[Comparative study on the degradation rate and regulatory effects of two resorbable collagen membranes during the in vivo implantation].

Objective: To explore the differences in the performance and tissue repair promotion effects of small intestinal submucosa membrane (SIS membrane) and Bio-Gide resorbable collagen membrane (Bio-Gide membrane) by performing the subcutaneous implantation models in mice. Methods: For in vivo studies, we stablished membrane implantation models using 6-8 week-old male C57BL/6 mice. The degradation rates were explored through HE staining analysis at different time points (1, 3, 5, 7, 14 and 28 d, 3 mice/group/time point). The influences of the two membranes on local macrophages and neovasculum were evaluated by immunofluorescence detection of F4/80 and CD31, and the mobilization effects of the two membranes on local stem cells were evaluated by immunohistochemical detection of Ki67 and CD146. For in vitro studies, mice periodontal ligament stem cells (mPDLSCs) were co-cultured with these two membrane materials, and the cell morphologies were observed by scanning electron microscopy. In addition, the gene expressions of Ki67, Cxcl1, Ccl1, Tnfa were investigated by real-time fluorescence quantitative PCR (RT-qPCR). Results: The results of in vivo studies showed that by day 28, there was no significant difference in degradation rate between these two membrane materials [SIS degradation rate: (16.84±4.00) %, Bio-Gide degradation rate: (24.07±3.97) %, P=0.090], illustrating that both of them could maintain the barrier effects for more than one month. In addition, there was no significant difference in the infiltration number of local F4/80 positive macrophages between these two groups by the day 3 after implantation [SIS: (20.67±5.69) cells/visual field, Bio-Gide: (25.33±2.52) cells/visual field, P=0.292]. However, compared with the Bio-Gide membrane, SIS membrane significantly promoted local CD31+vascular regeneration [SIS: (4.67±1.15) cells/visual field, Bio-Gide: (1.00±1.00) cells/visual field, P=0.015] and CD146+stem cell recruitment [SIS: (22.33±4.16) cells/visual field, Bio-Gide: (11.33±2.52) cells/visual field, P=0.025]. The RT-qPCR results also showed that SIS membrane promoted the gene expression of Cxcl1 (SIS vs Bio-Gide P<0.001) in mPDLSCs, but had no effect on the gene expression of Tnfa (SIS vs Bio-Gide P=0.885). Conclusions: SIS membrane showed a similar degradation rate compared with Bio-Gide membrane, and there was no significant difference in the effects of these two membranes on local inflammation or macrophages. Therefore, both of these membranes could meet the barrier effects required by guided tissue regeneration.

Oridonin alleviates the inhibitory effect of lipopolysaccharide on the proliferation and osteogenic potential of periodontal ligament stem cells by inhibiting endoplasmic reticulum stress and NF-κB/NLRP3 inflammasome signaling.

BACKGROUND: The aim of this study was to investigate the protective effect and mechanism of oridonin in an in vitro lipopolysaccharide (LPS)-induced human periodontal ligament stem cells (hPDLSCs) model of periodontitis. METHODS: Primary hPDLSCs were isolated and cultured, and then the expression of surface antigens CD146, STRO-1 and CD45 of hPDLSCs was detected by flow cytometry. The mRNA expression level of Runx2, OPN, Col-1, GRP78, CHOP, ATF4 and ATF6 in the cells was tested by qRT-PCR. MTT was taken to determine the cytotoxicity of oridonin at different concentrations (0-4 µM) on hPDLSCs. Besides, ALP staining, alizarin red staining and Oil Red O staining were utilized to assess the osteogenic differentiation (ALP concentration, mineralized calcium nodule formation) and adipogenic differentiation abilities of the cells. The proinflammatory factors level in the cells was measured by ELISA. The protein expression level of NF-κB/NLRP3 pathway-related proteins and endoplasmic reticulum (ER) stress-related markers in the cells were detected by Western blot. RESULTS: hPDLSCs with positive CD146 and STRO-1 expression and negative CD45 expression were successfully isolated in this study. 0.1-2 µM of oridonin had no significant cytotoxicity on the growth of hPDLSCs, while 2 µM of oridonin could not only greatly reduce the inhibitory effect of LPS on the proliferation and osteogenic differentiation of hPDLSCs cells, but also inhibit LPS-induced inflammation and ER stress in hPDLSCs cells. Moreover, further mechanism research showed that 2 µM of oridonin suppressed NF-κB/NLRP3 signaling pathway activity in LPS-induced hPDLSCs cells. CONCLUSIONS: Oridonin promotes proliferation and osteogenic differentiation of LPS-induced hPDLSCs in an inflammatory environment, possibly by inhibiting ER stress and NF-κB/NLRP3 pathway. Oridonin may have a potential role in the repair and regeneration of hPDLSCs.

Effects of Shendibushen on the Expression of CD146 and its Metabolic Pathways.

Context: Evidence from multiple studies has revealed that it's meaningful to evaluate the clinical significance of CD146 because it's related to an early diagnosis of chronic renal failure as well as to the severity of illness and the patient's prognosis. Objective: The current study intended to evaluate the therapeutic effects of Shendibushen on the clinical parameters of blood and urine and on fibrosis in the kidney in a rat model, using simulated renal tissue fibrosis that was surgically induced with unilateral ureteral obstruction (UUO). Also, our research team intended to analyze the metabolic pathway activated by Shendibushen both in rat and human kidneys through use of the Kyoto Encyclopedia of Genes and Genomes (KEGG) database and the GeneNetwork. The aim is to discover if a connection existed between CD146 and key genes in these pathways. Design: The research team conducted an animal study in Wistar rats. Intervention: The rats were divided into 5 groups of 14 animals each: (1) blank control group, (2) sham control group, (3) model group, (4) Niaoduqing group, and (5) Shendibushen group. Three groups had UUO surgically induced-the model, Niaoduqing, and Shendibushen groups. The sham control group received sham surgery, and the blank control group received no surgery. The Shendibushen and Niaoduqing groups received the relevant capsules once a day at a fixed time, for a total of 28 days. Outcome Measures: The levels of serum creatinine, blood urea nitrogen, microalbuminuria, serum soluble CD146, and urinary soluble CD146 were measured on the 14th and 28th days after modeling the rats. The degree of renal interstitial fibrosis was examined by hematoxylin and eosin (HE) staining and Masson trichrome staining. The changes at transcriptome level were obtained by target tissue sequencing. The KEGG database was used to analyze the potential pathway activated by the Shendibushen treatment. The GeneNetwork analysis was used to validate the correlation and identify the connections between CD146 and the key genes of the potential pathways. Results: Shendibushen capsules decreased the degree of renal interstitial fibrosis in the UUO rat model and reduced the serum creatinine, blood urea nitrogen, microalbumin, serum sCD146, and urinary sCD146 significantly compared to the model group (P < .05). Upon analysis of the metabolic pathways activated by Shendibushen, the study further verified, through use of the KEGG database, that CD146 activated the nuclear factor kappa B1 (NF-κB1) and transforming growth factor beta 1 (TGF-ß1)/ SMAD family member 2 (SMAD2) pathways. Conclusions: CD146 could become an early indicator in clinical monitoring. CD146 has a function related to the NF-κB1and TGF-ß1/ SMAD2 pathways under Shendibushen treatment.

exMCAM-Fc, an S100A8/A9-mediated-metastasis blocker, efficiently reduced the number of circulating tumor cells that appeared in the blood flow.

Metastasis is the major cause of treatment failure in cancer patients and of cancer-associated death so that therapeutic regulation of metastasis is very important subject for the cancer treatment. We have been reported that S100A8/A9, a heterodimer complex of S100A8 and S100A9, and its receptors play a crucial role in the lung tropic cancer metastasis, i.e., S100A8/A9 is actively secreted from the lung when cancer mass exists even at remote area from the lung and then functions to attract the distant cancer cells to the lung since cancer cells own the S100A8/A9 receptor(s) on their cell surface. Interestingly, one of the newly developed decoys, exMCAM-Fc, a Fc fusion protein with the extracellular region of melanoma cell adhesion molecule (MCAM), one of the S100A8/A9 receptors, that could prevent the interaction of S100A8/A9 with MCAM, efficiently suppressed the lung tropic cancer metastasis through exerting the several inhibitory effects on the S100A8/A9-mediated cancer cell events including enhanced mobility, invasion and attachment to the endothelial cells. However, it still remains to clarify if the decoy will reduce the number of circulating tumor cells (CTCs) that are defined as substantial cells in the context of organ tropic cancer metastasis. Here, we first show that exMCAM-Fc effectively reduces the number of CTCs in the blood flow of the melanoma bearing mice. The novel finding reinforces the suppressive role of exMCAM-Fc on the cancer metastasis. We therefore expect that exMCAM-Fc may greatly contribute to reduce treatment failure by the efficient blocking of the life threatening cancer metastasis.

Increased endothelial microparticles in obese and overweight children.

BACKGROUND: Obesity in children increases the risk of atherosclerosis. Endothelial dysfunction is an important factor in the pathogenesis of atherosclerosis, and endothelial microparticles (EMPs) are considered as markers of endothelial dysfunction. In this study, we aimed to evaluate circulating EMPs in obese and overweight children and to disclose the measure of obesity with the strongest relation with circulating microparticles and carotid atherosclerosis. METHODS: This prospective study included 55 obese and overweight children and 23 healthy controls. Insulin resistance was studied. Both in vivo and in vitro human umbilical vein endothelial cell evaluations were used for the study. Circulating EMPs (CD144 and CD146) were measured by flow cytometry. The carotid artery intima-media thickness (cIMT) and left ventricular mass index (LVMI) were measured using ultrasound and echocardiography, respectively. Study groups were compared for anthropometric measurement, insulin resistance, circulating EMP, cIMT, and LVMI. The relationship among overweight, obesity, and circulating EMPs were investigated. RESULTS: Blood pressure, CD144+EMP levels, and LVMI were statistically higher in the patients group than in the control group. The multiple logistic regression analysis and the backward elimination method showed that CD144+EMP and systolic blood pressure had a linear relationship with overweight and obesity. CONCLUSION: Our results suggest that endothelial damage starts in the early stage of childhood obesity and that obese and overweight children have increased circulating CD144+EMPs, showing that endothelial dysfunction and increased CD144+EMPs may be related to obesity.