Mesalazine vanishing time from rectal mucosa following its topical administration.

To investigate how long and how much Mesalazine (M) is available inside the rectal mucosa following its topical instillation, in patients (pts) with Ulcerative Colitis (UC). Two rectal biopsies for M concentration were obtained from 45 UC pts in clinical remission and on oral M treatment (OT), before a 4g enema randomly given to consentient pts every day (Group A, 15 pts), every 2 days (Group B, 15 pts) and every 3 days (Group C, 15 pts). Two additional biopsies were taken 1, 2 and 3 days after the last enema in group A, B and C respectively, at least 10 days later. All biopsies were immediately frozen at -80°C for later assay by means of high-performance light chromatography (HPLC). Data were analyzed using Student's t-test. Mean values±standard deviation of M mucosal concentration (ng/mg of tissue) were 1.32±1.41, 56.1±39.2, 9.65±6.60, and 6.39±5.03 in pts receiving OT alone, groups A, B and C, respectively. Values in Group A were statistically higher (p<0.001) than those in Groups B and C while no differences were found between Groups B and C. Values of OT were lower than groups A, B and C. M mucosal concentration rapidly decreases 2 days after a 4g enema, but after three days is still higher than OT alone. These results may provide data which would be useful to plan topical therapy and improve adherence to treatment.

Long-term oral plus topical mesalazine in frequently relapsing ulcerative colitis.

BACKGROUND: In cross-sectional studies, it was demonstrated that the therapeutic effect of mesalazine is closely related to its mucosal concentration. AIM: This study was carried out to verify in a longitudinal study if it was possible to improve the clinical course of ulcerative colitis at high risk of recurrence by increasing mucosal mesalazine concentration. METHODS: Eighteen consecutive ulcerative colitis patients on continuous oral 5-ASA treatment (2.4-3.2 g/day) in clinical remission who had had at least four moderate to severe relapses in the preceding 2 years (referred period) were assigned to assume oral (3.2-4.8 g/day) and topical (4 g/day) mesalazine in order to increase mucosal drug concentration and were followed up for 2 years (study period). The localisation of disease was 12 pancolitis, six left colitis. The number and severity of recurrences, number of visits and endoscopies, courses of steroids and days of hospitalisation were compared with those of the previous 2 years. Rank signed test for paired data was used for statistical analysis. RESULTS: The total number of recurrences was significantly lower during the study period in comparison with that of referred period (8 versus 80, respectively, p < 0.0001). No courses of steroids or hospitalisation were necessary during study period in comparison with those of referred period (0 versus 33, p < 0.0001; 0 versus 93, p = 0.03, respectively). A total number of 249 visits were done during the referred period and 116 during the study period (p < 0.0001) with a total of 87 endoscopies during referred period and 44 during study period (p < 0.0001). CONCLUSIONS: The continuous use of topical mesalazine associated with a high oral dosage significantly improves the clinical course of ulcerative colitis patients at high risk of relapse.

Infliximab in refractory pouchitis complicated by fistulae following ileo-anal pouch for ulcerative colitis.

AIM: To determine the efficacy of infliximab in the treatment of chronic refractory pouchitis complicated by fistulae following ileal pouch-anal anastomosis for ulcerative colitis. METHODS: This open study included seven patients (four females, three males) with chronic refractory pouchitis complicated by fistulae. Pouchitis was diagnosed by clinical, endoscopic and histological criteria. The sites of the fistulae were as follows: pouch-bladder in one, vaginal in three, perianal in two, and both vaginal and perianal in one. Extra-intestinal manifestations (erythema nodosum, arthralgia) were present in four patients. Crohn's disease was carefully excluded in all patients after re-evaluation of the history, re-examination of the original proctocolectomy specimen and examination of the proximal small bowel. All patients had been treated with antibiotics and three with steroids. Patients received infliximab, 5 mg/kg, at 0, 2 and 6 weeks. Azathioprine (2.5 mg/kg) was also started for all patients as bridge therapy. Clinical response was classified as complete, partial or no response. Fistulae closure was classified as complete (cessation of fistulae drainage and total closure of all fistulae), partial (a reduction in the number, size, drainage or discomfort associated with fistulae) or no closure. The pouchitis disease activity index and quality of life were also used as outcome measures. RESULTS: Clinically, all patients improved. At the 10-week follow-up, six of the seven patients had a complete clinical response, and five had complete fistulae closure. At the 10-week follow-up, the median pouchitis disease activity index decreased from 12 (baseline) (range, 10-15) to 5 (range, 3-8); the median quality of life decreased from 37 points (range, 33-40) to 14 (range, 9-18). Erythema nodosum and arthralgia showed complete remission soon after the first infusion of infliximab. CONCLUSIONS: These preliminary results indicate that infliximab may be recommended for the treatment of refractory pouchitis complicated by fistulae following ileal pouch-anal anastomosis for ulcerative colitis.

N-Butyrate rectal transport in cirrhotic patients.

Data about colonic mucosa transport of short-chain fatty acids in cirrhotic patients are still lacking. The aim of the present study was to compare the rectal mucosa transport of n-butyrate and its effect on transport of other electrolytes and endoluminal pH in normal subjects and in cirrhotic patients by using a rectal dialysis technique. Thirteen subjects with normal hepatic function tests and 17 cirrhotic patients were enrolled. Dialysis bags containing 80 mmol/liter of butyrate in a neutral pH (6.8) electrolyte solution were placed in the rectum of enrolled subjects for 60 min. Net transport rate was calculated for butyrate, sodium, chloride, potassium, and bicarbonate. The differences in pH between initial and final dialysis solutions was also evaluated in the two groups in the study. Net butyrate absorption was significantly lower in cirrhotic patients than in controls (65.2 +/- 38.6 vs 101.2 +/- 45.3 nmol/min/cm2, respectively; P = 0.02). Furthermore, cirrhotic patients showed a lower HCO3 secretion than controls (-26.9 +/- 19.9 vs -45.1 +/- 20.0, respectively; P = 0.01). No differences were found in transport of the other electrolytes. The pH in the final dialysis solution in cirrhotic patients was not significantly lower than in the controls (7.15 vs 7.35; P = 0.1). In conclusion, the impairment of butyrate absorption and the concurrent reduction of bicarbonate secretion observed in cirrhotic patients may suggest a selective hypoactivity of apical HCO3-/SCFA- antiport located at the colonocyte apical membrane.

Mucosal 5-aminosalicylic acid concentration inversely correlates with severity of colonic inflammation in patients with ulcerative colitis.

BACKGROUND AND AIM: The treatment of ulcerative colitis (UC) with 5-aminosalicylic acid (5-ASA) does not have the same therapeutic effect in all patients. We tested the hypothesis that the effectiveness of the drug is related to its mucosal concentration. PATIENTS: Twenty one UC patients receiving oral 5-ASA (2.4-3.2 g/day) were enrolled in the study. Four were also receiving topical treatment (2 g/day). METHODS: Six endoscopic biopsies were taken from the rectum for measurement of 5-ASA concentrations (ng/mg) by HPLC; soluble interleukin 2 receptor (sIL-2R) concentrations (U/ml) were measured by ELISA and histology. Endoscopic and histological appearance was graded on a four point scale (0-3). The Wilcoxon's rank test and Pearson's correlation coefficient were used for statistical analysis. RESULTS: Mucosal concentrations of 5-ASA were significantly higher (p=0.03) in patients with endoscopic scores of 0-1 compared with those with scores of 2-3 (16.1 (range 10.2-45) v 5. 5 (3.5-17.4), respectively) and in patients with lower histological inflammation compared with those with more severe scores (17.4 (10. 5-45) v 8.9 (3.5-17.2), respectively) (p<0.01). In contrast, mucosal sIL2-R concentrations were significantly lower in patients with slight endoscopic and histological lesions than in those with more severe disease. A significative inverse correlation (r=-0.85) was found between 5-ASA and sIL-2R mucosal concentrations (p=0.00008). CONCLUSIONS: In patients with UC, in the same area of the intestinal tract, we found that the higher the 5-ASA mucosal concentrations, the lower the IL-2R levels and endoscopic and histological scores. We hypothesise that maintenance of high mucosal 5-ASA concentrations in all colonic segments could contribute to improve clinical outcome in UC patients.

Anastomotic configuration and mucosal 5-aminosalicyclic acid (5-ASA) concentrations in patients with Crohn's disease: a GISC study. Gruppo Italiano per lo Studio del Colon e del Retto.

OBJECTIVE: Recurrence of Crohn's disease quite inevitably occurs after resection of distal small bowel and proximal colon, involving the neoterminal ileum close to the anastomosis. Oral 5-aminosalicylic acid (5-ASA) administered soon after surgery delays recurrence and reduces its severity. We recently observed that in operated patients submitted to prophylactic treatment with oral 5-ASA the rate of recurrence was significantly higher in those with end-to-end anastomosis than in those with other types of anastomosis (end-to-side, side-to-side). The hypothesis investigated in the present study was that patients with end-to-side or side-to-side anastomosis would benefit from a higher mucosal concentration of 5-ASA with respect to patients with end-to-end anastomosis. Therefore, the mucosal 5-ASA concentration was measured in the perianastomotic area of both groups. METHODS: The study was carried out in 19 patients submitted to radical surgery for Crohn's ileitis or ileocolitis, under oral prophylactic treatment with 5-ASA (Asacol). All patients were on regular endoscopic follow-up and were free of recurrence. Two biopsies were collected 3 cm from the anastomosis, in the neoterminal ileum, and two biopsies were collected at the colonic site 3 cm below the anastomosis. 5-ASA concentrations (ng/mg) were measured in tissue homogenates by high-performance liquid chromatography (HPLC) with electrochemical detection. RESULTS: The mucosal concentration of 5-ASA in the neoterminal ileum was significantly lower in patients with end-to-end anastomosis than in those with other types of anastomosis (median values: 29.4 ng/mg vs 92.9 ng/mg respectively; p < 0.001). Six of 10 patients (60%) with end-to-end anastomosis, but none of the nine patients with other types of anastomosis, showed 5-ASA mucosal concentrations <40 ng/mg at the neoterminal ileum. On the contrary, no patients with end-to-end anastomosis showed mucosal concentrations of 5-ASA >90 ng/mg, compared with the 57% of patients in the group with other types of anastomosis. No differences were observed for colonic biopsies. CONCLUSIONS: The different mucosal concentrations in these two groups may be explained by the difference in segmental transit time induced by the different anastomotic configurations. A slower preanastomotic transit time, demonstrated in patients with end-to-side or side-to-side anastomosis, could offer a prolonged contact time between the intestinal content and the mucosa, resulting in an increase in drug absorption.

Rectal and colonic mesalazine concentration in ulcerative colitis: oral vs. oral plus topical treatment.

AIM: To measure mucosal concentrations of mesalazine in ulcerative colitis patients treated with oral mesalazine alone, compared to patients treated with both topical and oral mesalazine. METHODS: Twenty-two patients with mild to moderate ulcerative colitis were randomized to receive 2.4 g/day of oral mesalazine (11 patients) or 2.4 g/day oral plus 4 g/day of topical mesalazine (11 patients). After 2 weeks of treatment, endoscopic biopsies specimens were taken from the rectum and in descending colon just distal of the splenic flexure and stored to -80 degrees C for later assay (HPLC). Wilcoxon's rank sum test for unpaired data was used for the statistical analysis. RESULTS: Mucosal levels of mesalazine in the rectum were significantly higher in patients who received oral plus topical treatment than in those who had oral treatment alone (52.1 ng/mg, range: 13.6-122.1 vs. 0.2 ng/mg, range: 0.2-9.7, respectively; P < 0.0001). Similarly, in the descending colon, the mucosal concentrations of mesalazine were significantly higher in patients who had oral plus topical treatment than in those receiving oral treatment alone (46.6 ng/mg, range: 6-112.6 vs. 15.9 ng/mg, range: 2.3-42.4, respectively; P=0.01). CONCLUSIONS: Topical treatment of mesalazine significantly increases mucosal concentrations of mesalazine up to the splenic flexure, supporting the rationale to treat left-sided ulcerative colitis with topical formulations of mesalazine.

Prevention of post-operative recurrence of Crohn's disease requires adequate mucosal concentration of mesalazine. Gruppo Italiano per lo Studio del Colon e del Retto.

BACKGROUND: Surgical resection of Crohn's disease is followed by early recurrence in a high percentage of patients. Mesalazine has been shown to be effective in the prevention of post-operative recurrence, but some 50% of patients under treatment recur at 3 years of follow-up. AIM: To establish whether the mucosal concentration of mesalazine might affect the development of post-operative recurrence. METHODS: Colon-ileoscopy was performed in 25 consecutive patients resected for Crohn's disease. The mean time from surgery was 14 months. After the operation, all patients were taking oral mesalazine (Asacol, 2.4 g/day). Ten patients showed signs of endoscopic recurrence (apthae, ulcers, narrowing of the lumen) in the neoterminal ileum, five of whom also showed juxta-anastomotic colonic involvement. Fifteen patients were free of recurrence. At endoscopy, four biopsies were taken from the perianastomotic area (two specimens at the ileal site and two specimens at the colonic site of the anastomosis). The specimens were weighed and immediately frozen at -80 degrees C. Mesalazine concentration (ng/mg) was measured in tissue homogenates by high- performance liquid chromatography with electrochemical detection. Fisher's exact test was used for the statistical analysis. RESULTS: The mean value of mucosal mesalazine concentration, expressed as ng/mg of tissue, was significantly lower in patients with recurrence than in those without recurrence both in the ileum (mean +/- s.d.: 21.6+/-28.3 vs. 70.9+/-47.4; P = 0.007) and in the colon (25.8+/-26.4 vs. 60.3+/-32.5; P = 0.010). CONCLUSIONS: The mucosal concentration of mesalazine in the juxta-anastomatic area is significantly lower in patients with recurrence than in those free of recurrence. These data could suggest an association between mucosal mesalazine concentrations and the clinical effectiveness of the drug.

Circulating soluble factor-inhibiting natural killer (NK) activity of fresh peripheral blood mononuclear cells (PBMC) from inflammatory bowel disease (IBD) patients.

This study was performed in order to assess the cytotoxic activity, both natural (NK) and antibody-dependent (ADCC), of PBMC from 38 IBD patients and correlate it with their clinical features. Cytotoxicity assays were performed using sensitive target cells for NK and ADCC activities. In some experiments, highly purified NK cells, obtained both by Percoll density gradient and by co-culturing non-adherent PBMC with RPMI 8866 feeder cells, were used as effector cells. Furthermore, we evaluated NK cell parameters such as number, surface expression of adhesion molecules (CD11a/CD18, CD49d and CD54) and response to different stimuli. We observed a decreased NK cytotoxicity of PBMC from IBD patients, both in ulcerative colitis (UC) and Crohn's disease (CD), independently of the clinical activity of disease. In contrast, the ADCC lytic activity was within normal range. The lower NK cytotoxic activity observed in our IBD patients cannot be related to a decreased number of NK cells, surface expression of adhesion molecules, defective response to IL-2 and maturative defect. Decreased NK activity was induced in PBMC of controls when serum of patients was added and this was unrelated to monocyte-derived modulating factor(s). Our data show a decreased natural killing by fresh PBMC from IBD patients. This lower activity seems to be unrelated to a primary NK cell defect, since purified NK cells exhibited normal levels of killing. It might be hypothesized that serum factors, possibly derived from lymphocytes, with inhibitory properties on NK activity, might be functionally active in the blood of IBD patients, thus modulating NK activity.

Effects of cisapride on abnormally prolonged endogastric alkalinity time and delayed gastric emptying in cirrhotic patients.

BACKGROUND/AIMS: An abnormally prolonged alkalinity time in endogastric long-term pH monitoring has been previously demonstrated in cirrhotic patients. Recently a growing body of evidence suggest the existence of severe abnormalities of gastric emptying in the same patients and more generally of gastrointestinal motility changes in many portal hypertensive animal models. Assuming that there was a good correlation between endogastric alkalinity and a delayed emptying of gastric bile reflux, the aim of this study was to evaluated the effect of a gastrokinetic drug, Cisapride, both on circadian gastric pH and on gastric emptying in cirrhotic patients compared with controls. MATERIALS AND METHODS: Ten in patients with chronic liver disease and portal hypertension (six males, four females, median age 49.5, range 28-59) were enrolled. The control group included twelve inpatients without cirrhosis (seven females and five males, average age 45 years, range 31-54) free from endoscopic esophagogastro-duodenal lesions. The subjects were submitted to a 24h endogastric pH monitoring and gastric emptying study before and after administration of Cisapride (10 mg tid for three days). To gastric emptying study we used an ultrasonographic method evaluating the ratio between the antropyloric region volume before and at a fixed time after a solid/liquid standard meal. RESULTS: Basal 24 h gastric pH monitoring in cirrhotic patients showed a significant prolonged time of alkalinity (pH conventionally over 4) during the entire registration and mainly in postprandial period vs controls. The same patients group showed also a delayed gastric emptying when compared to controls. Cisapride administration significantly reduced both the abnormally prolonged alkalinity time and delayed gastric emptying in cirrhotic group without affecting the same parameters in the control group. CONCLUSIONS: Cisapride significantly reduces both the delayed gastric emptying time and the abnormally prolonged alkalinity time in cirrhotic group. Taken together, the results offered an indirect evidence that abnormally prolonged alkalinity in cirrhotic patients may be due, at least in part, to changes in gastroduodenal motility leading to a reduced gastric clearance of potentially noxious duodeno-gastric alkaline reflux.

Simultaneous determination of 5-aminosalicylic acid, acetyl-5-aminosalicylic acid and 2,5-dihydroxybenzoic acid in endoscopic intestinal biopsy samples in humans by high-performance liquid chromatography with electrochemical detection.

A high-performance liquid chromatographic method for the simultaneous determination of 5-aminosalicylic acid (5-ASA), acetyl-5-aminosalicylic acid (Ac-5-ASA) and 2,5-dihydroxybenzoic acid (5-HSA) in human endoscopic intestinal biopsy with electrochemical detection has been developed and validated. A liquid-liquid extraction procedure was used to isolate these drugs from the biological material prior to analysis. The compounds were separated on an Erbasil S reversed-phase column using methanol-critic acid-sodium hydrogenphosphate-heptane-sulfonic acid-disodium ethylenediaminetetraacetate (pH 3) as mobile phase. The method was linear from 1.0 to 300 ng ml-1 for 5-ASA, from 10 to 1000 ng ml-1 for Ac-5ASA and from 0.1 to 10 ng m-1 for 5-HSA. The limit of detection for 5-ASA and for Ac-5-ASA was 1 ng ml-1 and that for 5-HSA was 0.1 ng ml-1. This procedure is suitable for pharmacological and clinical studies of 5-ASA.

Study of lipid changes in freeze-dried fish during storage. I. The interaction of relative humidity and tissue lipids.

Several studies on lipid stability of freeze-dried fish pointed out the relationship between relative humidity (RH) and autoxidation of highly unsaturated fatty acids. The present study shows the changes of tissue lipids in two different fish, freeze-dried and stored under various RH conditions. Fillets of Scomber scomber L and Sardina pilchardus sardina were freeze-dried and stored for 1, 3, 6 and 9 months. periodically, the sample of both fish were analyzed to evaluate the autoxidation of lipids. The results show that the autoxidation of lipids is quicker during 1st-3rd month of storage, while in a high humidity environment the tissue lipids change slower. According to these results both time of storage and relative humidity must be controlled to maintain the nutritive value of freeze-dried fish.