Ociated with decreasing levels of phosphorylated Smad-5. Transfection of those cells with gremlin siRNA plasmid resulted in substantially improved levels of phosphorylated Smad-5, whereas, there was no significant improve of BMP7 level soon after trasfection of gremlin siRNA plasmid. Taken together, our in vivo and in vitro information, as well because the functional research relating to BMP-7 and gremlin reported in the literature, help a model in which the important mechanism of therapeutic action of gremlin inhibition on DN is connected to the recovery of BMP-7 activity. Firstly, BMP-7 is involved in ameliorating renal damage as a result of mesangial proliferation by suppression of mesangial cell IL-1 Proteins MedChemExpress apoptoGremlin and Diabetic KidneyPLoS 1 www.plosone.orgGremlin and Diabetic KidneyFigure 3. Cell proliferation and apoptosis in diabetic mouse kidneys. (A) Detection of proliferating cell nuclear antigen (PCNA) by immunoperoxidase staining, within the kidneys of non-diabetic control mice (N), streptozotocin-induced diabetic mice treated with pBAsi mU6 Neo handle plasmid (STZ) or pBAsi mU6 Neo gremlin siRNA plasmid (Gremlin siRNA). (B and C) PCNA good cells in kidneys in the STZ group dramatically improve at week-1 and -2, and pBAsi mU6 Neo gremlin siRNA plasmid remedy drastically reduces PCNA optimistic cells both in glomeruli and tubules. Proliferating cells are barely seen in all three groups at week 12. (D) Co-immunostaining of diabetic kidney sections with antibodies against PCNA and Gremlin. Intensive Gremlin expression is often seen inside the cells with PCNA constructive signal. (E, F) In situ TUNEL assay. Apoptotic cells are observed predominantly in tubules inside the STZ group at week-12. The amount of apoptotic cells is significantly lowered by pBAsi mU6 Neo gremlin siRNA plasmid treatment. ( p,0.01 vs. non-diabetic control group, # p,0.01 vs. STZ group). Scale bars, 100 mm (A, B and E), and 10 mm (D). N = six mice per group. doi:10.1371/journal.pone.0011709.gsis. Accumulating proof suggests that early renal hypertrophy, partially resulting from cell proliferation, acts as a pacemaker for subsequent irreversible structural modifications, like glomerulosclerosis and tubulointerstitial fibrosis[31]. Secondly, maintenance of BMP-7 activity by inhibition of Gremlin expression could result in blockade of extracellular matrix (ECM) accumulation. It was reported that BMP-7 could reduce TGF-b-induced ECM protein accumulation in cultured mesangial cells by sustaining the levels and activity of MMP2, partially by means of prevention of TGF-bdependent upregulation of PAI-1[31,32,33]. Our data showed that treatment with gremlin siRNA plasmid resulted within a considerable reduction in mesangial areas and accumulation of collagen variety IV in diabetic mice, plus the reduced matrix metalloprotease (MMP-2) level in mesangial cells cultured beneath HG conditions was enhanced by transfection with gremlin siRNA plasmid. A precise query needs to be addressed whether or not Gremlin has BMP-7-independent effects on the pathogenesis of diabetic nephropathy. As shown in Figure 3D, the proliferative activity of mesangial cells is associated using the expression degree of Gremlin. It.
