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Comparison of IL-6, IL-8 Concentrations in H. Pylori- and Non-H. Pylori-associated Gastritis

BACKGROUND: Helicobacter pylori is a non-invasive microorganism causing intense gastric mucosal inflammatory and immune reaction. The gastric mucosal levels of the proinflammatory cytokines Interleukin 6 (IL-6) and IL-8 have been reported to be increased in H. pylori infection, but the serum levels in H. pylori infection is still controversial. The purpose of this study was to investigate the serum levels of IL-6 and IL-8 in H. pylori infection.METHODS: A cross sectional study was done on eighty consecutive gastritis patients admitted to endoscopy units at Adam Malik General Hospital and Permata Bunda Hospital, Medan, Indonesia from May-October 2014. Histopathology was performed for the diagnosis of gastritis. Rapid urease test for diagnosis of H. pylori infection. Serum samples were obtained to determine circulating IL-6 and IL-8. Univariate and bivariate analysis (independent t test) were done.RESULTS: There were 41.25% patients infected with H. pylori. Circulatory IL-6 levels were significantly higher in H. pylori-infected patients compared to H. pylori negative, but there were no differences between serum levels of IL-8 in H. pylori positive and negative patients.CONCLUSION: The immune response to H. pylori promotes systemic inflammation, which was reflected in an increased level of serum IL-6. Serum levels of IL-8 were not significantly different between H. pylori positive and negative.

RESULTS: There were 41.25% patients infected with H. pylori. Circulatory IL-6 levels were signiicantly higher in H. pylori-infected patients compared to H. pylori negative, but there were no differences between serum levels of IL-8 in H. pylori positive and negative patients.

Introduction
Helicobacter pylori is the most common bacterial infection of humans that is speciic for gastric epithelial cells. Depending on geographic location, H. pylori is inhabitant of half of the world population's stomach. H. pylori plays a crucial role in the development of gastrointestinal diseases such as chronic gastritis, gastric ulcer, and duodenal ulcer mucosa associated lymphoid tissue lymphoma and gastric adenocarcinoma. H. pylori is a non-invasive microorganism causing intense gastric mucosal inlammatory and immune reaction. (1,2) Cytokines are responsible for orchestrating the immune response. H. pylori-induced gastric mucosal cytokine overproduction has been clearly documented previously. Local production of proinlammatory cytokines such as interleukin 6 (IL-6) and IL-8 are thought to play a central role in the recruitment of inlammatory cells to the gastric mucosa in the presence of H. pylori. (3) Whether this inlammation is limited to gastric mucosa or causes systemic inlammation remains unclear.
The stomach has a large surface area and continuous spillover of locally produced cytokines into the blood stream is a possibility. There are few and conlicting data on circulatory cytokine levels in patients with H. pylori infection. (4) The purpose of the present study was to compare the serum levels of IL-6 and IL-8 as proinlammatory cytokines in H. pylori-and non-H. pylori-associated gastritis patients.

Patient selection
The present study was a cross sectional study on gastritis patients at Adam Malik General Hospital and Permata Bunda Hospital, Medan, Indonesia from May-October 2014. Eighty consecutive patients with gastritis admitted to endoscopy units were included in the study. All patients gave informed consent and the study was approved by the local ethic committee, No. 331/KOMET/FK USU/2014. None of the patients had received antibiotics, bismuth compounds, H2 antagonists, proton pump inhibitors or immune modulating drugs within the last four weeks before endoscopy. Patients with evidence of malignancy, immunosuppression, metabolic disorders, or gastrointestinal hemorrhage and patients who had a history of gastric surgery were excluded.

Diagnosis of gastritis
Diagnosis of gastritis was evaluated from biopsy of the mucosa of gastric antrum and body. The biopsy specimens were ixed in 10% formalin and embedded in parafin.
The samples were stained using Hematoxylin-Eosin and were evaluated by an anatomic pathologist from Faculty of Medicine, University of Sumatera Utara.

Diagnosis of H. pylori
H. pylori was considered positive from the positive results of rapid urease test. Result of the rapid urease test was read within 24 hours.

Serum levels of IL-6 and IL-8
Venous blood was drawn using a serum separator tube and allowed to clot for 30-45 minutes at room temperature before centrifugation for 15 minutes at approximately 1,000g. Serum was immediately stored frozen in aliquots at -20°C until assays for IL-6 and IL-8 were performed. IL-6 was assayed with Quantikine ELISA Human IL-6 Immunoassay (R&D System, Minneapolis, MN, USA), while IL-8 was assayed with Quantikine HS ELISA Human CXCL8/IL-8 Immunoassay (R&D System).

Statistical analysis
SPSS version 22 (SPSS Inc., Chicago) was used for the analysis. The data were analyzed using univariate and bivariate analysis with 95% conidence interval. Results are expressed as the mean +/standard deviation. Bivariate analysis was carried out using independent t test. p</0.05 was considered statistically signiicant.  Table 2. Serum level of IL-6 and IL-8 in H. pylori (+) and

H. pylori (-)
Mean age of gastritis subjects in this study was 49.3 + 13.4 (SD) years old, which is considered a productive age group.
In addition, the age groups with the most cases of gastritis were those in the age group of 46-60 and 30-45. This result is in accordance with the results from the previous studies such as that by Garg  H. pylori is the major recognized etiological agent inducing gastric inlammatory responses. These responses can be considered to have two components: chronic inlammatory responses associated with increased numbers of mononuclear cells in the lamina propria iniltration, including lymphocytes, monocytes/macrophages, and plasma cells; and acute inlammatory response characterized by intraepithelial and interstitial neutrophilic. These two responses may be regulated differentially following induction of cytokines involved in the inlammatory cascade. IL-6 may be involved in chronic inlammatory changes through its broad effects on growth and differentiation of mononuclear cells, including T and B lymphocytes and macrophages. IL-6 correlated strongly with the mononuclear cell density. with H. pylori. The majority of subject`s employment status was housewife (36.25%) and self-employed (32.5%).
The majority of the subjects had a normal nutrition status (53.75%).

Serum IL-6 and IL-8 levels in H. pylori infection
Mean serum levels of IL-6 and IL-8 were higher in H. pylori positive than H. pylori negative (Figure 1). Serum IL-6 level was signiicantly higher in infected group compared to H. pylori negative (p<0.05). Serum IL-8 level was higher in H. pylori positive than H. pylori negative (38.96 + 25.92 vs. 36.01 + 24.69), but not signiicant.