[Successful rituximab treatment of recurrent glomerulonephritis associated with antibodies against the glomerular basement membrane]. / Erfolgreiche Rituximabtherapie des Rezidivs einer Glomerulonephritis assoziiert mit Antikörpern gegen die glomeruläre Basalmembran.

This article presents a case of recurrent anti-GBM disease (with antibodies against the glomerular basement membrane [GBM]) in a 17-year-old patient successfully treated with rituximab. Kidney biopsy with detection of linear deposition of immunoglobulin G (IgG) along the basement membrane is the diagnostic gold standard, which should be accompanied by serological testing. However, standard assays for the detection of anti-GBM antibodies have a high rate of false-negative results. In this particular case, an increase in proteinuria despite standard therapy (plasmapheresis, steroids, cyclophosphamide) was the clinical correlate of relapsing disease. The use of rituximab completely resolved the recurrent anti-GBM disease.

MicroRNA-126 contributes to renal microvascular heterogeneity of VCAM-1 protein expression in acute inflammation.

Endothelial cells in different microvascular segments of the kidney have diverse functions and exhibit differential responsiveness to disease stimuli. The responsible molecular mechanisms are largely unknown. We previously showed that during hemorrhagic shock, VCAM-1 protein was expressed primarily in extraglomerular compartments of the kidney, while E-selectin protein was highly induced in glomeruli only (van Meurs M, Wulfert FM, Knol AJ, de Haes A, Houwertjes M, Aarts LPHJ, Molema G. Shock 29: 291-299, 2008). Here, we investigated the molecular control of expression of these endothelial cell adhesion molecules in mouse models of renal inflammation. Microvascular segment-specific responses to the induction of anti-glomerular basement membrane (anti-GBM), glomerulonephritis and systemic TNF-α treatment showed that E-selectin expression was transcriptionally regulated, with high E-selectin mRNA and protein levels preferentially expressed in the glomerular compartment. In contrast, VCAM-1 mRNA expression was increased in both arterioles and glomeruli, while VCAM-1 protein expression was limited in the glomeruli. These high VCAM-1 mRNA/low VCAM-1 protein levels were accompanied by high local microRNA (miR)-126 and Egfl7 levels, as well as higher Ets1 levels compared with arteriolar expression levels. Using miR-reporter constructs, the functional activity of miR-126 in glomerular endothelial cells could be demonstrated. Moreover, in vivo knockdown of miR-126 function unleashed VCAM-1 protein expression in the glomeruli upon inflammatory challenge. These data imply that miR-126 has a major role in the segmental, heterogenic response of renal microvascular endothelial cells to systemic inflammatory stimuli.

Sotrastaurin, a novel small molecule inhibiting protein-kinase C: randomized phase II study in renal transplant recipients.

Sotrastaurin, a selective protein-kinase-C inhibitor, blocks early T-cell activation through a calcineurin-independent mechanism. In this study, de novo renal transplant recipients with immediate graft function were randomized 1:2 to tacrolimus (control, n = 44) or sotrastaurin (300 mg b.i.d.; n = 81). All patients received basiliximab, mycophenolic acid (MPA) and steroids. The primary endpoint was the composite of treated biopsy-proven acute rejection (BPAR), graft loss, death or lost to follow-up at month 3. The main safety assessment was estimated glomerular filtration rate (eGFR); modification of diet in renal disease (MDRD) at month 3. Composite efficacy failure at month 3 was higher for the sotrastaurin versus control regimen (25.7% vs. 4.5%, p = 0.001), driven by higher BPAR rates (23.6% vs. 4.5%, p = 0.003), which led to early study termination. Median (± standard deviation [SD]) eGFR was higher for sotrastaurin versus control at all timepoints from day 7 (month 3: 59.0 ± 22.3 vs. 49.5 ± 17.7 mL/min/1.73 m(2) , p = 0.006). The most common adverse events were gastrointestinal disorders (control: 63.6%; sotrastaurin: 88.9%) which led to study-medication discontinuation in two sotrastaurin patients. This study demonstrated a lower degree of efficacy but better renal function with the calcineurin-inhibitor-free regimen of sotrastaurin+MPA versus the tacrolimus-based control. Ongoing studies are evaluating alternative sotrastaurin regimens.

FOXP3+ regulatory T-cells in renal allografts: correlation with long-term graft function and acute rejection.

BACKGROUND: The interpretation of a cellular infiltrate as cytotoxic or tolerogen represents an unsolved challenge in current transplantation. The so-called regulatory CD4+ CD25+ T-cells which express the FOXP3 gene have received increasing interest with respect to this question. The existing studies concerning the role of FOXP3+ Tregs for transplant tolerance yielded contradictory results. METHODS: We examined the numbers of the FOXP3+ Tregs in two groups of renal allograft biopsies both showing cellular infiltration, but either without (n=29) or with signs of acute cellular rejection (n=26), by means of immunofluorescence and correlated the amount of FOXP3+ Tregs to renal function at the time of biopsy and after 1 and 2 years of follow up. RESULTS: The number of FOXP3+ Tregs within infiltrates in non-rejecting biopsies did not correlate with renal function after 1 and 2 years. There were no significant differences in the numbers of FOXP3+ Tregs between biopsies with or without borderline infiltrates. Increased numbers of FOXP3+ Tregs were not associated with an ameliorated severity of graft rejection and did not correlate with outcome after the rejection episode and renal function after 1 and 2 years. CONCLUSIONS: The identification of the FOXP3+ regulatory cells within the allograft cannot be considered as an appropriate marker for the interpretation of infiltrates as cytotoxic or tolerogenic or as a prognostic marker for later transplant function.

B-cell activating factor BAFF as a novel alert marker for the immunological risk stratification after kidney transplantation.

The B cell activating factor BAFF has gained importance in the context of kidney transplantation due to its role in B cell survival. Studies have shown that BAFF correlates with an increased incidence of antibody-mediated rejection and the development of donor-specific antibodies. In this study, we analyzed a defined cohort of kidney transplant recipients who were treated with standardized immunosuppressive regimens according to their immunological risk profile. The aim was to add BAFF as an awareness marker in the course after transplantation to consider patient's individual immunological risk profile. Included patients were transplanted between 2016 and 2018. Baseline data, graft function, the occurrence of rejection episodes, signs of microvascular infiltration, and DSA kinetics were recorded over 3 years. BAFF levels were determined 14 d, 3 and 12 months post transplantation. Although no difference in graft function could be observed, medium-risk patients showed a clear dynamic in their BAFF levels with low levels shortly after transplantation and an increase in values of 123% over the course of 1 year. Patients with high BAFF values were more susceptible to rejection, especially antibody-mediated rejection and displayed intensified microvascular inflammation; the combination of high BAFF + DSA puts patients at risk. The changing BAFF kinetics of the medium risk group as well as the increased occurrence of rejections at high BAFF values enables BAFF to be seen as an awareness factor. To compensate the changing immunological risk, a switch from a weaker induction therapy to an intensified maintenance therapy is required.

Upregulation of TNF receptor type 2 in human and experimental renal allograft rejection.

An important role of TNF interacting with TNFR2 has been shown in different models of ischemic, nephrotoxic and immune-mediated renal injury. To systematically evaluate the expression of TNFR2 in renal allograft rejection, we investigated human renal allograft biopsies and, in addition, established an experimental transplantation model in rats to verify the human data under standardized conditions. The expression of TNFR2 was analyzed in 96 human renal allograft biopsies with different disease entities. In a 6-day and a 28-day experimental protocol, TNFR2 was examined in kidney specimens and in the urine of control, uni-nephrectomized and transplanted rats +/- cyclosporine treatment (n = 114). In human biopsies and in rat allografts on day 6 with acute allograft rejection, significantly elevated expression of TNFR2 was observed in tubular epithelial cells, podocytes, B cells and monocytes/macrophages. The expression level was associated with renal function. The TNFR2 expression level at day 28 was significantly lower compared to day 6. TNFR2 is markedly upregulated both in human and experimental acute renal allograft rejection. Our data are robust and consistent between different species, suggesting a role for TNFR2 in the early course of rejection.

Percutaneous transluminal angioplasty as first-line treatment of transplant renal artery stenosis.

BACKGROUND: Transplant renal artery stenosis (TRAS) is a frequent complication after renal transplantation, however long-term follow-up data after interventional treatment are rare. PATIENTS: In our transplant center 11 of 264 consecutive renal transplant recipients (4.17%) were diagnosed with TRAS. In addition, TRAS occurred in 2 renal transplant recipients that had been transplanted at other centers but who had their follow-up examinations in our center. Either a rise of the serum creatinine level and/or worsened systemic hypertension or routine examination with color Doppler sonography were indications for further diagnostic workup. METHODS: Direct angiography of the transplant renal artery was performed followed by percutaneous transluminal angioplasty (PTA) after the diagnosis of TRAS was confirmed in all of these patients. RESULTS: The immediate success rate for PTA was 92.3% (12/13). Only 1 patient with a severe kinking of the transplant renal artery had to undergo surgery to restore renal function. No complications occurred after the interventions. Thereafter the patients were monitored for a mean observation period of 33.15 months. Serum creatinine levels were significantly lower after the intervention, and estimated glomerular filtration rate (eGFR) increased accordingly. With regard to blood pressure there was only a trend for lower blood pressure levels and less antihypertensive use, whereas the dose of the prescribed drugs decreased significantly with time after interventional treatment of TRAS. In addition, a long-lasting rise of the hemoglobin levels could also be demonstrated. CONCLUSION: In summary, the beneficial effect of PTA of TRAS on renal function is long-lasting. Therefore, PTA, usually combined with stent placement, should be first-line treatment in TRAS in all patients. Surgical revascularization is only warranted, if PTA fails.

Severe renal vein stenosis of a kidney transplant with beneficial clinical course after successful percutaneous stenting.

A 51-year-old renal transplant recipient presented with marked renal function deterioration 13 months after renal transplantation. After exclusion of ureteral obstruction, transplant artery stenosis and acute rejection, the diagnosis of a severe renal vein stenosis was made by an MR scan. After angiographic confirmation of the stenosis, treatment was attempted with percutaneous stent angioplasty. The long-term clinical course was favorable, with marked improvement in renal function. Transplant renal vein stenosis is a rare, but potentially curable, cause of renal allograft functional deterioration.

Five-year follow-up after late conversion from calcineurin inhibitors to sirolimus in patients with chronic renal allograft dysfunction.

BACKGROUND: Chronic allograft nephropathy (CAN) is, among others, caused by nephrotoxic side effects of calcineurin inhibitors (CNI), which are to date still the mainstay of immunosuppressive therapy. Sirolimus (SIR), an immunosuppressive compound without effects on glomerular perfusion, has been used in CNI-sparing immunosuppressive protocols. We report the 5-year follow-up of a prospective, controlled conversion study from CNI to SIR in patients with moderately to severely impaired renal function. METHODS: Twelve renal transplant recipients with moderately to severely impaired renal function (estimated glomerular filtration rate of 17 to 35 mL/min according to the MDRD formula), enrolled in a prospective, controlled 1-year pilot study were followed for 5 years. RESULTS: Three renal grafts (25%) were lost during the 5-year follow-up. Graft loss was due to noncompliance in one patient and to CAN in the other two patients. These two patients returned to dialysis 43 and 59 months after conversion, corresponding to 86 and 75 months after transplantation, respectively. Six of nine patients had a stable or even better renal function compared to the baseline. The lipid profile increased initially, but then remained stable over time. CONCLUSION: Conversion of immunosuppressive therapy from CNI to SIR in patients with impaired renal function more than 1 year after transplantation is feasible and safe yielding improved renal function in the majority of patients, which was sustained at 5 years follow-up.

B-cell activating factor BAFF reflects patients' immunological risk profile after kidney transplantation.

The B-cell activating factor BAFF plays an important role in the development and maturation of B-lymphocytes, which can contribute to the generation of donor-specific antibodies and thus may influence graft function and graft survival. Inconsistent data on the role of BAFF levels after renal transplantation for the formation of donor-specific antibodies and the contribution for allograft rejection exist. The aim of the current study was to determine to what extent the degree of pre-immunization is reflected by each patient's BAFF levels before transplantation and in the follow-up. Furthermore, the impact of BAFF on allograft rejection frequency as well as severity and resulting allograft function over time was analyzed. Additionally, the impact of viral infections on BAFF levels after transplantation - as a potential confounder - was examined. For this purpose, a group of pre-sensitized patients (PRA>0%, (52±24% on average), n=40) was compared with non-sensitized patients (PRA=0%, n=62) and in a subsequent analysis stratification in accordance to the detected BAFF level was performed. Pre-sensitized patients had significantly higher BAFF levels before transplantation and suffered significantly more often from early steroid-resistant, mainly antibody-mediated rejections. A result which was confirmed also in highly sensitized patients with PRA levels >50%. Additionally, in the follow-up patients with either rising BAFF levels over time or BAFF levels above the median also had significantly more often antibody mediated rejections. Additionally, patients with BAFF levels above detected median even displayed impaired creatinine values as well as an induced eGFR slope up to month 48 after transplantation. The occurrence of viral infections (CMV, BKV) was only an additional influencing factor in the absence of concomitant allograft rejections. Therefore, the B-cell proliferation factor BAFF appears not only to reflect the immunological risk profile of patients in the context of kidney transplantation, it may possibly be further developed as a predictor of patients with an increased risk profile for subsequent allograft rejection and impaired allograft function.

Efficacy of Prolonged- and Immediate-release Tacrolimus in Kidney Transplantation: A Pooled Analysis of Two Large, Randomized, Controlled Trials.

BACKGROUND: Two large, prospective studies (12-03; OSAKA) compared the efficacy and tolerability of prolonged-release versus immediate-release tacrolimus in kidney transplant patients also receiving mycophenolate mofetil and low-dose corticosteroids (without induction therapy). METHODS: Data were combined into one database to compare results over 24 weeks using 3 alternative endpoints: biopsy-confirmed acute rejection (BCAR); the Food and Drug Administration composite endpoint (graft loss, BCAR, and loss to follow-up), and the European Medicines Agency composite endpoint (graft loss, BCAR, and graft dysfunction). The 95% confidence intervals were calculated (10% noninferiority margin). RESULTS: Overall, 633 patients received prolonged-release tacrolimus (12-03, n = 331; OSAKA, n = 302) and 645 received immediate-release tacrolimus (n = 336; n = 309). Baseline characteristics were comparable. Proportionately more patients receiving prolonged-release tacrolimus had trough levels of 5-15 ng/mL on day 1 (60.8%) and 2 (56.6%) versus immediate-release tacrolimus (42.5% and 43.9%, respectively, both P < .001). Efficacy of prolonged-release and immediate-release tacrolimus were similar as assessed by BCAR (13.9% vs 14.1%, respectively), European Medicines Agency composite endpoint (40.3% vs 38.3%) and US Food and Drug Administration composite endpoint (21.5% vs 19.8%). CONCLUSIONS: Novel efficacy endpoints as required by the European Medicines Agency and US Food and Drug Administration demonstrate noninferiority of prolonged-release versus immediate-release tacrolimus. Significantly more patients treated with prolonged-release tacrolimus versus immediate-release tacrolimus achieved trough levels of 5 to 15 ng/mL early after transplantation. ClinicalTrials.govNCT00189839; NCT00717470.

Exanthema and acute anuric renal failure.

A 15-year-old girl with a history of Kawasaki disease was admitted to our nephrological department due to acute renal failure. Despite antibiotic therapy because of fever and the symptoms of a pharyngitis in the last few days, the girl showed persisting fever and developed arthralgias, an exanthema and a rising serum creatinine as well as anuria. A wide variety of differential diagnoses has to be thought of because of the history of the Kawasaki disease (symptoms like fever, pharyngitis, exanthema and arthralgia), i.e. hemolytic-uremic syndrome, vasculitis, ascending infection, postinfection glomerulonephritis. In consideration of etiologically unclear "rapidly progressive renal failure" with anuria and thrombocytopenia an immediate renal biopsy was done and revealed a severe drug induced acute interstitial nephritis. Due to this diagnosis we treated the patient with corticosteroids. Within 4 weeks serum creatinine declined to 1.8 mg/dl but did not normalize.

Increase in renal vascular resistance after intake of cyclosporin A and tacrolimus and reversal by nitroglycerin spray: a study in patients with stable renal allograft function.

PURPOSE: Acute effects of drug administration on renal arterial resistance index (RI) are still discussed controversially. In our study we investigated the immediate effects of cyclosporin A (CyA) and tacrolimus (FK-506) on renal arterial resistance indices in patients with stable graft function after renal transplantation. Additionally we studied the effects of nitroglycerin spray on resistance indices. METHODS: RI was measured by color Doppler sonography at baseline, at 1 and 2 hours after intake of medication and 30 minutes after administration of nitroglycerin spray which followed the 2-hour measurement. 34 renal transplant recipients were examined. 16 patients received CyA, 18 patients received FK-506. Whole blood levels of calcineurin inhibitors were taken at each time point. Arterial blood pressure and heart rate were measured to assess possible systemic hemodynamic effects. RESULTS: Mean RI values increased significantly in both groups 1 hour after calcineurin inhibitor intake and remained still significantly elevated after 2 hours. There was no significant increase of mean arterial blood pressure nor was there any correlation between whole blood levels of calcineurin inhibitors and mean RI. 30 minutes after administration of nitroglycerin spray, mean RI values decreased significantly to a level even below baseline. Mean arterial blood pressure also decreased after administration of nitroglycerin. CONCLUSION: Renal RI values are markedly influenced by a recent intake of calcineurin inhibitors and vasoactive substances such as nitrates. This demonstrates the necessity of keeping standardized conditions when using RI as a tool in followup investigations after renal transplantation.

Pretransplant calcium levels have no predictive value for delayed graft function, long-term graft function, cardiovascular events, or graft and patient survival in renal transplantation.

BACKGROUND: Disorders of calcium homeostasis are one of the most common problems in patients with end-stage renal disease (ESRD). Elevated calcium levels increase the incidence of cardiovascular mortality in ESRD patients, and appear to be a risk factor for the occurrence of delayed graft function (DGF) after kidney transplantation. Therefore, we investigated the impact of pretransplant serum calcium levels on outcomes after kidney transplantation: DGF, acute rejection, graft function, and survival, as well as the incidence of cardiovascular events. METHODS: We studied 285 patients (96.9% of all transplanted patients) who underwent their first transplantation between 1995 and 2004. Demographic data were extracted from hospital records or were documented during follow-up; serum samples were collected at the time of transplantation. RESULTS: In our cohort the incidence of DGF was 16.5% and 35.4% of acute rejection episodes (ARE). However, pretransplant calcium levels were not related to DGF or ARE in our patient cohort. Furthermore, there was no correlation between pretransplant serum calcium level with the incidence of cardiovascular events or mortality, as well as graft function or survival. CONCLUSION: In our study population pretransplant calcium levels showed no effect on DGF, ARE rate, the occurrence of cardiovascular events or death, renal graft function, or survival. Therefore, pretransplant calcium level is not a helpful marker for risk stratification at the time of transplantation.

[The challenge of acute kidney injury : Cross-sectional study on prevalence and costs in a university intensive care unit]. / Die Herausforderung akute Nierenschädigung : Querschnittsstudie zu Prävalenz und Kosten auf einer universitären Intensivstation.

BACKGROUND AND OBJECTIVES: The prognosis of intensive care patients with acute kidney injury (AKI), which is associated with increased mortality is still poor. Current data on the prevalence and the resulting costs of AKI and an overview of the most common diagnoses associated with AKI in German intensive care units (ICU) are lacking. PATIENTS AND METHODS: In this retrospective study all adult admissions (> 18 Jahre) in the five ICUs at the University Clinic Regensburg (in total 78 beds) from1 January 2011 to 31 December 2013 were evaluated. The ICU diagnoses commonly associated with AKI were identified using the international classification of diseases 10 (ICD 10). The length of ICU and hospital stays and AKI-associated hospital costs in the diagnosis-related groups (DRG) based reimbursement system were compared. RESULTS: A total of 891 ICU patients with AKI were classified according to the ICD 10 code. Acute respiratory distress syndrome (ARDS), myocardial infarction (MI) and sepsis were the three most common ICU conditions associated with AKI. A total of 1103 patients were admitted with 1 of these 3 main diagnoses and 249 (22.6 %) of these patients developed AKI. Patients with AKI had significantly longer mean ICU and hospital stays compared to patients without AKI (18.6 vs 5.1 days and 23.8 vs. 10.4 days, respectively, p < 0.001). The presence of AKI in critically ill patients with ARDS, MI and sepsis resulted in additional costs of 2,019,120.42 € at the University Hospital of Regensburg in 2013. CONCLUSION: Acute kidney injury in critically ill patients represents a significant medical and socioeconomic burden. Early recognition of patients at risk, coordinated research into novel interventions and establishment of the National Acute Kidney Injury Network for implementation of evidence-based therapies may be the next steps to decrease the incidence and severity of AKI and save costs for the national healthcare system.

Analysis of the promoter of the human prostatic acid phosphatase gene.

From the analysis of two overlapping cosmid clones prepared from human genomic DNA libraries, a contig of 44 kb containing a 5' portion of the PAP gene and 17 kb of the upstream region was established. It was characterized by restriction mapping and sequence analysis of 2.5 kb upstream of the initiation codon. Two major transcription initiation sites were found to be located around 56 and 91 bp upstream of the initiation codon, as determined by nuclease S1 and primer extension mapping. Expression of the PAP gene was measured by Northern blots in the androgen responsive LNCaP cell line. It was found to be induced 2-3-fold by the addition of the synthetic androgen mibolerone to the cells. The induced mRNA levels were approx. 10-times lower than those for the prostate-specific antigen (PSA) in LNCaP cells.

Steroid withdrawal or steroid avoidance in renal transplant recipients: focus on tacrolimus-based immunosuppressive regimens.

Steroid-induced adverse effects after transplantation include cosmetic, metabolic, and cardiovascular complications. Steroid withdrawal or avoidance with cyclosporine-based regimens have been hampered by an unacceptably high rate of acute rejections and increased rates of graft loss. Recently the results of several large, randomized trials of steroid withdrawal/avoidance with tacrolimus-based immunosuppression in renal transplant recipients became available. A review of these trials appeared to be of clinical interest. Data from the THOMAS trial clearly indicate that steroid withdrawal from a regimen of tacrolimus, mycophenolate mofetil (MMF), steroids after 3 months after transplantation is safe with regard to acute rejection rate and graft survival. If an induction therapy with daclizumab is used in combination with tacrolimus and MMF (CARMEN trial), even steroid avoidance is safe with regard to acute rejection rate and graft survival. Finally, in the ATLAS trial, steroid avoidance with basiliximab in combination with tacrolimus (resulting in tacrolimus monotherapy) or alternatively with tacrolimus and MMF both resulted in similar graft survival, but higher rates of acute rejection. In conclusion, steroid withdrawal is safe from a triple-drug regimen of tacrolimus, MMF, and steroids after 3 months after transplantation, and steroid use may completely be avoided with tacrolimus, and MMF combined with daclizumab induction. Tacrolimus monotherapy may be achieved using basiliximab induction at the price of higher rates of acute rejection, but with unaffected graft survival. Thus tacrolimus-based immunosuppression with or without interleukin-2 receptor antagonist induction has made steroid withdrawal or avoidance a realistic option in renal transplantation.

Cardiovascular risk estimates and risk factors in renal transplant recipients.

Cardiovascular morbidity, including coronary artery disease and left ventricular hypertrophy, and mortality are high in patients following renal transplantation. Cardiovascular disease is thought to be due to traditional (hypertension, hyperlipidemia, diabetes mellitus and smoking) as well as nontraditional cardiovascular risk factors (microinflammation). Furthermore, immunosuppressive drugs, namely, calcineurin inhibitors, sirolimus, and steroids, have been reported to adversely affect cardiovascular risk factors (e.g., hypertension, hyperlipidemia, hyperglycemia). Evidence from comparative trials and from conversion studies suggest that blood pressure, hyperlipidemia, and hyperglycemia after renal transplantation may be differentially affected by the calcineurin inhibitors cyclosporine and tacrolimus. In the European Tacrolimus versus Cyclosporin A Microemulsion Renal Transplantation Study, 557 patients were randomly allocated to therapy with tacrolimus (n = 286) versus cyclosporine (n = 271). In addition, to blood pressure, serum cholesterol, HDL cholesterol, triglycerides, and blood glucose, we estimated the 10-year risk of coronary heart disease (Framingham risk score). Tacrolimus resulted in a significantly lower time-weighted average of serum cholesterol (P < .001), and mean arterial blood pressure (P < .05), but a higher time-weighted average of blood glucose (P < .01) than cyclosporine. Mean 10-year coronary artery disease risk estimate was significantly lower in men treated with tacrolimus, (10.0% versus 13.2%; P < .01) but was unchanged in women (4.7% versus 7.0%). Tacrolimus and cyclosporine microemulsion have compound-specific effects on cardiovascular risk factors that differentially affect the predicted rate of coronary artery disease.

Modulation of HIV-1 enhancer activity and virus production by cAMP.

The effect of cAMP on the transcriptional activity of the HIV-1 long terminal repeat/enhancer was investigated and compared to the effect of cAMP on virus replication. In culture cAMP repressed virus replication in vivo using different cell types. Transient transfection studies with HIV-1 enhancer-derived luciferase reporter gene constructs identified the minimal DNA sequence mediating the negative regulatory effect of cAMP on HIV-1 transcription. A single nuclear factor kappaB element from the HIV-1 enhancer mediates the repressive effect on transcription. AP-2 is not involved in cAMP repression. Stable transfection of Jurkat T cells with the co-activators CREB binding protein (CBP) and p300 completely abolished the cAMP repressive effect, supporting the hypothesis that elevation of intracellular cAMP increases phosphorylation of CREB, which then competes with phosphorylated p65 and Ets-1 for limiting amounts of CBP/p300 thereby mediating the observed repressive effect on transcription. These findings suggest an important role of cAMP on HIV-1 transcription.

Human muscle glyceraldehyde-3-phosphate dehydrogenase: reactivity of sulphydryl groups.

1. The kinetics of the reaction of thiol groups of glyceraldehyde-3-phosphate dehydrogenase from human muscle with 5,5'-dithiobis-2-nitrobenzoate (DTNB) was studied spectrophotometrically using the conventional and stopped-flow methods. 2. Each of the three thiol groups present in the enzyme subunit reacts with a different velocity. 3. The reaction with DTNB of the four sulphydryl groups of Cys-149, essential for the enzymatic activity, is biphasic, depends on the amount of NAD bound and is strongly slowed down when one mole of coenzyme is bound to the tetramer. NAD bound is only partially released from the holoenzyme treated with DTNB. 4. In the presence of borate, thiol groups of Cys-153 and Cys-244 do not react with DTNB.