Search PubMed⌕ Search

Biomedical subjects

S Ghosh

Publications and source records attributed to S Ghosh.

At least 199 records · Page 11Linked to original sources

Identification of canine helper T-cell epitopes from the fusion protein of canine distemper virus.

The fusion protein of canine distemper virus (CDV-F), a 662 amino-acid envelope protein, was used as the target molecule for identification of canine T helper (Th) epitopes. A library of 94 peptides, each 17 residues in length overlapping by 10 residues and covering the entire sequence of CDV-F, was screened using a lymphocyte proliferation assay with peripheral blood mononuclear cells (PBMC) obtained from dogs inoculated with canine distemper virus (CDV) vaccine. Initially we observed low and inconsistent proliferation of PBMC in response to these peptides, even when using cells obtained from dogs that had received multiple doses of CDV. Subsequently, the use of expanded cell populations derived by in vitro stimulation of canine PBMC with pools of peptides allowed the identification of a number of putative canine Th-epitopes within the protein sequence of CDV-F. There were two major clusters of Th-epitopes identified close to the cleavage site of the F0 fusion protein, while some others were scattered in both the F1 and F2 fragments of the protein. Some of these peptides, in particular peptide 35 (p35), were stimulatory in dogs of different breeds and ages. The identification of such promiscuous canine Th-epitopes encouraged us to assemble p35 in tandem with luteinising hormone releasing hormone (LHRH) a 10 amino-acid residue synthetic peptide representing a B-cell epitope which alone induces no antibody in dogs. The totally synthetic immunogen was able to induce the production of very high titres of antibodies against LHRH in all dogs tested. These results indicate that p35 could be an ideal candidate for use as a Th-epitope for use in outbred dogs.

Amino Acid Sequence↗

Clinical use of Infliximab in Crohn's disease: the Edinburgh experience.

UNLABELLED: Infliximab is an established treatment for steroid-resistant and fistulating Crohn's disease. Although efficacy has been shown in clinical trials, financial implications often limit its use and limited data exist regarding clinical practice. AIMS: To audit the clinical effectiveness of Infliximab. METHODS: We prospectively audited 50 consecutive patients [28 females; median age, 34 years (17-70 years)]. Disease activity and response rates were assessed by the Harvey-Bradshaw index. Clinical and disease data were collected and blood was taken for inflammatory markers, complement and double-stranded DNA antibodies. Patients received Infliximab at 5 mg/kg and were followed for 12 weeks. RESULTS: Indications for Infliximab were refractory Crohn's disease in 39 patients, fistulating Crohn's disease in six, pyoderma gangrenosum in one, pouchitis in two and coeliac disease in two. Thirty-one (79%) of the refractory Crohn's disease patients and four (66%) of the fistulating patients responded at 4 weeks. Twenty-one (54%) of the refractory Crohn's disease patients had a continued response at 12 weeks. Perianal disease was more prevalent in non-responders (7/8 vs. 12/31, P < 0.02). CONCLUSIONS: Response rates to Infliximab in our group are comparable to those of clinical trials. Despite the expense, it remains a useful adjunct to treatment in this otherwise difficult group of patients. Patients with perianal disease responded less well in our cohort.

Adolescent↗

Characterization of conditions for the primary culture of human small intestinal epithelial cells.

Intestinal epithelial cells (IECs) are important for many aspects of gut physiology and pathology. Different approaches have been tried for the primary culture of human IECs, with varying degrees of success, as apoptosis easily occurs. Our aim was to develop a method for primary culture of human IECs from biopsy material. IECs and Lamina propria (LP) cells were liberated from duodenal biopsies obtained from subjects undergoing routine endoscopy for clinical investigations, whose small bowel was macroscopically normal. IECs were cultured on collagen membranes in a 12-well tissue culture cluster, with LP cells and allogeneic Epstein-Barr Virus (EBV)-transformed B lymphocytes (allo-B cells) underneath, in the well. Cultured IECs were characterized by light and confocal microscopy. Cytokine levels in culture supernatants were measured by ELISA. Cells showed the columnar morphology of IECs, even after several days in culture. Best results were obtained from IECs cultured above both LP and allo-B cells. IECs did not form monolayers as do transformed epithelial cell lines, but they did preserve their original cell-cell contacts. Analysis of culture supernatants showed that IL-10 was produced by IECs initially, but IL-1ra was produced by LP cells in the underlying wells with increasing time in culture. Very little IL-1 beta was produced from any cultures. These results show that IECs can be isolated and maintained in primary culture for a short while, which could open new possibilities for research using patient material instead of cell lines.

Biomarkers↗

Outcomes of submandibular duct re-implantation for sialorrhoea.

Drooling is an important factor affecting the quality of lives of neurologically impaired children, and surgery often has to be performed. Submandibular duct relocation is a relatively simple procedure with a low complication rate, and has been carried out on 37 patients at the Sheffield Children's Hospital between 1986 and 1998. To determine the short-term and long-term outcome of this procedure, a questionnaire was sent to the parents/wards of the patients. Feedback was also obtained on complications, parents'/carers' perceptions and main concerns about the operation, particularly in those cases which did not improve. Both short-term (82.4%) and long-term (76.5%) control rates were high, and there were few complications, none of which had any long-term adverse effects. The most significant area of parental/carer dissatisfaction and concern in 'failed' cases was 'pain', and it was felt that proper preoperative counselling about analgesia and chances of failure is very important. We conclude that this is a safe, highly successful procedure which significantly improves the quality of lives of the majority of drooling children.

Adolescent↗

Comparison of the Moskowitz criteria and the pouchitis disease activity index (PDAI) for diagnosis of ileoanal pouch inflammation.

OBJECTIVE: To prospectively compare two indices for diagnosis of pouchitis: Moskowitz criteria and pouchitis disease activity index (PDAI). PATIENTS AND METHODS: Fifty-six consecutive patients with an ileoanal pouch for ulcerative colitis were recruited. A clinical questionnaire was composed and the Oresland functional score calculated. Pouchoscopy and biopsy were performed after cleansing of the pouch. RESULTS: Seven patients had pouchitis according to both Moskowitz and PDAI ('positive' group). Five patients had PDAI > or = 7, but did not fulfil the Moskowitz criteria ('negative' group). The pouchoscopy and Oresland functional scores were similar between the two groups. The biopsy neutrophilia and ulceration were more prominent in the 'positive' group (chi2=5.18, P < 0.05), whereas urgency (chi2=8.4, P < 0.001), evacuation difficulties (chi2=5.18, P < 0.05) and history of bleeding per pouch (chi2=4.18, P < 0.05) were more pronounced in the 'negative' group. CONCLUSION: Moskowitz criteria may be broadly in agreement with the PDAI; these, however, cannot be regarded as interchangeable.

Journal Article↗

Altered subjective appetite parameters in Crohn's disease patients.

BACKGROUND AND AIM: Altered appetite and early satiety may promote anorexia associated with Crohn's disease. The aim of this study was to assess the impact of disease activity on subjective appetite parameters in Crohn's disease patients. METHODS: Seventeen patients with Crohn's disease and 15 healthy controls (8 M: 7 F, 34 (20-35) years) were studied. Subjects rated their hunger, desire to eat, fullness and level of satiety using visual analogue scales after an overnight fast. Subjects were reassessed after ingestion of 500 and 1000 ml water. Anthropometry was used to determine percentage body fat. Serum leptin & TNF- alpha levels were assessed using immunoassay. Disease activity was determined using the Harvey-Bradshaw index. RESULTS: Hunger ratings for active Crohn's disease patients were significantly lower than controls at baseline (P<0.05). Desire to eat was lower in patients with active Crohn's disease than controls both at baseline (95% CI, 0.3 mm, 40.7 mm) and after ingestion of 500 ml water (95% CI, 1.25 mm, 51.9 mm) (P<0.05). Serum leptin concentrations were significantly associated with percent body fat (r=0.57;P<0.001) and, after correcting for body fat status, tended to be higher in patients with active Crohn's disease (mean 0.9 ng/ml/% body fat; SD 0.8 ng/ml/% body fat) compared with either patients with inactive disease (mean 0.4 ng/ml/% body fat; SD 0.3 ng/ml/% body fat) or healthy controls (mean 0.3 ng/ml/% body fat; SD 0.2 ng/ml/% body fat) (P=0.15, ns). Appetite parameters and serum leptin concentrations showed no significant correlation. CONCLUSIONS: Subjective appetite parameters were altered in patients with active Crohn's disease. At baseline, patients with active Crohn's disease were less hungry than healthy controls and had less desire to eat. After ingestion of 500 ml of water, desire to eat was significantly less in patients with active disease as compared with healthy controls. Serum leptin concentration corrected for percent body fat tended to be higher in patients with active Crohn's disease compared with inactive Crohn's disease and healthy controls, but the differences did not reach statistical significance.

Adult↗

Variant Creutzfeldt-Jakob disease (vCJD) and gastrointestinal endoscopy.

Variant Creutzfeldt-Jakob disease (vCJD) is a transmissible form of spongiform encephalopathy believed to be contracted from the consumption of bovine spongiform encephalopathy (BSE) infected beef products. To date over 100 individuals have developed this incurable disease. There have been no documented cases of iatrogenic infection, but there is a theoretical risk that surgical procedures could transmit the disease. This review describes the background of the disease and assesses the possible risks of transmission through endoscopic procedures. The risk of transmission by endoscopy is small and probably negligible if suitable procedures are followed. The greatest potential danger arises from healthy individuals who are incubating the disease. Pathological prions (PrP(sc)) may be found in lymphatic tissue of these individuals (particularly tonsils), but smaller amounts have been identified in the appendix and Peyer's patches. These prions are resistant to all forms of conventional sterilization. There is a theoretical risk that biopsy forceps and the operating channel of endoscopes could become contaminated. This review gives recommendations as to how these small risks can be minimized. They include the employment of single-use forceps for biopsies taken from the terminal ileum, greater attention to the maintenance of endoscopic equipment and accessories, more rigorous manual cleaning of endoscopic equipment and the use of well designed, disposable cleaning brushes for the operating channel of the endoscope.

Animals↗

A randomized trial of thermal ablative therapy versus expandable metal stents in the palliative treatment of patients with esophageal carcinoma.

BACKGROUND: Expandable metal stent insertion and thermal tumor ablation (TTA) both improve dysphagia in patients with advanced esophageal cancer, but no direct comparison study of their efficacy on health-related quality of life has been published. The aim of this study was to compare survival, relief of dysphagia, quality of life, and cost in patients treated by thermal ablation or stent insertion. METHODS: Sixty-five patients with histologically proven, inoperable esophageal and esophagogastric cancer were initially assessed by endoscopy, barium contrast radiography, and CT of the thorax and abdomen. Dysphagia and quality of life were serially assessed at monthly intervals. Patients were randomized to either repeated TTA or insertion of an expandable metal stent. RESULTS: Median survival was significantly longer for patients who underwent TTA; 125 days (17-546) versus 68 days (8-602) for those in whom a stent was inserted (p < 0.05), although relief of dysphagia was disappointing in both groups. Several patients in both groups had serious treatment-related complications and required further therapy. Median length of hospital stay and cost were greater for patients treated by TTA. Health-related quality of life was globally impaired in both groups at randomization and deteriorated significantly in the stent group. Pain was reported more commonly by patients with stents. CONCLUSIONS: The palliation of patients with advanced esophageal and esophagogastric cancer remains unsatisfactory. Health-related quality of life deteriorated in the stent group but not in the TTA group. Patients treated by TTA live longer than patients treated by stent insertion, but the cost of TTA is higher.

Aged↗

Modification of enteral diets in inflammatory bowel disease.

The provision of food is thought to promote the maintenance of gut integrity. Nutrients are able to elicit and affect both systemic and mucosal immune responses. Enteral diet therapy has long been known to be efficacious in inflammatory bowel disease (IBD), particularly in childhood Crohn's disease. However, the mechanisms of action of these diets are not clear. Nutritional repletion, direct effects on the gut mucosa or decreased intestinal permeability have all been postulated as being important in nutritional therapy. There is some evidence that the enteral diet has a direct effect on the gut mucosa by reducing cytokine production and the accompanying inflammation, thus leading to decreased intestinal permeability. Modifications of enteral diet composition have been evaluated in many studies. Such modifications include fat and/or protein content and the addition of bioactive peptides. The fatty acid composition of the enteral diet seems to have a much greater impact on its efficacy than modification of the N source. As specific fatty acids are precursors of inflammatory mediators derived from arachidonic acid, the reduction in these components may be beneficial in nutritional therapy for IBD. Addition of bioactive peptides to enteral diet formulas may also have a role; such peptides may have specific growth factor or anti-inflammatory actions. There is still much work to be done to define disease-specific enteral diet formulas that are effective as therapies for both Crohn's disease and ulcerative colitis.

Cell Membrane Permeability↗

Recognition and stabilization of a unique CPRI--structural motif in cucurbitaceae family trypsin inhibitor peptides: molecular dynamics based homology modeling using the X-ray structure of MCTI-II.

The high resolution crystallographic structure of MCTI-II complexed with beta trypsin (PDB entry 1MCT) was used to model the corresponding structures of the six inhibitor peptides belonging to Cucurbitaceae family (MCTI-I, LA-1, LA-2, CMTI-I, CMTI-III, CMTI-IV). Two model inhibitors, LA-1 and LA-2 were refined by molecular dynamics to estimate the average solution structure. The difference accessible surface area (DASA) study of the inhibitors with and without trypsin revealed the Arginine and other residues of the inhibitors which bind to trypsin. The hydration dynamics study of LA1 and LA2 also confirm the suitability of water molecules at the active Arg site. Moreover, the presence of a unique 3D-structural motif comprises with the four CPRI residues from the amino terminal is thought to be conserved in all the six studied inhibitors, which seems essential for the directional fixation for proper complexation of the Arg (5) residue towards the trypsin S1-binding pocket. The role of the disulphide linkage in the geometrical stabilization of CPRI (Cysteine, Proline, Arginine, Isoleucine) motif has also been envisaged from the comparative higher intra molecular Cys (3) -Cys (20) disulphide dihedral energies.

Amino Acid Motifs↗

Research strategies for design and development of NSAIDs: clue to balance potency and toxicity of acetanilide compounds.

Despite the fact that many modern drug therapies are based on the concept of enzyme inhibition, inhibition of several enzymes leads to pathological disorders. Clinically used nonsteroidal anti-inflammatory drugs (NSAIDs) bind to the active site of the membrane protein, cyclooxygenase (COX) and inhibit the synthesis of prostaglandins, the mediators for causing inflammation. At the same time, inhibition of hepatic cysteine proteases by some NSAID metabolites like NAPQI is implicated in the pathogenesis of hepatotoxicity. As a part of our efforts to develop new effective NSAIDs, a comprehensive investigation starting from synthesis to the study of the final metabolism of acetanilide group of compound has been envisaged with appropriate feedback from kinetic studies to enhance our knowledge and technical competency to feed the know-how to the medicinal chemist to screen out and design new acetanilide derivatives of high potency and low toxicity. Structure-function relationship based on the interaction of acetanilide with its cognate enzyme, cyclooxygenase has been studied critically with adequate comparison with several other available crystal structures of COX-NSAID complexes. Furthermore, to make the receptor based drug design strategy a novel and comprehensive one, both the mechanism of metabolism of acetanilide and structural basis of inhibition of cysteine proteases by the reactive metabolite (NAPQI) formed by cytochrome P450 oxidation of acetanilide have been incorporated in the study. It is hoped that this synergistic approach and the results obtained from such consorted structural investigation at atomic level may guide to dictate synthetic modification with judicious balance between cyclooxygenase inhibition and hepatic cysteine protease inhibition to enhance the potential of such molecular medicine to relieve inflammation on one hand and low hepatic toxicity on the other.

Acetanilides↗

Etiology and outcome of hydrops fetalis.

OBJECTIVES: To identify the etiology and pregnancy outcome of hydrops fetalis in a cohort of pregnancies referred to a tertiary maternal fetal medicine center in the UK. These data allow the review of a large series of pregnancies affected by hydrops fetalis and emphasize the importance of investigation and then treatment of individual cases. This provides parents with improved information and especially specific prognostic information. METHODS: A retrospective review of 63 consecutive cases of hydrops fetalis managed between September 1996 and March 1999. RESULTS: Of the pregnancies, 12.7% (n = 8) were associated with an 'immune' etiology. Of these, 62.5% (n = 5) had fetal anemia due to anti-D, 25% (n = 2) anti-Kell and 12.5% (n = 1) anti-c antibodies. The remaining 55 cases (87.3%) had a non-immune cause. Eight (14.5%) were due to human parvovirus B19 infection. Fourteen cases (25.5%) were associated with aneuploidy and, in four (7.3%), a primary hydrothorax was the cause of the non-immune hydrops fetalis. A cardiac cause was found in five (9.1%) cases. Three of these had supraventricular tachycardia and one had congenital complete heart block. Cystic hygroma was associated with hydrops fetalis in six cases. Twin-twin transfusion syndrome was the cause for hydrops in two cases. Massive transplacental hemorrhage was identified in one case. Fetal akinesia and muscular dystrophy caused hydrops in one case each. In 14.5% (8/55) of cases no obvious cause was identified and these were classified as 'idiopathic'. Three other cases could not be classified because parents declined investigations (unclassified). In the pregnancies with non-immune hydrops fetalis, the outcome was favorable in 27.3% (15/55) of cases. CONCLUSION: The prognosis of hydrops fetalis differs markedly between different etiological groups. Etiologies range from treatable causes with a good outcome and probably no long-term side-effects (as in case of parvovirus B19), to others which are incompatible with life or are associated with considerable perinatal morbidity and mortality.

Cohort Studies↗

Strychnos nux-vomica extract and its ultra-high dilution reduce voluntary ethanol intake in rats.

OBJECTIVES: To see whether Strychnos nux-vomica extract (mother tincture [MT]), its potency Nux 30c, and its principal alkaloid, strychnine, could reduce voluntary ethanol intake in rats. To analyze the solution structure of Nux MT, Nux 30c, 90% ethanol, and ethanol 30c by means of electronic (ES) and nuclear nuclear magnetic resonance (NMR) spectra. DESIGN: Potentially alcoholic rats were first given 20% ethanol and then kept on a two-choice bottle, one with 20% ethanol and another with tap water. These rats were given the following oral treatments for 15 days: group 1, control; group 2, strychnine at 0.36 mg/kg per day; group 3, ethanolic extract of S. nux-vomica seeds (Nux MT) at 3.6 mg/kg per day; and group 4, Nux 30c at 0.05 mL/d per rat. Nux 30c was prepared by successive dilution of Nux MT and 90% ethanol (1:100) and sonication at 20 kHz for 30 seconds in 30 steps. RESULTS: Both Nux MT and Nux 30c significantly reduced ethanol intake and increased water intake in rats. ES of two dilutions of Nux MT and Nux 30c showed intersections at more than one point suggesting existence of molecular complexes. ES of Nux MT in CCl4 showed a red shift when 90% ethanol was added indicating molecular complexation and charge transfer interaction between ethanol and Nux compounds. NMR spectra of Nux MT, 90% ethanol, ethanol 30c, and Nux 30c indicated a change in solution structure of the medium (90% ethanol) of Nux 30c. CONCLUSION: Nux MT and Nux 30c could reduce ethanol intake in rats. The altered solution structure of Nux 30c is thought to mimic Nux MT and produce ethanol aversion in rats.

Alcohol Deterrents↗

Redox imbalance in Crohn's disease intestinal smooth muscle cells causes NF-kappaB-mediated spontaneous interleukin-8 secretion.

Interleukin-8 (IL-8), a chemokine secreted by cells at injury sites, has recently been recognized as involved in the pathogenesis of Crohn's disease. However, the pathogenesis of enhanced spontaneous transcription of IL-8 by the bowel in patients with Crohn's disease is undefined. Although IL-8 is secreted primarily by neutrophils, macrophages, and endothelial and epithelial cells, we observed the involvement of mesenchymal cells in the inflammatory process. A smooth muscle cell line isolated from the ileum of a patient with Crohn's disease (CDISM) and maintained in culture exhibited spontaneous transcription and secretion of IL-8 when compared with intestinal smooth muscle cells obtained from a normal subject (NHISM). Furthermore, IL-8 transcription from CDISM cells was associated with remarkable spontaneous activation of the oxidant-sensitive transcription factor NF-kappaB, as assessed by transient transfection assays with an IL-8 promoter reporter construct, Western blot analysis, and electrophoretic mobility shift assays (EMSA). Finally, we report here that CDISM cells exhibit significantly altered redox balance. The antioxidant pyrrolidine dithiocarbamate (PDTC) restored the redox equilibrium by mechanisms that inhibit binding of NF-kappaB to its cognate site on the IL-8 promoter. These findings suggest that restoration of the redox balance could hold promise for therapeutic intervention in Crohn's disease.

Antioxidants↗

Dynamics of ankyrin-containing complexes in chicken embryonic erythroid cells: role of phosphorylation.

Chicken erythroid ankyrin undergoes a fairly rapid cycle of cytoskeletal association, dissociation, and turnover. In addition, the cytoskeletal association of ankyrin is regulated by phosphorylation. Treatment of erythroid cells with serine and threonine phosphatase inhibitors stimulated the hyperphosphorylation of the 225- and 205-kDa ankyrin isoforms, and dissociated the bulk of these isoforms from cytoskeletal spectrin. In vitro binding studies have shown that this dissociation of ankyrin from spectrin in vivo can be attributed to a reduced ability of hyperphosphorylated ankyrin to bind spectrin. Interestingly, a significant fraction of detergent insoluble ankyrin accumulates in a spectrin-independent pool. At least some of this spectrin-independent pool of ankyrin is complexed with the AE1 anion exchanger, and the solubility properties of this pool are also regulated by phosphorylation. Treatment of cells with serine and threonine phosphatase inhibitors had no effect on ankyrin/AE1 complex formation. However, these inhibitors were sufficient to shift ankyrin/AE1 complexes from the detergent insoluble to the soluble pool. These analyses, which are the first to document the in vivo consequences of ankyrin phosphorylation, indicate that erythroid ankyrin-containing complexes can undergo dynamic rearrangements in response to changes in phosphorylation.

Animals↗

The role of Asn14 in the stability and conformation of the reactive-site loop of winged bean chymotrypsin inhibitor: crystal structures of two point mutants Asn14-->Lys and Asn14-->Asp.

A double-headed chymotrypsin inhibitor, WCI, from winged bean seeds was cloned for structural and biochemical studies. The inhibitor was subjected to two point mutations at a conserved position, Asn14. This residue, known to have a pivotal role in stabilizing the first reactive-site loop (Gln63-Phe68) of the inhibitor, is highly conserved in the sequences of the other members of Kunitz (STI) family as well as in the sequences of Kazal family of serine protease inhibitors. The mutants, N14K and N14D, were subjected to biochemical assay and their characteristics were compared with those of the recombinant inhibitor (rWCI). Crystallographic studies of the recombinant and the mutant proteins are discussed. These studies were primarily aimed at understanding the importance of the protein scaffolding towards the conformational rigidity of the reactive-site loop. Our analysis reveals that, as the Lys14 side chain takes an unusual fold in N14K and the Asp14 side chain in N14D interacts with the loop residues by water-mediated hydrogen bonds, the canonical conformation of the loop has remained effectively intact in both the mutant structures. However, minor alterations such as a 2-fold increase in the inhibitory affinity towards the cognate enzyme were observed.

Amino Acid Sequence↗

Serum IgA tissue transglutaminase antibodies in coeliac disease and other gastrointestinal diseases.

We investigated the presence of IgA anti-tissue transglutaminase (tTG) antibodies in untreated coeliac disease (CD) and other gastrointestinal diseases, and compared IgA tTG concentrations with anti-endomysium (EMA) immunofluorescent findings. The study included 116 untreated CD patients (74 female, 42 male, age range 15-78 years, median 47 years), 82 treated CD patients, 65 patients with normal duodenal histology, 260 disease control samples and 29 healthy volunteers. IgA anti-tTG, EMA, and anti-gliadin (AGA) antibodies were measured. Serum total IgA was measured in the CD patients. Two IgA-deficient untreated CD patients were excluded. IgA EMA and IgA AGA were positive in 99 (87%) and 69 (61%), respectively, of the 114 untreated CD patients. Elevated IgA anti-tTG were found in 92/114 (81%) untreated coeliacs, 1/82 (1%) treated coeliacs, 2/65 (3%) non-coeliacs, 10/260 (4%) disease controls and 2/29 (7%) volunteers. Four of the untreated CD patients, with a normal serum total IgA concentration, were negative for all the serological tests. IgA anti-tTG concentrations were significantly higher in untreated coeliacs (median 10200 units/ml) than in other groups (Mann-Whitney, p<0.00001) and compared well with IgA EMA titres (r(2)=0.54; p<0.0001).

Adolescent↗