Anastassios C. Manolakis

A REVIEW OF THE POSTULATED MECHANISMS CONCERNING THE ASSOCIATION OF HELICOBACTER PYLORI WITH ISCHEMIC HEART DISEASE

Since its discovery, Helicobacter pylori has been implicated in the pathogenesis of several diseases, both digestive and extradigestive. Interestingly, the majority of the extradigestive‐related literature is focused on two vascular manifestations: stroke and ischemic heart disease. Potential mechanisms for the establishment of a H. pylori‐induced ischemic heart disease have been proposed with regard to chronic inflammation, molecular mimicry, oxidative modifications, endothelial dysfunction, direct effect of the microorganism on atherosclerotic plaques as well as changes regarding traditional or novel risk factors for ischemic heart disease or even platelet‐H. pylori interactions. A positive link between H. pylori infection and ischemic heart disease has been suggested by a series of studies focusing on epidemiologic evidence, dyslipidemic alterations, upregulation of inflammatory markers or homocysteine levels, induction of hypercoagulability, oxidation of low‐density lipoprotein, causation of impaired endothelial function, detection of H. pylori DNA in atherosclerotic plaques, and participation of certain antigens and antibodies in a cross‐reactivity model. There are studies, however, which investigated the relationship between H. pylori and ischemic heart disease with regard to the same parameters and failed to confirm the suggested positive association. Further studies in the direction of interaction between H. pylori and the hosts genotype as well as a quest for evidence towards novel risk factors for ischemic heart disease such as oxidative stress, vascular remodeling, vascular calcification, or vasomotor activity, may reveal a field of great interest, thus contributing to the determination of new potential mechanisms.

Calprotectin, Calgranulin C, and Other Members of the S100 Protein Family in Inflammatory Bowel Disease

BackgroundSince their discovery, S100 proteins have been associated with diverse diseases of inflammatory, degenerative, or malignant nature. Due to their participation in inflammation, they have also been studied with regard to inflammatory bowel disease (IBD).MethodTo provide a review of available literature, a PubMed, MEDLINE, and Embase-based literature search was performed, using all available nomenclature for each member of the S100 protein family, along with the terms inflammatory bowel disease, ulcerative colitis, Crohn’s disease, or indeterminate colitis.ResultS100A8/A9, also known as calprotectin, S100A12, or calgranulin C and in a lesser extent S100P, are involved in the pathogenesis, activity, diagnosis, and therapeutic management of IBD. The majority of available literature is focused primarily on S100A8/9, although there is growing evidence on the significance of S100A12. Most studies emphasize the potential merit of S100A8/A9 and S100A12, as markers for differential diagnosis, monitoring of activity, or disease relapse, in IBD. Limitations, regarding the diagnostic utility of these markers, seem to exist and are mainly related to the publication of conflicting results, i.e., for IBD activity, and to the fact that S100A8/A9 and S100A12 are not disease-specific.ConclusionsAlthough the existing data link specific S100 proteins with IBD, there are still several drawbacks in the use of these markers for diagnostic purposes. Thus, it seems that further research is mandatory in order to eliminate the impact of confounding factors but also to detect additional associations between S100 proteins and IBD or novel S100 proteins with a closer correlation with IBD.

Epidermal Growth Factor in the Treatment of Diabetic Foot Ulcers An Update

Management of diabetic foot ulcers remains a rather challenging task. Epidermal growth factor (EGF) plays a central role in wound healing. It acts on epithelial cells and fibroblasts promoting restoration of damaged epithelium. However, its bioavailability is impaired in chronic diabetic foot ulcers. Current evidence suggests that application of human recombinant EGF in addition to standard treatment is able to achieve both partial and complete healing and to prevent foot amputations. Its efficacy has been tested at various concentrations and by various administration routes (topical application and intralesional injection). Intralesional injection has better availability on the deep wound layers, but pain at the injection site is a common complaint. Generally, adverse events have been minor to mild. Finally, numerous issues need to be further clarified before widespread use of EGF becomes possible in everyday practice. Such issues include optimal dosage and administration route, characteristics of the ulcers most likely to heal (severity and ischemic/neuropathic or both), and cost-effectiveness.

The role of hyperbaric oxygen in the treatment of diabetic foot ulcers.

Diabetic foot ulcers are still extremely difficult to heal. Therefore, therapeutic options to improve healing rates are continuously being explored. Hyperbaric oxygen (HBO) has been used in addition to standard treatment of the diabetic foot for more than 20 years. Evidence suggests that HBO reduces amputation rates and increases the likelihood of healing in infected diabetic foot ulcers, in association with improved tissue oxygenation, resulting in better quality of life. Nonetheless, HBO represents an expensive modality, which is only available in few centers. Moreover, adverse events necessitate a closer investigation of its safety. Finally, it is not entirely clear which patients stand to benefit from HBO and how these should be selected. In conclusion, HBO appears promising, but more experience is needed before its broad implementation in the routine care of the diabetic foot.

Angiogenin, angiopoietin-1, angiopoietin-2, and endostatin serum levels in inflammatory bowel disease.

Background: Angiogenesis is a complex process, involving a great number of mediators. It is implicated in the pathogenesis of numerous diseases, holding a critical role in inflammatory bowel disease (IBD). The objective of this study was to assess serum levels of angiogenin, angiopoietin‐1, angiopoietin‐2, and endostatin in IBD patients. Methods: Measurement of all angiogenesis mediators was performed with a commercially available enzyme‐linked immunosorbent assay. Fifty‐two patients with ulcerative colitis (UC), 59 with Crohns disease (CD), and 55 healthy controls (HC) were included in the study. The values were analyzed with regard to disease and patients characteristics. Results: Angiogenin levels were significantly higher in IBD patients compared to HC (P < 0.001) and in UC and CD smoker patients compared to nonsmokers (P = 0.0121 and P = 0.005, respectively). Angiogenin levels were lower in UC patients receiving 5‐aminosalicylate (5‐ASA) alone, compared to those receiving combined therapy (P = 0.0478). Angiopoietin‐1 levels were significantly lower in IBD patients compared to HC (P < 0.0001) and increased in smokers compared to nonsmoker UC patients (P = 0.0085). IBD patients demonstrated increased angiopoietin‐2 levels compared to HC (P = 0.0131), while CD patients with disease restricted to the colon had significantly lower levels compared to other disease locations (P < 0.0001). Higher endostatin levels were recorded in UC patients with extensive colitis. Conclusions: Elevated serum angiogenin and angiopoietin‐2 levels and lower serum angiopoietin‐1 levels were shown in IBD patients, as well as a different pattern of angiogenic factor alterations related to location, treatment, smoking habits and gender. Inflamm Bowel Dis 2011

TLR4 gene polymorphisms: evidence for protection against type 2 diabetes but not for diabetes-associated ischaemic heart disease

OBJECTIVE Several factors either predisposing or protecting from the onset of diabetes mellitus type 2 (DM2) have been proposed. Two specific polymorphisms of toll-like receptor 4 (TLR4; Asp299Gly and Thr399Ile) have recently been identified either as candidate protector genes against DM2 and associated neuropathy or risk alleles for the manifestation of diabetic retinopathy. The impact of these alleles on the risk for ischaemic heart disease (IHD) is controversial while their role in diabetes-associated IHD has never been studied. DESIGN AND METHODS In order to clarify the potential impact of TLR4 polymorphisms on the predisposition for DM2 as well as on diabetes-related IHD vulnerability, the distribution of the mutant TLR4 Asp299Gly and Thr399Ile alleles in 286 DM2 patients and 413 non-DM2 controls with or without IHD, was examined. RESULTS Mutant alleles were predominantly detected in 79/413 non-diabetic individuals versus 15/286 DM2 patients (P<0.0001). The rates of positivity for mutant alleles were similar among diabetic patients with or without IHD (7/142 vs 8/144, P>0.1), whereas they proved different among non-diabetic individuals with or without IHD (39/145 vs 40/268, P=0.004). Following multivariate analysis, the difference between diabetic and non-diabetic subjects, with regard to TLR4 mutations alone, remained significant (P=0.04). CONCLUSIONS Mutant TLR4 alleles confer protection against DM2. However, their presence does not seem to play any role, protective or aggravating, in the manifestation of IHD either in diabetic or in non-diabetic individuals.

Downregulation of serum epidermal growth factor in patients with inflammatory bowel disease. Is there a link with mucosal damage

Background: Epidermal growth factor (EGF) is a multipotent peptide which contributes to epithelial development, inhibition of gastric acid secretion, acceleration of wound healing, and promotion of angiogenesis. The aim of this study is to evaluate serum EGF concentrations in inflammatory bowel disease (IBD) patients, with regard to disease and patients’ characteristics. Methods: EGF determination was performed by a commercially available enzyme-linked immunosorbent assay. Fifty-two patients with ulcerative colitis (UC), 59 with Crohns disease (CD), and 55 healthy controls (HC) were included in the study. Results: Mean ( ± SEM) serum EGF levels were 217.2 ( ± 30.40) pg/mL in UC patients, 324.6 ( ± 37.29) pg/mL in CD patients, and 453.1 ( ± 39.44) pg/mL in HC. Serum EGF levels were significantly lower in UC and CD patients compared to HC (P < 0.0001 and P = 0.0199, respectively). Lower serum EGF levels were observed in UC compared to CD patients (P = 0.0277). Extent of the disease was found to affect serum EGF levels in UC, demonstrating significant reduction in patients with left-sided colitis and pancolitis in comparison with those with proctitis (P = 0.0190 and P = 0.0024, respectively). EGF concentration was not influenced by other characteristics of patients and disease. Conclusions: Significantly, lower levels of serum EGF are observed in IBD patients compared to HC, while disease extent plays a key role in regulation of serum EGF in UC. Downregulation of serum EGF may be correlated with different patterns of bowel inflammation, epithelial development, and wound healing in IBD.

The implication of adiponectin and resistin in gastrointestinal diseases

Adiponectin and resistin, members of the adipokine family, are multi-task hormones involved in several disorders, including those of the alimentary tract. In the present review, eligible studies focusing on the role of adiponectin and resistin in gastrointestinal diseases are manifested together and classified according to anatomic criteria. In addition, similarities and common patterns have been recognized, ultimately revealing an inverse association: the down-regulation of adiponectin and up-regulation of resistin - both in vitro and in vivo - in gastrointestinal disorders, irrespective of their diverse nature - inflammatory, autoimmune or malignant - or anatomic position - esophageal, gastric, of the small intestine, colonic. Finally, a potential role for both adipokines in alimentary tract-related carcinogenesis has been identified, possibly representing a missing link between obesity and cancer.

Increased fetuin A levels in Helicobacter pylori infection: a missing link between H. pylori and insulin resistance?

To the Editor: Helicobacter pylori infection has been associated with diverse biological processes, including inflammation, metabolism, oncogenic transformation [1, 2]. In view of its effect on metabolic variables, H. pylori has been linked with both dyslipidaemia and insulin resistance (IR) [1, 2]. Among the various factors capable of inducing IR, the upregulation of α2-Heremans Schmid glycoprotein, also known as human fetuin A, has been linked with impaired insulin sensitivity, glucose metabolism and, subsequently, the onset of diabetes mellitus [3, 4]. Interestingly, certain pathogens have been shown to induce an increase in the level of fetuin A or fetuin A-like molecules such as Mycobacterium bovis in cattle and severe acute respiratory syndrome-coronavirus (SARSCoV) in humans [5]. Based on these data, we performed a study to investigate whether the H. pylori-induced IR is mediated through an upregulation of fetuin A levels. Determination of circulating fetuin A (by ELISA; BioSource Europe, Nivelles, Belgium), fasting insulin (by ELISA; DRG Instruments, Marburg, Germany) and glucose levels (hexokinase method, cat. no. OSR6521; Olympus Life Science Research Europa, Hamburg, Germany) was performed for 105 non-diabetic individuals (Table 1) undergoing oesophagogastroduodenoscopy owing to dyspeptic complaints. According to the results of a Campylobacter-like organism test and histology, study participants were classified into H. pylori-positive (Hp, n=72) and negative (Hp, n=33) groups matched for age, sex, BMI and smoking. For IR, the HOMA-IR (www.hepcnomads. co.uk/HOMACalc.htm, accessed 11 November 2010) was used. Details of lipoprotein, triacylglycerol and C-reactive protein (CRP) levels were also available from routine examinations. None of the participants was receiving any medication and none had any factor that could affect H. pylori diagnosis, fetuin A levels or glucose metabolism (family history of diabetes mellitus, obesity, proton pump inhibitor use, liver, kidney, systemic disorders, polycystic ovary disease). The study had been approved by the Ethics Committee of the University of Thessaly Medical School. Informed consent was obtained from all individuals prior to inclusion in the study. A. C. Manolakis : E. K. Tiaka :A. N. Kapsoritakis : F. Tsiopoulos : S. P. Potamianos (*) Department of Gastroenterology, University of Thessaly, School of Medicine, Mezourlo, 41110 Larissa, Greece e-mail: spotam@med.uth.gr

Impact of TLR‐4 Polymorphisms on Circulating Levels of Antibodies Against Helicobacter pylori

Dear Editor, Over the years, Helicobacter pylori infection has been associated with a variety of intestinal, as well as extraintestinal disorders [1], thus underlining the need for the development and application of successful diagnostic and therapeutic strategies [2]. Among the procedures used for the diagnosis of infection from this Gram-negative bacterium, serology antibodies against H. pylori is one of the most commonly applied in research and clinical practice [2]. On the other hand, various factors capable of modifying immune responses against pathogens are currently known, including polymorphisms of toll-like receptors (TLR) [3,4]. These molecules and their polymorphic alleles have been implicated in the recognition of several bacterial components i.e. lipopolysaccharides of Gram-negative bacteria from TLR-4 [3–5]. To investigate whether the presence of polymorphic alleles could influence the production of antibodies against H. pylori, antibody titers were compared between H. pylori-infected individuals, with either wild-type or mutant TLR-4 genotypes. The TLR-4 Asp299Gly and Thr399Ile polymorphic alleles were genotyped using a polymerase chain reaction (PCR) and direct sequencing, in 246 individuals with documented, via both campylobacter-like organism (CLO) test and histology, H. pylori infection. For the purposes of the PCR amplification, a forward, 5¢-TCTAGAGGGCCTGTGCAATT-3¢, and a reverse primer, 5¢-TGAAACTCACTCATTTGTTTCAA-3¢, were used. Using an enzyme-linked immunosorbent assay (ELISA, Enzygnost; Dade Behring Marburg GmbH, Marburg, Germany), IgG and IgA antibodies against H. pylori were determined. All study participants lacked any disorder that could alter the immune response and were not under immuno-modulatory or -suppressive medication. A total of 40 individuals were tested positive for both mutant alleles, heterozygotes, while the rest 206 were homozygotes for the wild-type allele. The simultaneous presence –cosegregationof TLR-4 Asp299Gly and Thr399Ile alleles, in our study was 100%. The mean ± SEM of anti-H. pylori IgG titers were 51.90 ± 4.927 U ⁄ mL, in individuals with the wild-type genotype, and 48.94 ± 9.878 U ⁄ mL, in the group carrying the mutant alleles (p > .05). On the contrary, a statistically significant difference was recorded when antiH. pylori IgA titers were compared between carriers of wild-type (11.61 ± 1.330 U ⁄ mL) and mutant polymorphic alleles (5.887 ± 0.6506 U ⁄ mL) (p < .001). Evidently, lower IgA levels were found in the group of TLR-4 Asp299Gly and Thr399Ile gene carriers. This observation does not come entirely as a surprise, because interactions between TLR-4 and Gram-negative bacteria have already been described [3–5]. The impact of the TLR-4 Asp299Gly and Thr399Ile polymorphic alleles, however, on immune responses and especially those triggered by pathogens, has been debated [3]. One of the major reasons for the lack of unanimity regarding this matter was that the simultaneous presence of the two alleles did not seem to increase susceptibility or alter the cytokine response to microorganisms [3]. The results from this study seem to contradict the notion that the cosegregated mutant alleles ‘‘behave’’ pretty much like the wild-type allele. As for the rather selective downregulation of IgA response against H. pylori, it could be hypothesized that it reflects a defect or a dysregulation in the mucosal defense against H. pylori. Besides, the production of IgAs, circulating and mucosal, which represent the most common immunoglobulin subclass in the mucosa, can be triggered by TLR-facilitated recognition of microorganisms, in the intestinal epithelium [4,5]. In addition, a similar blunt antibody production has been reported for anti-chitobioside (ACCA) and anti-outer membrane porin (Omp) IgA, in patients with inflammatory bowel disease carrying the TLR-4 Asp299Gly allele [6]. This, however, is the first attempt to record a potential interaction between widely studied polymorphisms, tightly linked to the host’s immune responses toward Gram-negative bacteria, and H. pylori. In view of the notion that the presence of these specific TLR-4 polymorphic alleles could reduce the diagnostic utility of H. pylori serology, our findings are not consistent with this remark, because the IgG response against H. pylori remained intact. All in all, it seems that the idea that the host’s genetic background can ‘‘carve’’ the reaction toward environmental challenges is once more verified this time via the suppression of anti-H. pylori Helicobacter ISSN 1523-5378