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Biomedical subjects

J M Gilbert

Publications and source records attributed to J M Gilbert.

At least 37 records · Page 2Linked to original sources
PubMed1996

Quantification of systemic and local immune responses to individual rotavirus proteins during rotavirus infection in mice.

The purpose of the present study was to develop a quantitative assay that could be used to measure the local and systemic immune responses to specific rotavirus proteins following rotavirus infection of adult mice. To measure these responses, we used an immunocytochemical staining assay of Spodoptera frugiperda (Sf-9) cells which were infected with recombinant baculovirus expressing selected rotavirus proteins. The specificity of the assay was documented by using a series of monoclonal antibodies to individual rotavirus proteins. We observed that the assay had high levels of sensitivity and specificity for a series of VP7- and VP4-specific neutralizing monoclonal antibodies which recognized conformation-dependent epitopes on their target proteins. We also studied immunoglobulin G (IgG) immune responses in serum and IgA immune responses in the stools of mice infected with wild-type murine rotavirus strain EHPw. In both sera and stools, the most immunogenic proteins were VP6 and VP4. VP2 was less immunogenic than VP6 or VP4, and the immune responses to VP7, NSP2, and NSP4 were very low in serum and undetectable in stools.

Animals↗
PubMed1996

Splenectomy in a general hospital.

Splenectomy is often performed in patients with malignant disease or trauma who are at a high risk of complications. In the long term, it increases the risk of infection by encapsulated bacteria. An audit was performed to determine the reasons for splenectomy in a district general hospital, to review the results and complications of surgery, and to see how often the patients were prescribed antibacterial prophylaxis. Twenty-eight patients underwent splenectomy in 3 years. The indication was haematological disease in 13 and trauma in four. In the remaining nine the spleen was removed either as part of a radical gastrectomy or during some other abdominal procedure. Six of the 28 patients had died, one within 30 days from disseminated intravascular coagulopathy following an emergency gastrectomy and splenectomy for haematemesis, two from progressive haematological malignant disease, two from non-haematological malignancy, and one from bronchopneumonia. Of the nine patients (32%) with complications, three required a further laparotomy. Most patients had been prescribed pneumococcal vaccine (85%) and prophylactic antibiotics (93%).

Adolescent↗
PubMed1995

Receptor-induced conformational changes in the subgroup A avian leukosis and sarcoma virus envelope glycoprotein.

We recently reported that Tva, the host cell receptor for subgroup A avian leukosis and sarcoma viruses, binds specifically to the subgroup A envelope glycoprotein (Env-A) (J.M. Gilbert, P. Bates, H. E. Varmus, and J. M. White, J. Virol. 68:5623-5628, 1994). Here we have tested the hypothesis that binding of Tva causes conformational changes in Env-A that correlate with its conversion from a fusion-inactive to a fusion-active state. Conformational changes were examined by both a proteolysis and an immunoprecipitation assay. A temperature-dependent conformational change, demonstrated by the generation of a specific thermolysin digestion product of the surface (SU) subunit, occurred when a soluble form of Tva (sTva) was incubated with Env-A. sTva did not induce this conformational change in Env-C or in a noninfectious precursor form of Env-A, Env-A CL. However sTva did induce the conformational change in Env-A CL that had been pretreated in vitro to produce the SU and transmembrane (TM) subunits. Moreover, interaction of Tva with Env-A at 25 degrees C, but not at 4 degrees C, appeared to reveal a previously buried segment of the putative fusion peptide of Env-A. Our results suggest that binding of Tva to Env-A results in specific conformational changes in the Env-A glycoprotein that are relevant to the activation of its fusion function.

3T3 Cells↗
PubMed1995

Rotavirus-induced fusion from without in tissue culture cells.

We present the first evidence of fusion from without induced in tissue culture cells by a nonenveloped virus. Electron micrographs of two strains of rotavirus, bovine rotavirus C486 and rhesus rotavirus, show that virally mediated cell-cell fusion occurs within 1 h postinfection. Trypsin activation is necessary for rotavirus to mediate cell-cell fusion. The extent of fusion is relative to the amount of virus used, and maximum fusion occurs between pHs 6.5 and 7.5. Fusion does not require virus-induced protein synthesis, as virus from both an empty capsid preparation and from an EDTA-treated preparation, which is noninfectious, can induce fusion. Incubation of rotavirus with neutralizing and nonneutralizing monoclonal antibodies before addition to cells indicates that viral protein 4 (VP4; in the form of VP5* and VP8*) and VP7 are involved in fusion. Light and electron micrographs document this fusion, including the formation of pores or channels between adjacent fused cells. These data support direct membrane penetration as a possible route of infection. Moreover, the assay should be useful in determining the mechanisms of cell entry by rotavirus.

Animals↗
PubMed1994

The receptor for the subgroup A avian leukosis-sarcoma viruses binds to subgroup A but not to subgroup C envelope glycoprotein.

The putative subgroup A avian leukosis-sarcoma virus (ALSV) receptor (Tva) was recently cloned by gene transfer (P. Bates, J. A. Young, and H. E. Varmus, Cell 74:1043-1051, 1993; J. A. T. Young, P. Bates, and H. E. Varmus, J. Virol. 67:1811-1816, 1993). Susceptibility to infection by subgroup A ALSV is conferred on cells upon transfection with cDNAs encoding tva. The hypothesis that tva encodes a specific receptor for subgroup A ALSV predicts that the Tva protein should bind to subgroup A, but not to subgroup C, envelope glycoprotein. In this study, we examined this prediction by using several biochemical assays. We established stable NIH 3T3 cell lines expressing either Tva, the subgroup A envelope glycoprotein (Env-A), or the subgroup C envelop glycoprotein (Env-C) and used them in conjunction with soluble forms of these molecules to demonstrate specific binding. When cell lysates containing Tva were mixed with lysates of either Env-A or Env-C, an immunoprecipitable complex formed between Tva and Env-A but not between Tva and Env-C. A soluble, oligomeric form, of Env-A, not Env-C, binds to cells expressing Tva. Reciprocally, a secreted form of Tva can bind to cells expressing Env-A but not to cells expressing Env-C. A specific and stable complex formed between soluble Env-A and secreted Tva as demonstrated by sucrose density gradient centrifugation. Thus, by three kinds of assays, Tva appears to bind specifically to Env-A, which is consistent with genetic evidence that it serves as the cell surface receptor of subgroup A ALSV and the main determinant of subgroup specificity.

3T3 Cells↗
PubMed1994

Simple method to purify chicken immunoglobulin G.

A new method for purifying IgG was adapted for use with chicken serum. Chicken serum was mixed with caprylic acid, precipitated with ammonium sulfate, and dialyzed against PBS. Analysis by SDS-PAGE and Western blot confirmed successful purification of chicken IgG. Large quantities of highly purified IgG were easily obtainable for use in immunological investigations or for labeling. This technique is rapid, inexpensive, and simpler to perform than traditional ion exchange or gel filtration chromatography.

Ammonium Sulfate↗
PubMed1993

Generation of a water-soluble oligomeric ectodomain of the Rous sarcoma virus envelope glycoprotein.

Sequences encoding the transmembrane domain of the Rous sarcoma virus envelope (Env) glycoprotein were deleted and replaced with sequences that signal addition of a glycosyl phosphatidylinositol (GPI) membrane anchor. Stable NIH 3T3 cell lines expressing either the wild-type transmembrane-anchored Env or the Env chimera with a GPI tail were established. The GPI-anchored envelope glycoprotein is expressed, oligomerized, and transported to the cell surface in a manner identical to that of its wild-type transmembrane-anchored counterpart. The GPI-linked protein is quantitatively removed from the cell surface by treatment with phosphatidylinositol phospholipase C. The phosphatidylinositol phospholipase C-released, water-soluble Env glycoprotein ectodomain retains the wild-type oligomeric structure and provides a useful tool for studying the subgroup-specific binding and fusion activities of a prototypic retroviral Env glycoprotein.

3T3 Cells↗
PubMed1992

Electrophoretic and immunologic characterization of proteins of merozoites of Eimeria acervulina, E. maxima, E. necatrix, and E. tenella.

Merozoites of Eimeria acervulina, Eimeria maxima, Eimeria necatrix, and Eimeria tenella were compared by gel electrophoresis, western-blotting with chicken antiserum, indirect fluorescent antibody reactions, and antiserum neutralization. Merozoites from the 4 species had dissimilar patterns of proteins and antigens in soluble and membrane fractions. Coomassie blue staining of SDS-PAGE gels revealed 16-22 protein bands depending on the species of merozoite but only 3 bands per species in the membrane fractions. Homologous and heterologous antisera recognized 5-12 soluble fraction bands and 3-7 membrane fraction bands on immunoperoxidase-stained western blots, depending on the species. When antisera from infected chickens were used in an indirect fluorescent antibody reaction, the merozoites of E. tenella and E. necatrix had a strong reaction with homologous and heterologous antisera. Merozoites of E. acervulina and E. maxima reacted with homologous antisera but had a weak or no reaction with heterologous antisera. Chicken antiserum against E. tenella had no effect on the viability of E. tenella merozoites when they were inoculated into chicken embryos.

Animals↗
PubMed1991

Prevention of postoperative renal dysfunction in patients with obstructive jaundice: a multicentre study of bile salts and lactulose.

The role of preoperative lactulose and bile salts in the prevention of postoperative renal failure in patients with obstructive jaundice has been evaluated in a prospective randomized trial. One hundred and two patients undergoing surgery for obstructive jaundice (bilirubin greater than 100 mumols/l) were randomized into three groups: those receiving preoperative oral lactulose (n = 35), those receiving oral sodium deoxycholate (n = 32) and a control group of patients receiving no specific treatment (n = 35). All patients received intravenous fluids commencing the night before surgery. One patient in the control group and none in the treatment groups developed postoperative renal failure. Postoperative deterioration of renal function in patients with normal preoperative function was significantly more common in the control group than in the treatment groups (chi 2 = 8.1, d.f. = 2, P less than 0.02). The incidence of renal failure and impairment was lower in this control group than that reported in previous studies. This may be due to the introduction of adequate preoperative hydration. Additional protection occurs by the preoperative administration of either lactulose or sodium deoxycholate.

Acute Kidney Injury↗
PubMed1991

Evaluation of anticoccidial drugs in chicken embryos.

Infections of Eimeria tenella in chicken embryos were used to compare the anticoccidial activity of ten drugs. The minimal inhibitory concentration (MIC) and minimal toxic concentration (MTC) were affected by the time of inoculation into the embryos and by the chemical nature of the compounds. Some compounds (nicarbazin, amprolium) had no effect on the development of coccidia when they were injected into embryos after the day of infection. Drugs that act early in the life cycle of coccidia (robenidine, clopidol, decoquinate, diclazuril, halofuginone, monensin, salinomycin, and lasalocid) were active at 5-125 micrograms/embryo when they were injected on the day of infection. The ionophores and halofuginone were highly toxic to embryos; most synthetic compounds were nontoxic. The incubation of merozoites in drug suspensions prior to the infection of embryos did not result in embryo toxicity, but the resultant MICs were much higher than those obtained when drugs were injected directly into the embryos. Several products were essentially inactive. Neither nicarbazin nor amprolium prevented oocyst formation. The widely divergent endpoints for the MIC and MTC of anticoccidials in embryos seriously limits the application of this technique as a screen for anticoccidial drugs.

Animals↗
PubMed1991

The aetiology and surgery of carcinoma of the anus, rectum and sigmoid colon in Crohn's disease. Negative correlation with human papillomavirus type 16 (HPV 16).

Anal and recto-sigmoid Crohn's disease may rarely be complicated by the development of local malignancy. Eight patients with this association were identified from the records of St. Mark's Hospital between 1947 and 1988 and two from The London Hospital. The aetiology of the malignancies and their surgical management have been examined. Eight patients had adenocarcinoma of the anus, rectum or sigmoid colon and two had squamous cell carcinoma of the anus. Synchronous dysplasia, adenomas and carcinomas were found in four of the eight patients with adenocarcinoma and a progression to malignancy which is analogous to that in ulcerative colitis is proposed for these cases via a 'dysplasia-carcinoma' or 'adenoma-carcinoma' sequence. In the four other patients with adenocarcinoma, the tumour arose within an area of Crohn's disease or in association with a chronic Crohn's fistula. In these four cases no dysplasia was found in the specimens at that time and long-standing infection is the only aetiological factor identified. Evidence for infection with human papillomavirus (HPV 16) was sought by DNA hybridisation of archival material but none was found in material from the Crohn's disease, fistulae or adeno- or squamous cell carcinomas. Anal and rectosigmoid Crohn's disease disguised the presence of malignancy, and diagnosis was usually delayed. All patients underwent excisional surgery and eight of the ten had all of the large bowel removed either as a single procedure or in stages. Only three of the eight cases of adenocarcinoma had early malignancies (Dukes' stage A) and the remaining five cases had locally advanced disease (Dukes' stage B).

Adult↗
PubMed1990

Surgical correction of the efferent ileal limb for disordered defaecation following restorative proctocolectomy with the S ileal reservoir.

The S ileal reservoir has been superseded in this unit but 76 patients had this operation between 1976 and 1983. Forty-one (54 per cent) patients had to catheterize the reservoir to evacuate faeces and this was primarily due to the long efferent ileal limb. In six patients, the need to catheterize and other problems with defaecation were such that surgical correction of the efferent ileal limb was undertaken. These six patients are reported. Presenting features were the need to catheterize the reservoir, difficulty in catheterizing, faecal incontinence, stenosis of the efferent ileal limb and transanal prolapse of the efferent ileal limb. All patients had an excessively long efferent ileal limb of 8 cm or more which was resected and reanastomosed to the anal canal. The resection was performed endoanally in three patients but was successful in only one. In the two patients in whom endoanal excision was unsuccessful and in the remaining three resection of at least 5 cm was performed transabdominally with endoanal reanastomosis. Three of these five patients were converted from catheterizing the reservoir to spontaneous evacuation, but two patients still needed to catheterize. All six patients benefited in terms of the need for, or frequency of, catheterization, or by improvement in continence. Excessive length of the efferent limb of an S ileal reservoir may cause unsatisfactory defaecation, which may be improved by partial resection. The transabdominal route is recommended for the resection, with endoanal reanastomosis. This surgery is potentially problematic and, although no anastomotic leakage was encountered, a covering loop ileostomy is recommended.

Adult↗
PubMed1990

A new method for purification of Eimeria tenella merozoites.

A rapid and simple method for purifying second generation merozoites of Eimeria tenella was developed using a host tissue digestion fluid, containing 0.25% trypsin and 0.5% taurodeoxycholic acid, to liberate merozoites grown in chick embryos or from parasitized ceca. After filtration, the digestion procedure yielded 1.4 x 10(7) or 8.33 x 10(7) merozoites per embryo or cecum, respectively. These yields were nine-fold for embryos and three-fold for ceca in comparison to previous reports. Viability of the merozoites was normal as assessed by their ability to reinfect embryos and cell cultures. The new method has advantages in that large numbers of pure, viable merozoites can be obtained quickly and easily, and the procedures require minimal effort and supplies.

Animals↗
PubMed1990

Ocular manifestations and impression cytology of anorexia nervosa.

A prospective age- and sex-controlled study of seven anorexia nervosa patients and seven normal control patients was conducted to determine the ocular manifestations of anorexia nervosa. Slit-lamp examination of the anorexic patients demonstrated a high incidence (4/7) of multiple episcleral capillary aneurysms and subconjunctival hemorrhages. Two of seven anorexia nervosa patients had bilateral superficial punctate keratopathy. Anesthetized Schirmer tear testing demonstrated a significantly (P less than 0.005) reduced mean tear production in the anorexia nervosa group (11.3 mm) compared with the control group (22.4 mm). Masked interpretation of conjunctival impression cytology demonstrated moderate to severe conjunctival squamous metaplasia in the majority of the anorexia nervosa group (5/7) compared to normal conjunctival epithelium in the majority of the control group (5/7). Absence of nyctalopia, Bitot's spots, and xerosis, and lack of conjunctival goblet cell loss indicate that the anorexia nervosa group did not have vitamin A deficiency.

Adult↗