularly attractive because CXCR4 signaling is important to many disease processes and is a direct result of aberrant chemokine signaling. Therefore, developing a model of WHIM syndrome in a system that allows the direct visualization of motility and chemotactic events in vivo would be a beneficial tool to understand disease pathogenesis. Our current understanding of WHIM syndrome is mostly derived from in vitro experiments8,9,13 or in vivo mouse models in which observing chemotactic events is difficult.12 The zebrafish, Danio rerio, is an ideal system to visualize neutrophil migration in vivo.16 In addition, there has been substantial recent progress in understanding how neutrophils develop17,18 and respond to inflammatory cues in zebrafish.16,19,20
Here, we describe the generation of a zebrafish model of the primary immunodeficiency syndrome, WHIM syndrome. WHIM zebrafish are neutropenic, and WHIM neutrophils show impaired recruitment to wounds and tissue inflammation, recapitulating the human disease. Exploiting the optical clarity of zebrafish larvae, we use real-time imaging to directly observe neutrophil motility in vivo. Expression of WHIM truncation mutations of CXCR4 in zebrafish neutrophils induces neutrophil retention in hematopoietic tissue and mediates a retention signal that is SDF1 dependent. These findings support the power of zebrafish to model primary immune disorders and provide a valuable tool to screen for agents that modify CXCR4-SDF1 retention signals.
Transient expression in zebrafish and generation of CXCR4b– GFP and WHIM–green fluorescent protein transgenic lines
All protocols that used zebrafish in this study were approved by the University of Wisconsin-Madison Research Animal Resources Center. All DNA expression vectors contain either the zebrafish myeloperoxidase (MPO) promoter for neutrophil expression16 or a cytomegalovirus promoter, minimal Tol2 elements for efficient integration,21 and an SV40 polyadenylation sequence (Clontech Laboratories Inc). Constructs with each of the following in the backbone were constructed: zCXCR4b–green fluorescent protein (GFP), zCXCR4b-WHIM-GFP, mCherry, SDF1a-2A-mCherry, and Dendra2 (Evergen). Expression of constructs was obtained by injection of 0.5-1 nL of 25 ng/μL plasmid with 45 ng/μL in vitro transcribed (Invitrogen) Tol2 transposase mRNA into the yolk of 1-cell–staged embryos just below the cell. Injected embryos were raised to sexual maturity and screened by crossing with wild-type fish to identify founders. F1 embryos were screened at 2-3 days postfertilization (dpf) for GFP expression in the caudal hematopoietic tissue (CHT) and raised to sexual maturity. Experiments were done on progeny of F1 incrosses. Transgenic lines from ≥ 2 separate founders were generated for each injected construct. All results reported here are from experiments performed on one of the transgenic lines, but many experiments were replicated on other lines to ensure consistency between founders. All experiments were done ≥ 3 times, and images of fixed larvae are representative of ≥ 20 larvae.
Tailfin wounding, whole-mount immunolabeling, and Sudan Black staining
For wounding assays larvae were anesthetized in E3 containing 0.1 mg/mL tricaine and wounded in the ventral tailfin with a 25-gauge needle, or their tails were transected with a surgical blade. After wounding, live larvae were embedded in 1% low-melt agarose for imaging or fixed for further processing. Larvae were fixed in 1% formaldehyde in phosphate-buffered saline overnight at 4°C and double-immunolabeled for MPO and L-plastin as described22 or fixed in 4% formaldehyde overnight at 4°C and stained with Sudan Black as reported.18
Plasmids containing mpo, sdf1a, and sdf1b were linearized with restriction enzymes (SalI for mpo and NotI for sdf1a and sdf1b) to serve as template for transcription of antisense probes. Digoxigenin-labeled RNA probes were transcribed for sdf1a and sdf1b with the use of T3 RNA polymerase. For double whole-mount in situ hybridization (WISH), fluorescein-labeled antisense mpo RNA probe was generated with T7 RNA polymerase. WISH was performed as described.23 After staining, embryos were placed in 80% glycerol for imaging.
MO microinjection
Morpholino oligonucleotides (MOs; Gene Tools) were resuspended in 1× Danieau buffer at a stock concentration of 1mM. Final MO concentrations were injected (0.5-1 nL) into the yolk of 1- to 2-cell–staged embryos.
Experimental results were analyzed with the Prism version 4 (GraphPad Software) statistical software package. Statistical significance was determined with the unpaired Student t test (to compare 2 groups) or 1-way analysis of variance followed by Tukey multiple comparison post test (to compare multiple groups) at a 95% confidence interval. The resulting P values are included in the figure legends for each experiment; n = the number of larvae or neutrophils quantified.
Hematopoietic expression of CXCR4b and SDF1a in zebrafish larvae
The zebrafish genome encodes 2 SDF1 ligands, SDF1a and SDF1b, and 2 SDF1 receptors, CXCR4a and CXCR4b. The expression pattern of CXCR4 in zebrafish hematopoietic cells has not previously been defined. To determine whether CXCR4 is expressed in zebrafish neutrophils, we performed reverse transcription–polymerase chain reaction for both CXCR4a and CXCR4b on RNA isolated from neutrophils and macrophages sorted from MPO:Dendra2 transgenic larvae at 3 dpf by fluorescence-activated cell sorting (supplemental Figure 1A, available on the Blood Web site; see the Supplemental Materials link at the top of the online article).22,24,25 As previously reported, cells with low levels of fluorescence represent macrophage-like cells and express the receptor CSF1R. In contrast, cells with high levels of fluorescence represent neutrophil-like cells and express MPO (Figure 1A).22 We found that both macrophages and neutrophils express CXCR4b but not CXCR4a. In contrast, both CXCR4a and CXCR4b are expressed in whole kidney marrow isolated from adult zebrafish (Figure 1A). These findings suggest that the primary SDF1 receptor in neutrophils from zebrafish larvae is CXCR4b.
To determine the spatial distribution of the CXCR4 ligand, SDF1, in zebrafish larvae, we performed WISH for SDF1a or SDF1b mRNA at 2 and 3 dpf. Interestingly, we observed concentration of SDF1a mRNA in regions of neutrophil production, including in the head (Figure 1B), the CHT (region of larval neutrophil development located in the anterior ventral tailfin), and along the pronephric duct (Figure 1C; supplemental Figure 1F-G), as previously reported.26 These areas of high SDF1a expression correlated with 2 distinct populations of neutrophils observed in zebrafish during this stage of development (Figure 1D). SDF1b was also expressed in the head and ventral body at 2 dpf, but staining was generally more diffuse (supplemental Figure 1B-C). Taken together, these findings suggest that zebrafish neutrophils develop or accumulate in regions with high SDF1a expression in zebrafish larvae.
Generation of WHIM-GFP transgenic zebrafish line
To model the human disorder, WHIM syndrome, in zebrafish we expressed the homologous CXCR4 receptor truncation mutations in zebrafish neutrophils with the use of the MPO promoter. Alignment of the C-terminal tail of zebrafish CXCR4b with human CXCR4 showed a high level of conservation, especially in serine residues that are critical for the proper internalization of CXCR4 after binding to SDF1 (Figure 2A).27 By truncating 19 amino acids from the C-terminus of CXCR4b, we generated a truncation mutation analogous to mutations described in patients with WHIM syndrome7 (Figure 2A). To determine whether this mutation affects receptor internalization we expressed both wild-type CXCR4b-GFP and the truncation mutant (hereafter referred to as WHIM-GFP) in human embryonic kidney cells and treated the cells with human SDF1. As expected both CXCR4b-GFP and WHIM-GFP localized to the cell membrane of transfected human embryonic kidney cells (supplemental Figure 2A). Treatment with SDF1 induced internalization of CXCR4b-GFP into intracellular vesicles, but not GFP or WHIM-GFP, indicating that the WHIM truncation impairs CXCR4b internalization after ligand binding (Figure 2B).
To develop a zebrafish model of WHIM syndrome we generated a stable transgenic line by expressing WHIM-GFP specifically in neutrophils with the use of the MPO promoter (Figure 2C).16 The MPO:WHIM-GFP transgenic line will hereafter be referred to as WHIM-GFP. In accordance with our findings reported with the MPO:GFP transgenic line, we observed GFP+ neutrophils in the CHT in WHIM-GFP larvae (Figure 2D-E; supplemental Figure 2B). WHIM-GFP localized to the cell membrane in individual cells in the CHT (Figure 2G), compared with the diffuse localization of GFP alone (Figure 2F). In contrast to the previously published MPO:GFP16 transgenic, we did not observe ectopic expression of WHIM-GFP in tissues other than hematopoietic cells (supplemental Figure 2B).
Neutrophil development in WHIM-GFP larvae
To determine whether expression of the WHIM-GFP receptor affects the distribution of neutrophils during zebrafish development, wild-type or WHIM-GFP larvae were fixed at 2, 3, 7, or 13 dpf and stained with Sudan Black to highlight neutrophils.18 At 3 dpf, we observed normal neutrophil development in the CHT in both wild-type and WHIM-GFP larvae (Figure 3A-B). However, WHIM-GFP larvae showed neutrophil aggregates at 3 dpf (Figure 3B arrows), which were even more apparent at 7 dpf (Figure 3C-D arrows). Neutrophils were not detected ventrally over the gut (Figure 3C arrows and 3D) or along the dorsal ridge of WHIM-GFP larvae as seen in controls (Figure 3C arrowheads and 3D). Strikingly, at 13 dpf, WHIM-GFP larvae showed persistent neutrophil aggregates in the CHT (Figure 3F arrows) and in the midline region (Figure 3F arrowheads), whereas wild-type zebrafish no longer had neutrophils in the CHT (Figure 3E).
Sudan Black staining also showed neutrophil aggregates in the heads of 2- and 3-dpf WHIM-GFP larvae (supplemental Figure 3C-D; Figure 3K and 3L arrows) in contrast to wild-type larvae that show a diffuse distribution of neutrophils (supplemental Figure 3A-B; Figure 3G-H). These neutrophil aggregates persisted on the ventral side of the head at 7 (supplemental Figure 3H arrow) and 13 (supplemental Figure 3J arrow) dpf. Interestingly, less prominent neutrophil aggregates were also observed in the head of transgenic larvae that ectopically express wild-type CXCR4-GFP in neutrophils (supplemental Figure 4A-B). This is consistent with a previous study that reported similar phenotypes with overexpression of wild-type and WHIM-truncated CXCR4 in mouse models.12 These findings indicate that the WHIM truncation is a gain-of-function mutation in vivo.
By 7 dpf, neutrophil development was also apparent in the kidney in wild-type zebrafish (Figure 3M), which is the primary site of adult hematopoiesis. Interestingly, in WHIM-GFP transgenic larvae the hematopoietic tissues of the developing kidney had greater neutrophil accumulation compared with control larvae (Figure 3N arrow). This difference remained prominent at 13 dpf (Figure 3O-P). However, no differences in neutrophil development were observed among wild-type, MPO:GFP, or GFP− siblings of WHIM-GFP larvae. In addition, no differences in apoptosis of neutrophils were observed by staining with terminal deoxynucleotidyl-transferase–mediated deoxyuridine triphosphate nick-end labeling in WHIM-GFP and control larvae (K.B.W., unpublished data, January 2010). Together, these findings suggest that WHIM-GFP larvae have neutrophil aggregates at sites of SDF1a expression, including the CHT and ventral head. Furthermore, WHIM-GFP larvae have neutrophil aggregates in the kidney, the site of neutrophil development in adult zebrafish, reminiscent of the human disorder WHIM syndrome.
CXCR4-SDF1 signaling mediates neutrophil arrest and aggregation in the head
To determine whether neutrophils aggregate in regions of high SDF1a expression in the head, we compared SDF1a expression by WISH with Sudan Black staining. At 2 dpf a ventral view of the head showed 2 bilateral patches of SDF1a expression above the yolk sac (supplemental Figure 3E-F arrows) that converge at 3 dpf to form an area of concentrated SDF1a expression on the ventral side of the head (Figure 3I-J; supplemental Figure 1Gii arrow). The areas of SDF1a expression closely paralleled the location of neutrophil aggregates in the head of WHIM-GFP larvae at both 2 (supplemental Figure 3C-F) and 3 (Figure 3I-L) dpf. In contrast to SDF1a, at 3 dpf SDF1b was expressed on the ventral side of the head (supplemental Figure 1D-E) in areas without neutrophil aggregation, suggesting that neutrophils accumulate in areas with high SDF1a but not SDF1b expression. To confirm this in the context of an individual larva, we performed double WISH with probes for SDF1a (purple) and mpo (red). Indeed, in WHIM-GFP larvae at 3 dpf, we found that mpo expression colocalized with a patch of SDF1a expression on the ventral head (supplemental Figure 3K-L).
The formation of neutrophil aggregates in areas of high SDF1a expression suggests that WHIM-GFP neutrophils are retained by CXCR4b-SDF1a interactions in vivo. To determine whetherSDF1a mediates neutrophil aggregation in the head, endogenous SDF1a was depleted with MOs. Injection of high concentrations of SDF1a MO (between 1mM and 200μM) resulted in gross morphologic and neutrophil developmental defects (supplemental Figure 5A-B), whereas injection of a lower concentration (110μM) allowed for near normal neutrophil development (supplemental Figure 5C). Injection of the lower concentration of SDF1a (Figure 4C-D) but not SDF1b (Figure 4E-F) or control (Figure 4A-B) MO restored a diffuse distribution of neutrophils in the head, indicating that CXCR4b-SDF1a signaling mediates neutrophil aggregate formation. To characterize how CXCR4b-SDF1a interactions regulate neutrophil aggregation in the head, we monitored neutrophil motility in vivo with the use of live fluorescence microscopy in uninjected and SDF1a morphant MPO:GFP or WHIM-GFP transgenic larvae. Control neutrophils (MPO:GFP) in the head exhibit spontaneous, random migration in zebrafish larvae at 2-3 dpf.28 Live imaging shows rapid polarization and persistent migration of GFP+ control neutrophils in the head at 3 dpf (Figure 4H; supplemental Video 1). Knockdown of SDF1a in MPO:GFP larvae did not affect migration of MPO:GFP neutrophils (Figure 4I; supplemental Video 1). Strikingly, in WHIM-GFP larvae, neutrophils in the head show random protrusions but impaired persistent motility resulting in arrested migration and retention in regions of high SDF1a expression in the head (Figure 4J; supplemental Video 1). Injection of the SDF1a MO partially restored directed neutrophil motility and decreased neutrophil aggregation in WHIM-GFP larvae (Figure 4K; supplemental Video 1), indicating that SDF1a mediates the neutrophil retention signal. The 3-dimensional velocity of neutrophils in WHIM-GFP larvae was significantly impaired compared with control neutrophils (Figure 4G). Injection of the SDF1a MO, however, partially rescued the migration velocity of WHIM-GFP neutrophils (Figure 4G). Taken together these results suggest that CXCR4 WHIM mutations impair persistent neutrophil motility in the head of zebrafish larvae through interactions with endogenously expressed SDF1a.
To determine whether expression of SDF1a is sufficient to attract WHIM-GFP neutrophils and induce neutrophil aggregation we transiently expressed SDF1a-2A-mCherry in a mosaic fashion in WHIM-GFP larvae. The self-cleaving, viral 2A peptide29 allows for expression of untagged, secreted SDF1a while marking the secreting cells with mCherry. WHIM-GFP neutrophils could be found in close association with SDF1a-expressing cells in a variety of different tissues of the head, body, and yolk sac (Figure 4L), with occasional clusters of WHIM-GFP neutrophils forming in regions with ectopic expression of SDF1a (Figure 4Liii). In contrast, expression of mCherry alone failed to attract WHIM-GFP neutrophils (Figure 4L). Live imaging showed that the association of WHIM-GFP neutrophils and SDF1a-expressing cells was dynamic and persistent, often with long periods of close contact between the cells (supplemental Video 2). Together these results suggest that regionalized expression of SDF1a is necessary and sufficient to form WHIM-GFP neutrophil aggregates.
WHIM-GFP transgenic larvae are neutropenic
A hallmark of the human disorder, WHIM syndrome, is peripheral neutropenia. To determine whether WHIM-GFP larvae are neutropenic in the vasculature, we used live imaging to scan blood flow for GFP+ cells. In MPO:GFP larvae, neutrophils can be observed in the blood stream by 3 dpf. By quick acquisition of a small section of the dorsal aorta we quantified the number of bright GFP+ cells per minute of blood flow (Figure 5A). In 3- and 4-dpf MPO:GFP transgenic larvae the number of neutrophils in the blood varied from 6 neutrophils/min to 32 neutrophils/min with an average of approximately 21 neutrophils/min of blood flow (Figure 5B; supplemental Video 3). In contrast, GFP+ cells were absent in the blood of WHIM-GFP transgenic larvae at 3-4 dpf (Figure 5B; supplemental Video 3) and 7 dpf (K.B.W., unpublished data, June 2009). These findings show that we have generated a neutropenic model of WHIM syndrome in zebrafish.
Impaired chemotaxis of WHIM-GFP neutrophils to wounds
Wounding the fins of zebrafish larvae results in the rapid recruitment of neutrophils,30 and this recruitment can be visualized by live imaging.16 Interestingly, we observed impaired neutrophil chemotaxis to wounds in the ventral tailfin of WHIM-GFP larvae (Figure 5C; supplemental Video 4). To quantify the recruitment defect we used a fixed wounding assay with Sudan Black staining to visualize neutrophils at the wound. At 2 hours after wound there was robust recruitment of neutrophils to the wounds in control larvae with few detectable neutrophils recruited to the wounds of WHIM-GFP larvae (Figure 5D-F). A time course showed impaired neutrophil recruitment to wounds at all time points between 1 and 24 hours after wound (Figure 5G).
To determine whether WHIM-GFP neutrophils could respond to stronger inflammatory stimuli we examined neutrophil recruitment in response to tail transection and in the context of chronic inflammation. We have previously reported 2 mutant lines, spint1ahi2217 and clint1hi1520, that show epidermal hyperproliferation and robust chronic neutrophil recruitment into the tailfin.19,31 Using a MO targeting clint1 to recapitulate the clint1hi1520 phenotype, we observed neutrophil recruitment into the tailfins of control larvae but not in WHIM-GFP larvae (Figure 6A; supplemental Video 5). Similarly, transient expression of WHIM-GFP in neutrophils of spint1ahi2217 larvae19 impaired neutrophil recruitment to the tailfin (supplemental Video 6). WHIM-GFP neutrophils also failed to respond to tail transections (Figure 6B; supplemental Video 7). Interestingly, although neutrophil chemotaxis to tail transections was impaired, wounding induced increased protrusion and localized motility of WHIM-GFP neutrophils in the CHT (supplemental Video 7) of transgenic larvae. Taken together, these findings suggest that, even in the presence of robust inflammatory stimuli, neutrophils show impaired mobilization and recruitment into inflamed tissues in the WHIM transgenic.
To determine whether ectopic expression of wild-type CXCR4 receptor also affected neutrophil recruitment to wounds, we quantified neutrophil numbers at wounds in transgenic larvae that express wild-type CXCR4-GFP in neutrophils. Although neutrophil recruitment to wounds was reduced with ectopic expression of wild-type CXCR4b (supplemental Figure 4F), neutrophils remained responsive to wounding in contrast to WHIM-GFP larvae (supplemental Figure 4C-F; supplemental Videos 8-9). It is possible that this difference in recruitment may be due to variation in receptor expression levels or alternatively increased activity of the WHIM truncation compared with wild-type receptor. In any case, the findings indicate that the WHIM truncations induce a gain of function in CXCR4 signaling that impairs neutrophil motility in response to inflammatory signals in vivo.
Previously, we reported that larval inflammatory macrophages, a subset of macrophages, express low levels of GFP in MPO:GFP transgenic larvae, are highly phagocytic, have an elongated morphology, respond to wounds in the tailfin, and express L-plastin but not MPO.22 This population of cells will hereafter be referred to as macrophages. In WHIM-GFP larvae, macrophages do not express detectable levels of the WHIM transgene (K.B.W, unpublished data, April 2009). To determine whether macrophage recruitment was also affected in the WHIM-GFP transgenic, we examined macrophage chemotaxis to the wound by quantifying macrophage number with the use of double immunolabeling for MPO and L-plastin as recently reported.22 In control larvae, both L-plastin+/MPO+ neutrophils (Figure 6C bottom row arrows) and L-plastin+/MPO− macrophages (Figure 6C bottom row arrowheads) were recruited to the wounds. In 100% of WHIM-GFP larvae examined, L-plastin+/MPO− macrophages (arrowheads) were present at the wounds, whereas no L-plastin+/ MPO+ neutrophils (arrows) were detected (Figure 6C top row). This is an interesting observation because neutrophils are the first cells recruited to wounds in vivo. Our findings suggest that impaired neutrophil recruitment does not affect the later recruitment of macrophages.
Depletion of SDF1a rescues neutrophil motility in WHIM-GFP larvae
Previous studies have suggested that enhanced CXCR4-SDF1 signaling mediates neutrophil retention in the BM in patients with WHIM syndrome. However, current models are not amenable to directly testing this hypothesis. To determine whetherSDF1a mediates neutrophil retention in the zebrafish model of WHIM syndrome, we depleted endogenous SDF1a with the use of MO. After injection of a control MO, WHIM-GFP neutrophils showed impaired chemotaxis to wounds (Figure 7B) or tail transection (Figure 7E). In contrast, we found that after injection of the SDF1a MO, WHIM-GFP neutrophils showed robust recruitment to wounds (Figure 7A,C; supplemental Video 10) and tail transections (Figure 7F). Quantification showed a significant rescue in the ability of neutrophils to respond to wounding (Figure 7D) or tail transection (Figure 7G) after injection of the SDF1a MO. Taken together, the findings provide the first direct evidence showing that SDF1a is necessary for the neutrophil retention signal induced by CXCR4 WHIM truncations. Moreover, the findings suggest that intrinsic cell motility in the absence of SDF1a is not affected by truncation of CXCR4, but that the mutation increases the sensitivity of neutrophils to endogenous SDF1a, thereby inducing a retention signal that impairs neutrophil recruitment.
Localized activation of Rac is sufficient to direct migration of WHIM-GFP neutrophils
Rac guanosine triphosphatases are critical regulators of SDF1-induced directed migration in hematopoietic stem cells, T cells, cancer cells, and endothelial cells.32-35 Interestingly, previous studies have reported altered SDF1-induced F-actin polymerization in neutrophils from patients with WHIM, suggesting that WHIM neutrophils may have defects in polarized F-actin polymerization.8 To determine whether polarized, Rac-induced, F-actin polymerization is sufficient to rescue the directed migration of neutrophils in WHIM-GFP larvae, we used a genetically encoded Rac1 guanosine triphosphatase, which can be photoactivated reversibly and repeatedly by a 458 nm light,36 and can direct the migration of neutrophils in vivo.28 Photoactivation of Rac at the cell edge of WHIM-GFP neutrophils induced pseudopod protrusion and directed cell migration in WHIM-GFP larvae (Figure 7H-I) similar to control neutrophils.28 Repeated photoactivation of Rac was sufficient to direct the migration of WHIM-GFP neutrophils away from cell aggregates in the CHT (Figure 7J; supplemental Video 11) and into the tailfin (∼ 85μm in this example). After photoactivation was stopped, the neutrophil rapidly migrated back to the cell aggregates in the CHT (Figure 7J; supplemental Video 12). Together the findings suggest that polarized F-actin polymerization induced by Rac activation is sufficient to rescue the directed migration of WHIM-GFP neutrophils in vivo.
In this study we report a zebrafish model of a primary immunodeficiency syndrome, WHIM syndrome. We used the optical transparency of zebrafish to visualize how CXCR4-SDF1 signaling affects neutrophil trafficking in vivo. We have shown that expression of WHIM mutations in zebrafish neutrophils induces peripheral neutropenia and neutrophil retention in hematopoietic tissues, reminiscent of the human disorder. Live imaging shows that constitutive CXCR4-SDF1 signaling impairs persistent, directed neutrophil motility, thereby retaining neutrophils in regions of high SDF1a expression and inducing neutrophil aggregates. The neutrophil retention signal induced by WHIM truncation mutations is SDF1a dependent, because depletion of SDF1a with the use of MO restores neutrophil mobilization. The finding that SDF1a is required for neutrophil retention in the WHIM transgenic provides the first in vivo evidence to suggest that constitutive signaling through CXCR4-SDF1 mediates neutrophil retention in hematopoietic tissue contributing to peripheral neutropenia. Taken together, these findings support the utility of the zebrafish system to understand chemokine signaling through CXCR4-SDF1 and to identify putative targets that modify CXCR4-SDF1 functions in vivo.
CXCR4 and SDF1 have been studied during developmental cell migration, including migration of primordial germ cells, development of the lateral line, and movements of endodermal cells in gastrulation.2,37,38 Here, our findings suggest that neutrophils are located in areas where SDF1a mRNA is highly expressed, including the CHT and regions of the head in wild-type larvae. Previous studies have shown that endogenous SDF1a is essential for normal targeting of primordial germ cells in zebrafish.2 In contrast, we found that depletion of endogenous SDF1a mRNA by MO did not affect the targeting of neutrophils in control zebrafish, suggesting that there may be redundancy with other factors that also mediate neutrophil retention in hematopoietic tissues in vivo. However, when higher concentrations of SDF1a MO were used, we found that neutrophil development was impaired with fewer neutrophils present in the CHT, suggesting that endogenous SDF1a may affect targeting of hematopoietic stem cells that mediate neutrophil development in zebrafish, consistent with previous reports in the literature.39
This is, to our knowledge, the first study to visualize the consequences of altered chemokine signaling on neutrophil trafficking in vivo with the use of live imaging. Previous studies have reported that truncations of the CXCR4 receptor that impair receptor internalization do not alter chemotaxis of primordial germ cells, but rather affect the precision of migration by inducing prolonged "runs" and less tumbling-type migration in vivo.15 In contrast to the findings with primordial germ cell migration, we found that neutrophils in the WHIM transgenic displayed prominent random membrane protrusions but impaired persistent motility in vivo, resulting in retention within areas of high SDF1a expression. Our findings indicate that truncation of the C-terminal tail increases CXCR4 sensitivity to endogenous SDF1, supporting previous reports that suggest WHIM truncations block proper internalization and desensitization of the CXCR4 receptor.8,9,13 Taken together, the findings suggest that constitutive CXCR4-SDF1 signaling induces neutrophil stopping or retention within areas of concentrated SDF1a, including hematopoietic tissues, thereby impairing neutrophil trafficking into the vasculature or to tissue wounds.
There has been substantial recent interest in defining mechanisms that stop cell motility and contribute to retention within tissues. Perhaps the best studied example of stop signals has been in the context of T-cell motility stopping in response to contact with antigen-presenting cells with cognate antigen.40 Neutrophils have also been reported to display stopped migration in response to specific factors, including tumor necrosis factor-α.41 Stop signals have also been implicated in chemokine-mediated signaling in vitro. For example, exposure of neutrophils to one chemokine can impair the migration to other chemokines in vitro.42 Here, we demonstrate the effects of competing signals on neutrophil motility in vivo. Constitutive signaling through CXCR4-SDF1 impairs neutrophil recruitment to wounds despite competing gradients of hydrogen peroxide produced at the wound.43 Interestingly, polarized Rac activation was sufficient to overcome this retention signal and induce directed migration of WHIM-GFP neutrophils. Taken together, our findings suggest that neutrophil retention signals probably play an essential role in affecting neutrophil trafficking in vivo.
Zebrafish provide a powerful system to model human disease and to allow unique insight into disease pathogenesis because events can be visualized in vivo by real-time imaging. This has been shown in studies observing leukocyte trafficking in the context of chronic inflammation mutant lines,19,31,44 bacterial infection,45 and during granuloma formation in zebrafish mycobacterial infection.46 In addition, depletion of WASp, the protein mutated in the immune disorder Wiskott-Aldrich Syndrome, has shown defects in wound inflammation and clot formation.47 The zebrafish model of WHIM syndrome presented here provides novel insight into the pathogenesis of neutropenia in patients with WHIM syndrome. The findings show that the WHIM truncations induce a gain of CXCR4 function that requires the presence of SDF1a. Human studies and mouse models of WHIM syndrome supported the hypothesis that the WHIM mutation induced neutrophil retention in the BM, causing neutropenia. We now provide direct evidence that WHIM-induced neutrophil retention is mediated by altered signaling that can be rescued by depletion of SDF1a and blocking CXCR4-SDF1 signaling. Patients with WHIM are commonly treated with granulocyte colony-stimulating factor (G-CSF), which is thought to mobilize neutrophils from the BM by down-regulating expression of CXCR4 on neutrophils and reducing SDF1 expression in the BM.48,49 Because the zebrafish homolog of G-CSF has been identified and characterized,50 the zebrafish model of WHIM syndrome provides an ideal system to study how G-CSF affects neutrophil trafficking in vivo in WHIM syndrome.
Here, we used the optical transparency of zebrafish to visualize how CXCR4-SDF1 signaling affects neutrophil trafficking in vivo. Our work has demonstrated that constitutive signaling through the CXCR4 receptor induces neutrophil retention in hematopoietic tissue that impairs neutrophil motility and recruitment to inflamed tissues. The neutrophil retention signal is mediated by CXCR4-SDF1 signaling, because depletion of SDF1 restores neutrophil motility and wound recruitment. We propose that the WHIM transgenic zebrafish will provide a powerful in vivo tool to screen for agents that modify CXCR4-SDF1 signaling and may therefore have therapeutic potential.
An Inside Blood analysis of this article appears at the front of this issue.
The online version of this article contains a data supplement.
The publication costs of this article were defrayed in part by page charge payment. Therefore, and solely to indicate this fact, this article is hereby marked "advertisement" in accordance with 18 USC section 1734.
We thank Erez Raz for generous gifts of plasmids; J.R. Mathias, M.E. Dodd, Q. Deng, P.-Y. Lam, and E.A. Harvie for zebrafish feeding and maintenance; A.J. Wiemer, S.A. Wernimont, K.T. Chan, T.W. Starnes, and C.L. Cortesio for critical reading of the manuscript; and J.J. TeSlaa for help with sectioning.
This work was supported by the National Institutes of Health (grant GM074827; A.H.).
National Institutes of Health
Contribution: K.B.W. designed and performed experiments, analyzed data, and wrote the paper; J.M.G., J.C.S., and S.K.Y. performed experiments; and A.H. designed experiments, analyzed data, and wrote the paper.
Conflict-of-interest disclosure: The authors declare no competing financial interests.
Correspondence: Anna Huttenlocher, University of Wisconsin-Madison, 1550 Linden Dr, Madison, WI 53706; e-mail: firstname.lastname@example.com.


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Sausages can be cooked in different ways - boiled, grilled - and served with different vegetables. Source: Lori / Legion Media
I don't think I can move along any further with this blog without acknowledging the meal that is so Soviet that it should have been banned by the new government in 1991.
It's sausages. Not the organic, turkey-with-a-bunch-of-herbs type, where you all but get the bird's name on the label. No, the ones I mean are the brown-grey ones, with names like "delicious," or "milky," or just the name of the manufacturer, like "ostankinskie."
Nothing about these sausage packages would give any indication of what the sausage is actually made of.
I think the producers are trying to distract you from the very fact that the ingredients are…well, you don't want to know what they are.
It doesn't take long to distract the sausage-buyer though – "Ah, it's so easy to deceive me!..I'm grateful to be deceived!" – as a line in a poem by Russia's most famous poet, Alexander Pushkin, reads.
Foodies, skip the next line, or go eat a turkey sausage, because you're not going to like this: I, too, occasionally close my eyes and buy a packet of sausages, bring them home, cook them, and then eat and enjoy them, too.
When I was a kid, granny would sometimes make sausages with stewed cabbage: she'd cut up the sausages and distribute them among the cabbage in the faint hope that we might accidentally eat some cabbage, too. We were no fools, and never did.
After digging and consuming all the delicious sausage bits, we would spread the cabbage thinly on the plate and, content with the immaculate execution of the "cunning plan," would put our heads on our arms and say in a low voice, slowly: "bol'she ne mogu!" (I can't eat any more!). Then granny would try and convince us to have some of the cabbage, we would make puppy eyes and look all cute, and she would eventually give in.
This time though, I have to make and eat not just the sausage, but the cabbage, too, as that's what the recipe calls for. Worse still, I have to somehow feed it to my husband. I fear failure.
1 kg cabbage; 2 onions;
2 tablespoons tomato paste;
1 tablespoon vinegar;
1 tablespoon sugar;
1 tablespoon flour;
3 tablespoons butter;
salt and pepper to taste.
Lucky for him, the cabbage shrunk a lot during cooking so the sausage/cabbage ratio worked in his favor, and I got away with it.
My grandmother has a very similar view on sausages:
"Sometimes I would come in to the cafeteria and think to myself: 'I feel like some sausages,' so I'd buy two, some white bread and as soon as I finished eating I would feel disgusted and didn't eat them again for a couple a months – then I would do the same thing again. Today there is a choice of bad sausages, but back then there only one type. Also, I didn't buy sausages for you in Moscow – only in Estonia in summer as they were really good there."
We also discovered that the Soviet sausages didn't have any MSG in them. What's the point of junk food without MSG in it, I ask? That would be the one ingredient I would look for on the sausage cover. But that won't be for another year or so.
The recipe from the Soviet Cook Book, page 213. Click to enlarge the image
Shred cabbage and put in a saucepan with a tablespoon of butter and ½ cup water or broth. Cover and let simmer 40 minutes. In the meantime, slice the onions and brown them in another tablespoon of butter. After the cabbage has cooked 40 minutes, add onions, tomato paste, vinegar, sugar, salt, petter and 1 bay leaf. Simmer until tender, about 10 minutes. Melt last tablespoon of butter in a skillet and add flour until it is toasted. Add cabbage to the toasted flour, and bring to a boil.
Sauerkraut can be used in place of fresh cabbage, but in this case, you do not need to add vinegar when cooking it.
The recipe from the Soviet Cook Book, page 71. Click to enlarge the image
There are several different ways to prepare sausages. They can be boiled in lightly salted water and served with mustard or grated horseradish.
They can also be fried with tomatoes. To do this, cut sausages crosswise into 3-4 pieces, fry them in a pan with butter for 2-3 minutes, then add tomatoes that have been thinly sliced and sprinkled with salt and pepper and fry for two more minutes. Grated garlic can be added as well, and the fresh tomatoes can be replaced with canned tomatoes or tomato paste.
The recipe from the Soviet Cook Book, page 158. Click to enlarge the image
Sausages can also be taken out of their skins, sliced and then fried.
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Week 6 Daily Fantasy Football Value Picks
Ugh, what a rough outing for me last week. Fantasy Football can be just as frustrating and annoying as it can be fun and rewarding. All signs pointed toward a big week for Owen Daniels, who I hyped up all week. Because of this, he predictably fell flat on his face against Oakland. However, the beauty of daily fantasy is that every week allows you the opportunity to choose new players and get back in the game.
With that, let's dust ourselves off and get to the best values for week 6. Again, all prices come from DraftKings.
QB: Andy Dalton – $5700 STEAL OF THE WEEK
Here's a list of the quarterbacks averaging more points than Andy Dalton in games played: Tom Brady. That's the list. In DraftKings scoring Andy Dalton is averaging about 25 points per game and is absolutely on fire right now. He has a large collection of weapons to throw the ball to, and is playing the best football of his career. At this point, you have to roll out Dalton until his price rises to a point where it doesn't make anymore sense from a value perspective. Quick addition: Joe Flacco is also a great play this week. Both QBs are playing lower ranked passing defenses this week and cost less than $6000. Play either and you'll do well.
Honorable Mentions: Joe Flacco ($5900), Sam Bradford ($6000)
RB: Charcandrick West – $4000
Another running back pick that could blow up in my face like Boobie Dixon did last week, but the value is far too enticing. Completely unproven in Kansas City, West is in line to get the bulk of the work with Jamaal Charles injured. When you can get a running back who is going to start for this cheap and pair him with top-tier guys, you can do well in daily.
Honorable Mentions: Danny Woodhead ($4700), Darren Sproles ($3800)
WR: Donte Moncrief -$5000
Nothing too exciting with this pick and he isn't super cheap either. However, I have my reasons for choosing him. It's more likely that Andre Johnson's performance from last week was a flash in the pan than a consistent trend. With the return of Andrew Luck this week, expect him to look Moncrief's way quite a bit. Plus, the Colts are playing New England who will likely force the Colts to play catchup all game.
Honorable Mentions: Eric Decker ($4900), Mike Wallace ($4500)
TE: Tyler Eifert – $4900
Nothing too exciting from the tight ends this week in terms of value, so I went with a guy who won't break the bank but has the potential to top the list when all is said and done. Like Dalton, Eifert has a surprisingly tasty matchup on the road this week against Buffalo, who rank 27th against tight ends in DraftKings scoring. Unlike the Owen Daniels pick last week, Eifert is an integral part of the Bengals offense and often sees plenty of targets. Plus, this brings the popular daily fantasy strategy of stacking into the discussion, where someone plays a QB and a skill position player (often WR or TE) from the same team if they feel like they could have big weeks. With Dalton and Eifert both being relatively cheap, a huge game from the Bengals likely means huge games from both of them.
Honorable Mentions: Martellus Bennett ($4800), Zach Ertz ($2900)
DST: Minnesota Vikings – $2900
Playing at home against a struggling Chiefs team who just lost their best player and coming off of a bye, I think the Vikings have tons of potential at their current cost. They've averaged 8.8 points per game, which isn't terrible, and generate an average amount of turnovers. However, I do have a huge disclaimer. Earlier, I recommended Charcandrick West as a value play based on the sheer workload that he was going to get as the new starter. DO NOT start the Vikings defense and West in the same Daily Fantasy lineup. Although not impossible, it is highly unlikely that both will score a lot of points this weekend, as one will likely do well at the other's expense. You can get away with this in normal lineups, but not in daily ones.
Honorable Mentions: Detroit Lions ($2500), Chicago Bears ($2400)
Kyle Kirby covers Fantasy Football for MFST. You can follow him on Twitter @couch_qb
Kyle Kirby
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