To determine the feasibility of liver biopsy immunotyping to characterize hepatic lymphoid malignancies, we employed a panel of monoclonal antibodies on snap-frozen hepatic tissue from 18 patients. Six patients proved to have a histologic diagnosis of lymphoid malignancy. By using free avidin and biotin-blocking reagents to block endogenous biotin, followed by standard immunochemistry, immunotyping was successful in all six cases. Serial section typing allowed delineation of complex B cell phenotypes. Furthermore, architecture was preserved allowing discernment of disease patterns (e.g., sinusoidal, hairy cell leukemia vs. portal, follicular small-cleaved cell lymphoma. Unexpectedly, we found striking expression of common ALL antigen in normal bile canaliculi, which may prove of diagnostic or therapeutic relevance. This study establishes the utility of immunohistochemical techniques applied to hepatic biopsies as a valuable adjunct to histologic diagnosis as well as a tool in revealing the immunobiology of the liver.

Biopsy↗

PubMedDate not supplied

Process options in hepatitis B surface antigen extraction from transgenic potato.

The process conditions for recombinant hepatitis B surface antigen (HBsAg) extraction from transgenic potato were examined. The effects of temperature, the reducing agent beta-mercaptoethanol (BME), and proteinase inhibitors on the level of antigenic activity of recovered HBsAg were determined. Sedimentation profiles were performed to characterize HBsAg assembly into virus-like particles. Increasing the temperature of the sample for about 1 min increased the measured HBsAg antigenic activity. The optimum temperature was around 50 degrees C. A 3-fold enhancement of the antigenic activity was obtained in extract from transgenic potato expressing HBsAg, when monoclonal antibodies were used to assay for HBsAg. When antigenic activity was determined by polyclonal antibodies, no enhancement in the antigenic activity was obtained. Temperature may affect the conformation of the a epitope to which the monoclonal antibodies bind or alter the fluidity of surface lipid regions. BME increased the antigenic activity of HBsAg up to 4-fold when monoclonal antibodies directed against the a determinant were used, but there was no increase with polyclonal antibodies. This observation suggests that BME affects the structure or presentation of the a epitope. In the presence of BME and leupeptin, a proteinase inhibitor, higher antigenic activity was obtained. Leupeptin might protect the antigen, which might become more susceptible to proteolytic degradation after reduction, as a result of stimulation of sulfhydryl proteases. Although both temperature and BME increased the antigenic activity of HBsAg individually, when combined their interaction was antagonistic, resulting in reduced antigenic activity. Different proteinase inhibitors, including leupeptin, aprotinin, E-64, pefabloc, and pepstatin, had no significant effect on HBsAg from potato extract in a 2 h period in the absence of BME. The sedimentation profile of potato-produced HBsAg was determined in 5-30% sucrose gradients. Yeast-derived recombinant HBsAg was used as a positive control. The HBsAg from transgenic potato showed sedimentation and density properties that are very similar to the yeast-produced antigen, indicating assembly into virus-like particles. BME treatment did not change the sedimentation profile.

Hepatitis B Surface Antigens↗

PubMedDate not supplied

Cell hydration and mTOR-dependent signalling.

Insulin- and amino acid-induced signalling by the mammalian target of rapamycin (mTOR) involves hyperphosphorylation of the p70 ribosomal S6 protein kinase (p70S6-kinase) and the eukaryotic initiation factor 4E (eIF4E) binding protein 4E-BP1 and contributes to regulation of protein metabolism. This review considers the impact of cell hydration on mTOR-dependent signalling. Although hypoosmotic hepatocyte swelling in some instances activates p70S6-kinase, the hypoosmolarity-induced proteolysis inhibition in perfused rat liver is insensitive to mTOR inhibition by rapamycin. Likewise, swelling-dependent proteolysis inhibition by insulin and swelling-independent proteolysis inhibition by leucine, a potent activator of p70S6-kinase and 4E-BP1 hyperphosphorylation, in perfused rat liver is insensitive to rapamycin, indicating that at least rapamycin-sensitive mTOR signalling is not involved. Hyperosmotic dehydration in different cell types produces inactivation of signalling components around mTOR, thereby attenuating insulin-induced glucose uptake, glycogen synthesis, and lipogenesis in adipocytes, and MAP-kinase phosphatase MKP-1 expression in hepatoma cells. Direct inactivation of mTOR, stimulation of the AMP-activated protein kinase, and the destabilization of individual proteins may impair mTOR signalling under dehydrating conditions. Further investigation of the crosstalk between the mTOR pathway(s) and hyperosmotic signalling will improve our understanding about the contribution of cell hydration changes in health and disease and will provide further rationale for fluid therapy of insulin-resistant states.

Adipocytes↗

PubMedDate not supplied

Platelet activating factor in stool from patients with ulcerative colitis and Crohn's disease.

BACKGROUND/AIMS: Platelet activating factor (PAF) is a potent endogenous mediator in inflammatory processes. The role of this mediator, especially in connection with the unknown etiology of chronic inflammatory bowel diseases, remains poorly understood. A determination of PAF in stool may be helpful in recognizing quiescent inflammations in chronic inflammatory bowel diseases. A simple and reliable method for the determination of PAF in stool seems to be necessary to achieve this goal. METHODOLOGY: PAF analysis was performed with the help of a commercial PAF radioimmunoassay (RIA) kit after solid phase extraction (SPE) of ethanolic stool extracts. PAF was determined in stool from 10 healthy volunteers (m = 4; f = 6), 13 patients with ulcerative colitis (m = 7; f = 6) and 15 patients with Crohn's disease (m = 9; f = 6). Fecal PAF concentrations were compared with activity index of disease, endoscopic index, localization of lesions, leukocyte count, erythrocyte sedimentation rate (ESR), C-reactive protein (CRP), medical prednisolone treatment, sex and age of the patients. RESULTS: In healthy volunteers, no PAF was detectable in stool. In patients with Crohn's disease 319.2 +/- 143.5 pg PAF/g stool and in patients with ulcerative colitis 824.9 +/- 408.7 pg PAF/g stool could be determined. A significant correlation (p < 0.05) was found between PAF-content in stool and the endoscopical index and intestinal localization of inflammatory lesions. No further correlations could be detected in our patients. CONCLUSIONS: Fecal PAF assessment may be used clinically as a non-invasive method to estimate severity of mucosal inflammation in patients with inflammatory bowel disease (IBD).

Adult↗

PubMedDate not supplied

Cardiopulmonary resuscitation.

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Humans↗

PubMedDate not supplied

Guidelines for supervised wound care by emergency nurse practitioners.

Nearly 10 million patients with traumatic lacerations are treated annually in Emergency Departments in the United States. Since these wounds do not usually pose a threat to the patient's life, they assume a lower level of priority than emergent conditions. Consequently, treatment of patients with lacerations is often delayed until the emergent patient is resuscitated. In the event that wound care is inadvertently delayed, bacteria may proliferate to levels that result in infection. In selected patients, supervised wound care by an emergency nurse practitioner (ENP) can safely eliminate this delay in treatment. During their graduate training in the University of Virginia Nursing School, the ENPs are instructed to treat traumatic wounds with physician supervision. The criteria for patient selection and care are outlined in comprehensive guidelines reported herein. The bases for these treatment guidelines were clinical and experimental studies conducted at our medical center which examined the influence of various therapeutic decisions on the ultimate fate of the wound.

Asepsis↗

PubMedDate not supplied

Evaluation of supervised wound care guidelines.

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Evaluation Studies as Topic↗