CXCR5 is critically involved in progression of lupus through regulation of B cell and double-negative T cell trafficking.

The recruitment of immune cells to sites of tissue inflammation is orchestrated by chemokine/chemokine receptor networks. Among these, the CXCL13/CXCR5 axis is thought to be involved critically in systemic lupus erythematosus (SLE) and lupus nephritis pathogenesis. Beyond B cell abnormalities, another hallmark of SLE disease is the occurrence of aberrant T cell responses. In particular, double-negative (DN) T cells are expanded in the peripheral blood of patients with SLE and in lupus-prone mice. DN T cells induce immunoglobulin production, secrete proinflammatory cytokines and infiltrate inflamed tissue, including kidneys. We aimed to investigate how CXCR5 deficiency changes immune cell trafficking in murine lupus. We therefore crossed CXCR5(-/-) mice with B6/lpr mice, a well-established murine lupus model. B cell numbers and B cellular immune responses were diminished in CXCR5-deficient B6/lpr mice. In addition, we observed reduced accumulation of DN T cells in spleen and lymph nodes, paralleled by reduced splenomegaly and lymphadenopathy. In-vivo migration assays revealed reduced migration of CXCR5-deficient DN T cells into lymph nodes, and ex-vivo-activated CXCR5-deficient DN T cells failed to infiltrate kidneys of recipients. Moreover, DN T cells and B cells of CXCR5-deficient B6/lpr mice failed to migrate towards CXCL13 in vitro. We propose that CXCR5 is involved critically in B cell trafficking and germinal cell (GC) formation in murine lupus and in guiding pathogenic DN T cells into lymphoid organs and kidneys, and we therefore describe new pathomechanisms for the CXCL13/CXCR5 axis in SLE.

VEGF(165) mediates glomerular endothelial repair.

VEGF(165), the most abundant isoform in man, is an angiogenic cytokine that also regulates vascular permeability. Its function in the renal glomerulus, where it is expressed in visceral epithelial and mesangial cells, is unknown. To assess the role of VEGF(165) in glomerular disease, we administered a novel antagonist - a high-affinity, nuclease-resistant RNA aptamer coupled to 40-kDa polyethylene glycol (PEG) - to normal rats and to rats with mesangioproliferative nephritis, passive Heymann nephritis (PHN), or puromycin aminonucleoside nephrosis (PAN). In normal rats, antagonism of VEGF(165) for 21 days failed to induce glomerular pathology or proteinuria. In rats with mesangioproliferative nephritis, the VEGF(165) aptamer (but not a sequence-scrambled control RNA or PEG alone) led to a reduction of glomerular endothelial regeneration and an increase in endothelial cell death, provoking an 8-fold increase in the frequency of glomerular microaneurysms by day 6. In contrast, early leukocyte influx and the proliferation, activation, and matrix accumulation of mesangial cells were not affected in these rats. In rats with PHN or PAN, administration of the VEGF(165) aptamer did not influence the course of proteinuria using various dosages and administration routes. These data identify VEGF(165) as a factor of central importance for endothelial cell survival and repair in glomerular disease, and point to a potentially novel way to influence the course of glomerular diseases characterized by endothelial cell damage, such as various glomerulonephritides, thrombotic microangiopathies, or renal transplant rejection.

Gene-polymorphisms of angiotensin converting enzyme and endothelial nitric oxide synthase in patients with primary glomerulonephritis.

UNLABELLED: The ACE D- and the ecNOS a-allele have been associated with an adverse prognosis in patients with glomerulonephritis (GN). Using genomic DNA we investigated by RT-PCR whether the two polymorphisms are useful prognostic markers in GN patients. In patients with primary GN (IgA-GN n = 70, membranous GN n = 23, FSGS n = 17, MPGN n = 6) neither the whole group nor disease-specific subgroups exhibited any alterations from the normal ACE genotype distribution. No significant associations were detected between the ACE genotype and the development of hypertension, antihypertensive therapy required, progression rate of the disease, age of diagnosis and the antiproteinuric response to ACE-inhibition. In 40 IgA-GN patients with ESRD no increased prevalence of the D-allele was noted. The distribution of the ecNOS-alleles in the above patients (a-allele 22%, b-allele 78%) was comparable to that of the normal controls. No association with any of the parameters mentioned above were detected in the case of the ecNOS-alleles. CONCLUSIONS: In our Caucasian patients neither the determination of the ACE nor the ecNOS genotype offered any diagnostic or prognostic help.

[Aptamers: a novel approach to intervention studies and development of novel therapeutic approaches]. / Aptamere: ein neuer Ansatz für Interventionsstudien und zur Entwicklung zukünftiger Therapieansätze.

A rapidly growing number of factors is identified that might contribute to disease. To characterize the pathogenetic relevance of a particular factor, specific intervention studies in vivo appear necessary. The present discussion deals with a new class of inhibitors, i.e. aptamers. Aptamers (derived from the latin word "aptus" = fitting) are short DNA or RNA oligomers which can bind to a given ligand with high affinity and specificity due to their particular three-dimensional structure and which may thereby, for example, antagonize the biological function of the ligand. Aptamers have been generated against a large variety of molecules ranging from amino acids to complex proteins and even disaccharides. Using platelet-derived growth factor (PDGF) and an experimental mesangioproliferative glomerulonephritis as a model, we describe the in vivo effects of an antagonistic aptamer against PDGF-B. Such studies will greatly aid the identification of the biological role of particular mediators and ultimately the design of novel therapeutic strategies.

CCN3 is a novel endogenous PDGF-regulated inhibitor of glomerular cell proliferation.

CCN proteins affect cell proliferation, migration, attachment, and differentiation. We identified CCN3 as a suppressed gene following platelet-derived growth factor (PDGF)-BB or -DD stimulation in a cDNA-array analysis of mesangial cells. In vitro growth-arrested mesangial cells overexpressed and secreted CCN3, whereas the addition of the recombinant protein inhibited cell growth. Induction of mesangial cell proliferation by PDGF-BB or the specific PDGF beta-receptor ligand PDGF-DD led to downregulation of CCN3 mRNA, confirming the array study. Specific PDGF alpha-receptor ligands had no effect. CCN3 protein was found in arterial smooth muscle cells, the medullary interstitium, and occasional podocytes in the healthy rat kidney. Glomerular CCN3 was low prior to mesangial proliferation but increased as glomerular cell proliferation subsided during mesangioproliferative glomerulonephritis (GN). Inhibition of PDGF-B in mesangioproliferative disease led to overexpression of glomerular CCN3 mRNA. CCN3 localized mostly to podocytes in human glomeruli, but this expression varied widely in different human glomerulonephritides. Glomerular cell proliferation negatively correlated with CCN3 expression in necrotizing GN. Our study identifies CCN3 as an endogenous inhibitor of mesangial cell growth and a modulator of PDGF-induced mitogenesis.

Upregulation of group IB secreted phospholipase A(2) and its M-type receptor in rat ANTI-THY-1 glomerulonephritis.

Treatment of rat glomerular mesangial cell (GMC) cultures with pancreatic secreted phospholipase A(2) (sPLA(2)-IB) results in an enhanced expression of sPLA(2)-IIA and COX-2, possibly via binding to its specific M-type sPLA(2) receptor. In the current study, we have investigated the expression and regulation of sPLA(2)-IB and its receptor during glomerulonephritis (GN). In vivo we used the well-established rat model of anti-Thy 1.1 GN (anti-Thy 1.1-GN) to study the expression of sPLA(2)-IB and the M-type sPLA(2) receptor by immunohistochemistry. In addition, in vitro we determined the interkeukin (IL)-1beta-regulated mRNA and protein expression in primary rat glomerular mesangial and endothelial cells as well as in rat peripheral blood leukocytes (PBLs). Shortly after induction of anti-Thy 1.1-GN, sPLA(2)-IB expression was markedly upregulated in the kidney at 6-24 h. Within glomeruli, the strongest sPLA(2)-IB protein expression was detected on infiltrated granulocytes and monocytes. However, at the same time, the M-type receptor was also markedly upregulated on resident glomerular cells. In vitro, the most prominent cytokine-stimulated secretion of sPLA(2)-IB was observed in monocytes isolated from rat PBLs. Treating glomerular endothelial cells (GECs) with cytokines elicited only weak sPLA(2)-IB expression, but treatment of these cells with exogenous sPLA(2)-IB resulted in a marked expression of the endogenous sPLA(2)-IB. Mesangial cells did not express sPLA(2)-IB at all. The M-type sPLA(2) receptor protein was markedly upregulated on cytokine-stimulated mesangial and endothelial cells as well as on lymphocytes and granulocytes. During anti-Thy 1.1 rat GN, sPLA(2)-IB and the M-type sPLA(2) receptor are induced as primary downstream genes stimulated by inflammatory cytokines. Subsequently, both sPLA(2)-IB and the M-type sPLA(2) receptor are involved in the autocrine and paracrine amplification of the inflammatory process in different resident and infiltrating cells.

Biological responses to PDGF-BB versus PDGF-DD in human mesangial cells.

Platelet-derived growth factor (PDGF)-BB and PDGF-DD mediate mesangial cell proliferation in vitro and in vivo. While PDGF-BB is a ligand for the PDGF alpha- and beta-receptor chains, PDGF-DD binds more selectively to the beta-chain, suggesting potential differences in the biological activities. Signal transduction and regulation of gene expression induced by PDGF-BB and -DD were compared in primary human mesangial cells (HMCs), which expressed PDGF alpha- and beta-receptor subunits. The growth factor concentrations used were chosen based on their equipotency in inducing HMCs proliferation and binding to the betabeta-receptor. Both growth factors, albeit at different concentrations induced phosphorylation and activation of extracellular signal-regulated kinase 1 (ERK1) and ERK2. In addition, PDGFs led to the phosphorylation and activation of signal transducers and activators of transcription 1 (STAT1) and STAT3. HMCs proliferation induced by either PDGF-BB or -DD could be blocked by signal transduction inhibitors of the mitogen-activated protein kinase-, Janus kinase (JAK)/STAT-, or phosphatidyl-inositol 3-kinase pathways. Using a gene chip array and subsequent verification by real-time reverse transcriptase (RT)-polymerase chain reaction, we found that in HMC genes for matrix metalloproteinase 13 (MMP-13) and MMP-14 and, to a low extent, cytochrome B5 and cathepsin L were exclusively regulated by PDGF-BB, whereas no exclusive gene regulation was detected by PDGF-DD. However, at the protein level, both MMP-13 and -14 were equally induced by PDGF-BB and -DD. PDGF-BB and -DD effect similar biological responses in HMCs albeit at different potencies. Rare apparently differential gene regulation did not result in different protein expression, suggesting that in HMCs both PDGFs exert their biological activity almost exclusively via the PDGF beta-receptor.

Cytokines and glomerular injury.

Since reports of the first data on cytokines in renal diseases about 10 years ago, the field has seen an explosive growth over the last decade. It is now realized that cytokines can be synthesized by many different types of cells, including glomerular cells, upon adequate stimulation and exert a wide range of actions mostly in an autocrine or paracrine fashion. Cytokines not only contribute to developmental processes, but also to the maintenance of normal tissue integrity and in particular to the initiation and modulation of tissue injury, healing, or scarring. This review summarizes some recent data on the role of cytokines in glomerular disease and describes some of the first experimental approaches to therapeutically interfere with cytokine actions.

[Mutations in the PIG-A gene lead to GPI-deficiency in paroxysmal nocturnal hemoglobinuria]. / Mutationen im PIG-A-Gen führen zur GPI-Defizienz bei der paroxysmalen nächtlichen Hämoglobinurie.

Paroxysmal nocturnal hemoglobinuria (PNH) is characterized by the deficiency of glycosylphosphatidylinositol-(GPI-)-anchored surface molecules on blood cells. The biochemical basis of this deficiency is the lack of the first GPI biosynthesis intermediate GlcNAc-PI in the deficient cells corresponding to that in Thy-1- mouse lymphoma mutants of the class A. Recently, the responsible gene (PIG-A gene) has been cloned. Here, PIG-A transcripts in T-, NK- and EBV-transformed B cell lines of different PNH patients have been analyzed. In contrast to the uniform biochemical defect, these molecular analyses reveal heterogenous mutations of the PIG-A gene in different PNH patients.

Biology of GPI anchors and pathogenesis of paroxysmal nocturnal hemoglobinuria.

Recently, a number of leading scientists gathered in Hanover, Germany, to discuss glycosylphosphatidylinositol (GPI)-anchor structures. Here, Jörg Schubert and colleagues report on this highly interesting field.

Characterization of a dam mutant of Serratia marcescens and nucleotide sequence of the dam region.

The DNA of Serratia marcescens has N6-adenine methylation in GATC sequences. Among 2-aminopurine-sensitive mutants isolated from S. marcescens Sr41, one was identified which lacked GATC methylation. The mutant showed up to 30-fold increased spontaneous mutability and enhanced mutability after treatment with 2-aminopurine, ethyl methanesulfonate, or UV light. The gene (dam) coding for the adenine methyltransferase (Dam enzyme) of S. marcescens was identified on a gene bank plasmid which alleviated the 2-aminopurine sensitivity and the higher mutability of a dam-13::Tn9 mutant of Escherichia coli. Nucleotide sequencing revealed that the deduced amino acid sequence of Dam (270 amino acids; molecular mass, 31.3 kDa) has 72% identity to the Dam enzyme of E. coli. The dam gene is located between flanking genes which are similar to those found to the sides of the E. coli dam gene. The results of complementation studies indicated that like Dam of E. coli and unlike Dam of Vibrio cholerae, the Dam enzyme of S. marcescens plays an important role in mutation avoidance by allowing the mismatch repair enzymes to discriminate between the parental and newly synthesized strands during correction of replication errors.

Peripherally inserted central venous catheters for autologous blood progenitor cell transplantation in patients with haematological malignancies.

BACKGROUND: In contrast to the high risk of haemorrhage associated with the implantation of a central venous catheter (CVC) via the internal jugular or subclavian access, the use of a peripherally inserted catheter (PICC) offers the advantage of a lower risk of bleeding complications. However, the rate of phlebitis is higher with the PICC and its use has been declining. We have studied the benefits and adverse events of a new type of PICC and a common type. METHODS: From October 1999 to October 2001, 70 PICCs (Olimpicc, Vygon, Germany, n=40; and LIFECATH-PICC(PUR)5FR Vygon, Germany, n=30) were inserted into 66 patients with haematological malignancies and used for high-dose chemotherapy, total parenteral nutrition and autologous blood stem cell transplantation. While removing the catheter, central and peripheral blood cultures were taken. The catheter tip was investigated by the semi-quantitative roll-out method of Maki. RESULTS: Sixty-five PICCs were removed after a median of 8.9 days. In five cases a catheter-associated significant colonisation with coagulase-negative staphylococci occurred. In two instances catheter-related bacteraemia was found. CONCLUSION: In our study this catheter system was inserted in 94% of patients without problems and showed a low incidence of phlebitis (5/65). Because of the high rate of catheter malfunction reported during and after our study, the Olimpicc catheter is no longer available. The PICC system, and the LIFECATH-PICC(PUR)5FR in particular, offers a safe and effective alternative for central venous access to the internal jugular vein.

C-type natriuretic peptide inhibits mesangial cell proliferation and matrix accumulation in vivo.

Local C-type natriuretic peptide (CNP) production and CNP receptor expression have been demonstrated in glomeruli. However, the glomerular (patho-)physiological functions of CNP are largely unknown. We therefore investigated the effects of CNP on mesangial cell proliferation and matrix accumulation in the rat mesangioproliferative anti-Thy 1.1 model. Over seven days rats received a continuous infusion (1 microgram/kg/min) of either CNP (N = 6), an irrelevant control peptide (N = 3) or buffer alone (N = 6). Kidney biopsies were performed on days 2, 4 and 8. Few significant differences between the groups were noted on days 2 and 4. Compared to buffer treated rats on day 8, those receiving CNP showed a 35% reduction of glomerular mitoses, a 62% reduction of glomerular uptake of the thymidine analogue BrdU and a significant reduction in glomerular expression of PDGF B-chain. Double immunoperoxidase staining also revealed blunting of proliferating, activated mesangial cells (515 reduction of alpha-smooth muscle actin-/BrdU-positive cells) and macrophage influx. Moreover, there was a marked reduction of mesangial collagen IV and fibronectin accumulation at the protein and mRNA level. Rats receiving the control peptide were indistinguishable from buffer treated rats. Systemic blood pressure was reduced by 10 to 20% in both CNP and control peptide treated rats on day 8, excluding that the findings were due to hemodynamic effects of CNP. Our findings demonstrate that CNP is involved in the regulation of mesangial cell proliferation and matrix production in vivo. The data suggest the existence of a glomerular natriuretic peptide system that may regulate tissue homeostasis and contribute to resolution of mesangioproliferative diseases.

Treatment of experimental mesangioproliferative glomerulonephritis with non-anticoagulant heparin: therapeutic efficacy and safety.

Treatment with conventional heparin is effective in experimental mesangioproliferative glomerulonephritis. However, the long-term effects and safety of this therapy, in particular in the presence of mesangiolysis, have not been assessed. In addition, this therapy has been hampered by bleeding complications. In the present study, therefore, we investigated the long-term effects of a short course of non-anticoagulant (NA) heparin treatment in the anti-Thy 1.1 mesangioproliferative glomerulonephritis, in which early immune-mediated mesangiolysis subsequently leads to mesangial hyperproliferation. Rats received continuous ip NA-heparin or vehicle during the active mesangioproliferative phase (Days 2 to 9; early treatment) or during the early resolution phase (Days 10 to 17; late treatment). Whereas NA-heparin in the early treatment group did not affect the glomerular macrophage, lymphocyte, or platelet influx, it did lead to significantly decreased glomerular cellularity, mesangial cell proliferation, alpha-smooth muscle actin, desmin expression (ie, markers of activated mesangial cells), and matrix accumulation as well as to persistent mesangiolytic lesions including microaneurysms. Despite this latter finding, at Day 120, NA-heparin-treated rats of the early treatment group showed significantly better renal function and less proteinuria and glomerulosclerosis than vehicle-infused rats. In contrast, late therapy with NA-heparin neither accelerated resolution of the nephritis or otherwise affected the course of the disease. We conclude that transient NA-heparin therapy is effective in mesangioproliferative glomerulonephritis, both acutely and long term, when it is initiated during the active phase of the disease. Also, NA-heparin therapy is safe even in glomerular diseases accompanied by mesangiolysis.

Heterogeneous PIG-A mutations in different cell lineages in paroxysmal nocturnal hemoglobinuria.

Paroxysmal nocturnal hemoglobinuria (PNH) is an acquired clonal defect of hematopoietic stem cells in which affected cells are characterized by the lack of glycosylphosphatidylinositol (GPI)-anchored proteins. The lesion in PNH lies in the defective synthesis of N-acetyl-D-glucosaminyl-phosphatidylinositol (GlcNAc-Pl), the first intermediate in GPI biosynthesis. Reintroduction of the PIG-A gene into GPI(-) patient cells reportedly complements this defect. We have analyzed here PIG-A transcripts of six PNH patients. GPI+ and GPI- cell lines from each individual were used, ie, Epstein-Barr virus-transformed B-lymphoblastoid cell lines, T-cell lines, and natural killer cell clones. Reverse transcriptase polymerase chain reaction and sequencing showed three different PIG-A splicing variants in GPI+ cell lines, in which the largest transcript contained the wild-type PIG-A coding region sequence. GPI-deficient cell lines showed abnormal splicing variants. Sequencing of PIG-A complementary DNA and genomic DNA showed heterogeneous mutations ranging from different point mutations to small deletions. Two lymphocyte cell lines (T- and B-cell lines) of one patient presented with the same mutation. For another patient, two different mutations were detected in one natural killer cell line. Therefore, different cell lineages have somatic mutations in PIG-A that lead to PNH.

Early events leading to renal injury in obese Zucker (fatty) rats with type II diabetes.

UNLABELLED: Early events leading to renal injury in obese Zucker (fatty) rats with type II diabetes. BACKGROUND: More than half of the new patients admitted to dialysis therapy in some centers are diagnosed with type IIb diabetes, that is, diabetes associated with obesity. This study searched for a common final pathway of renal damage in this progressive renal disease. METHODS: The evolution of biochemical and morphological renal changes was examined in 6- to 60-week-old Zucker rats (fa/fa-rats), a model of obesity associated with type II diabetes. RESULTS: fa/fa-rats exhibited pronounced hyperinsulinemia and hyperlipidemia at 6 weeks and became diabetic after 14 weeks of age. Significant focal segmental glomerulosclerosis was first noted in 18-week-old fa/fa-rats and tubulointerstitial damage and proteinuria in 40-week-old fa/fa-rats. A comparison of kidneys of six-week-old fa/fa-and lean control (Fa/?) rats by immunohistology revealed a 1.8-fold increase in glomerular monocyte/macrophage counts in fa/fa-rats and a significant increase in de novo desmin expression in podocytes. Electron microscopy demonstrated an increase in the number of podocyte mitochondria and intracytoplasmic protein and fat droplets. Podocyte desmin scores markedly increased until week 18 in fa/fa-rats, whereas glomerular monocyte/macrophage counts peaked at 3.2-fold at week 14. Podocyte desmin expression, but not glomerular macrophage infiltration, correlated with damage in adjacent tubular cells, as evidenced by their de novo expression of vimentin. Progressive glomerular hypertrophy was detected in fa/fa-rats after 10 weeks. GBM width was significantly increased in 14-week-old fa/fa-rats as compared with lean controls. Mesangial cell activation (de novo expression of alpha-smooth muscle actin) and proliferation was low to absent throughout the observation period in fa/fa-rats. Renal cell death counts (TUNEL) remained unchanged in 6- to 40-week-old fa/fa-rats. Tubulointerstitial myofibroblast formation and matrix accumulation occurred late during the study duration in fa/fa-rats. CONCLUSION: These data suggest that early progressive podocyte damage and macrophage infiltration is associated with hyperlipidemia and type IIb diabetes mellitus, and antedates both the development of glomerulosclerosis and tubulointerstitial damage.

Improved survival and amelioration of nephrotoxic nephritis in intercellular adhesion molecule-1 knockout mice.

Intercellular adhesion molecule-1 (ICAM-1) expression is upregulated in nephrotoxic nephritis, a model of human rapidly progressive glomerulonephritis. To evaluate the pathogenetic relevance of ICAM-1 in this model, nephrotoxic nephritis was induced in ICAM-1 knockout mice and genetic controls. Mice were preimmunized with rabbit IgG in complete Freund's adjuvant. Seven days later they received rabbit anti-mouse glomerular basement membrane IgG. The early humoral immune responses (levels of circulating mouse anti-rabbit IgG, glomerular deposition of rabbit and mouse IgG and mouse C3c) were not altered in ICAM-1 knockout mice. During 28 d of follow-up, 3 of 19 control nephritic mice and 0 of 16 ICAM-1 knockout mice died. Proteinuria was high in nephritic control mice (means 10 to 12 mg/24 h at all time points investigated) and significantly reduced in nephritic ICAM-1 knockout mice (means <4.4 mg). Mean serum creatinine rose from 29 micromol/L at day -7 to 48 micromol/L (day 28) in nephritic control mice. This increase in serum creatinine was significantly lower in ICAM-1 knockout mice: 27 (day -7) and 36 micromol/L (day 28). Histologic analysis at day 28 revealed that ICAM-1 deficiency in nephrotoxic nephritis mice led to significantly reduced glomerular crescent formation (2+/-3% in ICAM-1 knockout mice versus 13+/-8% in nephritic controls) and tubulointerstitial injury (score 0.4+/-0.4 versus 2.0+/-1.1). By immunohistochemistry, ICAM-1 deficiency in nephritic mice led to significantly reduced (peri-)glomerular and/or interstitial macrophage influx, alpha-smooth muscle actin expression, and type IV collagen accumulation. These data indicate that ICAM-1 is a central mediator of glomerular and tubulointerstitial injury in murine nephrotoxic nephritis.

Novel approach to specific growth factor inhibition in vivo: antagonism of platelet-derived growth factor in glomerulonephritis by aptamers.

Mesangial cell proliferation and matrix accumulation, driven by platelet-derived growth factor (PDGF), contribute to many progressive renal diseases. In a novel approach to antagonize PDGF, we investigated the effects of a nuclease-resistant high-affinity oligonucleotide aptamer in vitro and in vivo. In cultured mesangial cells, the aptamer markedly suppressed PDGF-BB but not epidermal- or fibroblast-growth-factor-2-induced proliferation. In vivo effects of the aptamer were evaluated in a rat mesangioproliferative glomerulonephritis model. Twice-daily intravenous (i.v.) injections from days 3 to 8 after disease induction of 2.2 mg/kg PDGF-B aptamer, coupled to 40-kd polyethylene glycol (PEG), led to 1) a reduction of glomerular mitoses by 64% on day 6 and by 78% on day 9, 2) a reduction of proliferating mesangial cells by 95% on day 9, 3) markedly reduced glomerular expression of endogenous PDGF B-chain, 4) reduced glomerular monocyte/macrophage influx on day 6 after disease induction, and 5) a marked reduction of glomerular extracellular matrix overproduction (as assessed by analysis of fibronectin and type IV collagen) both on the protein and mRNA level. The administration of equivalent amounts of a PEG-coupled aptamer with a scrambled sequence or PEG alone had no beneficial effect on the natural course of the disease. These data show that specific inhibition of growth factors using custom-designed, high-affinity aptamers is feasible and effective.

Role of interleukin-6 in mediating mesangial cell proliferation and matrix production in vivo.

Mesangial cell proliferation and matrix overproduction characterize many progressive glomerular diseases. Based on currently available data, the role of interleukin-6 (IL-6) in mediating mesangial cell proliferation and matrix production is controversial. The present study attempts to clarify this issue by showing that: (1) IL-6 knock out mice develop a normal glomerular architecture and in particular a normal mesangium. (2) Mesangioproliferative glomerulonephritis induced by Habu snake venom is equally severe in IL-6 knock out mice as in control mice. (3) A continuous seven-day intraperitoneal infusion of 50 micrograms recombinant human IL-6 into rats with a prior minimal (subnephritogenic) injury to mesangial cells does not induce glomerular cell activation, cell proliferation, matrix production, leukocyte influx, platelet influx or proteinuria. (4) A continuous seven-day IL-6 infusion into rats with mesangioproliferative nephritis (anti-Thy 1.1 nephritis) increases matrix protein transcription in the absence of detectable effects on matrix protein accumulation and otherwise has no effect on the natural course of the disease. We conclude from these findings that IL-6 is not an important mediator of mesangial cell proliferation and matrix overproduction in vivo, and that currently little rationale exists to advocate anti-IL-6 therapy in mesangioproliferative disease states.