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S Adler

Publications and source records attributed to S Adler.

At least 73 records · Page 4Linked to original sources
PubMed1998

Site-directed mutagenesis identifies amino acid residues associated with the dehydrogenase and isomerase activities of human type I (placental) 3beta-hydroxysteroid dehydrogenase/isomerase.

3beta-hydroxysteroid dehydrogenase/steroid delta5-->4-isomerase (3beta-HSD/isomerase) was expressed by baculovirus in Spodoptera fungiperda (Sf9) insect cells from cDNA sequences encoding human wild-type I (placental) and the human type I mutants - H261R, Y253F and Y253,254F. Western blots of SDS-polyacrylamide gels showed that the baculovirus-infected Sf9 cells expressed the immunoreactive wild-type, H261R, Y253F or Y253,254F protein that co-migrated with purified placental 3beta-HSD/isomerase (monomeric Mr=42,000 Da). The wild-type, H261R and Y253F enzymes were each purified as a single, homogeneous protein from a suspension of the Sf9 cells (5.01). In kinetic studies with purified enzyme, the H261R mutant enzyme had no 3beta-HSD activity, whereas the Km and Vmax values of the isomerase substrate were similar to the values obtained with the wild-type and native enzymes. The Vmax (88 nmol/min/mg) for the conversion of 5-androstene-3,17-dione to androstenedione by the Y253F isomerase activity was 7.0-fold less than the mean Vmax (620 nmol/min/mg) measured for the isomerase activity of the wild-type and native placental enzymes. In microsomal preparations, isomerase activity was completely abolished in the Y253,254F mutant enzyme, but Y253,254F had 45% of the 3beta-HSD activity of the wild-type enzyme. In contrast, the purified Y253F, wild-type and native enzymes had similar Vmax values for substrate oxidation by the 3beta-HSD activity. The 3beta-HSD activities of the Y253F, Y253,254F and wild-type enzymes reduced NAD+ with similar kinetic values. Although NADH activated the isomerase activities of the H261R and wild-type enzymes with similar kinetics, the activation of the isomerase activity of H261R by NAD+ was dramatically decreased. Based on these kinetic measurements, His261 appears to be a critical amino acid residue for the 3beta-HSD activity, and Tyr253 or Tyr254 participates in the isomerase activity of human type I (placental) enzyme.

Amino Acid Sequence↗
PubMed1998

Identification of glial cell proliferation in early multiple sclerosis lesions.

Multiple sclerosis (MS) is a chronic inflammatory disease of the central nervous system which leads to destruction of myelin sheaths. The patterns of cell proliferation in the early course of the disease are largely unknown. The present study used immunohistochemical identification of proliferating glial cells in stereotactic brain biopsy material of eight patients with early chronic MS. Double-labelling with the proliferation marker MIB-1 detected proliferating oligodendrocytes (MOG), astrocytes (GFAP) and microglia/macrophages (Ki-M1P). The majority of proliferating cells were macrophages/microglia when compared with oligodendrocytes (P > 0.005) or astrocytes (P > 0.0005); only a minor proportion of microglia/macrophages, however, proliferated in situ. Astrocytic and oligodendroglial proliferation was sparse to absent and showed significant variations between different patients. There were statistically significant differences when comparing the amount of proliferation between lesions of different demyelinating activity: highest numbers of proliferating cells were found in early active lesions compared with demyelinated and early remyelinated lesions (P > 0.05) or the periplaque white matter (P > 0.01). MOG-positive oligodendrocytes proliferated occasionally in the early stages of lesion formation; this proliferation occurred in four cases but was independent of the stage of the disease. Since MOG is expressed by mature oligodendrocytes, and not by immature precursors, this might suggest a potential role for the proliferation of mature surviving oligodendrocytes with subsequent remyelination.

Adult↗
PubMed1998

Insulin-like growth factor system and the kidney: physiology, pathophysiology, and therapeutic implications.

The insulin-like growth factor (IGF) system, consisting of IGF-I and IGF-II, their binding proteins, and their receptors, is expressed in a spatial organization in the nephron, but circulating IGFs also affect the kidney. Renal and systemic IGF-I and the binding proteins are regulated by growth hormone and nutritional status. In the kidney, IGF-I dilates the resistance-regulating microvasculature, increases glomerular filtration rate, and promotes tubular phosphate and possibly sodium absorption. IGF-I contributes to compensatory renal growth in a variety of experimental models and may modestly contribute to progressive glomerular sclerosis. In chronic renal failure and the nephrotic syndrome, there are several abnormalities in the IGF system. In chronic renal failure, IGF-I increases renal function and may improve nutritional status due to its anabolic effects. IGF-I accelerates the recovery of renal function in animal models of acute renal failure, but results from clinical trials were less salutary. Several questions regarding the role of the IGF system in normal and abnormal renal biology and potential therapeutic applications in kidney diseases remain unanswered.

Acute Kidney Injury↗
PubMed1998

Comparative effects of the selective estrogen receptor modulators (-)-, (+)- and (+/-)-Z bisdehydrodoisynolic acids on metabolic and reproductive parameters in male and female rats.

Doisynolic acids are non-steroidal estrogenic compounds originally obtained from alkali fusion of estrone and equilenin. Z-bisdehydrodoisynolic acids (Z-BDDA) exhibit a low binding affinity accompanied by a disproportionately high biologic activity. Two experiments were designed to investigate the chronic effects of (+)-, (-)- and (+/-)-Z-BDDA and (+)-17beta-estradiol (E2) in male and female rats. The (+)-, (-)- and (+/-)-forms Z-BDDA were prepared and injected, daily for four to six weeks into male and female rats and changes in body weight, food intake, metabolic parameters, and reproductive parameters were investigated. Results from both experiments demonstrate that in male and female rats, (+)- and (+/-)-Z-BDDA had similar estrogenic effects on reproductive organ weight. Surprisingly, (-)-Z-BDDA did not induce the increase in uterine weight observed with (+)- and (+/-)-Z-BDDA and E2, demonstrating selective estrogen receptor modulation (SERM). Beneficial metabolic effects, although compound- and gender-specific, included a significant weight repression, reduction in cholesterol, reduction in blood glucose, and positive alterations in body fat distribution. Future research defining the optimal dosages of (-)-Z-BDDA that will maximize beneficial effects and minimize undesirable effects on reproductive tissues will lead to more efficacious treatment options for endocrine-responsive conditions in males and females.

Animals↗
PubMed1998

Glomerular endothelial cell injury mediated by shiga-like toxin-1.

The hemolytic uremic syndrome is an important cause of acute renal failure and is often associated with prior infection with enterotoxigenic strains of Escherichia coli. Significant pathologic changes in the glomerulus include endothelial cell swelling, detachment, and intravascular coagulation. We investigated the potential pathogenetic mechanisms of this disease by exposing cultured rat glomerular endothelial cells to shiga-like toxin 1 (SLT-1). Glomerular endothelial cell viability and protein synthesis were not affected by 10(-9) M SLT-1. Synthesis and release of thromboxane (Tx) A2, measured as the stable metabolite TxB2, and of 12-(S)-HETE were each increased 1.6 +/- 0.1 fold (p < 0.05). No change was observed in the production of PGE2 or 6-keto-PGF1 alpha, the active metabolite of PGI2. SLT-1 (10(-9) M) significantly increased cell retraction and the formation of gaps in the glomerular endothelial cell monolayer for up to 6 h following exposure. A similar effect was seen with 0.1 microM 12-(S)-HETE. SLT-1 and 0.01 microM 12-(S)-HETE also significantly decreased adhesion of glomerular endothelial cells to fibronectin (75 +/- 4 and 65 +/- 2% of control, respectively; p < 0.001) and laminin (81 +/- 5 and 67 +/- 4% of control, respectively; p < 0.01). These results support a role for SLT-1 in the production of the pathologic changes seen in the hemolytic uremic syndrome through stimulation of production of procoagulant, vasoconstrictor arachidonic acid metabolites with potential to produce endothelial injury and through alterations in glomerular endothelial cell adhesion to components of the glomerular basement membrane.

12-Hydroxy-5,8,10,14-eicosatetraenoic Acid↗
PubMed1998

Characterization of a human-specific regulator of placental corticotropin-releasing hormone.

The hypothalamic hormone CRH is also expressed in the placentas of humans and higher primates and may play an important role in the regulation of labor. In choriocarcinoma cell lines, activation of cAMP-dependent pathways increases human (h)CRH reporter gene expression. A cAMP-responsive region distinct from the cAMP response element at -220 bp, has been identified between -200 and -99 bp, and a candidate transcription factor was identified in nuclear extracts of human, but not rodent, choriocarcinoma cell lines. This region, which does not contain a canonical cAMP response element (CRE), transfers protein kinase A responsiveness to a heterologous promoter. Electromobility shift assays and methylation and uracil interference studies localized factor binding to a 20-bp region from -128 to -109 bp of the hCRH promoter. This 20-bp fragment exhibited a similar shift in nuclear extracts from both human term placenta and from human JEG-3 cells. Base contacts, identified in interference studies, were confirmed as critical for binding, as a mutation of these bases abolished factor binding. Furthermore, a CRH promoter containing this mutation exhibited a diminished response to forskolin. UV cross-linking demonstrated the protein in nuclear extracts from human, but not rodent, choriocarcinoma cell lines and estimated its size as 58 kDa. Although this factor participates in cAMP-regulated gene expression, competition electrophoretic mobility assays demonstrated that the factor does not bind to a CRE. Furthermore, neither anti-CREB nor anti-ATF2 antibodies alter factor binding. These data identify this 58-kDa protein as the human-specific CRH activator previously identified as a candidate factor contributing to the species-specific expression of CRH in human placenta.

Activating Transcription Factor 2↗
PubMed1998

CD59 protects glomerular endothelial cells from immune-mediated thrombotic microangiopathy in rats.

CD59 is a cell membrane-bound complement regulatory protein on glomerular cells that inhibits C5b-9 assembly and insertion. This report describes a recently developed model of immune thrombotic microangiopathy (TMA) induced by the renal artery perfusion of anti-glomerular endothelial cell (anti-GEN) antibody. To examine the role of CD59 in protecting the GEN from immune-mediated injury, rats underwent selective renal artery perfusion with F(ab')2 fragments of anti-CD59 monoclonal antibody to block CD59 activity or control mouse IgG followed by anti-GEN antibody or control goat IgG. Neutralization of CD59 in normal rats did not result in any significant functional or histologic changes. Perfusion with anti-CD59 did not change deposition of the pathogenic anti-GEN IgG used to induce the TMA model. However, neutralization of CD59 in the TMA model resulted in more C5b-9 formation in glomeruli, accompanied by increased platelet and fibrin deposition, more severe endothelial injury, and reduced renal function compared with the animals perfused with control F(ab')2 fragments. These results demonstrate directly that CD59 serves a protective role for GEN in this TMA model of rats, and confirm that C5b-9 formation has a critical pathogenic role in the mediation of the disease. CD59 may play an important role in protecting glomerular endothelium from other complement-mediated types of injury.

Animals↗
PubMed1997

Transcriptional regulation of estrogen-responsive genes by non-steroidal estrogens: doisynolic and allenolic acids.

Estrogen receptor (ER), a member of the nuclear receptor superfamily, exerts prominent physiological roles in both humans and other species by acting directly as a transcription factor, altering nuclear gene expression. One peculiarity of estrogenic regulation is that it is affected by a wide variety of non-steroidal compounds in addition to the natural hormone, estradiol. Doisynolic and allenolic acid compounds are non-steroidal compounds that act as potent estrogens in animal studies, yet bind to ER extremely poorly in competitive binding assays, raising the possibility of alternative molecular mechanisms for the observed estrogenic effects. In this work we demonstrate that (+/-)-Z-bisdehydrodoisynolic acid, (+/-)-Z-bisdehydrodoisynolic acid 3-methyl ether, and (-) allenolic acid can interact directly with ER. These compounds all serve as ligands for ER in mechanism-specific tissue culture-based reporter gene assays for both positive and negative gene regulation. We have also used a novel assay based on electromobility shift by ER for directly determining relative binding affinities for ER. In addition, we show cell-type-specific activity differences for (+/-)-Z-bisdehydrodoisynolic acid 3-methyl ether, supporting clinical observations indicating a higher potency of this compound in female animals than in humans.

Animals↗
PubMed1997

TNF-alpha regulation of C3 gene expression and protein biosynthesis in rat glomerular endothelial cells.

Glomerular endothelial cells are an important site of interaction with the cellular and soluble components of inflammation. To investigate the capacity of these cells to synthesize complement they were cloned from isolated rat glomeruli. Messenger RNA (mRNA) was extracted from the cells, reverse transcribed and used as the template to identify specific gene transcripts with the polymerase chain reaction (PCR). mRNA coding for the third component of the complement cascade (C3) was detected in unstimulated endothelial cells, whereas no message for the fourth component (C4) could be demonstrated. Using a semiquantitative method of PCR, we found that the expression of C3 is up-regulated by the cytokine tumor necrosis factor-alpha (TNF-alpha), but not by the cytokines interferon-gamma (IFN-gamma) and interleukin 1 alpha (IL-1 alpha). The increase in levels of C3 mRNA occurred in a time and dose dependent manner. This increase was dependent on new protein synthesis. Production of the C3 protein was demonstrated by radiolabeling and immunoprecipitation, and this also was stimulated by TNF-alpha. In conclusion, we demonstrate the production of C3 by microvascular endothelium of glomerular origin and its stimulation by TNF-alpha. We believe that this local synthesis could have a role in the pathogenesis of disease, however, the nature of this role at present remains unclear.

Animals↗
PubMed1997

Characterization of C3 receptors on cultured rat glomerular endothelial cells.

In this study we characterized C3 receptors on cultured rat glomerular endothelial cells (GEnC), using immunochemical and molecular techniques. GEnC membrane proteins were immunoprecipitated with a polyclonal antibody directed towards mouse complement receptor 2 (CR2). This anti-MCR2 immunoprecipitated GEnC proteins of 120 and 150 kDa. By immunohistochemistry, anti-MCR2 stained GEnC in rat glomeruli in vivo. Given the presence of CR2-like proteins on GEnC, subsequent studies were done to determine whether GEnC had C3-binding proteins. GEnC proteins of 80, 200, and 300 kDa specifically bound to columns of rat C3d-Sepharose and C3b-Sepharose, illustrating that these proteins were binding to the C3d portion of C3. The 80, 200, and 300 kDa C3d-binding proteins were distinct from the 120 and 150 kDa anti-MCR2 reactive proteins, as shown by immunoabsorption studies. Next, a specific cDNA probe for rat CR2 was generated by RT-PCR. Oligonucleotides were chosen from highly conserved regions in mouse and human CR2 spanning 224 bases, with the rationale that these would also be conserved in the rat. A 224 bp PCR product was generated from both rat GEnC and rat kidney cDNA, illustrating the presence of CR2 mRNA in these tissues. By Northern analysis, the CR2 PCR product hybridized to mRNA of 2 and 5 kb from GEnC. The 5 kb transcript was also identified in rat kidney mRNA. Therefore, proteins immunologically related to mouse CR2 are present in GEnC in vitro and in vivo. C3d-binding proteins of 80, 200, and 300 kDa are also present on rat GEnC, yet these appear to be immunologically distinct from the proteins identified by anti-MCR2. Whether the GEnC CR2 mRNA transcripts of 2 and 5 kb are translated into the 80 and 200 kDa C3d-binding proteins or the 120 and 150 kDa mouse CR2-like proteins remains to be defined.

Animals↗
PubMed1997

Predictors of the progression of renal disease in the Modification of Diet in Renal Disease Study.

The Modification of Diet in Renal Disease (MDRD) Study examined the effects of dietary protein restriction and strict blood pressure control on the decline in glomerular filtration rate (GFR) in 840 patients with diverse renal diseases. We describe a systematic analysis to determine baseline factors that predict the decline in GFR, or which alter the efficacy of the diet or blood pressure interventions. Univariate analysis identified 18 of 41 investigated baseline factors as significant (P < 0.05) predictors of GFR decline. In multivariate analysis, six factors--greater urine protein excretion, diagnosis of polycystic kidney disease (PKD), lower serum transferrin, higher mean arterial pressure, black race, and lower serum HDL cholesterol--independently predicted a faster decline in GFR. Together with the study interventions, these six factors accounted for 34.5% and 33.9% of the variance between patients in GFR slopes in Studies A and B, respectively, with proteinuria and PKD playing the predominant role. The mean rate of GFR decline was not significantly related to baseline GFR, suggesting an approximately linear mean GFR decline as renal disease progresses. The 41 baseline predictors were also assessed for their interactions with the diet and blood pressure interventions. A greater benefit of the low blood pressure intervention was found in patients with higher baseline urine protein. None of the 41 baseline factors were shown to predict a greater or lesser effect of dietary protein restriction.

Blood Pressure↗
PubMed1997

A new model of renal microvascular endothelial injury.

Although the importance of injury with consequent activation of endothelium is well-recognized in diseases affecting the glomerular endothelial cell (GEN), research on GEN injury in vivo has been hampered by the lack of adequate animal models. Here we report the establishment and characterization of a new GEN injury model in rats. This model was induced by selective renal artery perfusion with anti-GEN IgG and resulted in the severe acute renal failure with marked platelet deposition and development of a thrombotic microangiopathy involving glomeruli. Peritubular capillary endothelial cells were also damaged that was associated with severe tubular necrosis. Although the glomerular changes were severe, half of the glomeruli recovered by day 10, while interstitial changes remained throughout our observation time course. Proliferation of GEN was observed during the recovery phase. An increased expression of endothelial nitric oxide synthase in GEN was also observed, and may be an adaptive mechanism to counteract the thrombosis and ischemia. This model should be useful to investigate the pathophysiology of renal microvascular diseases and the mechanisms of GEN injury, activation and recovery in vivo.

Acute Kidney Injury↗
PubMed1997

Renal microvascular injury induced by antibody to glomerular endothelial cells is mediated by C5b-9.

We have recently developed a model of thrombotic microangiopathy with injury to the glomerular endothelial cell (GEN) induced by heterologous antibody to rat GEN. In addition to GEN injury rats developed glomerular platelet aggregation and fibrin deposition, acute renal failure, and acute tubular necrosis with interstitial inflammation. To study the role of complement in mediating this lesion, we induced the disease in normal complement PVG rats and measured the effects of generalized complement depletion with cobra venom factor (CVF) and of selective C6 deficiency using genetically C6 deficient PVG animals. Complement sufficient rats developed severe endothelial injury accompanied by platelet aggregation, fibrin deposition, decrease in endothelial cells assessed by antibody staining in the glomerulus, and macrophage infiltration. These changes were associated with marked reduction in renal function. These features were either absent or markedly diminished in complement depleted or C6 deficient rats. This demonstrates that C5b-9, the terminal product of activation of the complement cascade, plays an important role in the pathogenesis of this immune renal microvascular endothelial injury model. Thus, the complement system may play a pathogenic role in renal microvascular diseases such as thrombotic microangiopathy.

Acute Kidney Injury↗
PubMed1997

[Doubtful mammographic findings: the value of negative MR mammography for tumor exclusion].

PURPOSE: To determine whether the addition of MR mammography (MRM) is useful in excluding malignant lesions and how reliable negative MRM findings are. METHODS: Amongst 694 MRM's, those originally regarded as normal were retrospectively reappraised. 239 female patients were involved. In all these patients there were clinical, sonographic and/or mammographic findings which were not entirely normal but there was no urgent indication for histological clarification. In 46 patients there were, however, histological examinations since the patients themselves insisted on it. In the remaining patients there was clinical, sonographic, mammographic and/or MRM follow-up after 12 to 18 months. RESULTS: In 95.4% (200/239) a carcinoma could be excluded by means of MRM, in 7 patients a carcinoma in situ and in two patients an invasive carcinoma was demonstrated histologically which had not been demonstrated by MRM. Even in retrospect, no abnormality could be found. CONCLUSION: Because of the only moderate sensitivity of MRM in the recognition of carcinoma in situ, doubtful lesions which can be localised, should be biopsied by a stereotactic method. In cases where evaluation is difficult on clinical, sonographic and mammographic findings, MRM is of value in excluding tumours, particularly in patients with increased carcinoma risk.

Adolescent↗
PubMed1997

Transcriptional regulation of human placental corticotropin-releasing factor by prostaglandins and estradiol.

The mechanism of labor initiation in humans has not been completely elucidated. Prostaglandins, estrogens, and corticotropin-releasing factor (CRF) have all been shown to affect uterine myocytes and enhance uterine contractility. There are also indications that these uterine regulators have additional effects on other sites involved in labor and that they may act in concert or, perhaps, by regulating each other. Therefore, we evaluated the CRF promoter for transcriptional regulation by prostaglandins and estrogens. Human placental choriocarcinoma cell lines were transfected with CRF-luciferase reporter genes and treated with prostaglandins. Prostaglandin E2 (PGE2), but not prostaglandin F2alpha (PGF2alpha), stimulated CRF-luciferase expression in choriocarcinoma cell lines via a cAMP-dependent pathway. A combination of transfections and in vitro binding studies tested for potential regulation of CRF by estrogen receptor (ER). ER neither regulated the CRF promoter nor interacted with steroid response half-sites from the CRF promoter. Our results provide a molecular regulatory link between PGE2 and CRF, two compounds that enhance uterine contractile function. Combined with the stimulation of prostaglandin release by CRF, these data support a potentially important "feed-forward" regulatory loop involving CRF and PGE2 in parturition. In contrast, we found no evidence for direct effects of estrogens or PGF2alpha on CRF transcription.

Animals↗