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Non celiac gluten sensitivity (NCGS) is a syndrome characterized by a cohort of symptoms related to the ingestion of gluten-containing food in subjects who are not affected by celiac disease (CD) or wheat allergy. The possibility of systemic manifestations in this condition has been suggested by some reports. In most cases they are characterized by vague symptoms such as 'foggy mind', headache, fatigue, joint and muscle pain, leg or arm numbness even if more specific complaints have been described. NCGS has an immune-related background. Indeed there is a strong evidence that a selective activation of innate immunity may be the trigger for NCGS inflammatory response. The most commonly autoimmune disorders associated to NCGS are Hashimoto thyroiditis, dermatitis herpetiformis, psoriasis and rheumatologic diseases. The predominance of Hashimoto thyroiditis represents an interesting finding, since it has been indirectly confirmed by an Italian study, showing that autoimmune thyroid disease is a risk factor for the evolution towards NCGS in a group of patients with minimal duodenal inflammation. On these bases, an autoimmune stigma in NCGS is strongly supported; it could be a characteristic feature that could help the diagnosis and be simultaneously managed. A possible neurological involvement has been underlined by NCGS association with gluten ataxia, gluten neuropathy and gluten encephalopathy. NCGS patients may show even psychiatric diseases such as depression, anxiety and psychosis. Finally, a link with functional disorders (irritable bowel syndrome and fibromyalgia) is a topic under discussion. In conclusion, the novelty of this matter has generated an expansion of literature data with the unavoidable consequence that some reports are often based on low levels of evidence. Therefore, only studies performed on large samples with the inclusion of control groups will be able to clearly establish whether the large information from the literature regarding extra-intestinal NCGS manifestations could be supported by evidence-based agreements.

Non Celiac Gluten Sensitivity (NCGS) is characterized by immunological, morphological or symptomatic manifestations precipitated by gluten ingestion in individuals without celiac disease (CD). The most important challenge in NCGS is the diagnosis, currently based only on clinical observation. The “Salerno criteria” have been pointed out to achieve a reliable diagnosis even if they lack immediacy and practicality, thus making questionable patient’s adherence. Therefore, biological indicators supporting the clinical diagnosis of NCGS are advisable. For these reasons, many attempts have been performed in order to identify possible serological, immunological, histopathological, immunohistochemical and pathophysiological aspects characterizing this condition with the aim of using them for diagnostic purposes. In the present narrative review, we carried out an update of the current scenario of potential markers of NCGS. The main fault of available studies is that, in most cases investigations have been pointed out towards molecules, which cannot be searched in the current laboratories of clinical analysis. Therefore, the matter has been confined within basic research. Additionally, in these studies, sensitivity and specificity of biological markers were not computable. This is a relevant limit, since an ideal test for NCGS should have a good discriminative power against both CD and other causes of microscopic enteritis. Until now, serological tests have failed, while the search for a soluble marker indicative of activation of innate immune system as well as immunohistochemistry could be the promising bases for the development of appropriate investigations in the future.

In the present narrative review, we analyzed the relationship between seronegative celiac disease (SNCD) and immunoglobulin deficiencies. For this purpose, we conducted a literature search on the main medical databases. SNCD poses a diagnostic dilemma. Villous blunting, intraepithelial lymphocytes (IELs) count and gluten “challenge” are the most reliable markers. Immunohistochemistry/immunofluorescence tissue transglutaminase (tTG)-targeted mucosal immunoglobulin A (IgA) immune complexes in the intestinal mucosa of SNCD patients may be useful. In our experience, tTG-mRNA was similarly increased in seropositive celiac disease (CD) and suspected SNCD, and strongly correlated with the IELs count. This increase is found even in the IELs’ range of 15–25/100 enterocytes, suggesting that there may be a “grey zone” of gluten-related disorders. An immune deregulation (severely lacking B-cell differentiation) underlies the association of SNCD with immunoglobulin deficiencies. Therefore, CD may be linked to autoimmune disorders and immune deficits (common variable immunodeficiency (CVID)/IgA selective deficiency). CVID is a heterogeneous group of antibodies dysfunction, whose association with CD is demonstrated only by the response to a gluten-free diet (GFD). We hypothesized a familial inheritance between CD and CVID. Selective IgA deficiency, commonly associated with CD, accounts for IgA-tTG seronegativity. Selective IgM deficiency (sIgMD) is rare (

Background/Aim: In celiac disease (CD), there is increased mRNA coding for tissue transglutaminase (tTG) and interferon gamma (IFNα). In seronegative celiac patients, the mucosal immune complexes anti-tTG IgA/tTG are found. We assayed tTG- and IFNα-mRNA in the mucosa of patients with a clinical suspicion of seronegative CD and correlated the values with intraepithelial CD3 lymphocytes (IELs). Materials and Methods: Distal duodenum specimens from 67 patients were retrieved and re-evaluated for immunohistochemically proven CD3 IELs. Five 10 μm sections were used for the extraction and assay of tTG and IFNα coding mRNA levels using reverse transcriptase real-time polymerase chain reaction (RT-PCR). Samples from 15 seropositive CD patients and 15 healthy subjects were used as positive and negative controls. Results were expressed as fold-change. Results: Our series was divided into three groups based on IEL count: >25 (14 patients: group A), 15–25 (26 patients: group B), and 0–15 (27 patients: Group C). tTG-mRNA levels were (mean ± SD): CD = 9.8 ± 2.6; group A = 10.04 ± 4.7; group B = 4.99 ± 2.3; group C = 2.26 ± 0.8, controls = 1.04 ± 0.2 (CD = group A > group B > group C = controls). IFNα-mRNA levels were: CD = 13.4 ± 3.6; group A = 7.28 ± 3.6; group B = 4.45 ± 2.9; group C = 2.06 ± 1.21, controls = 1.04 ± 0.4. Conclusions: Our results suggest that tTG- and IFNγmRNA levels are increased in both seropositive and potential seronegative patients with CD, showing a strong correlation with the CD3 IEL count at stage Marsh 1. An increase in both molecules is found even when IELs are in the range 15–25 (Marsh 0), suggesting the possibility of a "gray zone" inhabited by patients which should be closely followed up in gluten-related disorders.

Background & Aims: Antibiotic resistance is the main reason for failure of Helicobacter pylori (H. pylori) treatment. Currently, guidelines recommend a treatment guided by antimicrobial susceptibility testing after two failures. However, microbial culture is not feasible everywhere, and the limited number of effective antibiotics against the bacterium narrows the options; thus a rescue therapy combining antibiotics with a low resistance may be fitting.Methods: Patients who have failed a first-line treatment (either prolonged triple or sequential regimens) and, successively, a levofloxacin-based triple therapy were considered for the study. Subjects underwent urea breath test (UBT), stool antigen test (ST) and endoscopy/histology to confirm the diagnosis. Cytopenia and impaired liver and kidney function were exclusion criteria. Fifty-four subjects were randomized 1:1 to two regimens: RMB Rabeprazole/Rifabutin/Minocycline/Bismuth sub-citrate or MTB Rabeprazole/Tinidazole/ Minocycline/Bismuth sub-citrate both for 10 days. The results were checked 6 weeks after the end of therapy with ST/UBT plus endoscopy when indicated.Results: RMB eradicated the bacterium in 21 patients. Two subjects dropped out. The eradication rate was 77.7% (CI 62.0-93.4%) at intention-to-treat and 84.0% (CI 69.6-98.4%) at per-protocol analysis. MTB was successful in 14 patients (51.9%, CI 33.1-70.7%). No patient withdrew from the treatment for adverse events. Drug-related side effects were reported only in 3 subjects, but in all cases the treatment was carried on.Conclusions: The association minocycline/rifabutin seems to have a synergic effect and a good therapeutic outcome in patients who have failed at least two previous regimens, although a trial on a large population is needed.

Microscopic enteritis (ME) is characterized by abnormal infiltration of intraepithelial lymphocytes in intestinal mucosa. It was described as duodenal lymphocytosis or lymphocytic duodenitis until the dedicated Consensus Conference of 2015. ME represents a common feature of several gluten-mediated and non-gluten related diseases; therefore, it is an umbrella term embracing several conditions. The most frequent causes of ME are gluten-related disorders (celiac disease, non-celiac gluten sensitivity, wheat allergy), Helicobacter pylori infection and drug-related damages. Less frequently, ME may be secondary to inflammatory bowel disease, some autoimmune conditions, immunoglobulin deficiencies, blood malignancies, infections and irritable bowel syndrome. Therefore, the differential diagnosis of ME may be challenging. The diagnosis of ME needs to be driven by predominant symptoms and patient history. However, it is often difficult to achieve an immediate identification of the underlying condition, and a broad variety of diagnostic tests may be required. Ultimately, long-term surveillance is needed for a final diagnosis in many cases, since a hidden or quiescent condition may be disclosed after a period of latency. In any case, strict collaboration between the clinician and the pathologist is pivotal. The treatment of ME should be personalized, depending on the underlying disease. For gluten-related conditions (celiac disease, gluten sensitivity, wheat allergy, dermatitis herpetiformis), a gluten-free diet may be proposed. For other conditions, a targeted etiologic treatment is necessary. In conclusion, ME represents a novel entity that is attracting increasing interest. The growing epidemiologic trend confirms that it will become a common condition in clinical practice.

Background and Aims. Nonceliac gluten sensitivity (NCGS) is a gluten-related emerging condition. Since few data about NCGS histopathology is available, we assessed the markers of lymphocyte and innate immunity activation. Materials and Methods. We retrieved duodenal biopsy samples of patients with NCGS diagnosis according to the Salerno criteria. We selected specimens of positive (seropositive celiac disease/Marsh 1-2 stage) and negative (normal microscopic picture) controls. Immunohistochemistry for CD3 (intraepithelial lymphocytes-IELs), CD4 (T helper lymphocytes), CD8 (T cytotoxic lymphocytes), and CD1a/CD117 (Langerhans/mast cells) was performed. ANOVA plus Bonferroni’s tests were used for statistical analysis. Results. Twenty NCGS, 16 celiac disease, and 16 negative controls were selected. CD3 in NCGS were higher than negative controls and lower than celiac disease (18.5 ± 6.4, 11.9 ± 2.8, and 40.8 ± 8.1 IELs/100 enterocytes; p<0.001). CD4 were lower in NCGS than controls and celiac disease (31.0 ± 22.1, 72.5 ± 29.5, and 103.7 ± 15.7 cells/mm2; p<0.001). CD8 in NCGS were similar to negative controls, but lower than celiac disease (14.0 ± 7.4 and 34.0 ± 7.1 IELs/100 enterocytes, p<0.001). CD117 were higher in NCGS than celiac disease and negative controls (145.8 ± 49.9, 121.3 ± 13.1, and 113.5 ± 23.4 cells/mm2; p=0.009). Conclusions. The combination of CD4 and CD117, as well as IEL characterization, may be useful to support a clinical diagnosis of NCGS.

Bacterial intestinal overgrowth syndrome (SIBO) treatment is based on antibiotics. Probiotics have been shown to give similar results, whilst no study is available about prebiotics. This study evaluated the addition of probiotics or prebiotics to antibiotics on SIBO symptoms in a 6-month follow-up. We enrolled 40 patients (14 males and 26 females) reporting abdominal compliant without gastrointestinal diseases/alarm symptoms. SIBO was diagnosed by the agreement of lactulose and glucose breath tests. Patients were randomly divided into two groups homogeneous for sex and age: group 1 received Rifaximin 400 mg/day for 7 days/month followed by Lactobacillus casei for 7 days more and group 2 antibiotic followed by short chain fructo-oligosaccharides. All patients recorded a questionnaire for subjective symptom evaluation according to Rome III criteria and Bristol scale for stool characters before the study and after 6 months. Statistics: Student’s t and Fisher’s exact tests. In group 1, a significant improvement was obtained in 5 out of 6 symptoms, whilst in group 2 in 4 out of 6 symptoms (nausea and number of bowel movements failed to improve). Despite we observed a trend of probiotics to be more effective than prebiotics, the difference in the percentage of improved symptoms was not significant (83,3% vs 66.6%; p= 0.57). Our preliminary data show a good outcome with sequential antibioticprobiotic/ prebiotic administration in patients with SIBO.

The diagnosis of Celiac Disease (CD) relies on the concordance of pathological, serological, genetic and clinical features. For this reason, the diagnosis of CD is often a challenge. Seronegative celiac disease (SNCD) is defined by the negativity of anti-tissue transglutaminase antibodies in the presence of a positive histology on duodenal biopsy samples, i.e. inflammatory infiltrate of intra-epithelial lymphocytes (IELs > 25/100 enterocytes), mild villous atrophy and uneven brush border associated to human leukocyte antigen (HLA) haplotype DQ2 and/or DQ8. SNCD is characterized by mucosal deposits of tissue transglutaminase (tTG)/anti-tTG immuno-complexes. These may counteract the passage of anti-tTG into the bloodstream, thus explaining seronegativity. Another reason for seronegativity may be found in an incomplete maturation of plasma cells with a consequent failure of antibodies production. This condition often characterizes immunoglobulin deficiencies, and, indeed, SNCD is common in subjects with immunoglobulin deficiencies. The management of SNCD still remains debated. The treatment option for SNCD may be represented by gluten free diet (GFD), but the usefulness and appropriateness of prescribing GFD are controversial. Some evidences support its use only in SNCD subjects showing CD clear clinical picture and compatible HLA status. The choice of GFD administration could be linked to an investigation able to diagnose SNCD in no doubt even if a reliable test is not currently available. On these bases, a test helping the diagnosis of SNCD is justifiable and desirable.

To assess the evolution of duodenal lymphocytosis (DL), a condition characterized by increased intraepithelial lymphocytes (IELs), over 2 years of follow-up.Consecutive patients undergoing upper endoscopy/histology for abdominal pain, diarrhea, weight loss, weakness or other extraintestinal features compatible with celiac disease (CD) were included. Evaluation of IELs infiltrate in duodenal biopsy samples was carried out by CD3-immunohistochemistry and expressed as number of positive cells/100 enterocytes. Diagnostic agreement on the IELs count was tested by calculating the weighted k coefficient. All patients underwent serological detection of autoantibodies associated with CD: IgG and IgA anti-tissue transglutaminase and endomysium. Each patient underwent further investigations to clarify the origin of DL at baseline and/or in the course of 2 years of follow-up every six months. Autoimmune thyroiditis, intestinal infections, parasitic diseases, bacterial intestinal overgrowth, hypolactasia and wheat allergy were detected. Colonoscopy and enteric magnetic resonance imaging were performed when necessary. Risk factors affecting the final diagnosis were detected by multinomial logistic regression and expressed as OR.Eighty-five patients (16 males, 69 females, aged 34.1 ± 12.5 years) were followed up for a mean period of 21.7 ± 11.7 mo. At baseline, endoscopy/duodenal biopsy, CD3 immunohistochemistry revealed:25 IELs/100 enterocytes in 22 subjects, 15-25 IELs in 37 and15 IELs in 26. They all had negative serum anti-transglutaminase and anti-endomysium, whilst 5 showed IgG anti-gliadin positivity. In the course of follow-up, 23 developed CD seropositivity and gluten sensitivity (GS) was identified in 19. Other diagnoses were: 5 Helicobacter pylori infections, 4 jejunal Crohn's disease, 1 lymphocytic colitis and 1 systemic sclerosis. The disease in the remaining 32 patients was classified as irritable bowel syndrome because of the lack of diagnostic evidence. At multivariate analysis, the evolution towards CD was associated with an IELs infiltrate25 (OR = 1640.4) or 15-25 (OR = 16.95), human leukocyte antigen (HLA) DQ2/8 (OR = 140.85) or DQA1*0501 (OR = 15.36), diarrhea (OR = 5.56) and weakness (OR = 11.57). GS was associated with IELs 15-25 (OR = 28.59), autoimmune thyroiditis (OR = 87.63), folate deficiency (OR = 48.53) and diarrhea (OR = 54.87).DL may have a multifactorial origin but the IELs infiltrate and HLA are strong predictive factors for CD development and a clinical diagnosis of GS.

Summary Background Helicobacter pylori clarithromycin resistance is increasing worldwide and different mutations are involved in its mechanisms. Recently, molecular methods have been proposed to assess these mutations. Aim To assess prevalence of primary clarithromycin resistance in two Italian areas, and the distribution of involved mutations, by using a novel method for real-time polymerase chain reaction. Methods Two hundred and thirty-two H. pylori-positive patients undergoing oesophagogastroduodenoscopy in two Italian towns (Rome, centre Italy; Foggia, south Italy) were enrolled. Helicobacter pylori infection was detected by histology, rapid urease and urea breath tests. Clarithromycin resistance was assessed by TaqMan real-time polymerase chain reaction on paraffin-embedded antral biopsies. Results Primary clarithromycin resistance was detected in 62 (26.7%) patients. Its prevalence did not differ between the two areas (31.5%, centre vs. 23.3%, south; P = 0.17) and between non-ulcer dyspepsia and peptic ulcer patients (28.4% vs. 20.7%, P = 0.2). The A2143G point mutation was detected in 35 (56.4%) patients, A2142G in 14 (22.6%), A2142C in eight (12.9%), whilst a double mutation (A2143G plus A2142C or A2142G) was present in the remaining five (8.1%) cases. Conclusions Our study found that primary clarithromycin resistance is highly prevalent in both central and southern Italy, and that A2143G is the most frequent point mutation involved in these areas.

Background & Aims: The currently recommended first-line eradication treatment of Helicobacter pylori in children is usually successful in about 75%. Recently, in adults, a novel 10-day sequential treatment has achieved an eradication rate of 95%. The aim of the study was to assess the H pylori eradication rate of the sequential treatment regimen compared with conventional triple therapy in children. Methods: Seventy-eight consecutive children with H pylori infection were randomized to receive either sequential treatment (omeprazole plus amoxicillin for 5 days, followed by omeprazole plus clarithromycin plus tinidazole for another 5 days) (n = 38; 15 boys [39.5%]; median age, 11.0 years [range, 3.3–16 years]) or triple therapy (omeprazole, amoxicillin, and metronidazole) for 1 week (n = 37; 15 boys [40.5%]; median age, 9.9 years [range, 4.3–16 years]). H pylori infection was based on 2 out of 3 positive tests results: 13C-urea breath test, rapid urease test, and histologic analysis. Eradication was assessed by 13C-urea breath test 8 weeks after therapy. Results: Seventy-four patients completed the study. H pylori eradication was achieved in 36 children receiving sequential treatment (97.3%; 95% confidence interval, 86.2–99.5) and 28 children receiving triple therapy (75.7%; 95% confidence interval, 59.8–86.7) (P 95%) in all. Conclusions: Our study shows, for the first time in children, that 10-day sequential treatment achieves a higher eradication rate than standard triple therapy, which is consistent with the results of adult studies.

Summary Background : A novel 10-day sequential treatment regimen recently achieved a significantly higher eradication rate than standard 7-day therapy in both peptic ulcer disease and non-ulcer dyspepsia. Its higher performance has recently been confirmed using a halved clarithromycin dose in peptic ulcer disease. Aims : To evaluate whether an acceptable eradication rate could also be obtained by halving the clarithromycin dose in dyspeptic patients and to assess the role of possible factors affecting the outcome of therapy. Methods : In a prospective, open-label study, 162 patients with non-ulcer dyspepsia and Helicobacter pylori infection, assessed by rapid urease test and histology, were enrolled. Patients were randomized to receive either 10-day sequential therapy, comprising rabeprazole 20 mg b.d. plus amoxicillin 1 g b.d. for the first 5 days, followed by rabeprazole 20 mg b.d., clarithromycin 250 mg b.d. and tinidazole 500 mg b.d. for the remaining 5 days (low-dose therapy), or a similar schedule with clarithromycin 500 mg b.d. (high-dose therapy). Four to six weeks after therapy, H. pylori eradication was assessed by endoscopy/histology. Results : A similar H. pylori eradication rate was observed following low- and high-dose regimens for both per protocol (94% vs. 95%; P = N.S.) and intention-to-treat (93% vs. 94%; P = N.S.) analyses. No major side-effects were reported. Halving the clarithromycin dose leads to a per patient saving in pharmaceutical costs of 24.6 euros. None of the variables examined affected the effectiveness of eradication of the sequential regimen. Conclusion : A reduction of the clarithromycin dose does not affect H. pylori eradication with the sequential regimen in non-ulcer dyspepsia and affords lower costs.

Association between Crohn's disease (CD) and lupus nephritis is very rare and, to the best of our knowledge, it has been described only once. We report here a clinical case of CD occurred in a young woman 8 years after a diagnosis of lupus nephritis according to clinical, laboratory and histological criteria. CD was unresponsive to steroids and immunosuppressants and, therefore, the patient was treated with anti-tumour necrosis factor alpha monoclonal antibody (Infliximab). This therapy led to the remission of both CD (50% of Crohn's Disease Activity Index--CDAI--decrease) and lupus nephritis (disappearance of pyuria in absence of infection, significant increase of serum albumin and improvement of renal function tests). The immunological background of both diseases has to be taken into account to explain either the association of the two disorders or the therapeutic response. Moreover, this clinical case confirms and extends the concept that in patients with CD a more accurate detection of autoimmune associated disorders is required.

Background/Aims : Alpha-interferon (α-IFN) and lamivudine are the two licensed drugs for patients with chronic hepatitis B, however, their efficacy in precore mutant chronic hepatitis B is limited. The aim of this study was to investigate the efficacy of 1 year α-IFN–lamivudine combination therapy for anti-HBe/hepatitis B virus- (HBV)-DNA positive patients. Methods : Between 1997 and 1999, 29 consecutive anti-HBe/HBV-DNA positive patients entered this prospective pilot study. Patients received 100mg lamivudine orally daily and α-IFN 6 million units (MU) three times weekly for 52 weeks. All patients were followed-up for 12 months after stopping therapy. Primary end points were loss of serum HBV-DNA and alanine transaminase normalization at week 52. Results : Overall, the end-treatment biochemical and virological response was 93% while the sustained response at week 104 was 14%. HBV-DNA negative patients did not experience a viral breakthrough during treatment; no tyrosine–methionine–aspartate–aspartate amino acid motif of HBV polymerase (YMDD) variant emerged. At week 52, 46% of patients with paired liver biopsies slides available, showed an histological improvement (histological activity index ≥2). Conclusions : Combination of lamivudine and interferon for 1 year is followed by high end-treatment virological and biochemical response rates, by improvement of liver histology and by the prevention of the emergence of YMDD mutation; however, the sustained response rate remains low.

INTRODUCTION: Autonomic neuropathy is thought to play a role in the pathogenesis of slow-transit constipation, but other gastrointestinal organs may also be involved, even if they are symptom-free. We investigated whether motility in gastrointestinal organs other than the colon was impaired in patients with slow-transit constipation and whether the autonomic nervous system was involved. METHODS: Twenty-one consecutive patients (18 females; median age, 46 years) with severe chronic constipation (≤2 defecations/week and delayed colonic transit time) were studied. Autonomic neuropathy function was tested with esophageal manometry, gastric and gallbladder emptying (fasting and postprandial motility) by ultrasonography, orocecal transit time (H2-breath test), colonic transit time (radiopaque markers), and anorectal volumetric manometry. The integrity of the autonomic nervous system was assessed by a quantitative sweat-spot test for preganglionic and postganglionic fibers, tilt-table test, and Valsalva electrocardiogram R-R ratio. RESULTS: Esophageal manometry showed gastroesophageal reflux or absence of peristalsis in five of the seven patients examined. Gallbladder dysmotility (i.e., increased fasting, postprandial residual volume, or both) was observed in 6 of 14 (43 percent) patients. Gastric emptying was decreased in 13 of 17 (76 percent) patients. Orocecal transit time was delayed in 18 of 20 (90 percent) patients; median transit time was 160 (range, 90–200) minutes. Median colonic transit time was 97 (range, 64–140) hours. Anorectal function showed abnormal rectoanal inhibitory reflex and decreased rectal sensitivity in 11 of 19 (58 percent) patients. Signs of autonomic neuropathy of the sympathetic cholinergic system were found in 14 of 18 (78 percent) patients. Only one of nine patients had vagal abnormalities detected with the Valsalva test and four of five patients with a history of orthostatic hypotension had a positive tilt-table test. CONCLUSIONS: Slow-transit constipation may be associated with impaired function of other gastrointestinal organs. More than 70 percent of patients with slow-transit constipation present some degree of autonomic neuropathy. Severe constipation may be the main complaint in patients with a systemic disease involving several organs and possibly involving the autonomic nervous system. This should be considered in the management of such cases.

With the aim of investigating a possible relationship between "objective" halitosis (established by sulfide levels in the breath) and Helicobacter pylori, we performed a study in 58 dyspeptic patients reported to suffer from "bad breath." Furthermore, we evaluated the effects on halitosis of eradication therapy (only for H. pylori-positive patients) and chlorhexidine antiseptic mouth rinses (in all patients). Sulfide compound assay indicated objective halitosis in 52/58 patients, 30 of whom were positive and 22 negative for H. pylori. In 19/30 eradication by double therapy provoked a decrease to below the cutoff value of sulfide levels in 15. In the other 11 of the 30 subjects, in whom H. pylori positivity persisted, halitosis parameters did not change. Chlorexidine reduced sulfides to below the cutoff value in 16/22 H. pylori-negative patients, but did not provoke any change in the 11 unsuccessfully treated H. pylori-positive subjects. In these, objective halitosis disappeared only after a successful eradication by triple therapy (9/11). Our results show a possible association between halitosis and H. pylori since bacterial eradication may resolve the symptom. Antiseptic mouthwashes may be effective only in absence of H. pylori, when halitosis may be due to oral putrefactive microbial activity. In a small number of subjects the cause and treatment of halitosis need to be clarified.

Helicobacter pylori (HP) infection is the main etiopathogenetic agent responsible for inflammatory and ulcerative changes in gastroduodenal mucosa and the basis for both intestinal and diffuse types of gastric carcinoma. In this latter case, intestinal metaplasia is the intermediary between gastritis and cancer. In this study we describe the proliferative activity of gastric epithelium in the progressive stages of HP infection. The expression of proliferating cell nuclear antigen (PCNA), which has proven to be a reliable method for this evaluation, was used as a marker. The study was performed on endoscopic biopsies of the gastric antrum of 40 patients, who were divided into five groups, eight in each group: normal histology and endoscopy, HP-; histological HP+ gastritis with normal endoscopy; histological HP+ gastritis with endoscopic evidence of chronic erosions; complete and incomplete intestinal metaplasia in a HP+ stomach. PCNA was detected by immunohistochemistry and expressed as labeling index, ie, percentage of positive nuclei either in the whole or upper third of foveolae. Our data show a progressive increase of epithelial proliferation in the successive stages of HP infection ranging from gastritis alone to the development of incomplete intestinal metaplasia, a well-known precancerous condition. The proliferative pattern tended to expand towards the upper foveolar third, which in normal conditions does not represent a site of epithelial renewal. These alterations may be related to the development of neoplastic transformations of gastric epithelium. It is well known that genetic mutations are facilitated in proliferating cells. Therefore, our results indicate that the high epithelial turnover, expressed by PCNA LI, may be an indicator of increased risk of neoplastic changes in long-standing untreated HP+ chronic gastritis.

AIM To evaluate mucosal baseline mRNA expression of tissue transglutaminase 2 (tTG2), interferon gamma (IFNγ), toll-like receptor 2 (TLR2) and Myeloid Differentiation factor 88 (MyD88) in patients with microscopic enteritis (ME). METHODS We retrospectively enrolled 89 patients with ME of different etiology, which was defined within a 2-year mean period of follow-up. Baseline histological examination was performed on Hematoxylin-Eosin stained sections and CD3 lymphocyte immunohistochemistry was used for intraepithelial lymphocyte count (IELs). ME was defined according to the criteria of Bucharest Consensus Conference. For each patient, formalin embedded biopsy samples of the duodenum referred to the period of ME diagnosis were retrieved. Real-time polymerase chain reaction (RT-PCR) was used to detect the amount of mRNA coding for tTG2, IFNγ, TLR2 and MyD88, and the quantity was expressed as fold change compared to controls. Control group was represented by duodenal normal specimens from 15 healthy subjects undergoing endoscopy for functional symptoms. Comparisons among continuous variables were performed by One way analysis of variance (ANOVA) and Bonferroni’s test. The χ2 test was used for categorical variables. Pearson’s test was used to evaluate correlations. Receiver operating curves were drawn for all four markers to estimate sensitivity and specificity in discriminating the development of CD and GS. RESULTS After a period of follow up of 21.7 ± 11.7 mo, the following diagnoses were achieved: gluten related disorders in 48 subjects (31 CD; 17 GS) and non-gluten related ones in 41 (29 Irritable Bowel Syndrome - IBS; 12 Others). CD patients had the highest tTG2 levels (8.3 ± 4.5). The ANOVA plus Bonferroni analysis showed that CD > Other ME > GS = IBS > negative controls. A cut off value of 2.258 was able to discriminate between CD and GS with a sensitivity of 52.94% and a specificity of 87.1%. Additionally, CD patients had the highest IFNγ levels (8.5 ± 4.1). ANOVA plus Bonferroni demonstrated CD > Other ME > GS = IBS > negative controls. A cut off of 1.853 was able to differentiate CD and GS with a sensitivity of 47.06% and a specificity of 96.77%. Patients with non gluten-related causes of ME exhibited the highest TLR2 levels (6.1 ± 1.9) as follows: Other ME > CD = GS = IBS > negative controls. TLR2 was unable to discriminate CD from GS. Patients with CD overexpressed MyD88 levels similarly to non gluten-related causes of DL (7.8 ± 4.9 and 6.7 ± 2.9), thus CD = Other ME > GS = IBS > negative controls. A cut off of 3.722 was able to differentiate CD from GS with a sensitivity of 52.94% and a specificity of 74.19%. IELs count (15-25 and more than 25/100 enterocytes) strongly correlated with mRNA levels of all tested molecules (P < 0.0001). CONCLUSION Our results confirm that a single marker is unable to predict a discrimination among ME underlying conditions as well as between CD and GS. Mucosal high levels of tTG and IFNγ mRNA may predict the development of CD more than GS with high specificity, despite an expected low sensitivity. TLR2 does not discriminate the development of CD from GS. MyD88 levels indicate that intestinal permeability is more increased when a severe intestinal damage underlies ME in both gluten related and unrelated conditions. Therefore, the results of the present paper do not seem to show a clear translational value.

The mammalian growth factor erv1-like (GFER) gene encodes a sulfhydryl oxidase enzyme, named Augmenter of Liver Regeneration (ALR). Recently it has been demonstrated that ALR supports cell proliferation acting as an anti-apoptotic factor. This effect is determined by ALR ability to support the anti-apoptotic gene expression and to preserve cellular normoxic conditions. We recently demonstrated that the addition of recombinant ALR (rALR) in the culture medium of H(2)O(2)-treated neuroblastoma cells reduces the lethal effects induced by the hydrogen peroxide. Similar data have been reported in the regenerating liver tissue from partially hepatectomized rats treated with rALR. The purpose of the present study was to evaluate the effect of the GFER inhibition, via the degradation of the complementary mRNA by the specific siRNA, on the behaviour of the apoptosis (apoptotic gene and caspase expression and apoptotic cell number) and of the oxidative stress-induced parameters (reactive oxygen species (ROS), clusterin expression and mitochondrial integrity) in T98G glioma cells. The results revealed a reduction of (i) ALR, (ii) clusterin and (iii) bcl-2 and an increase of (iv) caspase-9, activated caspase-3, ROS, apoptotic cell number and mitochondrial degeneration. These data confirm the anti-apoptotic role of ALR and its anti-oxidative properties, and shed some light on the molecular pathways through which ALR modulates its biological effects.

Anisakis simplex is a parasite that, if present in uncooked and contaminated saltwater fish, can invade the human gut. Two different clinical situations are recognized: the first, known as a gastrointestinal disease, varying from an asymptomatic episode to vomiting and diarrhea, and the second, classified as an adverse reaction to food, characterized by a wide spectrum of allergic reactions like rhinitis, conjunctivitis, or even anaphylaxis causing hypotension and/or shock. The intestinal epithelium, the major defense system against external molecules, represents an open gate for toxins and allergens if its protective function is compromised. Previous data have demonstrated a strict relationship between an altered intestinal permeability (I.P.) and worsening of the clinical manifestations in patients with adverse reactions to the food. In this article we evaluated the sensitization to A. simplex among patients who referred clinical symptoms of allergy. All subjects underwent commonly used alimentary skin prick test for food allergens, to which Ani s1, an A. simplex allergen, was added. In addition, in A. simplex-sensitized subjects, I.P. was determined upon their enrolment to the study (time 0) and after 6 months of consuming a raw fish-free diet (time 6). Five hundred and forty subjects were screened, and 170 had a positive skin prick test, 87 (51.2%) of whom were positive to Ani s1. Increased I.P. was evidenced in A. simplex-sensitized subjects with worse clinical symptoms, which receded after 6 months' elimination of raw seafood. With our data we demonstrated that the alimentary habit to eat raw fish represents a high risk for the integrity of the intestinal mucosa, and we suggest that this pathological situation may constitute an ideal, under-estimated, open gate for molecules that predispose to other, more important pathologies.

We describe the cases of two patients with Crohn's disease affected by severe perineal fistulae resistant to conventional therapies, successfully treated with FK 506, a new immunomodulatory drug. It is well absorbed from diseased bowel and preliminary experiences have indicated its short-term use in complicated Crohn's disease. The first patient was a 24-year-old male with perineal fistula and severe skin ulceration (8 cm of external opening diameter). He had undergone colectomy and ileostomy because of severe pancolitis refractory to medical treatment and had been treated with azathioprine and metronidazole. Two months after starting FK 506, a dramatic improvement made further surgical operation unnecessary. Local and general benefit was observed during the following 26 months, until FK 506 was withdrawn. The second patient was a 28-year-old male with a diagnosis of ulcerative pancolitis changed to Crohn's disease two months after the onset of a perineal fistula, recurring despite drainage procedures, steroid therapy, and total parenteral nutrition. FK 506 was administered for two months with a complete healing of fistula. Successively, it was stopped and corticosteroids (associated to enteral nutrition) were given because of recurrent rectal bleeding. Our experience encourages the use of oral FK 506 in complicated Crohn's disease and suggests the possibility of a long-term primary therapy other than the use as a "bridge" to other treatments.

Data in the literature suggest the possibility of using ultrasonography to diagnose sliding gastric hiatal hernia. The aim of this study was to confirm the diagnostic accuracy of transabdominal ultrasonography for the diagnosis of sliding gastric hiatal hernia, using endoscopy as the reference test. The latter procedure was used since it also recognizes the presence of esophagitis and/or varices, two pathological conditions that could per se influence ultrasonographic evaluation, based on measurement of the esophageal diameter.A total of 180 consecutive patients, admitted to our hospital for endoscopy, were examined. Of these, 12 patients were not included in the final study. After fasting, the remaining 168 patients were first evaluated by ultrasonography and later by endoscopy. Esophageal diameter was measured by ultrasonography at the level of the diaphragmatic hiatus.Using a diameteror = 18 mm as the sensitivity threshold, 29 patients potentially affected by hiatal hernia were identified by ultrasonography. Upper endoscopy examination confirmed the presence of hiatal hernia in 24 of these patients and documented 4 additional hernias in the group of patients with a transdiaphragmatic esophageal diameter18 mm (positive and negative predictive values were 82.7% and 97%, respectively). Analysis of the distribution of esophageal varices and esophagitis in all 168 excluded the possibility that these pathological conditions could influence the esophageal diameter.Ultrasonography represents a simple and well-tolerated diagnostic approach in sliding gastric hiatal hernia. The good diagnostic accuracy suggests its potential use in clinical and epidemiological settings.

Local immunosuppression within the liver and sex steroid changes, in both blood and tissue during liver regeneration, are well-recognized events. Dendritic cells (DC) play pivotal roles in the induction and regulation of immune responses. Their numbers are expanded markedly in vivo by fms-like tyrosine kinase 3 ligand (Flt3L) administration, without modification of their maturation state. Recent evidence suggests that estrogen can modulate DC function and promote a Th2-type immune response. Few data are available concerning the role of DC in liver regeneration. After 75% partial hepatectomy (PH) in male C57BL/6 mice, CD11c+ liver (L)DC increased significantly within 6 hours and maintained an immature phenotype. Numbers returned to pre-hepatectomy levels by 24 hours. The expanded LDC population showed increased IL-10 and reduced IFN-gamma gene transcription. Using these DC compared with control LDC as T cell stimulators in 72-hour mixed leukocyte cultures, IL-10 production was enhanced and IFN-gamma production reduced. LDC isolated 6 hours after 75% PH exhibited enhanced estrogen receptor (ER) expression, concomitant with increased serum estrogen levels. By contrast, spleen (S)DC isolated before and after PH showed no significant changes in their function (maturation state, T cell stimulatory activity, cytokine production, and ER expression). Increased liver regeneration (more than 50%) was observed 48 hours after 40% PH in the Flt3L-pretreated compared with the PBS group. In conclusion, interstitial LDC may play a key role in local immune regulation during liver regeneration, possibly linking estrogen-mediated immune modulation and hepatocyte proliferation.

Carpagnano GE, Resta O, Ventura MT,Amoruso AC, Di Gioia G, Giliberti T, Refolo L,Foschino-Barbaro MP (University of Foggia; andUniversity of Bari; Italy). Airway inflammationin subjects with gastro-oesophageal reflux andgastro-oesophageal reflux-related asthma. J InternMed 2006; 259: 323–331.Study objectives. Asthma and gastro-oesophagealreflux (GER) are both characterized by airwayinflammation.Design. The purposes of this work were (i) to studyairway inflammation in patients troubled by gastro-oesophageal reflux (GER) and GER associated withasthma, (ii) to ascertain whether GER can aggravateasthma by exacerbating the pre-existing airwayinflammation and oxidative stress and (iii) toestablish the validity of analysing breathcondensate and induced sputum when studyingthe airways of subjects affected by GER.Patients and methods. We enrolled 14 patientsaffected by mild asthma associated with GER (40 ±12 years), nine with mild but persistent asthma(39 ± 13 years), eight with GER (35 ± 11 years)and 17 healthy subjects (37 ± 9 years). Sputum cellcounts andconcentrationsofinterleukin-4 (IL-4), IL-6 and 8-isoprostane were measured in breathcondensate and supernatant.Measurements and results. GER-related asthma ischaracterized by an eosinophilic inflammation, asdetermined by elevated concentrations of IL-4 inbreath condensate and sputum supernatant, and bysputum cell analysis. GER alone presents aneutrophilic pattern of inflammation whendetermined by elevated concentrations of IL-6 insputum cell analysis. A concomitant increase hasbeen found in 8-isoprostane in GER associated (ornot associated) with asthma.Conclusions. We conclude that GER is characterizedby a neutrophilic airway inflammation and byincreased oxidative stress. GER does not howeveraggravate pre-existing airway inflammation inasthma patients. Determinations of inflammatoryand oxidant markers in the breath condensate ofsubjects with GER reflect these measured in theinduced sputum.Keywords: asthma, breath condensate, gastro-oesophageal reflux, induced sputum, airwayinflammation, oxidative stress.

Background An abnormal intestinal permeability could contribute to establish an altered sensitivity to food-allergen. Aim To evaluate the intestinal permeability in subjects with adverse reactions to food on allergen-free diet. Subjects Twenty-one patients with food allergy and 20 with food hypersensitivity on allergen-free diet were enrolled and divided in four groups according to the seriousness of their referred clinical symptoms when they were on a free diet. Methods Intestinal permeability was evaluated by Lactulose/Mannitol ratio urinary detection determined by anion-exchange chromatography. Results Statistically significant different Lactulose/Mannitol ratio was evidenced in subjects with food allergy (p = 0.003) or hypersensitivity (p = 0.0008) compared to control patients. The correlation between Lactulose/Mannitol ratio and the seriousness of clinical symptoms, by using Spearman test, was statistically significant for food allergy (p = 0.0195) and hypersensitivity (p = 0.005) patients. Conclusions The present data demonstrate that impaired intestinal permeability, measured in our conditions, is present in all subjects with adverse reactions to food. In addition, for the first time, we report a statistically significant association between the severity of referred clinical symptoms and the increasing of Intestinal Permeability Index. These data reveal that intestinal permeability is not strictly dependent on IgE-mediated processes but could better be related to other mechanisms involved in early food sensitisation, as breast-feeding, or microbial environment that influence the development of oral tolerance in early infancy.

Background: This is a report of the results of a multicenter study performed in children with dyspepsia from five pediatric centers in Puglia, a region in southern Italy. In the study, clinical features of Helicobacter pylori infection, the reliability of diagnostic techniques, and the involvement of bacterial strains were examined. Methods: Fifty-three outpatients with dyspepsia enrolled in our study and compiled a diary recording clinical symptoms in patients before they underwent the following diagnostic techniques : endoscopy, biopsy for histologic analysis, rapid urease test, 13 C urea breath test, serology specific for immunoglobulin (Ig) G and anti-CagA and VacA. Results: H. pylori showed a prevalence of 30.2% (n = 16). Histologic positivity was seen in all patients at the antral level (H. pylori-associated chronic gastritis). In the gastric body, bacterial chronic active gastritis was present only in six patients (H. pylori-associated chronic pangastritis). Clinical evaluation showed a significant difference in favor of subjects positive for H. pylori only for epigastric burning and/or pain (p < 0.001). The comparison of results of diagnostic tests, using histology as the gold standard, showed sensitivity and specificity of more than 93% for 13 C urea breath test and more than 85% for rapid urease test and serology. Anti-CagA antibodies were found in 64.3% and anti-VacA antibodies in 42.8% of H. pylori-positive patients. Conclusions: H. pylori prevalence in children with dyspepsia from the geographic area studied is comparable with that found in other developed countries. Approximately 50% of the studied patients were infected by cytotoxic strains. The urea breath test was the most reliable noninvasive diagnostic tool and is suitable for routine use, although endoscopy with histologic assessment remains the definitive investigation and is particularly important in patients with positive serology for CagA and VacA. Finally, the frequency of aggressive strains in our region seems to affect the clinical pattern; this emphasizes the importance of definitive diagnosis in children and offers a new role for serology.

Estrogen (E) and epidermal growth factors (EGF) receptors were assayed in the liver of nine patients with hepatocellular carcinoma (HCC). Total E and nuclear E receptors were decreased significantly in neoplastic tissue as compared to the levels found in surrounding nonneoplastic tissue. The EGF receptor was decreased also in neoplastic tissue. On the basis of binding data, a decrease in the number but not in affinity of both the E and EGF receptors was found.

(WHO-Grade 3-4). 31 patients died because of their basic disease. Tumor response was in total: complete response (n=l), partial response (n=l l), stable disease (n=70), progressive disease (n=30). Discussion: According to our prospective selection criteria palliative therapy in patients with inoperable HCC is safe and showing low rates of toxicity. In terms of low side effects and tumor response our selection criteria appears to be acceptable.

Background/Aims : Alpha-interferon (α-IFN) and lamivudine are the two licensed drugs for patients with chronic hepatitis B, however, their efficacy in precore mutant chronic hepatitis B is limited. The aim of this study was to investigate the efficacy of 1 year α-IFN–lamivudine combination therapy for anti-HBe/hepatitis B virus- (HBV)-DNA positive patients. Methods : Between 1997 and 1999, 29 consecutive anti-HBe/HBV-DNA positive patients entered this prospective pilot study. Patients received 100mg lamivudine orally daily and α-IFN 6 million units (MU) three times weekly for 52 weeks. All patients were followed-up for 12 months after stopping therapy. Primary end points were loss of serum HBV-DNA and alanine transaminase normalization at week 52. Results : Overall, the end-treatment biochemical and virological response was 93% while the sustained response at week 104 was 14%. HBV-DNA negative patients did not experience a viral breakthrough during treatment; no tyrosine–methionine–aspartate–aspartate amino acid motif of HBV polymerase (YMDD) variant emerged. At week 52, 46% of patients with paired liver biopsies slides available, showed an histological improvement (histological activity index ≥2). Conclusions : Combination of lamivudine and interferon for 1 year is followed by high end-treatment virological and biochemical response rates, by improvement of liver histology and by the prevention of the emergence of YMDD mutation; however, the sustained response rate remains low.

Recently reports have indicated that both cimetidine and ranitidine delay cell proliferation in rats following 70% partial hepatectomy and result in an increased mortality following this procedure. The present study was designed to determine whether three H2 blocking agents (cimetidine, ranitidine, famotidine) and a new, powerful antisecretory drug (omeprazole) specifically influence hepatocyte proliferation in primary culture. Hepatocytes were isolated from livers of normal male rats by the standard collagenase perfusion technique. Hepatic DNA synthesis and percent of labelled nuclei were determined after 48 h incubation. Hepatocytes in culture were incubated with the H2 blocking agents and omeprazole or with different concentrations of serum obtained from sham-operated or 70% hepatectomized rats treated or not with the same agents. Rats were injected intraperitoneally at 8:00 a.m. on two consecutive days. In hepatectomized rats, the first dose was injected at 8:00 a.m. immediately after surgery, the second, 24 h later. The serum of sham-operated or 70% hepatectomized rats that did not receive drugs served as control. No changes in DNA synthesis, percentage of labelled nuclei and transaminase were detected when the agents were added to the hepatocytes in culture at concentrations within the effective pharmacological dosage and 30 times higher. Similarly, no changes in these parameters were obtained when different concentrations of serum obtained from sham-operated rats treated with H2 blocking agents or omeprazole were added to the basal culture medium. However, a significant inhibition of DNA synthesis and of percentage of labelled nuclei was observed when hepatocytes were incubated in the presence of serum from 70% hepatectomized rats that had been treated with cimetidine or with ranitidine. The serum of 70% hepatectomized rats treated with famotidine and omeprazole had no effect on hepatocyte proliferation in vitro. No effect on transaminase was found in these conditions.

Cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP) were assayed in ascitic fluid from 27 patients with ovarian carcinoma and 23 patients with liver cirrhosis. The value of these cyclic nucleotides was correlated with standard methods for the clinical evaluation of tumors. No change in the cGMP levels was found in either of these groups. The cAMP content, however, was increased in 23 of the 27 cases of ovarian carcinoma. The high cAMP level was correlated with the cytological findings in only 13 (48.1%) of these cases.