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Biomedical subjects

M Schreiber

Publications and source records attributed to M Schreiber.

At least 91 records · Page 5Linked to original sources
PubMed1996

Mutational analysis of the ligand-binding domain of M-T2 protein, the tumor necrosis factor receptor homologue of myxoma virus.

The myxoma virus-encoded M-T2 protein shares extensive sequence homology with the ligand-binding domains of the TNF receptors (TNFRs) and has been shown to bind and inhibit rabbit TNF-alpha with affinities similar to those of TNF-alpha with cellular receptors. Here we show that M-T2 protein is secreted from infected cells as an N-linked glycoprotein, with both complex and hybrid or high mannose oligosaccharide chains. Since amino acid homology between M-T2 and cellular TNF receptors is limited to the four N-terminal cysteine-rich domains (CRDs), various M-T2 C-terminal truncations were created in recombinant vaccinia virus vectors. C-terminal deletions that include truncations up to the middle of the fourth CRD effectively bound and inhibited rabbit TNF-alpha. In contrast, removal of any one of the first three CRDs resulted in a mutant M-T2 protein incapable of binding or inhibiting rabbit TNF-alpha. The C-terminal portion of M-T2, which is not homologous to the cellular TNFRs, appears to be important for efficient secretion of M-T2 from infected cells, since all the C-terminal truncations, including a truncation removing only the last 24 amino acids, were effectively retained as intracellular proteins that were still capable of binding and inhibiting rabbit TNF-alpha. We conclude that the first three CRDs of M-T2 fulfill the same ligand-binding function as the cellular TNFRs, and the nonhomologous C-terminal region participates in protein trafficking of M-T2 in virus-infected cells.

Animals↗
PubMed1996

Myxoma virus T2 protein, a tumor necrosis factor (TNF) receptor homolog, is secreted as a monomer and dimer that each bind rabbit TNFalpha, but the dimer is a more potent TNF inhibitor.

The myxoma virus T2 (M-T2) gene expresses a secreted protein that contains significant sequence similarity to the ligand binding domains of the cellular tumor necrosis factor (TNF) receptors, specifically inhibits the cytolytic activity of rabbit TNFalpha and is an important virulence factor for myxoma virus infection in rabbits. M-T2 protein was overexpressed from vaccinia virus vectors, purified to apparent homogeneity, and found to specifically protect mouse and rabbit cells from lysis by rabbit TNFalpha at molar ratios comparable with the soluble versions of the host tumor necrosis factor receptors. M-T2 secreted from virus-infected cells is detected as both a monomer and a disulfide-linked dimer, both of which were shown by Scatchard analysis to bind rabbit TNFalpha (Kd values of 170 pM and 195 pM, respectively), values that are comparable with the affinities of mammalian TNFs with their receptors. In contrast to the rabbit ligand, M-T2 interacts with mouse TNFalpha with a much lower affinity, Kd of 1.7 nM, and was unable to inhibit the cytolytic activity of this ligand on mouse cells. Although both monomeric and dimeric forms bound rabbit TNFalpha with comparable affinity, the dimeric M-T2 protein was a far more potent inhibitor of rabbit TNFalpha, presumably because it can more effectively prevent dimerization of TNF receptors than can the M-T2 monomer.

Animals↗
PubMed1996

Expression of the myxoma virus tumor necrosis factor receptor homologue and M11L genes is required to prevent virus-induced apoptosis in infected rabbit T lymphocytes.

Myxoma virus is a leporipoxvirus that causes a highly lethal virulent disease known as myxomatosis in the European rabbit. An important aspect of myxoma virus pathogenesis is the ability of the virus to productively infect lymphocytes and spread to secondary sites via lymphatic channels. We investigated the infection of the CD4+ T lymphoma cell line RL-5 with myxoma virus and Shope fibroma virus, a related but benign leporipoxvirus, and observed that myxoma virus, but not Shope fibroma virus, was able to productively infect RL-5 cells. We also discovered that infection of RL-5 cells with Shope fibroma virus or attenuated myxoma virus mutants containing disruptions in either the T2 or the M11L gene resulted in the rapid induction of DNA fragmentation, followed by morphological changes and loss in cell integrity characteristic of cell death by apoptosis. Purified exogenous T2 protein was unable to prevent apoptosis, suggesting that T2 functions intracellularly. Thus, myxoma virus T2, originally described as a secreted homologue of the tumor necrosis factor receptor, and M11L, a novel transmembrane species with no known cellular homologue, function to extend virus host range for replication in rabbit T lymphocytes through the inhibition of apoptosis in infected T lymphocytes.

Animals↗
PubMed1996

Antikaliuretic action of trimethoprim is minimized by raising urine pH.

This study was designed to test the hypothesis that the antikaliuresis caused by trimethoprim could be diminished by alkalinizing the luminal fluid in the CCD, thereby converting trimethoprim from its cationic, active form to an electroneutral, inactive, form. Trimethoprim-induced inhibition of transepithelial Na+ transport was examined in A6 distal nephron cells by analysis of short circuit current. The voltage-dependence of the trimethoprim-induced block of Na+ channels was examined with patch clamp recordings of A6 cells. The antikaliuretic effect of trimethoprim was examined in vivo in rats pretreated with deoxycorticosterone and with NH4Cl to lower urine pH, and in rats also receiving acetazolamide to raise urine pH. We found that the concentration of trimethoprim required to inhibit the amiloride sensitive component of short circuit current by 50% (IC50) was 340 microM (at pH 8.2) and 50 microM (at pH 6.3). The IC50S of protonated trimethoprim were similar (34 microM at pH 8.2 and 45 microM at pH 6.3). The mean time open for the high selectivity, Na+ channel was reduced from 1679 +/- 387 msec to 502 +/- 98 msec with addition of 10-5 M trimethoprim to patch pipette solution at the resting membrane potential (-Vpipette = 0 mV). further decreases in mean time open were observed as -Vpipette was reduced (that is, apical membrane hyperpolarization) to -40 mV (mean time open = 217 +/- 85 msec) and to -80 mV (mean time open = 69 +/- 13 msec). In vivo, trimethoprim caused a > 50% reduction in potassium (K+) excretion due primarily to a fall in the [K+] in the lumen of the terminal CCD. This effect of trimethoprim was markedly attenuated in an alkaline urine induced by acetazolamide. We conclude that it is the charged, protonated species of trimethoprim which blocks epithelial Na+ channels. Increasing urinary pH decreases the concentration of the charged species of trimethoprim and minimizes its antikaliuretic effect.

Animals↗
PubMed1996

Influence of vaginal versus abdominal hysterectomy on perioperative glucose metabolism.

The aim of this study was to investigate the metabolic effects of abdominal versus vaginal hysterectomy with specific regard to perioperative glucose metabolism. Fourteen patients received either abdominal (AH, n = 7) or vaginal hysterectomy (VH, n = 7). Hepatic glucose production was measured before and 2.5 h after the operation by stable isotope technique ([6,6-2H2]-glucose). Metabolic substrates (glucose, lactate, nonesterified fatty acids [NEFA], beta-hydroxybutyrate) and hormones (insulin, glucagon, cortisol, catecholamines) were determined pre-, intra-, and postoperatively. VH induced a higher postoperative glucose concentration than the abdominal approach (VH, 148 +/- 25 mg/dL; AH, 111 +/- 16 mg/dL; P < 0.05). Since postoperative enhancement of hepatic glucose production was comparable in both groups, glucose clearance was lower after the vaginal procedure (VH, 1.7 +/- 0.3 mL.kg-1.min-1; AH, 2.1 +/- 0.3 mL.kg-1.min-1; P < 0.05). NEFA, beta-hydroxybutyrate, and catecholamines similarily increased after surgery. Cortisol levels were more increased after VH (VH, 80 +/- 26 micrograms/dL; AH, 37 +/- 14 micrograms/dL; P < 0.001). Lactate, glucagon, and insulin concentrations did not change perioperatively. The more pronounced hyperglycemic response to VH was due to lower peripheral glucose use caused by higher postoperative cortisol values. The mechanisms responsible for this marked cortisol enhancement after the vaginal operation as well as the clinical significance for patients with preexisting impaired carbohydrate tolerance, however, remained unclear and warrant further investigation.

3-Hydroxybutyric Acid↗
PubMed1996

Loss of antibody reactivity directed against the V3 domain of certain human immunodeficiency virus type 1 variants during disease progression.

We have previously shown that in AIDS patients a predominant species of infectious virus can be found which is not neutralized by homologous serum. The presence of the infectious virus was associated with the lack of type-specific antibody directed against the V3 domains of these virions. In contrast to this lack of V3-specific antibody, the other V3 domains of non-infectious virions were well recognized by antibody. To determine whether the lack of a V3-specific antibody response is due to a progressive loss of antibody during human immunodeficiency virus type 1 (HIV-1) infection, we monitored the anti-V3 antibody response in 90 patients over time. Anti-V3 antibodies were monitored by a V3-specific ELISA using 21 different V3 domains as a fusion with glutathione S-transferase (GST-V3) based upon sequences from 11 HIV-1 patient isolates and 10 sequences from an HIV-1 B subtype consensus-like GST-V3 expression library. This strictly heterologous screening showed a loss of V3-specific antibodies in 20 out of the 90 patients tested. To study the in vivo relevance of these findings we analysed V3 antibody loss in two patients. This strictly autologous antibody screening was performed based upon V3 sequences of the patients' cell-free virions. In both patients the loss of a V3-specific antibody could be detected in parallel to a decline of CD4+ T cells. Moreover, the escape of a distinct V3 variant was shown to correlate closely with the loss of the V3-specific antibody.

Amino Acid Sequence↗
PubMed1996

Can a severe degree of ketoacidosis develop overnight?

Insulin-dependent diabetes mellitus in poor control, alcohol intake associated with extracellular fluid volume contraction, or hypoglycemia may each lead to an increased rate of production of ketoacids. Generally, several days of illness are required before ketoacidosis becomes severe. Two clinical examples are presented to suggest that a severe degree of ketoacidosis may develop over a short period of time, literally overnight. In both examples, there was the ingestion of a modest amount of ethanol. From a quantitative analysis of factors that may influence the rate of production and removal of ketoacids, the following were deduced. Contributing factors to the very rapid development of maximal ketoacidosis could include the absence of a lag period for the conversion of ethanol to acetyl-coenzyme A in the liver and an impaired ability of the brain and kidneys to oxidize ketoacids, especially if these ketoacids are produced very rapidly and/or if less metabolic work is performed by these organs. In special settings, ketoacidosis may develop more rapidly than is generally appreciated.

Adult↗
PubMed1996

Is zinc protoporphyrin an indicator of iron-deficient erythropoiesis in maintenance haemodialysis patients?

BACKGROUND: Zinc protoporphyrin (ZPP), a metabolic intermediate generated in the red blood cell by incorporation of zinc instead of iron, has been suggested to be a sensitive and specific parameter of absolute iron deficiency in haemodialysis (HD) patients. METHODS: We studied 62 HD patients, 29-86 years old, with ZPP levels > 50 mumol/mol haeme (normal value of ZPP < 40 mumol/mol haeme) assessing the value of ZPP as a marker of functional iron deficiency at different cut-off points of ZPP. None of the patients had apparent inflammatory disease, infectious disease, or malignancy. ZPP, haemoglobin, iron and ferritin levels were determined before, and after a 24-week period of once-weekly i.v. administration of 40 mg iron, to determine whether ZPP levels return to normal during adequate iron supplementation (960 mg iron/ patient). RESULTS: There was no significant change in ZPP levels after iron supplementation in patients with a ZPP > 50 mumol/mol haeme (96.7 +/- 49.8 versus 88.4 +/- 43.5 mumol/mol haeme before and after iron administration respectively, P = n.s.). However, in patients with a ZPP > 90 mumol/mol haeme, there was a significant reduction in ZPP levels (141.2 +/- 54.5 versus 108.0 +/- 48.8 mumol/mol haeme, P < 0.001). Serum ferritin increased significantly in both groups. There was no correlation between ZPP and serum ferritin at any time during the study. There was also no correlation between serum aluminium levels and ZPP and no significant difference in changes in ZPP in patients receiving desferrioxamine therapy compared to those not receiving desferrioxamine therapy. We did find a significant correlation between moderately elevated total blood lead concentrations and ZPP levels at the end of the study. The ZPP levels were not significantly different in the range from 50-110 mumol/mol haeme before and after i.v. iron supplementation in the responders (10% increase of haemoglobin or 20% decrease of the recombinant human erythropoietin dose) compared with the non-responders. CONCLUSIONS: Our data indicate that ZPP cannot be used to predict the erythropoietic response to iron supplementation. However, ZPP levels may be an indicator of functional iron deficiency due to blockade of the reticuloendothelial iron release in haemodialysis patients.

Adult↗
PubMed1995

Fos is an essential component of the mammalian UV response.

Mouse 3T3 fibroblasts lacking c-fos were employed to demonstrate an essential function of the UV-inducible transcription factor AP-1 (Fos/Jun) in the response to the cytotoxic effects of short-wavelength ultraviolet (UVC) radiation. Clonogenic survival and proliferation of cells lacking c-fos were drastically reduced following UV irradiation. This UV hypersensitivity manifests itself primarily in increased cell death, partly by apoptosis, and prolonged recovery time from UV-induced cell cycle arrest. Co-culture with wild-type cells did not ameliorate the hypersensitivity of mutant cells. Transcriptional induction of the c-Fos target genes collagenase I, stromelysin-1 and stromelysin-2 by UV is almost absent in cells lacking c-fos which correlates with a reduced UV induction of AP-1 DNA-binding and transactivation activity. The repair of UV-induced DNA lesions was not affected, as shown by unscheduled DNA synthesis and host cell reactivation assays. These data demonstrate that c-Fos is involved in a novel protective function other than DNA repair against the harmful consequences of UVC.

3T3 Cells↗