Jürgen Steinhoff

The kidney as a second site of human C-reactive protein formation in vivo.

C‐reactive protein (CRP) is the main acute phase reactant in humans. Its production is presumably restricted to the liver but extrahepatic expression by inflamed tissue has not been studied indetail. By real‐time PCR and immunohistochemistry we here show that renal cortical tubular epithelial cells (TEC) express CRP mRNA and protein within 6 h after stimulation with conditioned medium (CM) or IL‐6, but not IL‐1α or TNF‐α. Western blot analysis with monoclonal anti‐CRP antibody that recognizes native CRP revealed protein secretion into supernatants of CM‐stimulated TEC cultures. While hepatoma‐derived Hep3B cells could be induced similarly, peripheral blood mononuclear cells could not. CRP mRNA transcripts were observed in nephrectomized renal allografts with severe acute rejection but not with chronic allograft nephropathy (CAN). Of 19 needle biopsies of acutely rejecting kidney transplants, 15 demonstrated CRP mRNA production with the relative expression levels increasing with the severity of rejection. On the other hand, none of 7 graft biopsies with acute tubular necrosis (ATN) or CAN showed CRP mRNA expression. By using monoclonal anti‐CRP antibody, cortical tubules as well as glomerular cells were shown to locally express CRP in rejecting, but not in ATN kidneys. We conclude that inflamed kidneys represent a so far unknown site of CRP formation in vivo. These data shed new light on the acute phase reaction not merely representing a systemic inflammatory pathway but probably being part of the local immune response.

Best supportive care and therapeutic plasma exchange with or without eculizumab in Shiga-toxin-producing E. coli O104:H4 induced haemolytic–uraemic syndrome: an analysis of the German STEC-HUS registry

BACKGROUND May 22nd marks the beginning of a Shiga-toxin-producing Escherichia coli (STEC) O104:H4 outbreak in Northern Germany. By its end on 27 July, it had claimed 53 deaths among 2987 STEC and 855 confirmed haemolytic-uraemic syndrome (HUS) cases. METHODS To describe short-term effectiveness of best supportive care (BSC), therapeutic plasma exchange (TPE) and TPE with eculizumab (TPE-Ecu) in 631 patients with suspected HUS treated in 84 hospitals in Germany, Sweden and the Netherlands using the web-based registry of the DGfN (online since 27 May). RESULTS Of 631 entries, 491 fulfilled the definition of HUS (median age 46 years; 71% females). The median (inter-quartile range) hospital stay was 22 (14-31) days. Two hundred and eighty-one (57%) patients underwent dialysis and 114 (23%) mechanical ventilation. Fifty-seven patients received BSC, 241 TPE and 193 TPE-Ecu. Treatment strategy was dependent on disease severity (laboratory signs of haemolysis, thrombocytopenia, peak creatinine level, need for dialysis, neurological symptoms, frequency of seizures) which was lower in BSC than in TPE and TPE-Ecu patients. At study endpoint (hospital discharge or death), the median creatinine was lower in BSC [1.1 mg/dL (0.9-1.3)] than in TPE [1.2 mg/dL (1.0-1.5), P < 0.05] and TPE-Ecu [1.4 mg/dL (1.0-2.2), P < 0.001], while need for dialysis was not different between BSC (0.0%, n = 0), TPE (3.7%; n = 9) and TPE-Ecu (4.7%, n = 9). Seizures were absent in BSC and rare in TPE (0.4%; n = 1) and TPE-Ecu (2.6%; n = 5) patients. Total hospital mortality in HUS patients was 4.1% (n = 20) and did not differ significantly between the TPE and TPE-Ecu groups. CONCLUSIONS Despite frequent renal impairment, advanced neurological disorders and severe respiratory failure, short-term outcome was better than expected when compared with previous reports. Within the limitations of a retrospective registry analysis, our data do not support the notion of a short-term benefit of Ecu in comparison to TPE alone in the treatment of STEC-HUS. A randomized trial comparing BSC, TPE and Ecu seems to be prudent and necessary prior to establishing new treatment guidelines for STEC-HUS.

Association Between Azithromycin Therapy and Duration of Bacterial Shedding Among Patients With Shiga Toxin–Producing Enteroaggregative Escherichia coli O104:H4

CONTEXT An outbreak of Shiga toxin-producing enteroaggregative Escherichia coli (STEC O104:H4) infection with a high incidence of hemolytic uremic syndrome (HUS) occurred in Germany in May 2011. Antibiotic treatment of STEC infection is discouraged because it might increase the risk of HUS development. However, antibiotic therapy is widely used to treat enteroaggregative E coli infection. In the German outbreak, a substantial number of patients received prophylactic azithromycin treatment as part of a therapeutic regimen with the C5 antibody eculizumab. OBJECTIVE To analyze the duration of bacterial shedding in patients with STEC infection who did and did not receive oral azithromycin therapy. DESIGN, SETTING, AND PATIENTS At a single center in Lübeck, Germany, 65 patients with STEC infection, including patients with HUS as well as STEC-infected outpatients without manifestation of HUS, were investigated between May 15 and July 26, 2011, and were monitored for a mean of 39.3 days after onset of clinical symptoms. MAIN OUTCOME MEASURE Carriage of STEC after azithromycin therapy. RESULTS Twenty-two patients received oral azithromycin and 43 patients did not receive antibiotic treatment. Among antibiotic-treated patients, long-term STEC carriage (>28 days) was observed in 1 of 22 patients (4.5%; 95% CI, 0%-13.3%) compared with 35 of 43 patients (81.4%; 95% CI, 69.8%-93.0%) who were not treated with antibiotics (P < .001). All 22 patients receiving azithromycin treatment had at least 3 STEC-negative stool specimens after the completion of treatment, and no recurrence of STEC was observed in these patients. As proof of principle, 15 patients who initially were not treated with antibiotics and were long-term STEC carriers were treated with oral azithromycin given for 3 days and subsequently had negative stool specimens. CONCLUSION Treatment with azithromycin was associated with a lower frequency of long-term STEC O104:H4 carriage.

Coincidence of Epstein-Barr virus reactivation, cytomegalovirus infection, and rejection episodes in renal transplant recipients

Reactivation of the Epstein-Barr virus was reported to occur frequently under immunosuppressive therapy following organ transplantation. However, little is known about the clinical significance of these EBV reactivations. Therefore, we searched for correlations among the treatment with various immunosuppressive drugs, the incidence of CMV infections, rejection crises, and serological signs of EBV reactivation. EBV-specific antibodies were measured with novel ELISAs, utilizing the recombinant antigens p72 (for anti-EBV nuclear antigen [EBNA]1-IgG), p54, and pl38 (anti-early antigen [EA]-IgM, -IgG, -IgA) in a follow-up study of 79 renal transplant recipients. Patients receiving antithymocyte globulin or antilymphocyte globulin therapy showed increasing anti-EA-IgG and -IgA more often than did patients not receiving antithymocyte globulin or antilymphocyte globulin therapy (P<0.05). In patients receiving OKT3 antirejection therapy, an-ti-EA-IgM seroconversion was found more frequently (P<0.01). A significant correlation was also found between groups of patients who had had at least one rejection episode versus patients without any sign of organ rejection, and the incidence of increasing anti-EA-IgG (P < 0.05). Since in most of these patients signs of EBV reactivation followed the appearance of the rejection episode, this may not be due to viral-induced rejection but may be caused by the reinforced immunosuppression during antirejection therapy. As opposed to patients with no signs of CMV infection and with nonsymptomatic CMV infection, patients undergoing symptomatic CMV infection showed anti-EA-IgM seroconversion (P< 0.01), increasing anti-EA-IgA (P < 0.01), and decreasing anti-EBNA-IgG (P < 0.01) more frequently. Our results confirm the role of immunosuppressive therapy in the pathogenesis of EBV reactivation. We further demonstrate a striking coincidence of EBV reactivation and symptomatic CMV infection.

Molecular Parameters for Precise Diagnosis of Asymptomatic Epstein-Barr Virus Reactivation in Healthy Carriers

ABSTRACT Asymptomatic Epstein-Barr virus (EBV) reactivations periodically occur in oral mucosa-associated lymphoid tissues. Until now, EBV reactivation has been diagnosed by serologic profiles that suggest virus replication. Serologic responses, however, are delayed and do not necessarily indicate ongoing replicative activity. The aim of the present study was to establish in healthy carriers parameters for a molecular diagnosis of reactivated EBV infection. Recent studies emphasized the association of an increase in peripheral-B-cell viral load with replicative activity at remote sites. Therefore, real-time PCR was used to quantitate EBV genomes in the peripheral blood mononuclear cells (PBMC) (viral load) and plasma samples (viremia) of 22 healthy EBV-seropositive blood donors over a period of 15 months. Furthermore, transcription of the immediate-early gene encoding BZLF1 was investigated in the PBMC of all volunteers. Serology suggested reactivation in nine donors, of whom all but one showed at least once a significant increase in viral load. Another five individuals also exhibited significant changes in viral load but no serologic response. Of the 13 volunteers with significant increases in viral load, 6 had a period of viremia accompanying the rise in viral load. A stable viral load without viremia and negative serology was seen in eight adults. BZLF1 mRNA was undetectable throughout. We conclude that for healthy subjects serology underestimates the frequency of asymptomatic EBV reactivations. Prospective examination of peripheral viral load and viremia is suitable for the exact diagnosis of EBV reactivation, which might be of advantage for immunocompromised patients in whom EBV reactivations are considerably harmful.

The long persistence of CMV DNA in the blood of renal transplant patients after recovery from CMV infection.

A total of 30–50% of all renal transplant recipients undergo infections caused by human cytomegalovirus. With the introduction of ganciclovir and foscarnet for specific antiviral therapy there is an increasing demand for diagnostic tools that allow the early and rapid identification of CMV as the causative agent of the observed disease. We and others previously showed the direct detection of pp65 antigen in peripheral blood leukocytes to be an excellent marker for active cytomegalovirus infection. In order to establish whether the detection of CMV DNA by the polymerase chain reaction (PCR) sup

Treatment of posttransplant hypertension by laparoscopic bilateral nephrectomy

BACKGROUND Hypertension is an important risk factor for the development of chronic graft failure and decreased graft and patient survival after renal transplantation. METHODS Between September 1994 and August 1996, 14 patients underwent laparoscopic bilateral nephrectomy for treatment of drug-resistant hypertension after successful renal transplantation. Common causes of hypertension were largely excluded before bilateral nephrectomy. A scoring system was developed for comparison of different antihypertensive regimes. In this system, points were given according to type and dosage of each antihypertensive drug. RESULTS At 6-month follow-up, all patients showed well-controlled blood pressure (median of mean arterial pressure: 104 vs. 130 mmHg preoperatively, P<0.001, n=14), and significantly fewer antihypertensive drugs were needed according to the scoring system (48.9+/-20.9 points vs. 105.9+/-23.5 points preoperatively, P<0.001, n=14). During laparoscopy, three conversions to open surgery were necessary. Postoperatively, four complications occurred. After laparoscopy, immunosuppression and other oral medication were given continuously. The hospital stay ranged between 3 and 6 days (median: 5 days). CONCLUSIONS The results indicate that bilateral nephrectomy using the laparoscopic technique can be an effective alternative method for a selected group of patients with severe hypertension, which is unresponsive to conservative management after successful renal transplantation with regard to improving the long-term graft survival.

Time course and frequency of Epstein-Barr virus reactivation after kidney transplantation: linkage to renal allograft rejection.

The onset and frequency of Epstein-Barr virus (EBV) reactivation after kidney transplantation are unknown. By use of quantitative real-time polymerase chain reaction measurements, evidence of early EBV reactivation, occurring within the first week after the initiation of immunosuppressive therapy (median, 3 days), was observed in 13 of 23 patients, of whom 10 subsequently developed rejection episodes after 2-45 days (median, 5 days). By contrast, rejection was only diagnosed in 1 of 10 patients who did not show signs of viral reactivation. We suggest that EBV reactivation may induce a T cell response that, through the phenomenon of allo-cross-reactivity, could play a critical role in graft rejection.

Nephrotoxicity of ifosfamide, carboplatin and etoposide (ICE) alone or combined with extracorporeal or radiant-heat-induced whole-body hyperthermia

Abstract Although whole-body hyperthermia combined with specific genotoxic chemotherapy can be shown to enhance neoplastic cell killing without a concomitant rise in bone marrow toxicity, nephrotoxicity can become treatment-limiting. This study compares the kidney toxicity to the kidney of ifosfamide, carboplatin and etoposide (ICE) chemotherapy alone, and ICE chemotherapy combined with either extracorporeal (e-WBH) or radiant-heat-induced hyperthermia (r-WBH) in 43 patients with refractory sarcoma. Within 3 days of ICE chemotherapy treatment there was a significant increase in urinary protein excretion and a reduction of the glomerular filtration rate. These effects were more pronounced if WBH was added. The use of immunoluminometric assays revealed a predominance of low-molecular-mass proteins. This increase in protein excretion persisted in the e-WBH-treated group, whereas it vanished within 3 weeks in both the group treated with ICE alone and that treated with r-WBH. Our findings suggest that ICE chemotherapy causes transient tubular and glomerular damage, which is enhanced by WBH. In terms of long-term nephrotoxicity e-WBH was more nephrotoxic than r-WBH. This finding is consistent with our clinical observations.

Insulin inhibits caspase-3 activity in human renal tubular epithelial cells via the PI3-kinase/Akt pathway.

Apoptotic mechanisms in proximal renal tubular epithelial cells (PTEC) are crucial in the pathogenesis of acute kidney injury. We investigated whether insulin alters anti-apoptotic signalling in human PTEC. Cells were deprived of insulin for 0, 24 or 48 h and then stimulated with insulin for 0 or 5 min. Apoptosis was induced by camptothecin incubation. Insulin receptor kinase (IR-kinase) activity, phosphorylation of insulin receptor substrate-1 (IRS-1), IRS-1-associated PI3-kinase (p85), Ser273-phosphorylation of Akt and caspase-3 activity (C3-activity) were determined. Insulin stimulation increased the activity of IR-kinase, IRS-1 phosphorylation, p85 association with IRS-1 and Ser273-phosphorylation of Akt by at least 250%, respectively and decreased the C3-activity by 45% (p < 0.01, respectively). Deprivation of insulin for 24 and 48 h reduced basal and insulin-stimulated IR-kinase activity, IRS-1 phosphorylation, p85 association with IRS-1 and Ser273-phosphorylation of Akt by 30-40% and increased C3-activity by 15-20% (p < 0.01, respectively). Incubation with camptothecin increased C3-activity by 250-300% (p < 0.001). Subsequent insulin stimulation reversed the camptothecin induced increase of C3-activity. Our data indicate that apoptosis in PTEC is regulated by the insulin dependent PI3-kinase/Akt pathway. The enhancement of tubular-specific cell survival signals might represent a potential therapeutic tool for the protection of renal function in acute kidney injury.