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To coincide with the release of our Product Pages Best Practice Guide, I've been looking around for examples of excellent pages from e-commerce sites.
Not every page in this list is perfect, but they all contain great examples of features that have been used to showcase and sell products, such as great use of video and imagery, presentation of product features, and user reviews.
Amazon's product pages are long and require lots of reading and scrolling (it would take screenshots to show the whole iPod Touch page) to digest the lot, but the pages clearly work well, and contain plenty of innovations that are now commonplace on e-commerce sites.
The most obvious of these innovations is user reviews, but the cross-selling options (customers who bought this also bought...) are also inspired. The pages may be cluttered, but if customers are looking for any information about a product, it's almost certain to be on the page somewhere.
Schuh is a great example of how product images can be used to overcome the fact that customers cannot see product at close hand or try shoes on.
One way it does this is through a prominently displayed 'easy 365 day returns' policy, which reassures customers that items can be returned, and the other is through product images.
Schuh provides eight images of these trainers so shoppers can see them from every conceivable angle, and there is also the option of a 360° rotating view of the product.
The product pages of gadget retailer Firebox are almost as cluttered as Amazon's, but they contain just about everything you need to know about the products.
I like the use of product videos sent in by customers which show the various gadgets in use.
Firebox pays £50 for each video submitted, and this looks to be value for money, as the videos show other shoppers that the product works, and that other customers have bought it without any problems.
ASOS gets many things spot on with its product pages. Key information on delivery and returns is provided in a prominent position above the fold, cross-selling is relevant, and a nice big call to action has been placed in the centre of the page.
ASOS also does well with product imagery, with several views provided, as well as the option of a catwalk video, which can be quickly viewed without leaving the page or opening a pop-up window.
Kiddicare's product pages are great at conveying everything customers will need to know about a product. Multiple images and videos are provided, handy for showing how to fold and unfold a pram for instance.
It also has plenty of product information, while showing both the countdown clock for next day delivery and the number of products left is a good way to create urgency in the customer's mind.
What I especially like about Kiddicare's product pages is the presentation of user reviews. Reviews are shown in a lightbox on the page, which saves space, while summarising pros and cons, and best uses of the products is very helpful for shoppers.
Hotel Chocolat
The product pages on this site do the basics reasonably well, but are most notable for the quality of product photography. High resolution images of chocolates like this are great for stimulating the taste buds of shoppers.
Etsy provides plenty of information on its product pages, and presents information about sellers in a useful way for shoppers:
Customers can see feedback information about the seller at a glance, or can contact them with any questions. I also like to two calls to action have been placed on the page so they can be seen above and below the fold.
Comet does an excellent job of presenting a lot of product information, technical specifications, and useful tools for customers without making its product pages too cluttered.
There is a lot of useful detail here; reviews and a summary of the average score, a summary of product features above the fold, clear delivery information, and even the option to enter a postcode and check stock at your local store.
Tabs are used to present the product details, user reviews, buyer's guides and FAQs, which keeps the page at a reasonable size. Nice clear calls to action too.
The product pages on this site are a great example of best practice. The page contains plenty of product information and reassurances for customers about delivery, returns etc, while still retaining enough white space to make the page easy to digest.
Webtogs uses icons to present key information in a way which stands out, so customers scanning the page can quickly see the dimensions of the tent, the fact that delivery is free, and the big add to basket button.
Naked Wines
I have included these product pages as they contain a couple of elements which are not used in the previous examples. One is the option to 'meet the winemaker' and get an idea about the story behind the wine.
The other is the use of user reviews and ratings to help shoppers make a decision about the wine. The wall underneath the image provides lots of comments from other users about the wine, which can be very useful.
The box on the right, which shows the percentage of users that would buy the wine again, and whether it represents value for money or not is a great idea, simply presented.
I'm sure there are some other great product pages that I haven't mentioned in this list. Let me know your suggestions below...
Comments (21)
Could I put in a word for Net-A-Porter, which demonstrates the roles engagement and inspiration can play in e-commerce? The sites you chose are high on function, but low on those key factors, surely high on the requirements of a long-term e-commerce success?
That's a really useful list. Co-incidentally we ran a usability review of Firebox.com yesterday which you can read about on our blog here:
Online Christmas Shopping – Firebox.com review
Hi Paul,
I nearly added Net-A-Porter to the list but the pages are similar to Asos so I left them out. A good example though.
I think the only thing this article is missing is some examples of how brands integrate social sharing into their product pages, such as with Tweet this and Facebook Like buttons.
Boden also deserve a mention for the way they've approached size selection on products- doing away with dropdowns (which hide out of stocks) and showing immediately what is available/ isn't available or indeed when it will be available.
Some good examples here. It's important to note though that although each of these elements are important it's also vital to make sure the page works as a whole and information is gradually revealed where needed, and in a logical order!
Looks like some incredible pages. i love the tent.
Amazon has been used as a good example for a while now by various people, From an SEO point of view they have a lot of information and content which is good, From a conversion point of view they have all the info there to reasure the buyer (or put them off)
And from a user point of view, everything can be done above the fold!
Great list. Getting all the relevant information above the fold is vital. Also, as a high percentage of visitors will enter at the product page and not see your competitive advantages it's important to have them on there. Such as http://www.business-accessories.co.uk/products/Wallee-%252d-iPad-Wall-Mount.html. Webtise
Oh dear! If these are the best.....I'd hate to see the worst. This is the best argument for getting some new blood into web design. Strikes me that UX people, planners and clients have had too much influence.
These product pages look great, used the Asos site before and always thought it was displayed very well. The Naked Wines page is very nicely done. Often I see shops and they can be really let down by their product pages but all these look great.
some really nice ones there, including one of my clients, so i'm really chuffed, it would have been good to see the numbers relating to the pages (coversion rates) though thats obviously wishful thinking. I'd also like to see how many of these are using CRO in this part of their site, are any dynamically optimising their product pages based on where users are coming from or what users have previously browsed, is that previous interaction affecting how the site is presented? really being a data monkey, i see this more as an opinion of whats best (not that there's anything wrong with that) rather than what is proven to be best with numbers and statistics.
Product page design is always a balance between users real-world needs, design principles and creativity, brand considerations and designing an optimal conversion path. What is sad is that so many companies still get that wrong. I agree, the list if far from complete, but it offers a snapshot. Thanks.
As a product photographer, I see a lot of product pages everyday. Some are good, some are bad and some don't work at all. The thing I always try and remind my customers is Keep it simple so your customer can navigate through it.
GREAT examples of good product focus, clean design (in some cases!), and how to visually channel the customer towards purchase.
Interesting that I didn't see examples of social shopping--but this probably because as a new trend, not as many are yet using it. But, like ratings & reviews, this is clearly going to be the next area of innovation.
Quorus is already leading in the social shopping area and has some exciting projects underway. Keep your eyes open for new customers taking advantage of this.
As the recent IF study noted on social shopping, "Word of mouth is nothing new--what is different is where it is taking place." Your customers will be taking you here (to social shopping), so best to start thinking about it now!
Thanks for these great examples of product detail from ecommerce sites. Look forward to more!
I noticed amazon mentioned, I think they are heading towards - how not to do a product page. Information overload in my book. I think for those in the web world we are patient and happy to find the end goal. My wife gives up after about 5 secs of not finding the checkout button and I am sure she is not alone. Enough info to help people through the buying process, but then clear and simple next steps.
Hi Michael - I take the point about Amazon, and I'm not sure that kind of clutter would work on other sites. I included Amazon mainly for the innovations in cross-selling and user reviews.
Nice little report. We aim to give our customers as much information as possible on the product. We also take our own pictures of the product as most supplier images can be very different from the finished product!
Also, (just in case we get a flood of customers asking for £50 each) we reward the best video with a £50 voucher, but all video entries get a £10 voucher.
The abel and cole ecommerce/shopping cart process is really strong - it holds the users hand and the design is reassuring - nice piece of work. I agree with previous poster about Amazon - losing some clarity with their interface - information overload - that said they must do oodles of conversion optimisation so it is probably working well for them despite its lack of aesthetic quality
Great post Graham, and the Econsultancy product pages best practice guide is well recommended in case readers/contributors haven't taken a look yet.
In response to one or two comments made on this post:
@Aaron Buckely - great example of how to encourage your customers at Firebox.com to get involved in generating content (and not just any content, content that will actually help to convert future visitors into making a purchase with the user-generated videos).
Looking beyond the product page, I've seen that you have moved to an enclosed checkout. The reason I mention this is that your website features in the e-commerce usability and best practice training course I deliver for Econsultancy. In addition around 18 months ago I wrote an blog post on Econsultancy looking at whether retailers are following best practice to increase their conversion rates, which looked specifically at the concept of enclosing your checkout. You will see that at the time your website featured as a 'checkout not enclosed' example.
Would you be able to share any insights into the (hopefully!) improved checkout conversion rate that you have seen since moving to an enclosed checkout?
@Kevin Barnes - I agree, Boden is a good example of ensuring that stock availability is made clear when choosing your size. For out-of-stock items perhaps they could look at providing more information such as if/when a particular size will be back in stock (and even allowing a visitor to get notified when it does) although this isn't currently on most e-commerce sites I work with.
@Graham - as you point out Amazon certainly isn't the model that all retailers should be following - my somewhat controversial blog post on Econsultancy - Amazon relying on brand credibility rather than good usability - talks in depth about the customer experience of both Amazon and The Book Depository. In summary, if you were new to online shopping and hadn't heard of Amazon (yeh I know, ticking both of these boxes would be a hard task!) then my word would Amazon be an overwhelming and potentially frustrating 1st time shopping experience.
@All readers - if you have benefitted from the advice and recommendations in this post, then you will be interested in the training course I mentioned earlier in this comment that I deliver:
Econsultancy Training - E-commerce Usability and Best Practice
Key Areas Looked At:
- product page best practice
- cross selling and up selling
- shopping basket best practice
- checkout best practice
- checkout form best practice
- advanced user experience techniques
The course includes live evaluation and competitor benchmarking which always proves to be one of the most popular parts of the training course.


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Recently, I was asked an interesting billing question many acupuncturists will be able to relate to. An acupuncturist who was new to billing had some concerns over how to bill. This acupuncturist had heard in addition to the acupuncture codes (97810-97814) that there was also a code to bill for the needles themselves.What is this code and as is the correct way to bill? Other questions included: Can I bill four units of acupuncture if I spend one hour with the patient? Finally, I am also curious about how the ANSI 5010 format for billing will affect me? I heard it will change billing formats in 2012 and I have no idea what it is.
The first question is one that is often confused by many acupuncturists as there are codes for needles (A4215 needle, sterile, any size, each) however, it is appropriate for this code to be used for acupuncture services. Using needles is integral and essential to have acupuncture and as such needles are considered part of the service and not separately billable. In other words, the value of the needles is included in the charges for the overall service.
Specifically, the needless are considered "incident to" the acupuncture service. In billing for acupuncture you may bill for the sets of needles provided with codes 97810 initial set, manual, 97811 additional set(s) manual or 97813 initial set, electrical, and 97814 additional set(s) electrical. And, no separate coding for the needles themselves.
The needle code is used for the patient to take home and use for home injections like what a diabetic patient would use for insulin.
The latter question is often a confusing one as well as many acupuncturists may read the codes and place an emphasis on the aspect that indicates time. While the acupuncture codes are listed as 15 minutes face-to-face time one cannot ignore that it also requires additional needle(s) to qualify. If you were to spend time face-to-face with the patient, insert a set of needles, they rest on the needles for a period of time (even up to an hour or more) the code that can be billed would be for only one set. As 97811 or 97814 must have reinsertion of one or more needles. To bill for 4 units there would have to 4 separate insertions of sets of needles to qualify.
The American National Standards Institute (ANSI) 5010 may have no immediate impact on offices not billing insurance or billing electronically. ANSI 5010 is the new version of HIPAA transaction standards that regulates the electronic transmission of healthcare transactions. Meaning the format of and how electronic claims are formatted and transmitted to the carrier. This is covered under HIPAA of course as the transactions must be secure. This 5010 standards will replace the existing 4010/4010A1 version of HIPAA transactions as if creates a fix to the shortcomings in the current version, including that the current format does not support forthcoming ICD-10 coding.
In many ways these standards are clearing the way and standardizing health care information into one standard which will allow a universal electronic health record system that is compatible for all health care providers and payers. The goals are to:
- Increase transaction uniformity
- Support pay for performance
- Streamline reimbursement transactions
- Support ICD10 codification
The implementation of HIPAA 5010 has substantial changes in the data submitted as well as the data you receive in response to your electronic inquiries. The implementation may require changes to the software, systems, and perhaps procedures that you use for billing Medicare and other payers. However, in most but a few instances these changes are the responsibility and format used, but the clearinghouse and may require no or an update to your current software. The older your version of billing software the greater the possibility of a more complex and potentially costly upgrade.
Based on this it would be a good time to evaluate your current system and whether or not an upgrade is necessary and if so, and the cost is high it may be a benefit to change to a newer system that is online and has no costs for upgrades. There are several services for electronic billing that are no cost to the provider and are compliant with the 5010 standard. However, any change must be thought out thoroughly. For example, consideration must be made to patient information already in your current database being compatible and easily transferred or mapped to the new system. Also an online version of information only may mean when the system or Internet is down, you will have no access to your data.
For electronic billing under the 5010, the date of implementation is January 1, 2012. Testing should have been or is being done by all clearinghouses and software manufacturers to ensure compliant and correct by the January deadline.
It is likely you have already received information from your current software manufacturer or clearinghouse for needed updates. While updates may be necessary on the provider side they can be from very little or none, to almost a new system with the commensurate costs attached. If you have had no information from your software or clearinghouse vendor take it upon yourself to contact them and assure that you have all that is needed to be compliant and they transmitting your information in the correct manner by January.
This 5010 standard is also setting the groundwork for the universal use of Electronic Health Records (EHR) as it creates a standardization of how the records are encoded, shared, and transmitted. The implementation of EHR is currently not mandated (date is set for 2014). There is an incentive bonus paid to some providers who use EHR in 2012, but unfortunately acupuncturists are not part of that group. Only providers who may bill and are paid by Medicare can receive the bonus.
The ANSI 5010 also affects the use of the upcoming ICD-10 (October 2013 implementation). The ICD-10 implementation will allow for greater specificity in defining diagnosis. For instance there is designation of right and left as well as initial encounter to subsequent encounter. The ICD9 codes number about 13,000 while the ICD10 is 68,000 with codes all having alpha beginning and variations of 3-7 digits codes and procedure codes. The codes are much more specific in the ICD-10 implementations of ICD10 in European countries have indications that its use will lead to improved healthcare services at lower costs with fewer errors and improved results. It appears the ICD10 will offer a greater level of codes that may be used by acupuncturists.
Click here for more information about Samuel A. Collins.

subset: commoncrawl


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9 October, K2 in Stockholm hosted the Nordic bouldering championships.
Among the men, the line-up was strong, but pretty much everyone expected the final to be Magnus Midtboe-show. The man has done both 9b and 8C this autumn, remember? The only possible threat, now that Nalle Hukkataival couldn't come, was expected to be Finnish prodigy Anthony Gullsten, who've climbed multiple 8B's as well as 8c+ this summer.
Daniel Andersson, competing in front of his home crowd, had other plans however.
Qualifying in a modest 4th spot, he turned it on in the final and flashed all four problems, thereby crushing all opposition. Great success!
Also, among the women, it was a Swedish success, and a double victory even, with Anja Hodann beating Matilda Söderlund into 2nd place by the slimmest of margins, with Hannah Midtboe taking the bronze.


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One of the questions I am asked most frequently is 'which email service provider or delivery platform is the best?'
The answer is 'how long is a piece of string?'
My company, Alchemy Worx use or have used many of the leading email delivery platforms on behalf of our client base, including Bluestreak, Cheetahmail, Digital Impact, Dreammail, Emailvision, Exacttarget, Ezmail, Lyris, Responsys, Silverpop and Twelve Horses.
But the right solution for a company can only be determined by evaluating a whole list of variables, including...
- Business focus - B2B, B2C or both?
- List size.
- Intended volumes.
- Intended frequency.
- Programme complexity.
- Reputation.
- The depth of integration with other channels.
Having said that, we have developed some general principles that should help you find the right ESP for your organisation. These guidelines also apply if you are looking for an in-house solution such as StrongMail, Message Systems or Lyris.
First of all, go for the larger companies. Their servers are going to be more robust, they're likely to have more staff, including a big ISP relations or reputation management team and have the ear of companies such as AOL, Microsoft and Yahoo!.
In particular, ask how many people they have working on ISP relations and reputation management. Ask whether their reporting can give you a breakdown by domain so you can see which ISPs are causing you problems.
For those that prefer it, there are some very good European ESPs to choose from, but don't be afraid to work with an American company as they dominate the email delivery space. This truly is a global business and experienced companies of scale are often US-based.
Establish how familiar they are with your needs. It makes sense to try to find out what percentage of their clients are of a similar industry, size and have similar needs to you. You can then select the provider with the most clients matching your scale and requirements.
Don't be seduced by functionality. Technology companies have a tendency to try and dazzle you with capability. The reality is that you are unlikely to get round to using most of that whizzy functionality in the short term and it will be many months, if not years, before you get round to sending highly-personalised messages built up from hundreds of dynamic content segments.
Focus on the functionality you need now or in the very near future. So what you're really looking for is ease of use and interface speed. The delivery service you choose should have a user-friendly, intuitive interface that makes setting up and automating repetitive tasks straightforward.
Support is also very important, particularly if you're going for a self-service solution. So ask how quickly they turn around queries, whether they offer telephone support or just email.
Email marketing is all about understanding how your customers are interacting with your communications. So the reporting interface is absolutely critical and there are significant differences between products.
For example, not all technologies allow you to group your mailings by campaign. In some ways, it's more important to look at the reporting interface than at deployment functionality. Good email marketing is a constant learning and feedback loop, which requires detailed reports.
Remember you know best when it comes to marketing your company's products and services and the ESPs can only provide you with technology that helps you achieve your objectives. Their tools alone will not turn a poor email marketing strategy or campaign into a good one.
Dela Quist is the CEO of
Alchemy Worx
© Alchemy Worx 2008
Related research: Email Marketing Buyer's Guide
Comments (7)
Saying "Chose a larger company because their servers are more robust"
is a bit like saying "Chose a larger bank as it is less likely to fail".
Server robustness is not at all linked to size -of course it is important. But equally important is delivery reputation and local support -ie can you call someone in the UK.
I was very happy reading this article, since it is years that we work on those points.
We built our system in a marketing perspective. These tools are for marketers and therefore has to work like marketers.
Lets face it, what is really important for a marketer.
A user-friendly interface (if possible in their own native language), clear and understandable reporting (with domainname reporting),
campaign management without the technical fuss and certainly full support (sometimes difficult if you work with a US based company and you work in Europe).
And you don't always have to be a big US company for that, as an example we are presenting our client case at MarketingSherpa Email Summit in Miami.
AND we are the only European speaker ;-)
Hi Chris
Thanks for your response, it is always good to get feedback - most of our clients are major brands such as The carphone warehouse Sainsbury's Bank and Intercontinental Hotels Group so that part of my advice is aimed at that kind of company.
I also qualified this point by making it clear you should find out what percentage of their clients are of a similar industry, size and have similar needs to you. You can then select the provider with the most clients matching your scale and requirements.
I totally agree that support is important and make that clear, however as per your comment on banks, being able to talk to someone in the UK does not guarantee good support or swift turnaround.
PS The only 2 banks I know that failed in the last 25 years are BCCI and Northern Rock both small ;-)
Hi Kenny
I agree Alchemy Worx is not a big US company either and I look forward to seeing you at the Insider Summit, I spoke there last year.
I think the first decision is whether it makes most sense to purchase a delivery system of your own or outsource the mail to an ESP. Larger organizations with existing custom designed data systems and applications may prefer the integration opportunities provided by a purchased - in house - solution. There is already a new wave of delivery platforms opening up that focus on deep integration with existing systems. For example, Hurricane Server, which is designed for Windows shops who would benefit from an MTA that integrates with their existing SQL databases and other applications. In Hurricane Server's case, developers plug their own .Net code into the process and handle delivery events in real time. For example, updating an address list at the time the message delivery fails. And Hurricane Server's web service API enables developers to automate configuration and administration from their own applications and interfaces.
If you do not have large existing systems however, it might make more sense for you to leverage what ESP's have to offer.
Exchanging information between external systems and internal databases are very crucial indeed. Therefore flexible webservices in a good platform are a must.
You benefit from both, the knowledge and deliverability monitoring of your ESP and the data-intelligence in your own infrastructure.
Actually you can do better! Connect and share an ESP platform with and external managed database with customer information.
You use your ESP system to collect and enrich online info, your external managed database to collect offline info. Synchronise both and use that intelligence to optimize both communication channels.
Couldn't agree more. The lines are blurring btween email marketing and marketing automation too. So consider the bigger picture before going just for email marketing, when marketing automation may get you further, faster.


text:
The best place to purchase a certified pre-owned Mercedes SUV is at a Mercedes-Benz dealership. Buyers can search for dealerships on the Mercedes-Benz website by entering their ZIP codes in the search box.Continue Reading
In April 2015, Mercedes-Benz has three different models of certified pre-owned SUVs for model years 2011 through 2014. The GLK-Class is a small SUV with a 3.6-liter V-6 engine and a seven-speed automatic transmission. The mid-size M-Class has a five-star safety rating. The GL-Class is Mercedes-Benz's largest SUV, and this model has off-road capabilities. Qualified buyers can apply for credit to purchase a certified pre-owned Mercedes SUV by filling out applications at Mercedes-Benz dealerships.Learn more about Car Makes & Models


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Hamilton-Fairfield Bike Trail is a 10.4 mile moderately trafficked out and back trail located near Hamilton, Ohio that features a river and is good for all skill levels. The trail offers a number of activity options and is accessible year-round. Dogs are also able to use this trail but must be kept on leash.
dogs on leash
kid friendly
mountain biking
nature trips
road biking
trail running
wild flowers
Hamilton monument on the river to Fairfield Waterworks Park
Nice trail very relaxing. Great for morning biking or evening.
nice ride
One of my favorites. This one continues north to Metro parks north of Hamilton.
Stay tuned, I'll map it next year.
The Hamilton bike trail now goes to rentschler forest. There and back 21 miles
Jon W.
This is a very nice trail for cycling. It's about 11 miles out and back, and scenic all the way. Starting in downtown Hamilton, it's only a few minutes before you forget you are in a city. The view is great along the Great Miami River is great and the trail is well kept. After you pass Miami University - Hamilton, you go through a wooded area, through Joyce Park, and then through a wildflower meadow before the turnaround.
Be on the lookout for families out for a stroll, and of course, be on the lookout for roller bladers, who seem oblivious to all around them, and take up the entire width of the trail!
Overall, a great little ride.


text:
1Cardiff University-Peking University Cancer Institute, Cardiff University School of Medicine, Heath Park, Cardiff, UK
2Department of Oncology, Wangjing Hospital of China Academy of Traditional Chinese Medicine, Hua Jia Di Street, ChaoYang District, Beijing 100102, P.R. China
3China Academy of Traditional Chinese Medicine, Nan Xio Jie, Dong Cheng District, Beijing, 100700, P.R. China
Received Date: January 17, 2014; Accepted Date: February 21, 2014; Published Date: February 28, 2014
Citation:Yin Y, Sanders AJ, Feng L, Jiang WG (2014) Knockdown of AAMP Impacts on HECV Cell Functions In Vitro and Affects the Expression of VECadherin. Angiol 2:125. doi: 10.4172/2329-9495.1000125
Copyright: © 2014 Yin Y, et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Visit for more related articles at Angiology: Open Access
Background: Angio-Associated Migratory Cell Protein (AAMP) is a 52 kDa protein expressed in a variety of human cell lines. Previous studies have demonstrated that AAMP is involved in endothelial cell adhesion, migration, and tube formation and plays roles in signalling pathways, such as RhoA-ROCK
Methods: In this study, we knocked down the expression of AAMP in HECV cells using hammerhead ribozymes and assessed the influence of AAMP on endothelial cells. In order to explore potential mechanisms, Wnt/β catenin inhibitors (FH535 or IWP2) were also used in a number of the functional assays and the immunofluorescent staining patterns of several key molecules were explored.
Results: Knockdown of AAMP expression was observed in the HECV cell line following transfection with the ribozyme transgene. Cellular migration and attachment, assessed using ECIS methods, and tubule formation were significantly inhibited by the knockdown of AAMP. Additionally, cell growth and cell matrix adhesion was also substantially reduced following AAMP knockdown, though this did not reach significance. Cell aggregation levels showed no statistical difference between AAMP knockdown cells and control cells. Interestingly, immunofluorescence staining showed AAMP knockdown cells had a reduced expression of VE-cadherin. Some overlap of function was seen between AAMP knockdown and the FH535 inhibitor.
Conclusion: AAMP appears to influence endothelial cell migration and tubule formation and potentially, to a lesser effect, cell matrix adhesion and growth and also affects the expression of VE-cadherin. This data suggests that AAMP may play a role in angiogenesis. Assays performed with the FH535 inhibitor suggest a potential relationship between AAMP and Wnt/β catenin signalling.
Angio-Associated Migratory Cell Protein AAMP; Angiogenesis; Endothelial; VE-cadherin; Wnt inhibitor
The angiogenesis process is vitally important for growth, development and maintenance of a normal physiological state, playing key roles in essential processes such as reproduction, wound healing, and development [1,2]. Despite this, imbalance of the angiogenic process is observed in a number of disease states. One key example is seen in cancer, where tumour angiogenesis is essential in facilitating advanced tumour growth and increased metastatic potential, a realisation that has led to the development of anti-angiogenic therapies [1,2].
In 1995, Beckner et al. [3] isolated a 52kDa protein from a human melanoma cell line, termed Angio-Associated Migratory Cell Protein (AAMP), during a search for motility associated cell surface proteins. The cytoplasmic and membrane located protein contains two immunoglobulin-like domains, a WD40 repeat and a heparin binding consensus sequence [3]. The WD40 repeat motif is found in a wide range of proteins with diverse roles in signal transduction, transcriptional activation, cytoskeletal regulation and cell cycle control [4]. The homology of AAMP with members of the immune globular super family suggests AAMP may have similar roles to these members, which include recognition and binding, cell-cell interaction and cell signalling [5-9].
AAMP has a wide expression pattern, being observed in numerous endothelial and aortic smooth muscle cells, activated T-lymphocytes, renal proximal tubular cells, dermal fibroblasts, glomerular mesangial cells, rat myocytes and a number of cancerous cell lines including human melanoma cells, prostate and breast carcinoma cells and benign mammary cells [3,10-16]. A number of studies have focused on this molecule in cancer, demonstrating an enhanced expression of AAMP in invasive gastrointestinal stromal tumours [9]. Enhanced expression of AAMP was also seen in breast Ductal Carcinoma in Situ (DCIS) with necrosis and found to be elevated in the T47D cell line subjected to hypoxia [17]. Recently, a study conducted in our laboratories has explored the importance of AAMP in a clinical cohort of breast cancer patients and the impact of targeting AAMP in a number of breast cancer cell lines [16]. Our data suggests that AAMP levels are elevated in tumour compared to normal samples and higher expression of AAMP is generally associated with a poorer patient prognosis [16]. Additionally, knockdown of AAMP in MCF-7 and MDA-MB-231 breast cancer cell lines brought about reductions in aggressive traits such as growth, matrix adhesion and cellular invasion, though the impact on these traits was different between the two cell lines [16,17]. Taken together, these studies implicate a role for AAMP in cancer progression.
A role for AAMP has also been implicated in the process of angiogenesis, where anti-recombinant AAMP can inhibit endothelial tube formation on Matrigel and the cellular motility of endothelial and smooth muscle cells [11,18-20]. Similarly, it has been suggested that the extracellular form of AAMP has a positive role in angiogenesis [18]. Further studies have implicated AAMP in the translocation and activation of RhoA in both endothelial and smooth muscle cells and subsequently, influence downstream signalling of RhoA through its effector Rho-Associated Kinase (ROCK) and hence cell migration [20]. AAMP has also been identified as a novel binding partner of thromboxane A2 receptor alpha and beta (TPα and TPβ), potentially implicating it in vascular pathologies [14].
In this study, we knocked down the expression of AAMP in HECV human endothelial cells, using hammerhead ribozyme transgenes, and assessed the influence of AAMP on endothelial cell growth, adhesion, migration, tubule formation and cell aggregation in vitro. Wnt/β catenin inhibitors (IWP2 and FH535) were also used in the functional assays and VE-cadherin and catenin (α, β and γ) expression patterns were examined to explore potential mechanism of action of AAMP.
Cell culture
The human HECV endothelial cell line was purchased from Interlab, (Interlab, Naples, Italy).Cells were routinely cultured in DMEM/Ham's F12 with L-Glutamine medium (Sigma, Dorset, UK), supplemented with antibiotics and 10% foetal calf serum (Sigma, Dorset, UK), and incubated at 37.0°C, 5% CO2 and 95% humidity. Low passage cells (passage 5) were used to generate stocks of control and transfected cells. Cells were frequently revived from initial stocks to ensure experimental consistency.
Generation of AAMP knockdown in HECV cell lines
Anti-human AAMP hammerhead ribozyme transgenes were designed and cloned into a pEF6/V5-His-TOPO plasmid vector (Invitrogen, Paisley, UK) as described previously [16]. Plasmids containing either AAMP specific ribozyme transgenes or empty pEF6 control plasmids were transfected into HECV cells using electroporation and subjected to a selective period (5 μg/ml Blasticidin) before being maintained in maintenance medium (0.5 μg/ml Blasticidin). Transfected HECV cells containing the ribozyme transgene plasmids were routinely tested to confirm knockdown of AAMP expression. Cells were respectively labelled as HECVWT (wild type), HECVAAMPrib (HECV cells containing the ribozyme transgene to AAMP) and HECVpEF6 (control cells containing empty pEF6 plasmid).
RNA extraction and reverse transcription-polymerase chain reaction (RT-PCR)
TRI-reagent (Sigma, Dorset, UK) was used to extract RNA from transfected and control cells. Subsequently, RNA was quantified and standardised to 250 ng before proceeding with reverse transcription (iScript™ cDNA synthesis kit; Bio-Rad, Hemel Hempstead, UK). PCR was then undertaken on the resulting cDNA using the following cycling conditions: 94°C for 40 sec, 55°C for 40 sec, 72°C for 60 sec. PCR was conducted over 34 cycles with an initial 5 min denaturing step (94°C) and a final 10 min extension step (72°C). PCR products were separated electrophoretically on an agarose gel before being stained in ethidium bromide and visualised under UV light. The result was captured and the band density was calculated using Image J software for semiquantitative analysis. Primer sequences are provided in Table 1.
|Gene of interest
|Primer name
|Primer sequence (5'-3')
|Optimal annealing temperature (°C)
|AAMP Ribozyme
|AAMP Ribozyme 1F
|AAMP Ribozyme 1R
Table 1: Primers/Ribozyme transgenes used in the present study
SDS-PAGE and Western blotting
Protein was extracted from transfected and control HECV cells grown in a 75 cm2 flask following detachment with a cell scraper, addition of lysis buffer (HCMF buffer containing 0.5% SDS, 1% Triton X-100, 2 mM CaCl2, 100 μg/ml phenylmethylsulfonyl fluoride, 1 mg/ ml leupeptin, 1 mg/ml aprotinin and, 10 mM sodium orthovanadate) and incubation on a rotor wheel for 1 hour. Samples were subsequently micro-centrifuged at 13,000g to remove insolubles, before being quantified (Bio-Rad DC Protein assay kit; Bio-Rad laboratories, Hemel Hempstead, UK), standardised to 2 mg/ml, diluted in Laemmli 2x concentrate sample buffer (Sigma, Dorset, UK) and boiled for 5 minutes. Samples were electrophoretically separated on a 10% acrylamide gel before being transferred onto a Nitrocellulose membrane (Santa Cruz Biotechnologies, Sant Cruz, CA, USA). The membrane was subsequently probed using the respective primary antibodies, (AAMP and GAPDH; Santa Cruz Biotechnologies, Santa Cruz, CA, USA) at a concentration of 1:70 and specific peroxidase conjugated secondary antibodies (Sigma, Dorset, UK) at a concentration of 1:350, in line with the Snap ID protein system guidelines (Millipore, Watford, UK). Protein bands were documented using a gel documentation system (UVI Tech, Cambridge, UK).
In vitro growth assay
The impact of AAMP knockdown on HECV cells was assessed using an in vitro growth assay. As previously described, three 96-well plates were set up and labelled as Day 1, Day 3, and Day 5 and cells were seeded into each well at a density of 2,000 cells/well [16]. After incubating for 1, 3 and 5 days, media was removed and cells were fixed in 4% formaldehyde (v/v) and stained with 0.5% (w/v) crystal violet. Acetic acid (10%, v/v) was used to extract the crystal violet stain taken up by the cells and cell density was determined by measuring the absorbance of this extracted crystal violet solution at a wavelength of 540 nm using an ELx 800 spectrophotometer (Bio-Tek Instruments Inc., Winooski, VT, USA). The percentage increase in cell density between day 1 and either day 3 or day 5 was then calculated.
In vitro matrigel adhesion assay
In vitro Matrigel adhesion assays were used to compare any differences in cell adhesion abilities of AAMP knockdown cells and control cells as described previously [21]. Briefly, a 96-well plate was pre-coated with 5 μg of Matrigel in 100 μl serum free media per well. After being dried and re-hydrated, 45,000 cells were seeded into each well and incubated for 45 minutes. Following incubation, nonadherent cells were removed by vigorous washing with BSS. Adherent cells were then fixed in 4% formaldehyde (v/v), and stained with 0.5% (w/v) crystal violet. Images of adherent cells were then captured under X20 objective magnification and counted using the Image J software.
Electric cell-substrate impedance sensing (ECIS) based cellular attachment assay and motility assay
The ECIS model system (Applied Biophysics Inc, NJ, US) was used for analysis of cellular attachment and migration as described previously [22]. A 96W1E plate was used in the present study. Following stabilisation of the 96W1E array, the same number of HECVWT, HECVAAMPrib and HECVpEF6 (100,000 cells per well) were seeded in 200 μl of medium per well. Impedance and resistance of the cell layer in the first 3 hours was recorded for the attachment assay. Following attachment of seeded cells to the array and establishment of a confluent monolayer within the well (10 hours after seeding), the monolayer was electrically wounded at 6 V for 30 seconds. Electrical wounding of the monolayer within the array results in a disruption of the monolayer (250 μm diameter wound). Subsequently, the impedance and resistance of the cell layer were immediately recorded over a period of 4 hours for detection of cellular migration as surrounding cells migrate to recolonise the wound.
In vitro matrigel tubule formation assay
Matrigel endothelial cell tubule formation assays were set up, based on a previously described method [23], to assess the involvement of AAMP knockdown on HECV angiogenic potential, in the presence and absence of a series of treatments with IWP2 (2.5 nM, 25 nM, 250 nM) and FH535 (0.1 μM, 1 μM, 10 μM). Briefly, 500 μg of Matrigel in serum-free medium was added to each well of a 96-well plate and placed in an incubator for a minimum of 40 minutes to set. Once the Matrigel had set, 35,000 HECV control or transfected cells were seeded onto the Matrigel layer, and either normal media or media containing inhibitors was added to the appropriate wells. Cells were incubated for 4-5 hours. Following incubation, any tubules that had formed were visualized under the microscope and images captured. Total tubule perimeter/ field in these images was later quantified using Image J software.
In vitro cell aggregation assay
Cell aggregation was assessed using a previously described method [24]. Cells were first briefly washed in BSS and then detached in HCMF buffer (160 mM NaCl, 0.6 mM Na2HPO4, 0.1% (w/v) glucose, and 0.01 M HEPES, pH 7.4) containing 0.01% trypsin and 2 mM CaCl2 for 30 mins at 37°C incubation. Cells were pelleted and washed in HCMF buffer containing CaCl2 before being counted. Each tube was seeded with 1ml of cells at a concentration of 1×105/ml in HCMF buffer containing CaCl2 and a different concentration of Wnt inhibitors were added (IWP2 at 25 nM and FH535 at 1 μM) before being placed on a shaker. Fifty micro litres of medium from each tube was taken and added to 50 μl of 4% Formalin to fix at the following time points: 0 min, 30 min, 60 min and 90 min. Cells were counted and aggregation index was calculated using the formula (N0-Nt)/N0×100%, where N0 is the number of particles at the 0 min time point, and Nt is the number at the chosen time.
Immunofluorescence staining
Immunofluorescence staining was used to assess any alterations in the expression patterns of VE-cadherin, α-catenin, β-catenin and γ-catenin brought about by AAMP knockdown and any subsequent alterations caused by the addition of Wnt inhibitors (IWP2 at 25 nM and FH535 at 1 μM). Briefly, 20,000 cells per well were seeded into a 16 well Chamber slide (LAB-TEK Fisher Scientific UK, Loughborough, Leics, UK) together with the inhibitor (IWP2 at 25 nM or FH535 at 1 μM) or normal medium. Cells were incubated at 37°C, 5% CO2 overnight. Following incubation, the medium was aspirated and cells were fixed in ice-cold ethanol at -20°C for at least 20 minutes before being rehydrated in BSS. Cells were then permeabilized with 0.1% TritonX100 for 2 minutes. Cells were placed in TBS buffer containing 5% horse serum for 20 minutes to block non-specific binding. Following this blocking step, cells were washed with TBS buffer twice. The primary antibody was then added at a concentration of 1:100, made up in TBS buffer containing horse serum for 1 hour. The primary antibody was then removed through three washes in TBS buffer. FITC conjugated secondary antibodies at a concentration of 1:100, made in TBS bufferwith horse serum, were added for 1hour in the dark. Cells were then washed three times with TBS buffer before being mounted with Fluorosave (Calbiochem-Nova Biochem ltd) Nottingham, UK). Images were viewed and captured using a fluorescent microscope. Antibodies used in the above study were as follows: anti-VE-cadherin (sc-101580, anti-mouse; Santa Cruz Biotechnologies), anti-α-catenin (606-259-1550, anti-mouse, Sigma), anti-β-catenin (059K4754, antirabbit, Sigma), anti-γ-catenin (P-8087, anti-mouse, Sigma), FITC antirabbit (F1262, Sigma), FITC anti-mouse (F3008, Sigma).
Statistical analysis
Statistical analysis of data was carried out using Sigma Plot 11 statistical software package. Experimental procedures were independently repeated a minimum of three times and data was analysed using one way ANOVA Holm Sidak post hoc test, ANOVA on Ranks Tukey post hoc test or a two way ANOVA test. Data was considered to be significant at p <0.05.
Ribozyme transgene manipulation of AAMP expression
RT-PCR indicated that transcript expression of AAMP was successfully knocked down in HECVAAMPrib cells in comparison to the expression levels seen in wild-type cells (HECVWT) and in empty plasmid control cells (HECVpEF6) (Figure 1A). This trend was further confirmed through semi quantitative analysis of band density, demonstrating large reductions in expression following normalisation against GAPDH levels (Figure 1B). Additionally, Western blotting was used to assess AAMP protein expression in both the control and ribozyme transgene transfected HECV cell lines. Similar to the trends seen at the transcript level, AAMP protein expression was observed in the control cell line (HECVpEF6) and displayed large reductions in expression levels following transfection with the AAMP ribozyme transgene (HECVAAMPrib) (Figure 1C), which was again confirmed through semi-quantitative band densitometry (Figure 1D).
Figure 1: Ribozyme transgene knockdown of AAMP in HECV endothelial cells
(A/B) RT-PCR and semi-quantitative analysis of GAPDH normalised band densitometry, demonstrating substantial reductions in AAMP transcript levels, in comparison to wild type and plasmid control cells, following transfection with the AAMP ribozyme transgene.
(C/D) Similarly, a reduction in AAMP protein levels was also observed using Western blot analysis and quantification of protein bands. Representative images shown.
Knockdown of AAMP impacts on HECV cell growth
The effect of AAMP on the growth of HECV cells was examined using an in vitro cell growth assay. Knockdown of AAMP appeared to influence the growth rate of the HECV cell line. Close to significant differences were observed between the growth rates of control cells and those containing the ribozyme transgene (HECVpEF6, HECVWT and HECVAAMPrib; p=0.083 over 3 day incubation and p=0.06 over 5 day incubation periods) (Figure 2A).
Loss of endogenous AAMP dramatically reduces HECV cell adhesion
The cell-matrix adhesive capacity of HECV cells over a 45 minute incubation period was examined using an in vitro Matrigel adhesion assay. Loss of endogenous AAMP resulted in a dramatic decrease in matrix-adhesion levels in HECVAAMPrib cells compared to the control HECVWT and HECVpEF6 cells though statistical analysis between the groups did not quite reach significance (p=0.05) (Figure 2B).
Figure 2: Impact of AAMP knockdown on cell function
(A) Knockdown of AAMP brought about a general reduction in HECV cell growth rate over both 3 and 5 day incubation periods, this did not reach significance. (B) Targeting AAMP also reduced the capacity of HECV cells to adhere to an artificial Matrigel basement membrane, though again this did not reach significance. (C) However, HECVAAMPrib cells were less able to attach to the array substrate in an ECIS attachment model system in comparison to HECVpEF6 and HECVWT control cells over the experimental course (p < 0.001) and also demonstrated a significantly reduced migratory ability following electrical wounding of the monolayer using the ECIS system than HECVpEF6 and HECVWT control cells over the experimental course (p < 0.001) (D).
Knockdown of AAMP impacts HECV cell attachment and motility abilities
The effects of AAMP expression on cell attachment and motility were assessed using an ECIS assay. Knockdown of AAMP, similar to the Matrigel adhesion assay, dramatically reduced HECV cell attachment in comparison to control cell lines. Using this methodology, statistical differences in attachment can be seen between the knockdown cells and control cells over the course of the experiment (p<0.001; HECVAAMPrib vs. HECVpEF6, p<0.001; HECVAAMPrib vs. HECVWT, p<0.001) (Figure 2C). Knockdown of AAMP also impacted on the migratory capacity of HECV cells following electric wounding. Significant differences were observed between HECVAAMPrib cells and the wild type and pEF6 plasmid control cells over the course of the experiment (p<0.001; HECVAAMPrib vs. HECVpEF6, p<0.001; HECVAAMPrib vs. HECVWT, p<0.001)(Figure 2D).
Tubule formation of HECV cells was altered by AAMP knockdown
The impact of AAMP knockdown on tubule formation was examined in both the presence and absence of several concentrations of the Wnt/β catenin inhibitors, IWP2 and FH535 inhibitors. Significant differences in tubule formation levels were observed within the treatment groups (p<0.001). HECVpEF6 cells without treatment formed tubule-like structures when seeded onto Matrigel. AAMP knockdown cells were seen to possess only a weak capability to form tubules on Matrigel and significant decreases in tubule formation levels, following quantification of tubule perimeters, were observed between HECVAAMPrib and HECVpEF6 cells (p<0.001) (Figure 3A/B). Treatment with the IWP2 inhibitor, at the range of concentrations tested in this study did not seem to have any substantial effects on the level of tubule formation in either the control HECVpEF6 or HECVAAMPrib cell lines, with no significant differences being observed in comparison to the respective untreated cells. A similar trend was seen with the FH535 inhibitor at the lower concentrations (0.1 μM and 1 μM). However, at the highest tested concentration of 10 μM, the FH535 inhibitor appeared to have differential effects on HECVpEF6 and HECVAAMPrib cells. At this concentration, FH535 could significantly reduce the level of tubule formation in the HECVpEF6 control (p<0.001 vs. untreated HECVpEF6) whilst having no further effect on HECVAAMPrib cells (p=0.642 vs. untreated HECVAAMPrib).
Figure 3: Impact of AAMP knockdown on HECV tubule formation
Targeting AAMP in the HECV cell line significantly reduced the angiogenic tubule formation capacity in comparison to control HECVpEF6 cells (p < 0.001). The addition of the IWP2 and FH535 Wnt/β catenin signalling inhibitors had no significant effects on either control or transfected cells except at the highest 10μM concentration of FH535 where, at this concentration, FH535 brought about a significant decrease in HECVpEF6 tubule formation in comparison to untreated HECVpEF6 cells (p < 0.001).
(A) Representative images of tubule formation assays for each group.
(B) Mean quantified total tubule perimeter of all tubules within the microscopic field
AAMP knockdown had no impact on cell aggregation
An aggregation assay was performed to assess the involvement of AAMP in cell-cell adhesion. No significant differences in cell-cell aggregation, over the experimental course, were observed between HECVWT, HECVpEF6 and HECVAAMPrib cells (p=0.63) (Figure 4A). In addition, neither IWP2 (25nM) or FH535 (1 μM) treatment was seen to have any impact on cell aggregation in either the control or the ribozyme transfected cell line over the course of the experiment (p=0.054 within the HECVpEF6 treatment groups, Figure 4B and p=0.565 within the HECVAAMPrib treatment groups, Figure 4C).
Figure 4: AAMP knockdown does not alter HECV cell aggregation
Aggregation assays indicate that knockdown of AAMP did not significantly alter the rates of cell aggregation and no significant differences were observed between HECVpEF6, HECVWT and HECVAAMPrib cells (A). Additionally, treatment of HECVpEF6 cells (B) or HECVAAMPrib cells (C) with either of the IWP2 or FH535 inhibitors did not have any significant impact on cell aggregation rates of either cell line.
VE-Cadherin expression is reduced following AAMP knockdown
Immunoflourescent staining was used to examine the staining profile and localisation of α, β and γ catenin, together with VE-cadherin in control and ribozyme transfected HECV cells (Figure 5). Expression patterns and staining intensities of the catenins were similar in both the HECVpEF6 and HECVAAMPrib cells and few differences in staining intensities were observed in the images and following quantification of staining intensity (Figure 5A-F). The results of the immunofluorescence staining indicated that AAMP knockdown cells had a lower level of VE-cadherin expression compared with its pEF6 control which was further confirmed following quantification of staining intensity (Figure 5G and H). A statistically significant difference in VE-cadherin staining was discovered within the knockdown and treatment groups (p=0.004). Levels of VE-cadherin expression were significantly reduced in HECVAAMPrib cells compared to HECVpEF6 cells (p<0.05). Similarly, treatment of HECVpEF6 cells with FH535 resulted in a similar reduction in VE-cadherin, and differences in VE-cadherin staining patterns could be observed between HECVpEF6 cells treated with or without FH535 (p<0.05). However, treatment with IWP2 did not change VE-cadherin staining intensity in either HECVAAMPrib or HECVpEF6 cells compared to the respective untreated control (p>0.05).
Figure 5: Immunoflourescent staining demonstrating the relationship between AAMP and cadherin/catenins Staining profiles (A,C, E,G) and quantified staining intensities (B,D,F,H) for α, β, γ catenin and VE-cadherin in HECVpEF6 and HECVAAMPrib in the presence and absence of the IWP2 and FH535 inhibitors. Similar staining profiles and intensities were observed for α, β and γ catenin in both HECVpEF6 and HECVAAMPrib cells. HECVAAMPrib cells displayed lower levels of VE-cadherin staining that HECVpEF6 cells and quantification of intensities showed a significantly decreased level of VE-cadherin in HECVAAMPrib cells compared to HECVpEF6 control cells (P < 0.05). The staining profile of VE-cadherin in HECVpEF6 cells was also affected following treatment with FH535 inhibitor, which similarly brought about a significant decrease in staining intensity compared to untreated HECVpEF6 cells (p < 0.05). Arrows indicate example of staining.
The AAMP gene was originally identified during research into motility-associated proteins and was found to be expressed extensively in all the tested types of endothelial cells and other different cell lines and tissues [3,10-16]. Previous studies show that loss of AAMP result in a reduction in endothelial cell adhesion, migration and tube formation. In the present study, we verified the expression of AAMP in the human HECV endothelial cell line and subsequently used a ribozyme transgene system to knockdown AAMP expression in this cell line. Our results indicate that knockdown of AAMP, whilst not significant, brings about a general reduction in proliferation rates of HECV endothelial cells over 3 and 5 day growth periods. Migration and tubule formation are significantly inhibited when AAMP expression is reduced. Similarly, cell-matrix adhesion rates are inhibited following AAMP knockdown, with significant reductions observed using the ECIS model and substantial reductions observed using the Matrigel adhesion assay, though these did not quite reach significance (p=0.05). These results are in line with previous research reports [3,13,15,20]. Our study further explored the relationship between AAMP and Wnt/β catenin signalling using the FH535 (suppresses both Wnt/β-catenin and Peroxisome Proliferator-Activated Receptor (PPAR) signalling) and IWP2 (an inhibitor of Wnt processing and secretion) compounds. In the tubule formation assays, knockdown of AAMP significantly reduced the formation of tubule like structures compared to the control HECVpEF6 cells. Further treatment with the IWP2 inhibitor had little additional effects on either HECVpEF6 or HECVAAMPrib cells. Treatment of these cells with FH535 did bring about differential effects between the two lines, where treatment with the highest (10 μM) concentration could significantly reduce HECVpEF6 tubule formation (vs. untreated HECVpEF6 cells) but had no further effect on AAMP knockout HECV cells. This result may suggest some relationship between AAMP and these pathways, however this trend was only observed at the higher inhibitor concentration and further work will be required to fully clarify this relationship.
Perhaps the most important observation in the present study is the relationship between the expression of AAMP and VE-cadherin seen using immunofluorescence staining. Reduced expression of VE-cadherin was observed in the HECVAAMPrib cells compared to the HECVpEF6 cells. VE-cadherin plays a vital role in endothelial cell binding. Endothelial cell homotypic adhesion is mediated by two types of adhesive structures, tight junctions and adherens junctions, where VE-cadherin, an endothelial-specific member belonging to the cadherin family, plays a key role in mediating the adhesion through adherens junctions [25]. VE-cadherin is linked to a large number of intracellular partners which mediate its anchorage to the actin cytoskeleton from one side and from the other transfer intracellular signals. This potential relationship, between AAMP and VE-cadherin expression, may in part be responsible for the change in endothelial adhesion and motility caused by the knockdown of AAMP. Previous studies have also reported that VE-cadherin may form a multi-protein complex with Vascular Endothelial Growth Factor (VEGF) receptor 2 (VEGFR2) and limit its internalization and signalling activity [26]. Similarly, VE-cadherin can interact with transforming growth factor beta (TGFβ) receptor complex, platelet derived growth factor β receptor or Fibroblast Growth Factor (FGF) receptor 1 and others [27-29]. These interactions contribute to the control of endothelial proliferative signals and induction of vascular stability. This may again partly explain the impact on HECV cell growth following AAMP knockdown in our study. VE-cadherin directly binds inside the cells to β-catenin, an intracellular protein which is a crucial downstream element of the so-called canonical Wnt pathway, but it is also firmly bound to VEcadherin [30]. β-catenin transcriptional activity is usually associated to cell proliferation. Our immunofluorescence staining data suggests that VE-cadherin was not only reduced with AAMP knockdown, but also by FH535, however, the mechanism of how FH535 impacts on VE-cadherin needs further investigation. It is again interesting to note that at the higher concentration, FH535 inhibition was able to reduce tubule formation levels of HECVpEF6 control cells to a similar level to HECVAAMPrib cells and again may suggest some relationship. Whether AAMP has some functional relationship with Wnt/β-catenin requires further investigation.
Consistent with recent reports, AAMP knockdown reduces HECV endothelial cell adhesion, migration and tubule formation. In our recent study, in vitro growth ability may also be linked with AAMP. Our study also demonstrated that reduced AAMP is accompanied with low expression of VE-cadherin, and may also have some link with Wnt/β-catenin. The mechanism of how AAMP plays a role in these signalling pathways needs further research. The results presented here implicate a role for AAMP in a number of traits essential for the process of tumour angiogenesis and potentially implicate a relationship with Wnt/β-catenin signalling. Further elucidation of this relationship and mechanism may highlight potential anti-angiogenic therapeutic strategies.
The authors are grateful to Cancer Research Wales (CRW) and the Albert Hung Foundation for supporting this work. Dr Yin is a recipient of the Cardiff University China Medical Scholarship.


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In Ireland in the 1840's, people ate from 7 to 15 pounds of potatoes apiece every day. In 1845, a blight attacked the potatoes, turning them black and rotten, and causing a famine in Ireland in which over one million people died from starvation and disease. More than two million people emigrated because they could no longer make a living in Ireland.
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Reef Bay Trail is a 4.4 mile lightly trafficked out and back trail located near Saint John, Saint John that features a great forest setting. The trail is rated as difficult and primarily used for hiking and birding.
historic site
no dogs
Trail goes through two forests and remains of deserted sugar mill. It ends at the beach. The trail is mostly down hill.
Amazing hike!! Stay hydrated!! It's actually almost 6 miles from start to finish, longer if you take other trails off of this trail, but stay on the trail leading to the factory, you'll be so glad you did!
great trail with lots of wildlife, flowers and ruins. many trail options off this one to enhance your hike (petroglyphs, lameshur, lesparance). it's mostly easy, downhill one way, the uphill back is only difficult in the last .5 miles, make lots of stops, beach is naturally beautiful but not for swimming
I presume ranked hard because of the hike back...quite a climb. So many hermit crabs as you get closer to water. This was our most challenging hike on St John's but not the favorite.
Beautiful trail. A lot to see. The way down was really steep in the beginning so you can imagine what the way up was like. Bring lots of water to hydrate. Would definitely rate it moderate-hard.
This was a very easy trail down to the beach and back. You can hike over the rocks and go to a private beach and snorkel. The trail ends at a Rum Still and it is pretty cool. There are many little trails off of Reef bay to run off to. You can most definitely make a nice day of all of these little ones. Trails is maintained and the big huge things that look like massive bee hives are actually termites. You will see them all over and they are harmless.
Groomed trail. (We actually started near Maho Bay hiking up the Maria Hope Trail (rugged, trailhead not marked) then crossed Centerline Road to the Reef Bay Trailhead.) Side trip to petroglyphs a must. Right before you reach the ruins there is an amazing population of large land crabs. Sizeable bat population in one of the ruins. The actual beach was crawling with ants so we didn't stay long. We took the L'esperance trail out which is a tad longer, but less steep. Would rate both Reef Bay Trail and L'Esperance Trail at moderate.
Great hike. Well maintained trail with lots to look at along the way. Beautiful beach at the bottom to cool you off for the return trip. We loved it!
Difficult Hike, bring plenty of water.


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Would You Hit It?
June 12, 2010 / Posted by: Michael K
Believe it or not, this is not a picture of Tommy Girl trolling The Ramble for a piece to feed to his Scientolohole. This is Tommy on the set of a commercial for ESPN showing us that when he waxes the dead Thetans off his body his chest follicles come off too. And to answer my own question, XENU NO! If you hit that, you would wake up in a Scientology sauna barfing the glib out of your system. That might sound sexy on paper, but you know it's not.
Here's more pictures of Tommy and Cameron Diaz on the set yesterday. In case you need a palate cleanser, I threw in some pictures of Simon Baker summoning the panty pudding in Monte Carlo. Oh, how I just want to slide around one of his golden locks.
Our commenting rules are pretty simple: If you make any overly offensive comment (racist, bigoted, etc..) or go way off topic when not in an Open Post, your comments will be deleted and you will be banned. If you see an offensive or spammy comment you think should be deleted, flag it for the mods and they'll be forever grateful and give you their first born (although, you probably don't want that).


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82nd Annual Diadeloso Includes Old Traditions, New AdditionsApril 8, 2014
Follow us on Twitter: @BaylorUMedia
Media contact: Lori Fogleman, (254) 710-6275
WACO, Texas (April 8, 2014) – It's a little hard to explain just what Diadeloso is to someone who isn't an Oso, but it's more than a day of frivolity and shenanigans on a random Thursday in spring.
"This cherished tradition, which is over 80 years old, provides our community a momentary break from classroom and other daily routines to enjoy entertaining acts and performances, engage in friendly competitions and take pleasure in being outdoors together on the beautiful Baylor campus," said Kevin P. Jackson, Ph.D., vice president for student life at Baylor University, in an email to Baylor students.
The 2014 Diadeloso, or "Day of the Bear," will be held throughout the day on Thursday, April 10, on Fountain Mall, 1360 S. Fifth St.
The first Diadeloso took place in 1932 with the same purpose it has today: to give students a break from classes with food, games, entertainment and beautiful spring weather. It was reorganized by the Baylor Chamber of Commerce in 1935 and has undergone several name changes over the years, including "All University Day," "Physical Fitness Day" and "May Day." Finally, in 1966 students voted to name the annual occasion "Diadeloso" or "Dia" for short.
Events and activities on Fountain Mall are open to the Baylor and Waco communities. Student organizations will host free activity booths; dogs will have the opportunity to participate in the Dia Dog Show, judged by the dog daycare center Dogtopia; children can participate in kid-friendly activities from 11 a.m. to 2 p.m.; and individuals and teams can compete in athletic tournaments throughout the day.
Unique to the 82nd annual Diadeloso is the Second Annual Fun Run. Participants will be covered in colored foam as they make their way through a three-station course that ends with a colorful pit of foam.
"We were actually testing it out this morning and having quite a bit of fun with it," said Kelcie Evans, Diadeloso chair for the Chamber of Commerce. "Most people don't want to come up at 10 in the morning and run. This is a way we can make it a little more interesting."
Registration for the Fun Run ends tonight, Tuesday April 8, at 11:59 p.m., or once the run reaches 500 entries.
Several student organizations and Uproar artists will perform on stage throughout the day, with a culminating performance at 8 p.m. by the band NEEDTOBREATHE, who has been touring the country and have performed at music festivals Bonaroo and Austin City Limits and throughout Europe. Click here for a complete stage schedule.
No Dia would be complete without exotic animals right here on campus. Past Diadelosos have seen camels and capuchin monkeys; this year, Fountain Mall will play host to a micro mini pig, a kinkajou, a marmoset, a hedgehog and a giant turtle.
For more details and a complete schedule of Diadeloso events, visit www.baylor.edu/diadeloso/.
by Rachel Miller, student newswriter, (254) 710-6805
Baylor University is a private Christian university and a nationally ranked research institution, characterized as having "high research activity" by the Carnegie Foundation for the Advancement of Teaching. The university provides a vibrant campus community for approximately 15,000 students by blending interdisciplinary research with an international reputation for educational excellence and a faculty commitment to teaching and scholarship. Chartered in 1845 by the Republic of Texas through the efforts of Baptist pioneers, Baylor is the oldest continually operating university in Texas. Located in Waco, Baylor welcomes students from all 50 states and more than 80 countries to study a broad range of degrees among its 11 nationally recognized academic divisions. Baylor sponsors 19 varsity athletic teams and is a founding member of the Big 12 Conference.
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Frequently Asked Question
What are your store hours?
How much does it cost to pick you own fruit?
How many pieces of fruit are in a tray?
What are your shipping costs?
What shipping carrier do we use?
What is your return policy?
12 to 15 Oranges
9 to 12 Grapefruit
20 to 25 Tangerines
Each tray weighs approximately 10 lbs.
Ship up to four trays per box!
1/2 Bushel = 5 to 6 Pomelos
1 Bushel= 10 to 12 Pomelos
Haines City Location - 6000 Polk City Road, Haines City, FL 33844
Monday - Saturday, Closed Sunday
Monday - Saturday, Closed Sunday
Lakeland Downtown Farmers Market - 200 N Kentucky Ave, Lakeland, FL 33801
*Saturdays Only
100% Satisfaction Guarantee
We take great pride in shipping Florida's Freshest Citrus right to your doorstep and want our customers to be satisfied. If there is any problem with one of our shipments we like to be the first to know, so we can fix it promptly. Contact us toll free at 1-800-49-FRESH. Thank you!


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Mexican indie band The Plastics Revolution created a stop-motion music video for their single "Invasión" using 1,905 Instagram photos that were taken on an iPhone.
"Every single frame of this music video is an actual picture that we ran through Instagram. We never shot any video. We only shot still photography," said the band on Vimeo.
No more words. Just hit play and watch the video below!
Source: Mashable


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Asphalt walking paths & trails
The selection of surface material for trails and paths is primarily based on anticipated type and intensity of trail use, as well as terrain, climate, design life, maintenance, cost, and availability. Soft surface materials are low cost, but require substantial maintenance and are not suitable for many of the recreational activities today's trails and paths are used for. Hard surface materials, specifically concrete and asphalt, provide years of service with low maintenance.
The following are the key elements to consider in building quality asphalt trails:
Bike paths and walking trails in Northeast Ohio should be constructed to match the existing topography as closely as possible however, longitudinal slopes should not exceed five percent and a cross slope of two percent is desirable to provide adequate drainage away from the pavement surface. Proper drainage is one of the most important factors affecting pavement performance.
Proper drainage entails efficient removal of excess water from the trail. Surface water runoff should be handled using swales, ditches and sheet flow. Catch basins, drain inlets, culverts and underground piping may also be necessary. These structures should be located off of the pavement path.


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Virginia Highlands Community CollegeHome Menu
Employee Directory
Hamed, Kevin
- Departments:Biology
- Divisions:Arts & Sciences
- Title:Professor of Biology
- Phone:(276) 739-2431
- Email:email@example.com
- Office LocationOTC 220
Return to full list >>
URL: http://vhcc2.vhcc.edu/khamed
For summer appointments, please email the instructor directly.
Please contact via VHCC email at firstname.lastname@example.com for an appointment.
For immediate assistance, please contact the Arts & Sciences Division at 739-2433.


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'You must make women count as much as men; you must have an equal standard of morals; and the only way to enforce that is through giving women political power so that you can get that equal moral standard registered in the laws of the country' - Emmeline Pankhurst, 1913.
Eighty years later…
'You grow up with your father holding you down and covering your mouth so another man can make a horrible searing pain between your legs' - Catharine MacKinnon, 1993.
What's gone wrong with feminism?
Feminism used to be about equality, not female superiority
Many modern-day feminists engage in man-bashing rather than making dignified demands for equality, as the feminists of the early twentieth century did. So Robin Morgan unashamedly admits, 'I feel that man-hating is an honourable and viable political act'. The view many modern feminists seek to spread, in the words of Marilyn French, is that 'all men are rapists and that's all they are'. A Feminist Dictionary goes so far as to define 'male' as 'a degeneration and deformity of the female'. This is an effort to make men seem inferior, not to elevate the position of women. It starkly contrasts with the goal of feminists of old, which was to make women be viewed as equal, not superior, to men.
Feminism used to be about freedom, not censorship
Recently, feminists have supported moralistic wars on porn, the Sun's Page 3, and misogynistic Twitter messages. They are essentially advocating more government intervention into our private lives and our freedom of speech. Yet 43 years ago, Germaine Greer wrote in the The Female Eunuch that 'freedom is fragile and must be protected. To sacrifice it, even as a temporary measure, is to betray it'. Whereas feminism was once concerned with increasing the freedoms of women, and by extension all inhabitants of society, now it tends towards demanding the reduction of freedoms.


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Angler Waylon Bush, along with Captain Douglas Ebanks on 'Blue Moon', reeled in a 170.4 pound yellow fin tuna. This was the largest fish caught on the 50-pound test class, beating the previous largest which was 146-pounds, this was also the 3rd largest fish of its kind (tuna) caught in the history of Cayman.
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Bush reels in record 170.4 lb tuna
February 10, 2017
1 Min Read
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About the author
Jordan Armenise
From Hamilton, Ontario, Canada, Jordan Armenise began his sports broadcasting journey with the Hamilton Tiger-Cats of the Canadian Football League. It was in this role where he was able to merge his best attributes: sports knowledge and personality. While with the Ticats, Jordan provided live analysis and interviews while developing a digital portfolio of one-on-one interviews, episodic sports comedy and full length sports & news features. He has also worked with CBC Sports for Hockey Night in Canada, the 2014 Sochi Olympics and Special Olympics as well as roles with Cineflix and Cream Productions as an Assistant Director.
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text: SOURCES: Byron Cryer, MD, spokesman, American Gastroenterological Association; associate professor of medicine, University of Texas Southwestern Medical Center, Dallas. Nieca Goldberg, MD, spokeswoman for the American Heart Association; chief of women's cardiac care, Lennox Hill Hospital, New York; author, Women Are Not Small Men: Lifesaving Strategies For Preventing And Healing Heart Disease In Women . John Klippel, MD, president and CEO, Arthritis Foundation, Atlanta. Scott Zashin, clinical assistant professor, University of Texas Southwestern Medical Center; author of Arthritis Without Pain . American College of Rheumatology web site. Arthritis Foundation web site. American Heart Association web site. American College of Gastroenterology web site. American Gastroenterological Association web site. American Academy of Family Physicians web site. American Academy of Allergy, Asthma, and Immunology web site.


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Are all candles the same?
Old Factory Candle Review & Giveaway
By Jennifer Schultz for the MommyBrief.com
Disclosure: This blog may received products mentioned for free and/or accept compensation Regardless, all reviews and opinions are my own. I am disclosing this in accordance with the Federal Trade Commissions 16 CFR, Part 255 Guides.
I was so excited to be chosen to review the Old Factory Candles, because I L-O-V-E candles. But, not all candles are the same – some don't smell anything like their scents when they are burning, some burn so unevenly that the wick gets covered and you have to throw the candle away half way through, and others burn so quickly that you wonder why you paid top dollar for a candle that lasted 2 hours.
I am happy to say that my experience with the Old Factory Candles was wonderful. First, I had selected the "Happy Holidays" assortment that included the scents: Christmas Tree, Candy Cane & Gingerbread. I always love the smells of Christmas through the house so I can hold on to the season a little longer. Candy Cane was my favorite of the three candles as it left the house smelling so sweet! When I burned the Gingerbread candle everyone was asking what I was baking, and I would secretly giggle to myself knowing it was a candle. The Christmas Tree scent was a little too heavy for me, but, did smell like the real thing. If you want to keep Christmas in your home a little longer this would be the candle set for you!
About Old Factory Candles
Old Factory candles are hand-poured in the USA with natural soy wax, self-trimming cotton candle wicks, and premium fragrance oils. The Old Factory Candle Gift Sets each contain 3 different scented candles. Each candle burns clean and even for 20 hours. The subtle jar design makes it the perfect complement for any decor, and any room of the house – kitchen, bedroom, bathroom, family room, great room, and basement. Each gift set is themed around a group of related fragrances. So everyone is sure to find something perfect for every occasion, every room in the house, and all the special someones in your life.
Here is your chance to win your own set of Old Factory Candles! ($25 value)
Giveaway ends on 1/10/15 at 12am.
1. One winner will be selected at random from all entries received.
2. By entering this giveaway, you agree to all terms and conditions noted below and on the entry form.
3. How to enter – use the form to like us on Facebook and get an extra entry for following us on Twitter.


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Watch how to make this recipe.
In a mixing bowl, whisk together the egg whites, sesame oil, and 1 tablespoon of the cornstarch until foamy; season with salt and pepper. Add the beef and chives, toss to coat in the marinade and stick it in the refrigerator for 30 minutes.
Pour the peanut oil in a large skillet or wok and place over high heat. When the oil is smoking hot, add 1/2 of the beef. (If you fry all the meat at once the oil temperature will drop and the beef will stew instead of crisp.) Fry for 2 minutes then remove to a paper towel to drain; repeat with the remaining beef. Very carefully drain all but 2 tablespoons of the hot oil into a safe container.
Stir-fry the ginger, garlic, green onions, and chile in the remaining oil until they perfume. Add the green beans; season with salt and pepper. Stir-fry the beans for 2 minutes until crisp-tender; remove from the pan. Mix the chicken broth with the remaining tablespoon of cornstarch to make a slurry and add it to the pan. Add the soy sauce, vinegar, sugar, peanut butter, and orange peel. Simmer until the sauce is thick, about 5 minutes. Return the beef and green beans to the pan, tossing to coat in the sauce. Garnish with chopped peanuts and serve with rice.
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|Fashion Grilles Limited
|4000 Weston Road, Weston, ON
|Indal Products Limited
|350 Clayson Road, Weston, ON
|Mcknight Window Industries Limited
|4000 Weston Road, Weston, ON
|Easy Up Shelving (canada) Inc.
|30 Penn Drive, Weston, ON M9L2A9
|Roamin' Roses Mktg. Ltd.
|301 Dixon Road Suite 1410, Weston, ON
|Carlson Marketing Group Ltd.
|100 Marmora Street, Weston, ON M9M2X9
|Meyers Studios Limited
|165 La Rose Avenue Apt 107, Weston, ON M9P3S9
|Donlee Manufacturing Industries Limited
|1945 Wilson Avenue, Weston, ON M9M1A9
|Brand Leather & Hide Canada Ltd.
|222 Dixon Road Suite 103, Weston, ON M5P3S5
|Solignum Inc.
|200 Norelco Drive, Weston, ON M9L 1S4
|Innovation Canada Inc.
|2 Woodpark Road, Weston, ON M9P 1M1
|Golden Star Trade Development Inc.
|1137 Barmac Drive, Weston, ON M9L1X4
|Donlee Manufacturing Industries Limited
|430 Signet Drive, Weston, ON M9L2T6
|Canada Electric (1977) Limited
|59 Penn Drive, Weston, ON
|Racine Hydraulics (canada) Limited
|2424 Finch Avenue West, Weston, ON
|El-mot International Ltd.
|390 Dixon Road Suite 303, Weston, ON
|Amara Machines Ltd.
|115 Weston St, Weston, ON
|Truline Plastics (canada) Ltd.
|113 Toryork Rd, Weston, ON
|Blackwood Hodge Quebec Sales Ltd.
|10 Suntract Rd, Weston, QC M9N3N5
|Blok-lok, Limited
|30 Millwick Drive, Weston, ON M9L1Y3
|Lormac Warehousing Limited
|27 Holgate St, Weston, ON M9R1C6
|Canadian Council On Electrical Maintenance Inc.
|42 Gaydon Avenue, Weston, ON M9M1G7
|Canadian Promotions for Newspapers Ltd.
|2010 Islington Avenue Suite 1005, Weston, ON M9P3S8
|Chemplas Limited
|10 Hartsdale Dr, Weston, ON
|Civic Aersols Limited
|22 Strathdee Dr, Weston, ON
|137376 Canada Inc.
|135 Millwick Dr., Weston, ON M9L1Y7
|A.p.v. - Crepaco of Canada Ltd.
|1250 Ormont Rd., Weston, ON M9L2V4
|Douglas Campbell Limited
|265 Dixon Road Suite 305, Weston, ON M9R1R7
|Ajax Precision Manufacturing Limited
|37 Penn Drive, Weston, ON M9L2A6
|Duomat Machinery Ltd.
|10 Suntract Rd, Weston, ON M9N3N5
|Indal Limited
|4000 Weston Road, Weston, ON M9L2W8
|Elken Ltd.
|120 Widdicombe Place Apt. 908, Weston, QC
|Euclid Equipment Rentals Ltd.
|10 Suntract Rd, Weston, ON
|Ferry-morse Seed Company Canada Limited
|44 Clayson Rd., Weston, ON M9M2G9
|Fleer Ltd. -
|77 Rivalda Road, Weston, ON M9M2M6
|Elegant Lighting Ltd.
|5707 Steeles Avenue West, Weston, ON
|The Holes-webway Co. of Canada Ltd.
|116 Rivalda Rd, Weston, ON
|Jellicoe Resource Associates Ltd.
|390 Dixon Road Suite 1812, Weston, ON
|Byron King Inc.
|94 Millwick Drive, Weston, ON M9L1Y3
|Kinney Canada Inc.
|100 Mainshep Road, Weston, ON M9M1L5
|Faraday (canada) Limited
|339 Ormont Drive, Weston, ON M9L1N8
|Macmillan Bloedel Building Materials Limited
|50 Oak St, Weston, ON M9N1S1
|Meyers Studios (canada) Limited
|165 La Rose Avenue Apt 107, Weston, ON M9P3S9
|Memorial Society Association of Canada
|Station A Box 96, Weston, ON M9N3M6
|Champion Parts Rebuilders (canada) Ltd.
|224 Milvan Dr., Weston, ON M9L2A5
|97885 Canada Limited
|2424 Finch Avenue West Unit 1, Weston, ON
|Pylon Electronics Inc.
|50 Bartor Road, Weston, ON M9M2G5
|Ram Partitions Limited
|4000 Weston Road, Weston, ON
|Cesare Chrome Furniture Mfg. Ltd.
|126 Millwick Drive, Weston, ON M9L1Y6
|Reuland Productions Limited
|4 Landigo Dr, Weston, ON
|Richardson-vicks Limited
|2 Norelco Dr, Weston, ON M9L1R9
|Riddell Products (canada) Limited
|22 Elrose Ave, Weston, ON
|Orion Acceptance Corporation Ltd.
|Station "a" P.o.box 1004, Weston, ON M9N3N5
|Standard Auto Glass Western (alta) Limited
|116 Milvan Drive, Weston, AB M9L1Z9
|Tilt-n-load Service of Canada Limited
|50 Church Street, Weston, ON M9N1M8
|Tobin Tractors (1957) Limited
|10 Suntract Road, Weston, ON M9W3N9
|Watersearch Limited
|3650 Weston Road, Weston, ON
|Werby Industries (canada) Ltd.
|368 Signet Drive, Weston, ON M9L1V2
|Ccc Solvents Ltd.
|79 Fenmar Drive, Weston, ON M9L1M3
|Etc Energy Technology Consulting Canada Inc.
|19 Muirdale Avenue, Weston, ON M9R2V3
|Canadian White Water Association
|12 Wendell Avenue, Weston, ON M9N3K5
|Canadian Paralegal Association
|50 Lockerbie Ave., Weston, ON M9N 3A2
|R. Laidlaw Lumber Co. Limited
|50 Oak Street, Weston, ON
|West York Construction International (qatar) Limited
|270 Toryork Drive, Weston, ON M9L1Y1
|Spaceflex Inc.
|136 Rivalda Road, Weston, ON
|Etraco Marketing Ltd.
|Station A P.o.box 185, Weston, ON M9N3M7
|Phoenix Paper Products Limited
|21 Fenmar Dr., Weston, ON M9L1L7
|The Bartenders' Association of Canada
|330 Dixon Road Apt. 1907, Weston, ON M9R1S9
|Lewis Shoes Canada Limited -
|100 Mainshep Road, Weston, ON M9M1L5
|90922 Canada Limited
|430 Signet Drive, Weston, ON M9L2T6
|104793 Canada Inc.
|390 Dixon Road Suite 1612, Weston, ON M9R1T4
|105064 Canada Inc.
|200 Norelco Drive, Weston, ON M9L 1S4
|Vantage Sports (ont) Inc.
|3650 Weston Road Unit 17, Weston, ON M9L1W2
|Zuron Marketing Systems Inc.
|301 Weston Road Suite 101-102, Weston, ON
|Spaco Marketing Services Ltd.
|53 Widdicombe Hill Blvd Suite 1805, Weston, ON M9R1Y3
|103918 Canada Ltd.
|445 Fenmar Drive, Weston, ON M9L2R6
|Landex Commercial Interiors Inc.
|150 Rivalda Road Unit 1, Weston, ON
|Tvm Video Distributors Inc.
|430 Signet Drive Suite C, Weston, ON M9L1V5
|Aggregate Machinery Ltd.
|10 Suntract Road, Weston, ON M9N3N9
|Aggregate Machinery Manufacturing Ltd.
|10 Suntract Road, Weston, ON M9N3N9
|Calvin Seminary of Canada
|2157a Weston Road, Weston, ON M9N 1X8
|Allied (eastern) Woodworking Machinery Ltd.
|453 Signet Drive, Weston, ON M9L1V5
|Canadian Organization of Porcelain Art
|960 Scarlett Road, Weston, ON M9P2V7
|Berenson Inc.
|4101 Weston Road, Weston, ON M9L1W6
|Turner and Burns Limited
|4101 Weston Road, Weston, ON M9L1W6
|Candad Export Inc.
|4101 Weston Road Unit 12, Weston, ON M9L1W6
|6268609 Canada Inc.
|5125 Steeles Avenue West, Weston, ON M9L 1R5
|6268668 Canada Inc.
|5125 Steeles Avenue West, Weston, ON M9L 1R5
|6269150 Canada Inc.
|5125 Steeles Avenue West, Weston, ON M9L 1R5
|6299385 Canada Incorporated
|33 King St Apt 311, Weston, ON M9N 3R7
|Hi Speed Truck Transportation Service Inc.
|81 Navenby Cr, Weston, ON M9L 1B3
|Modern Album of Canada, Ltd.
|2450 Finch Avenue West Unit 7, Weston, ON M9M2E9
|Canadian Humanitarian Cultural Foundation
|219 Toryork Drive, Weston, ON M9L1Y2
|Talk of The Town Hair Centre Inc.
|741 Fenmar Drive, Weston, ON M9L1C8
|Pelsue Inc.
|127 Milvan Drive, Weston, ON M9L1Z8
|Bronco Gym Equipment Incorporated
|7 Arkley Cres., Weston, ON M9R3S2
|Ireton International Barter Inc.
|34 Newstead Road, Weston, ON M9P3G3
|Pandit Jaitly Astrologer Inc.
|390 Dixon Road Apt. 301, Weston, ON M9R1T4
|Sih Construction Ltd.
|25 Duncanwoods Drive Apt. 504, Weston, ON M9L2C5
|Jaitly's Self-consciousness Society Inc.
|390 Dixon Road Apt. 301, Weston, ON M9R1T4


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We are in the midst of a paradigm shift where cities and municipalities are looking to strengthen local energy security [...] the Community Energy Guide for Canada will help city builders champion this important transformation in their communities.
Boston, MA (PRWEB) November 14, 2013
The International District Energy Association (IDEA) announced the release of its newest publication for the Canadian market, Community Energy: Planning, Development and Delivery – Strategies for Thermal Networks, which aims to support mayors, planners, community leaders, and private developers who are interested in planning, designing and deploying district energy systems for more efficient and resilient infrastructure in their cities, communities and towns.
The new guidebook is designed to provide Canadian city-builders with experience-based insight to the energy project development process to help them in making informed decisions regarding the analysis, planning, development and delivery of district energy systems. The work builds on previously released UK and US Community Energy Guides, this time with a Canadian focus on provincial and municipal policies and market conditions.
IDEA, a 104-year old non-profit industry association based outside of Boston, MA, engaged UK-based district energy specialist Michael King, author of the UK and US guidebooks by the same name, to collaborate with IDEA's own Brad Bradford. King is a principal with District Energy Development LTD in the UK and a board member of Aberdeen Heat & Power of Scotland, a recent IEA District Energy Climate Award Winner. Bradford is IDEA's Community Energy Planner and holds a Masters of Planning degree from the University of Waterloo. Together with industry leaders, King and Bradford updated the guide for the Canadian market to reflect provincial policies, the specific nuances and drivers of system deployment, and leverage the emerging momentum around the community energy planning process and integration of community-based energy systems within broader planning frameworks.
The Canadian edition of the guidebook is designed to equip key decision makers with the knowledge and understanding to make informed decisions on energy, environmental and economic matters relevant to local energy implementation. IDEA will disseminate this best practice approach across Canada in conjunction with partner organizations and agencies to better inform the marketplace, build greater awareness of the advantages of district energy, CHP and waste energy recovery, and engage public/private partnerships in industry growth.
Brad Bradford, coauthor of the guidebook states, "We are in the midst of a paradigm shift where cities and municipalities are looking to strengthen local energy security and reduce reliance on a centralized electricity generation model where power supplies are distant and remote from the customer base. Canadian urban centers are experiencing tremendous growth and district energy systems are being deployed to ensure the delivery of efficient, robust and resilient heating, cooling, and power. The Community Energy Guide for Canada will help city builders champion this important transformation in their communities."
On Monday, November 11, IDEA hosted a workshop titled "Community Energy: Planning, Development and Delivery" in conjunction with the QUEST annual conference in Markham, Ontario. This one day workshop featured: Pernille M. Overbye, Market Manager at Ramboll Group; Richard Damecour, President at FVB Energy; Robert Thornton, President & CEO of IDEA; and coauthors of the guide, Michael King and Brad Bradford. Panelists included: Bruce Ander, President & CEO of Markham District Energy; Dennis Fotinos, President & CEO of Enwave Energy Corporation; and Pat Bohan, Director of Business Development for District Energy at ENMAX. The workshop discussed best practices and reviewed a step-by-step approach to develop a successful community energy plan and deploy district energy. You can view the workshop program here for more detailed information.
IDEA gratefully acknowledges the sponsoring companies who provided financial and technical support and editorial contributions to the Canadian Guide: Ramboll Group; FVB Energy Inc.; Dalkia Canada; DIALOG; ENMAX; Golder Associates; Kerr Wood Leidal; Markham District Energy Inc.; QUEST; Rovanco Piping Systems; Solar Turbines; and Urecon.
A free on-line version of the guide is available here.
The International District Energy Association (IDEA) is a nonprofit 501(c)(6) industry trade association founded in 1909 and governed by a 22-member Board of Directors. IDEA represents approximately 1600 members who are district heating and cooling executives, managers, engineers, consultants and equipment suppliers from 26 countries. Association members operate district energy systems owned by utilities, municipalities, hospitals, military bases and airports throughout North America and around the world. Visit http://www.districtenergy.org for more information.


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Lamb Steak Frites
Whole and butterflied leg of lamb are familiar cuts, but steaks cut from the leg are much less common. They're worth trying, especially because they're easy and they cook so quickly. Here, the steaks are basted with a flavorful herb-infused butter, then served with fries as a clever variation on steak frites.
- 2 1 1/4-pound lamb leg steaks, cut about 1 1/4 inches thick
- 1 cup chicken stock
- 1 cup dry red wine
- 1 shallot, minced
- 6 Tbsp unsalted butter
- 2 garlic cloves
- 4 small thyme sprigs
- 2 Tbsp canola oil
- Freshly ground pepper
- Kosher salt
- 0.5 tsp ground fennel seeds
- 0.5 tsp ground cumin
- French fries, for serving
- Rub the lamb with the cumin and fennel; season with salt and pepper. In a cast-iron skillet, heat the oil until smoking. Add the lamb and cook over high heat, turning once, until browned. Reduce the heat to moderate. Add the thyme, garlic and 4 tablespoons of the butter and cook for 5 minutes, basting the lamb with the butter. Turn the steaks and cook, basting, until an instant-read thermometer inserted in the thickest part registers 130°, 5 minutes; transfer to a carving board and let rest for 10 minutes. Thickly slice against the grain.
- Pour off all but 1 tablespoon of the fat. Cook the shallot over moderately high heat until softened. Add the wine and simmer until syrupy. Add the stock and simmer until slightly reduced, 3 minutes. Off the heat, whisk in the remaining 2 tablespoons of butter; season with salt and pepper. Serve with the lamb and French fries.
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Recipe Credit: Tony Maws
Image Credit: John Kernick