Lukas Kairaitis

PARTIAL DEPLETION OF MACROPHAGES BY ED7 REDUCES RENAL INJURY IN ADRIAMYCIN NEPHROPATHY

Background:  Because macrophages are considered to be possible effectors of disease in Adriamycin (ADR) nephrosis, we hypothesized that depletion of macrophages might protect against the initiation of renal injury. In the present study, a monoclonal antibody (ED7) directed against CD11b/CD18 integrin, which is expressed by macrophages, was used to investigate the pathogenetic effects of macrophages in ADR nephropathy.

Molecular Mechanisms by Which Iron Induces Nitric Oxide Synthesis in Cultured Proximal Tubule Cells

Nitric oxide (NO) levels are increased after exposure of cultured proximal tubule cells (PTC) to non-haem iron, potentially contributing to PTC injury in disease states associated with increased iron exposure, including proteinuric renal disease. The mechanisms underlying this observed increase were investigated. After 3 h exposure to 400 µM nitrilotriacetate (NTA)-Fe, inducible nitric oxide synthase (iNOS) mRNA expression was significantly increased, with a corresponding increase in iNOS protein after 12 h. The nuclear binding activity of NFĸB with 400 µM NTA-Fe was increased, and pyrrolidine dithiocarbamate (PDTC), an antioxidant inhibitor of NFĸB, prevented both activation of NFĸB and NO production in response to NTA-Fe. Inhibition of protein tyrosine kinase reduced iNOS mRNA, iNOS protein levels and NO production in response to NTA-Fe. The effect of tyrosine kinase inhibition on NFĸB activation was variable, with herbimycin but not genistein having an inhibitory effect. Activation of either protein kinase A or C increased iNOS mRNA and protein levels, and NO production in response to NTA-Fe, whereas only the protein kinase C activator phorbol dibutyrate (PDBu) had a stimulatory effect on NFĸB activation. The protein kinase A activator forskolin did not alter iron-induced activation of NFĸB. These data suggest that the observed increase in NO production by PTC in response to iron is due to increased transcription of iNOS. The transcriptional regulation of this response is complex and involves NFĸB, protein tyrosine kinase and the protein kinases A and C.

Defining a Significant Stenosis in an Autologous Radio‐Cephalic Arteriovenous Fistula for Hemodialysis

The current definition of a significant stenosis in an autologous arteriovenous fistula (aAVF), the percentage narrowing compared with the adjacent “normal” vessel, is inaccurate. We believe a significant stenosis in the aAVF is an absolute minimal luminal diameter determined by the requirements of the hemodialysis pump. To determine what absolute diameter constitutes a hemodynamically significant stenosis in a radio‐cephalic autologous arteriovenous fistula (RC aAVF), the minimal luminal diameter of dysfunctional RC aAVF was compared to that of functional RC aAVF using grayscale and color ultrasound. There were 93 fistulas in study group and 77 in control group. The mean minimum luminal diameter in study group was significantly lower than in control group (2.19 vs. 4.71 mm, p 0.001). With a cutoff value of 2.7 mm, there was 90% sensitivity and 80% specificity in distinguishing functional fistula from dysfunctional fistula. The area under the receiver–operator curve was 90% (CI 84–94%), indicating that a 2.7 mm diameter is accurate in discriminating functional from dysfunctional fistulas. An absolute minimal luminal diameter of 2.7 mm, as determined with grayscale and color ultrasound, is a useful cutoff for defining significant stenosis in a RC aAVF.

The CD40–CD154 co-stimulation pathway mediates innate immune injury in adriamycin nephrosis

BACKGROUND Blockade of CD40-CD40 ligand (CD154) interactions protects against renal injury in adriamycin nephropathy (AN) in immunocompetent mice. To investigate whether this protection relied on adaptive or cognate immunity, we tested the effect of CD40-CD154 blockade in severe combined immunodeficient (SCID) mice. METHODS SCID mice were divided into three groups: normal, AN + hamster IgG (ADR+IgG group) and AN + anti-CD154 antibody (MR1) (ADR+MR1 group). AN was induced by tail vein injection of 5.2 mg/kg of adriamycin (ADR). Hamster IgG (control Ab) or MR1 was administered intraperitoneally on days 5, 7, 9 and 11 after ADR injection. Histological and functional data were collected 4 weeks after ADR injection. In vitro experiments tested the effect of soluble and cell-bound CD154 co-cultured with CD40-expressing cells [macrophages, mesangial cells and renal tubular epithelial cells (RTEC)]. RESULTS All experimental animals developed nephropathy. Compared to the ADR+IgG group, ADR+MR1 animals had significantly less histological injury (glomerulosclerosis and tubular atrophy) and functional injury (creatinine clearance). Kidneys of ADR+MR1 animals had significantly less macrophage infiltration than those of ADR+IgG animals. Interestingly, expression of CD40 and CD41 (a platelet-specific marker) was significantly less in ADR+MR1 animals compared to ADR+IgG animals. In vitro, CD154 blockade significantly attenuated upregulation of CCL2 gene expression by RTEC stimulated by activated macrophage-conditioned medium. In contrast, platelet-induced upregulation of macrophage and mesangial cell proinflammatory cytokine gene expression were not CD154-dependent. CONCLUSION CD40-CD154 blockade has a significant innate renoprotective effect in ADR nephrosis. This is potentially due to inhibition of macrophage-derived soluble CD154.

Effects of proactive iron and erythropoiesis-stimulating agent protocol implementation on achieving clinical guideline targets for anaemia in a satellite haemodialysis patient cohort.

Aim:  Anaemia management with erythropoiesis‐stimulating agents (ESA) and i.v. iron replacement in haemodialysis patients poses several clinical challenges, including maintaining stable haemoglobin (Hb) levels within target ranges while achieving lowest effective ESA dose. This manuscript describes the effect of implementing proactive protocol‐driven adjustments for iron and ESA in maintenance haemodialysis patients.

Prevention of recurrent calcium nephrolithiasis

Date written: May 2005

Tubular‐interstitial interactions in proteinuric renal diseases

SUMMARY: Progressive chronic renal disease of all types is characterized by a relatively uniform set of tubular (atrophy, hypertrophy, hyperplasia) and interstitial (inflammatory cell infiltration, fibrosis) pathological changes, the severity of which correlates well with the degree of proteinuria and the decline in glomerular filtration. Complex and redundant pathophysiological events underlie the relationship between proteinuria, tubular injury and interstitial damage, forming potential targets for therapy. The outcome of these events is also determined by additional factors not limited to proteinuric diseases, including tissue hypoxia, complement activation and mediators produced in response to glomerular damage. In response to proteinuria and these additional factors, tubular cells are activated to produce a large number of chemoattractants, proinflammatory and profibrotic cytokines and matrix proteins, which cause, at least in part, interstitial inflammation and fibrosis. In turn, soluble products of interstitial inflammatory cells (predominantly T lymphocytes and macrophages), dendritic cells and fibroblasts are at least partially responsible for altering the phenotype of tubular cells. Abnormal tubular‐interstitial interactions may also depend on direct cell contact rather than soluble mediators, as evidenced by up‐regulation of integrins and cell adhesion molecules. Myofibroblasts, a key cell in the production of interstitial fibrosis, may arise in response to the above mediators by trans‐differentiation of fibroblasts, perivascular cells and tubular cells. In addition, tubular cells have been shown to be capable of the induced expression of the signalling molecules necessary for them to behave as antigen‐presenting cells and thereby potentially initiate interstitial inflammation through the classic interactions of cognate immunity. It is probable that several, but not all, of this bewildering number of pathophysiological events are of prime importance to the development of tubulointerstitial damage in proteinuria. The immediate challenge for investigators is to define the relative importance of these events and, thus, the most appropriate targets for new treatments.

Protein Z is reduced in chronic kidney disease and not elevated in patients on haemodialysis.

Protein Z (PZ) is a vitamin K dependent serine protease inhibitor, which along with its cofactor Protein Z-dependant protease inhibitor (ZPI) has anti-Xa activity. PZ has previously been reported to be elevated in patients with end-stage chronic kidney disease (CKD) on haemodialysis but not in non-dialysed CKD patients raising the possibility that PZ may have a role in the bleeding diathesis of patients on haemodyalisis. PZ was measured in controls (n = 18), CKD on haemodialysis (n = 23) and CKD not on dialysis (n = 23). Patients on vitamin K antagonists, with acute inflammatory conditions (as measured by CRP) or liver dysfunction were excluded PZ levels were reduced in CKD patients when compared to levels in the control group (1.35 μg/mL vs 1.80 respectively, p = 0.022). Subgroup analysis revealed a trend toward reduction in mean PZ in the CKD subgroups (non-dialysed CKD group 1.37 (p = 0.08); haemodialysis 1.33 (p = 0.12)). There were no significant differences in the PZ levels for different stages of non-dialysed CKD patients when stratified by the level of glomerular filtration rate (GFR). These data show that PZ levels were reduced in patients with CKD, and not elevated in patients on haemodialysis, arguing against a role for PZ in the bleeding diathesis of renal failure. This finding is in contradiction to a previous report which found PZ to be elevated in patients on haemodialysis.

Diameter of inflow as a predictor of radiocephalic fistula flow

Introduction: The optimal method for vascular access surveillance is largely unknown. A previous case–control study suggested a simplified anatomical measure obtained by Doppler ultrasound—the narrowest segment of the circuit or “minimal luminal diameter” may identify patients with a dysfunctional radiocephalic arteriovenous fistula. The relationship between minimal luminal diameter and access flow (Qa) in the radiocephalic arteriovenous fistula has not previously been studied. Methods: Patients undergoing Doppler ultrasound of a radiocephalic arteriovenous fistula over an 8-month period were identified retrospectively. Minimal luminal diameter was identified and demographic and clinical data were collected. Qa was estimated by Doppler estimation of brachial artery flow. The relationship between minimal luminal diameter and Qa was examined by correlation and using different levels of minimal luminal diameter as a simplified measure to detect or exclude low Qa (<600 mL/min). Results: A total of 81 Doppler ultrasound scans were performed. In all, 26 scans demonstrated brachial artery flow <600 mL/min. Simple logistic regression indicated a weak statistical relationship between the minimal luminal diameter and Qa (R2 = 0.27, p < 0.01). Minimal luminal diameter performed poorly as a marker of low Qa with low specificity, however, showed high negative predictive value for ruling out low Qa at a minimal luminal diameter of 3.2 mm or higher (94%). Qa estimated by brachial artery flow correlated well with Qa estimated by indicator dilution (R2 = 0.83, p < 0.01) without significant mean difference or proportional bias. Conclusion: Minimal luminal diameter correlates weakly with Qa. Low minimal luminal diameter values should not be used in isolation to determine low Qa for a radiocephalic arteriovenous fistula. Conversely, minimal luminal diameter >3.2 mm largely excludes a low-flow radiocephalic arteriovenous fistula in this cohort. Brachial artery flow is a reasonable measure of Qa in comparison with indicator dilution.

Multicentre, randomised, blinded, control trial of drug-eluting balloon vs Sham in recurrent native dialysis fistula stenoses

Background: Endovascular treatment of autogenous arteriovenous haemodialysis fistula stenosis has high reintervention rates. We investigate the effect of drug-eluting balloons in the treatment of recurrent haemodialysis fistula stenosis. Methods: This is a randomised, controlled, investigator-initiated and run, prospective, blinded, multicentre trial. Patients with recurrent autogenous arteriovenous haemodialysis fistula stenosis received standard endovascular treatment plus drug-eluting balloon or standard endovascular treatment plus uncoated balloon (Sham). Primary endpoint was late lumen loss in trial area on ultrasound at 6 weeks, 3, 6 and 12 months. Secondary endpoints were freedom from reintervention to the Index Trial Area and decline in fistula flow (Qa). Interim analysis was performed at 6 months (unblinded due to timeliness). Results: Patients with 132 recurrent stenoses (48% in bare Nitinol stents) were randomised with 70 receiving drug-eluting balloon and 62 Sham. At 6 months, decline in late lumen loss was 0.23 ± 0.03 mm/month for Sham and 0.045 ± 0.03 mm/month for drug-eluting balloon arm, a significant difference (0.18 mm, p = 0.0002). At 12 months, this difference persisted at 0.12 mm (p = 0.0003). At 6 months, significant difference in late lumen loss for instent restenoses (p = 0.0004) was observed, with non-significant difference for unstented restenoses (p = 0.065). Mean time for freedom from reintervention was 10.14 months for Sham versus 42.39 months for drug-eluting balloon (p = 0.001). The same was shown for instent (p = 0.014) and unstented (p = 0.029) restenoses. Qa decline rate at 6 months was 36.89 mL/min/month (Sham) and 0.41 mL/min (drug-eluting balloon). The difference was significant (36.48 mL/min; p = 0.02) and persisted to 12 months (p = 0.44). Conclusion: Paclitaxel drug-eluting balloon significantly delays restenosis after angioplasty for recurrent autogenous arteriovenous haemodialysis fistula stenosis, persisting to 12 months. Drug-eluting balloon significantly increases freedom from reintervention at 12 months with these effects true in stented and unstented fistulas.