Timo Jahnukainen

Alpha-haemolysin of uropathogenic E. coli induces Ca2+ oscillations in renal epithelial cells.

Pyelonephritis is one of the most common febrile diseases in children. If not treated appropriately, it causes irreversible renal damage and accounts for a large proportion of end stage renal failures. Renal scarring can occur in the absence of inflammatory cells, indicating that bacteria may have a direct signalling effect on renal cells. Intracellular calcium ([Ca2+]i) oscillations can protect cells from the cytotoxic effects of prolonged increases in intracellular calcium. However, no pathophysiologically relevant protein that induces such oscillations has been identified. Here we show that infection by uropathogenic Escherichia coli induces a constant, low-frequency oscillatory [Ca 2+]i response in target primary rat renal epithelial cells induced by the secreted RTX (repeats-in-toxin) toxin α-haemolysin. The response depends on calcium influx through L-type calcium channels as well as from internal stores gated by inositol triphosphate. Internal calcium oscillations induced by α-haemolysin in a renal epithelial cell line stimulated production of cytokines interleukin (IL)-6 and IL-8. Our findings indicate a novel role for α-haemolysin in pyelonephritis: as an inducer of an oscillating second messenger response in target cells, which fine-tunes gene expression during the inflammatory response.

Bacteremic urinary tract infection in children.

Objective. To assess the clinical characteristics of bacteremic urinary tract infection (UTI) in children. Design. Clinical data of Finnish children with bacteremic UTI (n = 134) from 1985 to 1994 were analyzed. Their symptoms, laboratory and imaging findings were compared with those of age‐ and sex‐matched patients hospitalized for blood culture negative UTI. Results. Generally, no major differences were seen in clinical findings between bacteremic and nonbacteremic patients. Bacteremic children had more frequent feeding problems (P = 0.02), and children ≥12 months of age tended more often to have abdominal pain and vomiting than did nonbacteremic patients. Fever was the major initial symptom in both study groups, but no significant difference occurred in the mean highest temperature or in the mean of duration of fever before admission to the hospital. The mean concentration of serum C‐reactive protein on admission was significantly higher in bacteremic patients (116 vs. 76 mg/l;P < 0.01). After onset of antimicrobial treatment fever lasted significantly longer in bacteremic patients than in control patients (means, 2.3 vs. 1.1 days;P < 0.01). Anatomic or functional abnormalities in the urinary tract were detected in 51%vs. 46%, respectively. Obstruction of the urinary tract (9%vs. 1%, P < 0.01) and Grade 3 to 5 vesicoureteral reflux (30%vs. 16%, P = 0.02) were significantly more frequent in bacteremic patients with UTI. Obstruction or vesicoureteral reflux was found in 46% of children with bacteremic UTI caused by Escherichia coli vs. 89% of children with non‐E. coli infection (P < 0.01). Conclusions. Clinical symptoms do not significantly distinguish bacteremic from nonbacteremic children with UTI. Outcome of bacteremic UTI was comparable with that of nonbacteremic UTI. Bacteremic children, especially those with non‐E. coli UTI, more often had anatomical or functional abnormalities in the urinary tract.

Mechanisms of renal damage owing to infection.

Urinary tract infection (UTI) is a common bacterial illness in children. It is known to be associated with an increased risk of permanent renal cell damage and scarring which may lead to generation of pathological conditions such as hypertension, pre-eclampsia during pregnancy, renal insufficiency, and end-stage kidney disease. The pathophysiology of renal scarring is still obscure, which makes the prevention of renal damage difficult. During acute infection, there are numerous factors that may contribute to tissue damage. Inflammatory responses are activated by host defense mechanisms as well as by specific bacterial virulence factors. Understanding of these complex mechanisms would be helpful to better identify children at high risk of developing renal scarring following UTI.

Renal manifestations of Henoch–Schönlein purpura in a 6-month prospective study of 223 children

Objective To assess the risk factors for developing Henoch–Schönlein purpura nephritis (HSN) and to determine the time period when renal involvement is unlikely after the initial disease onset. Design A prospective study of 223 paediatric patients to examine renal manifestations of Henoch–Schönlein purpura (HSP). The patients condition was monitored with five outpatient visits to the research centre and urine dipstick testing at home. Results HSN occurred in 102/223 (46%) patients, consisting of isolated haematuria in 14%, isolated proteinuria in 9%, both haematuria and proteinuria in 56%, nephrotic-range proteinuria in 20% and nephrotic-nephritic syndrome in 1%. The patients who developed HSN were significantly older than those who did not (8.2±3.8 vs 6.2±3.0 years, p<0.001, CI for the difference 1.1 to 2.9). Nephritis occurred a mean of 14 days after HSP diagnosis, and within 1 month in the majority of cases. The risk of developing HSN after 2 months was 2%. Prednisone prophylaxis did not affect the timing of the appearance of nephritis. The risk factors for developing nephritis were age over 8 years at onset (OR 2.7, p=0.002, CI 1.4 to 5.1), abdominal pain (OR 2.1, p=0.017, CI 1.1 to 3.7) and recurrence of HSP disease (OR 3.1, p=0.002, CI 1.5 to 6.3). Patients with two or three risk factors developed nephritis in 63% and 87% of cases, respectively. Laboratory tests or blood pressure measurement at onset did not predict the occurrence of nephritis. Conclusion The authors recommend weekly home urine dipstick analyses for the first 2 months for patients with HSP. Patients with nephritis should be followed up for more than 6 months as well as the patients with HSP recurrence.

Clinical course of extrarenal symptoms in Henoch–Schönlein purpura: a 6-month prospective study

Objective To describe the extrarenal symptoms and clinical course of Henoch–Schönlein purpura (HSP). Design A prospective national multicentre trial with 6-month follow-up. Patients A total of 223 newly diagnosed paediatric HSP patients. Results Purpura was the initial symptom in 73% of the patients and was preceded by joint or gastrointestinal manifestations in the rest by a mean of 4 days. Joint symptoms, abdominal pain, melena, nephritis and recurrences occurred in 90%, 57%, 8%, 46% and 25% of the patients, respectively. Orchitis affected 17/122 (14%) of the boys. Seven patients developed protein-losing enteropathy characterised by abdominal pain, oedema and serum albumin under 30 g/l, and an additional 49 patients had subnormal albumin levels without any proteinuria. Positive fecal occult blood (26/117, 22%) and α1-antitrypsin (7/77, 9%) suggested mucosal injury even in the patients without gastrointestinal symptoms. HSP was often preceded by various bacterial, especially streptococcal (36%) and viral infections. Previous streptococcal infection did not induce changes in the level of complement component C3. Recurrences were more frequent in patients >8 years of age (OR 3.7, CI 2.0 to 7.0, p<0.001) and in patients with nephritis (OR 4.6, CI 2.3 to 8.9, p<0.001). Patients with severe HSP nephritis had more extrarenal symptoms up to 6 months. There was no difference in the clinical course between the prednisone-treated and non-treated patients during the 6-month follow-up. Conclusions Serum albumin is often low in HSP patients without proteinuria, due to protein loss via the intestine. Although corticosteroids alleviate the symptoms, they seem not to alter the clinical course of HSP during 6 months of follow-up.

Uropathogenic Escherichia coli Toxins Induce Caspase-Independent Apoptosis in Renal Proximal Tubular Cells via ERK Signaling

Background: Pyelonephritis is a risk factor for renal tubular epithelial cell damage. Recent studies have shown that Escherichia coli and/or its toxins may stimulate apoptotic cell death in renal tubular cells, but the underlying molecular mechanisms remain to be elucidated. Methods: Confluent LLC-PK1 cells were exposed to E. coli toxins from overnight cultures of the uropathogenic O6K13H1 (O6) and the nonpathogenic W3110. The cell death was studied with morphological and biological assay. Results:E. coli soluble toxins from uropathogenic O6:K13:H1(O6) strain were found to induce apoptosis in a dose- and time-dependent manner in LLC-PK1 cells. The expression of FasR and the phosphorylation of ERK1/2 were significantly upregulated by O6 soluble toxins in a time-dependent manner. Cell death was completely inhibited by two specific ERK1/2 inhibitors, but not by a broad caspase inhibitor, zVAD-fmk, implicating a caspase-independent pathway via ERK. Moreover, we found that lysophosphatidic acid could trigger a survival signal through G-proteins and PI3K. Conclusion: We demonstrate that apoptosis induced by uropathogenic E. coli toxins is dependent on ERK1/2. Caspases, although being activated, are not necessary for cell death, and they act after the ERK signaling at which point cells become committed to cell death or can be rescued.

A Case of Primary JC Polyomavirus Infection–Associated Nephropathy

A 15‐year‐old boy with a posterior urethral valve received a deceased donor kidney transplant (KT) in March 2011. Basiliximab induction followed by tacrolimus‐based triple medication was used as immunosuppression. Eleven months after KT, the graft function deteriorated and the biopsy demonstrated interstitial nephritis suggestive of acute rejection. BK polyomavirus (BKPyV) surveillance in urine and plasma was negative. The patient received methylprednisolone pulses and anti‐thymocyte globulin. Immunohistochemistry was positive for simian virus 40 (SV40) large T‐antigen (LTag) in the biopsies, and quantitative polymerase chain reaction for JC polyomavirus (JCPyV) indicated high viral loads in urine and borderline levels in plasma. Immunosuppression was reduced and follow‐up biopsies showed tubular atrophy and interstitial fibrosis. Two years after KT, antibody‐mediated rejection resulted in graft loss and return to hemodialysis. Retrospective serologic work‐up indicated a primary JCPyV infection with seroconversion first for IgM, followed by IgG, but no indication of BKPyV infection. In the SV40 LTag positive biopsies, JCPyV deoxyribonucleic acid (DNA) with archetype noncoding control region was detected, while BKPyV DNA was undetectable. To the best of our knowledge, this is the first reported case of primary JCPyV infection as the cause of PyV‐associated nephropathy in KT.

Long‐term follow‐up of renal function after high‐dose methotrexate treatment in children

High‐dose methotrexate (HD‐MTX) is commonly used in treatment of pediatric leukemias and lymphomas. Transient deterioration in renal function is frequently noted during HD‐MTX treatment, but possible long‐term changes are less well known. In this study we aimed to study long‐term renal prognosis after HD‐MTX treatment, and to find possible underlying risk factors for reduced renal function.

Comparison of glomerular function tests in children with cancer.

Evaluation of renal function should be performed as part of the follow-up during and after chemotherapy in pediatric cancer patients. The aim of this study was to compare an isotope clearance method [isotope glomerular filtration rate (iGFR)] with alternative methods to determine GFR in such patients. Isotope GFR [99mTc-labeled diethylene triaminopentoacetic acid (DTPA) or 51Cr-labeled ethylenediaminetetra-acetate (EDTA)] was measured in 36 children (112 studies) and compared with simultaneously measured creatinine clearance (CrCl), serum creatinine (SCr), and cystatin C (CysC) concentrations, as well as the results of Schwartz, Counahan–Barratt, and Cockroft–Gault formulae, using general linear mixed models. Our results showed a significant association between iGFR and CysC concentrations (p < 0.001). No linear relationship was observed between CrCl and iGFR (p = 0.7). As expected, the results of height-based formulae (Counahan–Barratt and Schwartz) had significantly (p = 0.004) better correlation to iGFR than the results of a formula based on weight (Cockroft–Gault) (p = 0.19). Despite significant linear correlation, intraclass correlation coefficients showed poor agreement. Tests of similarity between iGFR estimates showed differences between average values of GFR. Therefore, determination of iGFR remains the method of choice in estimation of GFR in cancer patients. In our study population, assay of serum CysC was the most reliable alternative method to measure glomerular function.

Carbon monoxide prevents apoptosis induced by uropathogenic Escherichia coli toxins

Urinary tract infections (UTIs) are often caused by Escherichia coli (E. coli). Previous studies have demonstrated that up-regulation of heme oxygenase-1 (HO-1) may trigger a survival mechanism against renal cell death induced by E. coli toxins. The present study analyses the role of carbon monoxide (CO), an end product of HO-1, in the survival mechanism. Moreover, we identified hemolysin as a putative pro-apoptotic toxin in the E. coli supernatant. Tubular cells were incubated with CO in the presence or absence of E. coli toxins. Uropathogenic or transformants of non-pathogenic strains expressing hemolysin were used. We found that the survival pathway during E. coli infection might be activated by HO-1-derived production of CO. The protection by CO was also associated with up-regulation of p21 protein expression. Furthermore, we found that in children with pyelonephritis, all the E. coli strains expressing hemolysin induced apoptosis. In E. coli strains not expressing hemolysin, only 45% of the strains could induce apoptosis. In conclusion, generation of CO elicited by HO-1 could promote survival signaling in renal cells. Hemolysin is one of the secreted toxins that are involved in inducing apoptosis during UTI.