Ramipril can alleviate the accumulation of renal mesangial matrix in rats with diabetic nephropathy by inhibiting insulin-like growth factor-1.

PURPOSE: To investigate the impact of Ramipril (RAM) on the expressions of insulin-like growth factor-1 (IGF-1) and renal mesangial matrix (RMM) in rats with diabetic nephropathy (DN). METHODS: The Sprague Dawley rats were divided into normal control (NC) group (n = 12), DN group (n = 11), and DN+RAM group (n = 12). The ratio of renal weight to body weight (RBT), fasting blood glucose (FBG), HbA1c, 24-h urine protein (TPU), blood urea nitrogen (BUN), creatinine (Cr), renal pathological changes, the levels of IGF-1, fibronectin (FN), type IV collagen (Col-IV), and matrix metalloproteinases (MMP)-2 were compared among the groups. RESULTS: Compared with NC group, the RBT, FBG, HbA1c, TPU, BUN, Cr, and RMM in DN group were significantly increased (P < 0.05), the IGF-1, FN, and Col-IV were significantly upregulated (P < 0.05), while MMP was significantly downregulated (P < 0.05). Compared with DN group, the indexes except for the FBG and HbA1c in DN+RAM group were significantly improved (P < 0.05), among which IGF-1 exhibited significant positive correlation with TPU(r=0.937), FN(r=0.896) and Col-IV(r=0.871), while significant negative correlation with MMP-2 (r=-0.826) (P<0.05). CONCLUSION: RAM may protect the kidneys by suppressing IGF-1 and mitigating the accumulation of RMM.

Effects of PI and PIII Snake Venom Haemorrhagic Metalloproteinases on the Microvasculature: A Confocal Microscopy Study on the Mouse Cremaster Muscle.

The precise mechanisms by which Snake Venom Metalloproteinases (SVMPs) disrupt the microvasculature and cause haemorrhage have not been completely elucidated, and novel in vivo models are needed. In the present study, we compared the effects induced by BaP1, a PI SVMP isolated from Bothrops asper venom, and CsH1, a PIII SVMP from Crotalus simus venom, on cremaster muscle microvasculature by topical application of the toxins on isolated tissue (i.e., ex vivo model), and by intra-scrotal administration of the toxins (i.e., in vivo model). The whole tissue was fixed and immunostained to visualize the three components of blood vessels by confocal microscopy. In the ex vivo model, BaP1 was able to degrade type IV collagen and laminin from the BM of microvessels. Moreover, both SVMPs degraded type IV collagen from the BM in capillaries to a higher extent than in PCV and arterioles. CsH1 had a stronger effect on type IV collagen than BaP1. In the in vivo model, the effect of BaP1 on type IV collagen was widespread to the BM of arterioles and PCV. On the other hand, BaP1 was able to disrupt the endothelial barrier in PCV and to increase vascular permeability. Moreover, this toxin increased the size of gaps between pericytes in PCV and created new gaps between smooth muscle cells in arterioles in ex vivo conditions. These effects were not observed in the case of CsH1. In conclusion, our findings demonstrate that both SVMPs degrade type IV collagen from the BM in capillaries in vivo. Moreover, while the action of CsH1 is more directed to the BM of microvessels, the effects of BaP1 are widespread to other microvascular components. This study provides new insights in the mechanism of haemorrhage and other pathological effects induced by these toxins.

Resveratrol ameliorates diabetes-induced renal damage through regulating the expression of TGF-beta 1, collagen IV and Th17/Treg-related cytokines in rats / El resveratrol mejora el daño renal inducido por la diabetes a través de la regulación de la expresión de TGF-beta 1, colágeno IV y las citocinas relacionados con Th17/Treg en ratas

OBJECTIVES: This study aimed at detecting the protective effects of resveratrol on diabetes-induced renal damage and on the expression of transforming growth factor-beta 1 (TGF-β1), collagen IV and Th17/Tregrelated cytokines in streptozotocin-induced diabetic rats. METHODS: Twenty diabetic rats were further randomly divided into diabetic model group (DM group) and resveratrol group with 10 animals in each group. Another 10 non-diabetic rats served as control. The dia-betic rats in the resveratrol group were administered resveratrol for eight consecutive weeks (via gavage, 50 mg/kg daily, dissolved in saline). Rats in the control group and DM group received the same volume of saline only (via gavage). Renal function was measured. Histopathology changes of the kidney tissue were observed using haematoxylin and eosin staining. Levels of TGF-β1 and collagen IV in kidney homogenate were measured with enzyme-linked immunosorbent assay (ELISA). The level of Th17-related cytokines (IL-17A, IL-25) and Treg-related cytokines (IL-35, IL-10) in serum and in the supernatant of the kidney homogenate were determined using ELISA. RESULTS: Diabetic rats had damaged renal function, higher levels of TGF-β1, collagen IV, IL-17A and IL-25, as well as lower levels of IL-35 and IL-10, when compared to the control rats. Compared to the diabeticrats without resveratrol treatment, application of resveratrol to the diabetic rats ameliorated the renal function, inhibited the expression of TGF-β1, collagen IV, IL-17A and IL-25, and increased the expression IL-35 and IL-10. CONCLUSION: Resveratrol might ameliorate diabetes-induced renal damage through mediating the balance of Th17/Treg-related cytokines and inhibiting the expression of TGF-β1 and collagen IV.

OBJETIVOS: Este estudio estuvo encaminado a detectar los efectos protectores del resveratrol en el daño renal inducido por diabetes y en la expresión del factor de crecimiento transformante beta-1 (TGF-β1), el colágeno IV, y las citocinas relacionados con Th17/Treg en ratas con diabetes inducida por estreptozotocina. MÉTODOS: Veinte ratas diabéticas fueron divididas aleatoriamente en un grupo modelo diabético (Grupo MD) y un grupo de resveratrol, con 10 animales en cada grupo. A las ratas diabéticas en el grupo de resveratrol se les administró resveratrol durante ocho semanas consecutivas (mediante sonda nasogástrica, 50 mg/kg diarios, disuelto en suero salino). Las ratas en el grupo control y el grupo MD recibieron el mismo volumen de solución salina solamente (vía sonda nasogástrica). Se midió la función renal. Se observaron cambios en la histopatología del tejido del riñón usando tinción con hematoxilina y eo-sina. Se midieron los niveles de TGF-β1 y colágeno IV en un homogeneizado de riñón con ensayo por inmunoabsorción ligado a enzimas (ELISA). El nivel de las citocinas de Th17 (IL-17A, IL-25) y las citocinas de Treg (IL-35, IL-10) en suero y en el sobrenadante del homogeneizado de riñón, se determinaron mediante ELISA. RESULTADOS: Las ratas diabéticas tuvieron daño de la función renal, niveles más altos de TGF-β1, colágeno IV, IL-17A y IL-25, así como niveles más bajos de IL-35 e IL-10, en comparación con las ratas control. En comparación con las ratas diabéticas sin tratamiento con resveratrol, la aplicación de resveratrol en las ratas diabéticas mejoró la función renal, inhibió la expresión de TGF-β1, colágeno IV, IL-17A y IL-25 y aumentó la expresión de IL-35 y IL-10. CONCLUSIÓN: El resveratrol podría mitigar el daño renal inducido por la diabetes mediante la mediación con el equilibrio de las citocinas relacionados con Th17/Treg, e inhibiendo la expresión de TGF-β1 y colágeno IV.

Effects of cyclosporin, phenytoin, and nifedipine on the synthesis and degradation of gingival collagen in tufted capuchin monkeys (Cebus apella): histochemical and MMP-1 and -2 and collagen I gene expression analyses.

BACKGROUND: The purpose of this experimental study was to evaluate the collagen fiber distribution histologically after phenytoin, cyclosporin, or nifedipine therapy and to correlate it with collagen I and matrix metalloproteinase (MMP)-1 and -2 gene expression levels. METHODS: Gingival samples from the canine area were obtained from 12 male monkeys (Cebus apella). The mesial part of each sample was assessed by reverse transcription-polymerase chain reaction, whereas the distal part was processed histologically for picrosirius red and hematoxylin and eosin stainings, as well as for collagen IV immunostaining. One week after the first biopsy, the animals were assigned to three groups that received daily oral dosages of cyclosporin, phenytoin, or nifedipine for 120 days. Additional gingival samples were obtained on days 52 and 120 of treatment from two animals from each group on the opposite sides from the first biopsies. RESULTS: Picrosirius red staining showed a predominance of mature collagen fibers in the control group. Conversely, there was an enlargement of areas occupied by immature collagen fibers in all groups at days 52 and 120, which was not uniform over each section. There was a general trend to lower levels of MMP-1 gene expression on day 52 and increased levels on day 120. Phenytoin led to increased levels of MMP-2 and collagen I gene expression on day 120, whereas the opposite was observed in the nifedipine group. CONCLUSION: Cyclosporin, phenytoin, and nifedipine led to phased and drug-related gene expression patterns, resulting in impaired collagen metabolism, despite the lack of prominent clinical signs.

Sec61alpha synthesis is enhanced during translocation of nascent chains of collagen type IV in F9 teratocarcinoma cells after retinoic acid treatment.

Nascent procollagen peptides and other secretory proteins are transported across the endoplasmic reticulum (ER) membrane through a protein-conducting channel called translocon. Sec61alpha, a multispanning membrane translocon protein, has been implicated as being essential for translocation of polypeptide chains into the cisterns of the ER. Sec61alpha forms a protein complex with collagen and Hsp47, an ER-resident heat shock protein that binds specifically to collagen. However, it is not known whether Sec61alpha is ubiquitously produced in collagen-producing F9 teratocarcinoma cells or under heat shock treatment. Furthermore, the production and utilization of Sec61alpha may depend on the stage of cell differentiation. Cultured F9 teratocarcinoma cells are capable of differentiation in response to low concentrations of retinoic acid. This differentiation results in loss of tumorigenicity. Mouse F9 cells were grown in culture medium at 37 degrees C and 43 degrees C (heat shock treatment) treated or not with retinoic acid, and labeled in certain instances with 35S-methionine. Membrane-bound polysomes of procollagen IV were then isolated. Immunoprecipitation and Western blot analysis were performed using polyclonal antibodies against collagen IV, Hsp47 and Sec61alpha. Under retinoic acid-untreated conditions, F9 cells produced undetectable amounts of Sec61alpha. Sec61alpha, Hsp47 and type IV collagen levels were increased after retinoic acid treatment. Heat shock treatment did not alter Sec61alpha levels, suggesting that Sec61alpha production is probably not affected by heat shock. These data indicate that the enhanced production of Sec61alpha in retinoic acid-induced F9 teratocarcinoma cells parallels the increased synthesis of Hsp47 and collagen type IV.

Sec61alpha synthesis is enhanced during translocation of nascent chains of collagen type IV in F9 teratocarcinoma cells after retinoic acid treatment

Nascent procollagen peptides and other secretory proteins are transported across the endoplasmic reticulum (ER) membrane through a protein-conducting channel called translocon. Sec61alpha, a multispanning membrane translocon protein, has been implicated as being essential for translocation of polypeptide chains into the cisterns of the ER. Sec61alpha forms a protein complex with collagen and Hsp47, an ER-resident heat shock protein that binds specifically to collagen. However, it is not known whether Sec61alpha is ubiquitously produced in collagen-producing F9 teratocarcinoma cells or under heat shock treatment. Furthermore, the production and utilization of Sec61alpha may depend on the stage of cell differentiation. Cultured F9 teratocarcinoma cells are capable of differentiation in response to low concentrations of retinoic acid. This differentiation results in loss of tumorigenicity. Mouse F9 cells were grown in culture medium at 37ºC and 43ºC (heat shock treatment) treated or not with retinoic acid, and labeled in certain instances with 35S-methionine. Membrane-bound polysomes of procollagen IV were then isolated. Immunoprecipitation and Western blot analysis were performed using polyclonal antibodies against collagen IV, Hsp47 and Sec61alpha. Under retinoic acid-untreated conditions, F9 cells produced undetectable amounts of Sec61alpha. Sec61alpha, Hsp47 and type IV collagen levels were increased after retinoic acid treatment. Heat shock treatment did not alter Sec61alpha levels, suggesting that Sec61alpha production is probably not affected by heat shock. These data indicate that the enhanced production of Sec61alpha in retinoic acid-induced F9 teratocarcinoma cells parallels the increased synthesis of Hsp47 and collagen type IV

Immunohistochemical changes in kidney glomerular and tubular proteins caused by rattlesnake (Crotalus vegrandis) venom.

Renal damage is an important cause of death in patients who have survived the early effects of severe crotalid envenomation. Extracellular matrix of renal tissue is altered by Crotalus toxin activities. The aim of this study was to describe how cytoskeletal proteins and basal membrane components undergo substantial alterations under the action of Crotalus vegrandis crude venom and its hemorrhagic fraction (Uracoina-1) in mice. To detect the proteins in question, the immunoperoxidase method with monoclonal and polyclonal antibodies was used. Cell types within renal lesions were characterized by phenotypic identification, by means of immunohistologic analysis of marker proteins using different primary antibodies against mesangial cells, endothelial cells, cytoskeletal proteins (intermediate filament), extracellular matrix and basal membranes. Samples for morphological study by standard procedures (biotin-streptavidin-peroxidase technique) using light microscopy were processed. Positive and negative controls for each antigen tested in the staining assay were included. After crude venom and hemorrhagic fraction inoculation of mice, the disappearance of cytoskeletal vimentin and desmin and collagen proteins in the kidney was observed. In extracellular matrix and basal membranes, collagen type IV from envenomed animals tends to disappear from 24 h to 120 h after venom injection.