BackgroundRecently, we reported that toll-like receptor (TLR)2 and TLR4 localized on the glomerular endothelium in the glomeruli of streptozotocin (STZ)-induced type 1 diabetic mice and high fat diet feed-induced type 2 diabetic mice, and that periodontal pathogen Porphyromonas gingivalis LPS (Pg-LPS) administration lowered the survival rate of diabetic mice. The present study aims to examine the effect of TLR4 blocking on the suppression of Pg-LPS-induced diabetic nephropathy.MethodsThe survival rate and morphological/biochemical features for streptozotocin-induced diabetic mice with Pg-LPS and TLR4 blocker eritoran administration were investigated by reporter gene assay, urine and blood analysis, immunohistochemistry, and real time-PCR.Results and ConclusionsAll of the diabetic mice administered Pg-LPS were euthanized within the survival period of almost all of the diabetic mice. The blood urea nitrogen and creatinine, expression of TLR2 and TGF-b, and type 1 collagen accumulation, in the diabetic mice increased significantly with the Pg-LPS administration. In spite of the limited TLR4 activation with Pg-LPS, the TLR4 blocker eritoran decreased blood urea nitrogen and creatinine, and raised the survival rate of the Pg-LPS-administered diabetic mice slightly. The high expression levels of TLR2, TGF-b, and type 1 collagen in Pg-LPS-administered diabetic mice decreased with eritoran. Nuclear STAT3 which enhances TLR2 expression was detected in the TLR2-expressing glomeruli of diabetic mice. The TLR2 and STAT3 gene expression increased by the Pg-LPS administration but decreased with eritoran. These may suggest that Pg-LPS-induced diabetic nephropathy is mainly dependent on TLR2 signaling on glomerular endothelial cells, and that TLR4 blocker eritoran may play a role to slow the progress of diabetic nephropathy.

Podoplanin is a mucin-type highly O-glycosylated glycoprotein identified in several somatyic cells: podocytes, alveolar epithelial cells, lymphatic endothelial cells, lymph node stromal fibroblastic reticular cells, osteocytes, odontoblasts, mesothelial cells, glia cells, and others. It has been reported that podoplanin-RhoA interaction induces cytoskeleton relaxation and cell process stretching in fibroblastic cells and osteocytes, and that podoplanin plays a critical role in type I alveolar cell differentiation. It appears that podoplanin plays a number of different roles in contributing to cell functioning and growth by signaling. However, little is known about the functions of podoplanin in the somatic cells of the adult organism because an absence of podoplanin is lethal at birth by the respiratory failure. In this report, we investigated the tooth germ development in podoplanin-knockout mice, and the dentin formation in podoplanin-conditional knockout mice having neural crest-derived cells with deficiency in podoplanin by the Wnt1 promoter and enhancer-driven Cre recombinase: Wnt1-Cre;PdpnΔ/Δmice. In the Wnt1-Cre;PdpnΔ/Δmice, the tooth and alveolar bone showed no morphological abnormalities and grow normally, indicating that podoplanin is not critical in the development of the tooth and bone.

Podoplanin is the only counter-receptor of platelet CLEC-2 and is expressing on mature osteoblast, but there is no report on the role of podoplanin and CLEC-2 in calcification. This study aimed to investigate the role of podoplanin binding to CLEC-2 in the calcification of osteoblasts carrying homozygously deleted Pdpn alleles (Pdpn Δ/Δ ) by heterozygously expressing collagen type I alpha 1 promoter (Col1a)-driven Cre recombinase. There were no macroscopic abnormalities in the bone and dentin of Col1a11-Cre;Pdpn Δ/Δ mice but the coccygeal bone medullary cavity was very narrow. In the quantitative analysis for alizarin red-stained products and alkaline phosphatase activities on the cultured calvarial osteoblasts, the amounts of calcified products and alkaline phosphatase activity of calvarial osteoblasts of both Pdpn fl/fl and Col1a11-Cre;Pdpn Δ/Δ mice were significantly higher in the calcification medium than in the α-mem. Both the amounts of calcified products and alkaline phosphatase activity of calvarial osteoblasts from Pdpn fl/fl mice were significantly lower in the calcification medium with CLEC-2 than without CLEC-2 while there were no significant differences in the amounts of calcified products and alkaline phosphatase activities of calvarial osteoblasts from Col1a11-Cre;Pdpn Δ/Δ mice with CLEC-2. Platelet CLEC-2 may play a role in regulating the calcification via binding to podoplanin on mature osteoblasts expressing podoplanin in the medullary cavity of a part of the bone.