Protective Effects of Cilastatin against Vancomycin-Induced Nephrotoxicity.

Vancomycin is a very effective antibiotic for treatment of severe infections. However, its use in clinical practice is limited by nephrotoxicity. Cilastatin is a dehydropeptidase I inhibitor that acts on the brush border membrane of the proximal tubule to prevent accumulation of imipenem and toxicity. The aim of this study was to investigate the potential protective effect of cilastatin on vancomycin-induced apoptosis and toxicity in cultured renal proximal tubular epithelial cells (RPTECs). Porcine RPTECs were cultured in the presence of vancomycin with and without cilastatin. Vancomycin induced dose-dependent apoptosis in cultured RPTECs, with DNA fragmentation, cell detachment, and a significant decrease in mitochondrial activity. Cilastatin prevented apoptotic events and diminished the antiproliferative effect and severe morphological changes induced by vancomycin. Cilastatin also improved the long-term recovery and survival of RPTECs exposed to vancomycin and partially attenuated vancomycin uptake by RPTECs. On the other hand, cilastatin had no effects on vancomycin-induced necrosis or the bactericidal effect of the antibiotic. This study indicates that cilastatin protects against vancomycin-induced proximal tubule apoptosis and increases cell viability, without compromising the antimicrobial effect of vancomycin. The beneficial effect could be attributed, at least in part, to decreased accumulation of vancomycin in RPTECs.

Gene Deficiency in Activating Fcγ Receptors Influences the Macrophage Phenotypic Balance and Reduces Atherosclerosis in Mice.

Immunity contributes to arterial inflammation during atherosclerosis. Oxidized low-density lipoproteins induce an autoimmune response characterized by specific antibodies and immune complexes in atherosclerotic patients. We hypothesize that specific Fcγ receptors for IgG constant region participate in atherogenesis by regulating the inflammatory state of lesional macrophages. In vivo we examined the role of activating Fcγ receptors in atherosclerosis progression using bone marrow transplantation from mice deficient in γ-chain (the common signaling subunit of activating Fcγ receptors) to hyperlipidemic mice. Hematopoietic deficiency of Fcγ receptors significantly reduced atherosclerotic lesion size, which was associated with decreased number of macrophages and T lymphocytes, and increased T regulatory cell function. Lesions of Fcγ receptor deficient mice exhibited increased plaque stability, as evidenced by higher collagen and smooth muscle cell content and decreased apoptosis. These effects were independent of changes in serum lipids and antibody response to oxidized low-density lipoproteins. Activating Fcγ receptor deficiency reduced pro-inflammatory gene expression, nuclear factor-κB activity, and M1 macrophages at the lesion site, while increasing anti-inflammatory genes and M2 macrophages. The decreased inflammation in the lesions was mirrored by a reduced number of classical inflammatory monocytes in blood. In vitro, lack of activating Fcγ receptors attenuated foam cell formation, oxidative stress and pro-inflammatory gene expression, and increased M2-associated genes in murine macrophages. Our study demonstrates that activating Fcγ receptors influence the macrophage phenotypic balance in the artery wall of atherosclerotic mice and suggests that modulation of Fcγ receptor-mediated inflammatory responses could effectively suppress atherosclerosis.

Peptide inhibitor of NF-κB translocation ameliorates experimental atherosclerosis.

Atherosclerosis is a chronic inflammatory disease of the arterial wall. NF-κB is a major regulator of inflammation that controls the expression of many genes involved in atherogenesis. Activated NF-κB was detected in human atherosclerotic plaques, and modulation of NF-κB inflammatory activity limits disease progression in mice. Herein, we investigate the anti-inflammatory and atheroprotective effects of a cell-permeable peptide containing the NF-κB nuclear localization sequence (NLS). In vascular smooth muscle cells and macrophages, NLS peptide specifically blocked the importin α-mediated nuclear import of NF-κB and prevented lipopolysaccharide-induced pro-inflammatory gene expression, cell migration, and oxidative stress. In experimental atherosclerosis (apolipoprotein E-knockout mice fed a high-fat diet), i.p., 0.13 µmol/day NLS peptide administration for 5 weeks attenuated NF-κB activation in atherosclerotic plaques. NLS peptide significantly inhibited lesion development at both early (age 10 weeks) and advanced (age 28 weeks) stages of atherosclerosis in mice, without affecting serum lipid levels. Plaques from NLS-treated mice contained fewer macrophages of pro-inflammatory M1 subtype than those from respective untreated controls. By contrast, the relative smooth muscle cell and collagen content was increased, indicating a more stable plaque phenotype. NLS peptide also attenuated pro-inflammatory gene expression and oxidative stress in aortic lesions. Our study demonstrates that targeting NF-κB nuclear translocation hampers inflammation and atherosclerosis development and identifies cell-permeable NLS peptide as a potential anti-atherosclerotic agent.

Suppressors of cytokine signaling abrogate diabetic nephropathy.

Activation of Janus kinase/signal transducers and activators of transcription (JAK/STAT) is an important mechanism by which hyperglycemia contributes to renal damage, suggesting that modulation of this pathway may prevent renal and vascular complications of diabetes. Here, we investigated the involvement of suppressors of cytokine signaling (SOCS) as intracellular negative regulators of JAK/STAT activation in diabetic nephropathy. In a rat model, inducing diabetes resulted in JAK/STAT activation and increased expression of SOCS1 and SOCS3. In humans, we observed increased expression of glomerular and tubulointerstitial SOCS proteins in biopsies of patients with diabetic nephropathy. In vitro, high concentrations of glucose activated JAK/STAT/SOCS in human mesangial and tubular cells. Overexpression of SOCS reversed the glucose-induced activation of the JAK/STAT pathway, expression of STAT-dependent genes (chemokines, growth factors, and extracellular matrix proteins), and cell proliferation. In vivo, intrarenal delivery of adenovirus expressing SOCS1 and SOCS3 to diabetic rats significantly improved renal function and reduced renal lesions associated with diabetes, such as mesangial expansion, fibrosis, and influx of macrophages. SOCS gene delivery also decreased the activation of STAT1 and STAT3 and the expression of proinflammatory and profibrotic proteins in the diabetic kidney. In summary, these results provide direct evidence for a link between the JAK/STAT/SOCS axis and hyperglycemia-induced cell responses in the kidney. Suppression of the JAK/STAT pathway by increasing intracellular SOCS proteins may have therapeutic potential in diabetic nephropathy.

Suppressors of cytokine signaling modulate JAK/STAT-mediated cell responses during atherosclerosis.

OBJECTIVE: Suppressors of cytokine signaling (SOCS) proteins are intracellular regulators of receptor signal transduction, mainly Janus kinase/signal transducers and activators of transcription (JAK/STAT). We investigated the effects of SOCS modulation on the JAK/STAT-dependent responses in vascular cells, and their implication in atherosclerotic plaque development. METHODS AND RESULTS: Immunohistochemistry in human plaques revealed a high expression of SOCS1 and SOCS3 by vascular smooth muscle cells (VSMCs) and macrophages in the inflammatory region of the shoulders, when compared to the fibrous area. SOCS were also increased in aortic lesions from apoE(-/-) mice. In cultured VSMCs, endothelial cells, and monocytes, SOCS1 and SOCS3 were transiently induced by proinflammatory cytokines, proatherogenic lipoproteins, and immune molecules. Furthermore, overexpression of SOCS suppressed STAT activation and reduced inflammatory gene expression and cell growth, whereas SOCS knockdown increased these cell responses. In vivo, antisense oligodeoxynucleotides targeting SOCS3 exacerbated the atherosclerotic process in apoE(-/-) mice by increasing the size, leukocyte content, and chemokine expression in the lesions. CONCLUSIONS: SOCS expressed in atherosclerotic lesions are key regulators of vascular cell responses. Activation of this endogenous antiinflammatory pathway might be of interest in the treatment of atherosclerosis.