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J B Klein

Publications and source records attributed to J B Klein.

At least 37 records · Page 2Linked to original sources
PubMed1998

Bacterial phagocytosis activates extracellular signal-regulated kinase and p38 mitogen-activated protein kinase cascades in human neutrophils.

The hypothesis that bacterial phagocytosis by human polymorphonuclear neutrophils (PMNs) stimulates MAPK cascades that regulate respiratory burst activation was tested. Extracellular response kinase (ERK) and p38 kinase, but not c-Jun NH2-terminal kinase, activities were increased within 5 min of phagocytosis of plasma-opsonized Staphylococcus aureus (S-SA), reached maximum at 20-30 min, and remained elevated through 60 min. The role of Fcy receptors was examined using gamma globulin-opsonized SA (IgG-SA), whereas CR3 receptors were activated by particulate beta-glucan. IgG-SA stimulated a maximal ERK activity at 30 min, whereas p38 activity was maximal at 5 min. Beta-glucan stimulated maximal ERK activity at 5 min and maximal p38 activity at 2 min. Non-opsonized bacteria were ingested at 10% of the level of S-SA and stimulated a minimal increase in ERK and p38 activity at 60 min. S-SA stimulation of ERK was inhibited by wortmannin, LY294002, and genistein, but not calphostin C; whereas p38 stimulation was inhibited by calphostin C and genistein, but not wortmannin and LY294002. Simultaneous measurement of phagocytosis and H2O2 production by flow cytometry was used to assess the role of ERKs and p38 kinase in phagocytosis. The MEK inhibitor PD098059 had no significant effect on phagocytosis or H2O2 production. The p38 kinase inhibitor SB203580 significantly attenuated H2O2 production, whereas phagocytosis was unaffected. In conclusion, bacterial phagocytosis stimulates ERK and p38 activation by distinct signal transduction pathways. Phagocytosis-stimulated p38 kinase activity is necessary for optimal H2O2 production.

Antigens, CD↗
PubMed1997

Formyl peptide receptors are coupled to multiple mitogen-activated protein kinase cascades by distinct signal transduction pathways: role in activation of reduced nicotinamide adenine dinucleotide oxidase.

Formyl peptide receptor activation of three mitogen-activated protein kinase (MAPK) cascades, extracellular signal-regulated kinases (ERKs), N-terminal kinases (JNKs), and p38 MAPK was examined in differentiated HL-60 granulocytes. FMLP stimulated a concentration- and time-dependent increase in ERK, JNK, and p38 MAPK activities, all of which were dependent on a pertussis toxin-sensitive G protein. Pharmacologic inhibitors were used to examine the roles of tyrosine kinases, phosphatidylinositol 3-kinase, protein kinase C, and phospholipase C. FMLP-stimulated ERK activity was dependent on tyrosine kinases, phosphatidylinositol 3-kinase, protein kinase C, and phospholipase C; p38 MAPK activation was dependent on phosphatidylinositol 3-kinase and phospholipase C; while JNK activation was independent of all of these signaling components. The mitogen-activated protein kinase/ERK kinase inhibitor PD098059 reduced ERK activation by 90%, while an inhibitor of p38 MAPK, SB203580, inhibited p38 MAPK activation by 80%. Both PD098059 and SB203580 inhibited FMLP-stimulated superoxide release, as did inhibitors directed against protein kinase C, tyrosine kinases, and phosphatidylinositol 3-kinase. We conclude that formyl peptide receptors are coupled to three MAPK cascades by Gi proteins. ERKs, p38 MAPK, and JNKs are each activated by distinct proximal signal transduction pathways. Activation of p38 MAPK is necessary for FMLP stimulation of respiratory burst activity; however, a second signal that may involve ERK is also required for this activity.

Calcium-Calmodulin-Dependent Protein Kinases↗
PubMed1997

Safety and tolerability of cyclosporine microemulsion versus cyclosporine: two-year data in primary renal allograft recipients: a report of the Neoral Study Group.

BACKGROUND: The new microemulsion formulation of cyclosporine (CsA-ME) is more bioavailable than cyclosporine (CsA) in de novo renal transplant patients. Therefore, it was of interest to compare the safety profile of each formulation in such patients. METHODS: In a multicenter, double-blind, parallel-group study, 101 renal transplant recipients were randomized after transplantation to receive either CsA (n=50) or CsA-ME (n=51) capsules twice daily for 2 years. Of these patients, 54 (CsA, n=26; CsA-ME, n=28) completed 1 year of the study and entered the second-year, double-blind extension. Initial dose at the time of transplantation was 5 mg/kg b.i.d.; doses were titrated to target trough levels. METHODS: The mean (+/- SD) doses at the end of 2 years were 4.6 +/- 1.8 and 3.8 +/- 1.1 mg/kg per day for CsA- and CsA-ME-treated patients, respectively. The mean (+/- SD) CsA trough levels at end point were 187 +/- 63 and 210 +/- 95 ng/ml for CsA- and CsA-ME-treated patients, respectively. At least one adverse event was reported by 25/26 (96%) of CsA- and 27/28 (96%) of CsA-ME-treated patients. No patient discontinued the study because of adverse events. No deaths occurred during the study. Renal function, as measured by serum creatinine levels, and blood pressure were comparable over time in both treatment groups. CONCLUSIONS: There was no significant difference in safety and tolerability between CsA- and CsA-ME-treated kidney recipients for 2 years after transplantation.

Adolescent↗
PubMed1997

TNF translationally modulates the expression of G1 protein alpha(i2) subunits in human polymorphonuclear leukocytes.

Priming of polymorphonuclear leukocyte responses to chemoattractants by TNF plays an important role in host defenses and inflammatory responses. TNF-induced priming is associated with an 80% increase in the membrane density of G alpha(i2) protein that is coupled to chemoattractant receptors. The present study examines the hypothesis that TNF stimulates increased synthesis of alpha(i2). Within 10 min of addition, TNF stimulated a significant increase in total cellular G alpha(i2), as determined by pertussis toxin-catalyzed ADP ribosylation, which was blocked by the translation inhibitor cycloheximide. Immunoprecipitation of biosynthetically labeled alpha(i2) showed that TNF increased alpha(i2) synthesis by about 20% at 10 min. Nuclear run-ons showed no change in alpha(i2) mRNA synthesis in TNF-treated cells; however, steady state alpha(i2) mRNA levels were reduced following a 10-min exposure to TNF. Pretreatment with cycloheximide prevented the TNF-induced reduction in steady state alpha(i2) mRNA levels. Therefore, TNF stimulates alpha(i2) protein synthesis and mRNA degradation in the same time frame as priming. The increased alpha(i2) synthesis results from increased translation, not transcription, of alpha(i2) mRNA. Simultaneous G alpha(i2) protein synthesis and mRNA degradation provide a mechanism by which TNF priming is associated with a rapid, self-limiting increase in G protein expression.

Biological Transport↗
PubMed1996

The pharmacokinetics of a microemulsion formulation of cyclosporine in primary renal allograft recipients. The Neoral Study Group.

This study was a randomized, double-blind, 12-week comparison of the pharmacokinetics, safety, and tolerability of two cyclosporine (CsA) formulations, cyclosporine emulsion capsules and oral solution for microemulsion and cyclosporine, in the postoperative management of renal transplant patients. Of the 101 patients, aged 18 to 65, who entered the study, 89 were evaluable for pharmacokinetics. Initial dosage was 10 mg/kg per day, administered twice daily in two equal doses. Dosages were adjusted to achieve target CsA concentrations. The pharmacokinetic (PK) parameters (dose-normalized) of greatest interest were maximum blood concentration (C(max)/dose), time to reach maximum concentration (t(max), area under the blood concentration-vs.-time curve (AUC/dose), and trough blood concentrations (Co h/dose). The relative CsA bioavailabilty was found to be significantly enhanced with cyclosporine emulsion compared with cyclosporine with a 16% to 31% increase in AUC and a 32% to 42% increase in C(max). Intrapatient variability of PK parameters was significantly lower with cyclosporine emulsion than with cyclosporine for AUC, C(oh), t(max), and C(max) in many instances. This indicates a more consistent, rapid, and more complete total absorption of CsA. Despite higher CsA C(max) levels and AUCs with cyclosporine emulsion, safety and tolerability (detailed in a parallel report) were comparable to those of cyclosporine. The PK advantages of cyclosporine emulsion over cyclosporine are either independent of food conditions or possibly reflective of more consistent absorption of CsA with cyclosporine emulsion. The findings suggest that de novo use of cyclosporine emulsion may simplify and improve management of organ transplant recipients and that the PK advantages of cyclosporine emulsion may translate into clinical benefits.

Adolescent↗
PubMed1996

The safety and tolerability of cyclosporine emulsion versus cyclosporine in a randomized, double-blind comparison in primary renal allograft recipients. The Neoral Study Group.

A 12-week, randomized, double-blind, multicenter pharmacokinetics study was conducted to compare the clinical safety and tolerability of cyclosporine capsules and oral solution for microemulsion and cyclosporine in 101 primary renal transplant recipients Cyclosporine emulsion has more complete absorption and improved bioavailability compared with cyclosporine, and dosing of both cyclosporine formulations was adjusted to achieve comparable whole-blood trough levels. Mean serum creatinine values were higher in the cyclosporine emulsion group at baseline, 8, and 12 weeks (P<0.05). The incidence of acute rejection was similar in both treatment groups although fewer patients required monoclonal antibody therapy in the cyclosporine group (31% vs. 82%, respectively). Despite the increased bioavailability of cyclosporine emulsion, no significant differences in the incidence of adverse events were observed; the safety, tolerability, and efficacy of cyclosporine emulsion and cyclosporine were comparable.

Administration, Oral↗
PubMed1996

Azotemia, TNF alpha, and LPS prime the human neutrophil oxidative burst by distinct mechanisms.

The oxidative burst of neutrophils from azotemic patients (AzoPMNs) is primed for an enhanced response compared to neutrophils from normal subjects (NorPMNs). The mechanism for this priming is unknown, although TNF alpha does not further prime AzoPMNs. The present study examines the hypothesis that azotemia and TNF alpha prime neutrophils by the same mechanism. Formyl peptide receptor expression and degranulation were not primed in AzoPMNs, but were primed by both LPS and TNF alpha. LPS was also able to prime the AzoPMN oxidative burst. Guanine nucleotide exchange by multiple guanine nucleotide binding proteins, including heterotrimeric G-proteins and low molecular weight GTP-binding proteins (LMWGs), was increased in AzoPMNs, as demonstrated by GTP gamma S binding and azidoanilide GTP photoaffinity labeling. The plasma membrane density of G-protein alpha i2, alpha i3, and alpha s subunits and the density in the cytosol of the LMWG, Rap1A, was present in significantly greater amounts on plasma membranes from AzoPMNs. FMet-Leu-Phe-stimulated phospholipase D activity, but not basal activity, was significantly greater in AzoPMNs. Finally, incubation of NorPMNs in plasma from azotemic patients resulted in a significant increase in basal GTP gamma S binding. These results demonstrate that priming of AzoPMNs is restricted to oxidative burst activity and that it occurs by a mechanism distinct from that utilized by TNF alpha and LPS. While the exact mechanism remains unknown, it appears to involve a plasma factor and changes in LMWG expression or activity.

Adult↗
PubMed1995

TNF-alpha stimulates increased plasma membrane guanine nucleotide binding protein activity in polymorphonuclear leukocytes.

TNF-alpha enhances the response of polymorphonuclear leukocytes (PMN) to chemoattractants: however, the mechanism by which this occurs is unclear. We addressed the hypothesis that TNF-alpha enhances the PMN response to chemoattractants by increasing chemoattractant receptor transmembrane signaling, using fMLP as the model chemoattractant. fMLP-stimulated guanine nucleotide binding (G) protein activation was significantly increased in plasma membranes isolated from PMNs exposed to TNF-alpha 100 U/ml for 10 minutes (TNF-M), compared to membranes from control cells (CM). Formyl peptide receptor number and affinity were not significantly different in CM and TNF-M. Gi and Gs content were increased in TNF-M as measured by pertussis toxin and cholera toxin (CT) catalyzed ADP-ribosylation, respectively. The increased Gi was coupled to the formyl peptide receptor as shown by receptor-specific CT labeling of Gi. Immunoblot analysis showed that both G alpha i2 and G alpha 3 were increased in TNF-M. The functional activity of the increased G protein content was demonstrated by increased NaF-stimulated phospholipase D activity in TNF-alpha-treated PMNs. We conclude that TNF-alpha rapidly stimulates increased PMN plasma membrane expression of G proteins that couple formyl peptide receptors to effector enzymes. Regulation of G protein expression may be a significant mechanism by which TNF regulates PMN function.

Cell Membrane↗
PubMed1995

Chemoattractant receptor-specific differences in G protein activation rates regulate effector enzyme and functional responses.

The hypothesis that disparate neutrophil functional responses to various chemoattractants are regulated by receptor-specific rates of G protein activation was examined in HL-60 granulocytes. The initial rates of G protein activation and the affinity of receptor-stimulated G proteins for GTP gamma S in HL-60 membranes stimulated by fMet-Leu-Phe, C5a, and leukotriene B4 (LTB4) differed significantly among the chemoattractants, with a rank order of fMet-Leu-Phe > C5a > LTB4. Equilibrium GTP gamma S binding showed that all three chemoattractants activated a common pool of G proteins. Stimulation of phospholipase D activation, measured as phosphatidylethanol generation, and superoxide release in intact cells also occurred with a rank order of fMet-Leu-Phe > C5a > LTB4. On the other hand, the rank order of receptor affinities for ligand and of the EC50 of chemoattractant stimulation of GTP gamma S binding was C5a > LTB4 > fMet-Leu-Phe. C5a and LTB4 receptor densities were similar but were less than formyl peptide receptor density. Graded pertussis toxin treatment proportionally reduced superoxide release and phospholipase D activation to all three chemoattractants. The results suggest that receptor-specific differences in G protein affinity for guanine nucleotides lead to different rates of guanine nucleotide exchange and, thereby, contribute to disparate effector enzyme and functional responses.

Amino Acid Sequence↗
PubMed1994

Effect of prenylcysteine analogues on chemoattractant receptor-mediated G protein activation.

The hypothesis that carboxylmethylation of gamma subunits plays a role in G protein activation was tested by examining the ability of N-acetyl-S-farnesyl-L-cysteine (AFC) and its methyl ester (AFC-ME) to inhibit G protein-mediated signalling in intact HL-60 granulocytes and isolated HL-60 plasma membranes. Incubation of HL-60 granulocytes with AFC or AFC-ME inhibited superoxide release stimulated by fMet-Leu-Phe, but not by opsonized bacteria. AFC-ME, but not AFC, inhibited NaF- and PMA-stimulated superoxide release. Addition of AFC to HL-60 membranes inhibited fMet-Leu-Phe-, leukotriene B4- (LTB4) and C5a-stimulated GTP gamma S binding and GTP hydrolysis more potently than it inhibited basal guanine nucleotide exchange. AFC-ME inhibited basal- and ligand-stimulated G protein activation with equal potency, but less potently than AFC. AFC also inhibited mastoparan-stimulated GTP gamma S binding. Binding of fMet-Leu-Phe and LTB4 to HL-60 membranes was completely inhibited by AFC, while AFC-ME inhibited ligand binding by less than 50%. Neither AFC nor AFC-ME inhibited pertussis toxin or cholera toxin-catalysed ADP-ribosylation of alpha i. It was concluded that AFC interrupts signal propagation in G protein-dependent pathways by multiple mechanisms, including inhibition of ligand-receptor interactions, of receptor-G protein coupling and of guanine nucleotide binding to G proteins. Carboxylmethylation alters the specificity of AFC interruption of signal propagation in intact cells and isolated membranes.

Acetylcysteine↗
PubMed1994

Influence of suspension on the oxidative burst by rat neutrophils.

The influence of spaceflight on the oxidative burst of neutrophils is not known. The present study was designed to evaluate the influence of antiorthostatic suspension, a ground-based modeling system designed to simulate certain aspects of weightlessness that occur after spaceflight, on the capacity of rat neutrophils to express the oxidative burst, an important host defense mechanism against microbial pathogens. Rats were suspended in whole body harnesses in the antiorthostatic orientation for a 3- or 7-day period. Control rats were suspended orthostatically or allowed to remain in vivarium cages without the attachment of any suspension materials. After suspension, peripheral blood was harvested and neutrophils were isolated by density gradient centrifugation. The enriched neutrophil preparations were stimulated with N-formyl-methionyl-leucine-phenylalanine and phorbol myristic acid to induce the oxidative burst. It was found that neutrophils isolated from suspended animals released the same levels of superoxide anion as did vivarium control animals that were not suspended, indicating that whole body suspension did not alter this aspect of rat neutrophil function.

Animals↗
PubMed1994

MR imaging diagnosis of central precocious puberty: importance of changes in the shape and size of the pituitary gland.

OBJECTIVE: Central precocious puberty occurs as a result of premature pituitary stimulation and increased secretion of gonadotropins. The aims of this study were to analyze MR imaging findings in the pituitary glands of children with central precocious puberty compared with matched control subjects, to define MR imaging-derived variables useful in the diagnosis of central precocious puberty, and to correlate MR imaging-derived variables with the hormonal profile and other imaging and clinical findings. MATERIALS AND METHODS: Twenty-six children with central precocious puberty (two boys and 24 girls) were divided into two subgroups according to MR imaging findings: idiopathic (21 patients) and nonidiopathic (five patients: three hypothalamic hamartomas, one pineal tumor, one empty sella syndrome). The control group consisted of 17 normal age- and sex-matched children (two boys, 15 girls). Analyzed parameters included pituitary height, length, width, midsagittal cross-sectional area, calculated volume, and shape. The shape was assessed by a pituitary grading system and two other shape indexes (length-to-height and length-to-width ratios). Pituitary grade was defined by the concavity of the upper pituitary surface (grade 1 = marked concavity, grade 2 = mild concavity, grade 3 = flat, grade 4 = mild convexity, grade 5 = marked convexity). RESULTS: Pituitary grade showed a highly significant difference among groups (p < .001). Area, height, and length-to-height ratio were significantly different (p < .05), whereas length, width, length-to-width ratio, and volume were not. There was no significant difference in any of the variables compared between idiopathic and nonidiopathic groups. When selected variables (pituitary grade, area, height, length) in the central precocious puberty group were stratified by bone age and findings on pelvic sonograms, patients with advanced bone age had a significantly higher pituitary grade (p < .01) and had a tendency toward a greater pituitary length. Pituitary size and shape correlated with the hormonal profile. CONCLUSION: Change in pituitary grade is the most helpful variable for the diagnosis of central precocious puberty in a prepubertal child. A high pituitary grade (4 or above) is highly predictive of central precocious puberty, with the highest specificity and positive predictive value, but with low sensitivity. The use of combinations of high pituitary grade with two other positive findings (height and area greater than 1 SD from the respective means in the control group) improves the sensitivity, specificity, and predictive value of MR imaging in the diagnosis of central precocious puberty.

Analysis of Variance↗
PubMed1993

Role of isoprenoid metabolism in chemotactic peptide receptor-mediated G protein activation.

The role of isoprenylation in formyl peptide receptor-mediated G protein activation was studied using plasma membranes isolated from normal HL-60 granulocytes and from cells in which isoprenylation was inhibited with mevastatin. Plasma membrane expression of formyl peptide receptors and G protein beta subunits, but not alpha i2 and alpha i3, was significantly reduced by inhibition of isoprenylation. This reduced expression resulted in impaired basal and fMet-Leu-Phe-stimulated G protein activation.

Adenosine Diphosphate Ribose↗
PubMed1993

Differential uncoupling of chemoattractant receptors from G proteins in retinoic acid-differentiated HL-60 granulocytes.

The ability of HL-60 cells differentiated with DMSO or retinoic acid (RA) to serve as models for polymorphonuclear leukocytes was examined. NaF and FMLP stimulated superoxide release from both groups of cells, but the response was attenuated in RA-differentiated cells. Isolated plasma membranes from DMSO-M and RA-differentiated (RA-M) cells were used to examine receptor and guanine nucleotide binding regulatory protein (G protein) expression. Ligand binding showed that formyl peptide and leukotriene B4 receptors were present in equal density on DMSO-M and RA-M. Formyl peptide receptors on RA-M failed to demonstrate high affinity binding sites. Both G alpha i2 and G alpha i3 were present in equal density on RA-M and DMSO-M by immunoblotting. FMLP, but not leukotriene B4, failed to stimulate guanosine 5'-(gamma-thio)triphosphate binding or GTP hydrolysis and guanine nucleotides failed to inhibit FMLP binding to RA-M. Addition of solubilized membranes containing normal formyl peptide receptors to solubilized RA-M reconstituted FMLP stimulated G protein activation, whereas addition of normal G proteins had no effect. We conclude that formyl peptide, but not leukotriene B4, receptors are uncoupled from G proteins in RA-differentiated HL-60 granulocytes. Although RA-differentiated cells are an inadequate model of neutrophil transmembrane signaling, they may provide a unique model by which to study the molecular requirements for formyl peptide receptor-G protein interactions.

Cell Differentiation↗
PubMed1993

Flow cytometric analysis of altered mononuclear cell transmembrane potential induced by cyclosporin.

Immunosuppression by the fungal metabolite cyclosporin A (CsA) is characterized by functional inhibition, rather than destruction of cells. Because activation of immune cells involves intracellular signalling events associated with modulations of cell transmembrane potential (TMP), we tested the ability of cyclosporin A (CsA) to modulate immune mononuclear cell TMP in vitro using a TMP sensitive cationic dye, dihexyloxacarbocyanine (DIOC6(3)). All analyses were performed by flow cytometry. CsA increased TMP in monocytes and lymphocytes isolated from the blood of healthy human volunteers. CsA-induced hyperpolarization was time and concentration dependent in monocytes while the lymphocyte hyperpolarization, although time dependent, was evident over the entire range of CsA concentrations tested. CsA-induced hyperpolarization of lymphocytes was dependent on potassium ion (K+) efflux as indicated by the absence of hyperpolarization in 154 mM KCl or with pretreatment with 100 microM quinine (an inhibitor of K+ channels). Monocyte hyperpolarization by CsA was not inhibited in either system. Dihydrocyclosporin C (DH-CsC), an immunosuppressive analog of CsA, also hyperpolarized mononuclear cells. The anionic TMP sensitive dye bis oxonal (diBA-C4) indicated that CsA treatment depolarized mononuclear cell plasma membranes. The mitochondrial poison carbonyl cyanide p-trifluoromethoxy-phenyl-hydrazine (FCCP) eliminated CsA induced hyperpolarization and also indicated that CsA caused plasma membrane depolarization. We conclude that brief in vitro exposure to cyclosporin alters the transmembrane electrical potential of human lymphocytes and monocytes.

Cyclosporine↗
PubMed1993

Extracorporeal shock-wave lithotripsy in transplanted kidney.

Urolithiasis is the least described urologic sequela of renal transplantation. We describe a renal transplant patient who presented with painless gross hematuria. An intravenous pyelogram demonstrated a 4 x 7-mm calculi in the region of the ureteropelvic junction, causing moderate hydronephrosis. The patient was treated successfully with extracorporeal shock-wave lithotripsy (ESWL). Serum creatinine and twenty-four-hour creatinine clearance were unchanged from levels prior to ESWL.

Adult↗
PubMed1993

The effect of interferon-gamma on rejection and neutrophil function following transplantation.

Rats were used as a model for a living heterotopic cardiac allograft organ transplant. Rats treated in this model with recombinant rat interferon-gamma (IFN-gamma) showed accelerated rejection in a dose-dependent fashion. However, rats treated with maintenance doses of cyclosporine and IFN-gamma expressed increased rejection at 20 days that had resolved completely by 45 days post-transplantation. Polymorphonuclear leukocytes (neutrophils) were isolated from the blood of rats, and their function was determined by treating the cells with f-Met-Leu-Phe (fMLP) and measuring superoxide produced. Results indicate that the neutrophils from rats treated with maintenance doses of cyclosporine and IFN-gamma still had increased IFN-gamma-modulated fMLP-induced respiratory burst and that maintenance cyclosporine therapy can inhibit the IFN-gamma-mediated accelerated rejection without compromising the antimicrobial effects of IFN-gamma treatment.

Abdomen↗