OBJECTIVE:To investigate the protective effect of NDUFA13 protein against acute liver injury and liver fibrosis in mice and explore the possible mechanisms.METHODS:BALB/C mice (7 to 8 weeks old) were divided into normal group, CCl4 group, CCl4+AAV-NC group and CCl4+AAV-NDU13 group (n=18). Mouse models of liver fibrosis were established by intraperitoneal injection of CCl4 twice a week for 3, 5 or 7 weeks, and the recombinant virus AAV8-TBG-NC or AAV8-TBG-NDUFA13 was injected via the tail vein 7-10 days prior to CCl4 injection. After the treatments, pathological changes in the liver of the mice were observed using HE and Masson staining. Hepatic expression levels of NDUFA13 and α-SMA were detected with Western blotting, and the coexpression of NDUFA13 and NLRP3, TNF-α and IL-1β, and α-SMA and collagen Ⅲ was analyzed with immunofluorescence assay.RESULTS:HE and Masson staining showed deranged liver architecture, necrotic hepatocytes and obvious inflammatory infiltration and collagen fiber deposition in mice with CCl4 injection (P < 0.001). NDUFA13 expression markedly decreased in CCl4-treated mice (P < 0.001), while a significant reduction in inflammatory aggregation and fibrosis was observed in mice with AAV-mediated NDUFA13 overexpression (P < 0.001). In CCl4+AAV-NDU13 group, immunofluorescence assay revealed markedly weakened activation of NLRP3 inflammasomes (P < 0.001), significantly decreased TNF-α and IL-1β secretion (P < 0.001), and inhibited hepatic stellate cell activation (P < 0.05) and collagen formation in the liver (P < 0.001).CONCLUSION:Mitochondrial NDUFA13 overexpression in hepatocytes protects against CCl4- induced liver fibrosis in mice by inhibiting activation of NLRP3 signaling.

Abstract Mage-D1 (MAGE family member D1) is involved in a variety of cell biological effects. Recent studies have shown that Mage-D1 is closely related to tooth development, but its specific regulatory mechanism is unclear. The purpose of this study was to investigate the expression pattern of Mage-D1 in rat tooth germ development and its differential mineralization ability to ectomesenchymal stem cells (EMSCs), and to explore its potential mechanism. Results showed that the expression of Mage-D1 during rat tooth germ development was temporally and spatially specific. Mage-D1 promotes the proliferation ability of EMSCs but inhibits their migration ability. Under induction by mineralized culture medium, Mage-D1 promotes osteogenesis and tooth-forming ability. In vitro, Mage-D1 not only binds to p75 neurotrophin receptor (p75NTR) but also to distal-less homeobox 1(Dlx1) and msh homeobox 1 (Msx1). These findings indicate that Mage-D1 is absolutely important in tooth germ development. p75NTR, Dlx1, and Msx1 seem to be closely related to the underlying mechanism of Mage-D1 action.

目的 探讨不同强度的低强度脉冲超声(LIPUS)对永生化根尖牙乳头干细胞(ISCAP)分化及增殖的影响.方法 采用CCK8细胞增殖计数法探究不同强度LIPUS对ISCAP细胞增殖的影响;通过碱性磷酸酶(ALP)、茜素红染色及Western Blot探究ISCAP的成骨/成牙本质分化,通过实时荧光定量聚合酶链反应技术(RT-q-PCR)检测不同强度LIPUS辐照ISCAP后ALP、抗runt转录因子2(RUNX2)、成牙本质基质蛋白1(DMP-1)、血管内皮生长因子(VEGF)的信使核糖核酸(mRNA)表达量探究细胞的成骨/成牙本质/成血管分化的定量分析.结果 60 mW/cm2强度的超声刺激LIPUS后ISCAP增殖速率最大,不同强度的LIPUS对其成牙本质/成骨方向的分化有促进作用,并且在一定范围内其促进分化的能力与强度呈正相关;LIPUS对ISCAP成血管方向的分化也有促进作用,但与强度无明显关联,其促分化能力呈现出时间依赖性趋势.结论 LIPUS作为非侵入性治疗方式具有促进牙源性干细胞的增殖及分化的潜能,可为牙髓再生提供治疗思路.

Mage-D1 (MAGE family member D1) is involved in a variety of cell biological effects. Recent studies have shown that Mage-D1 is closely related to tooth development, but its specific regulatory mechanism is unclear. The purpose of this study was to investigate the expression pattern of Mage-D1 in rat dental germ development and its differential mineralization ability to ectomesenchymal stem cells (EMSCs), and to explore its potential mechanism. Results showed that the expression of Mage-D1 during rat dental germ development was temporally and spatially specific. Mage-D1 promotes the proliferation ability of EMSCs but inhibits their migration ability. Under induction by mineralized culture medium, Mage-D1 promotes osteogenesis and tooth-forming ability. Furthermore, the expression pattern of Mage-D1 at E19.5 d rat dental germ is similar to p75 neurotrophin receptor (p75NTR), distal-less homeobox 1 (Dlx1) and msh homeobox 1 (Msx1). In addition, Mage-D1 is binding to p75NTR, Dlx1, and Msx1 in vitro. These findings indicate that Mage-D1 is play an important regulatory role in normal mineralization of teeth. p75NTR, Dlx1, and Msx1 seem to be closely related to the underlying mechanism of Mage-D1 action.

不可逆性牙髓炎是口腔科常见疾病,临床针对该类患者主要采用根管治疗术,包括根管预备、根管消毒、根管充填3个阶段,其中根管充填是最主要的阶段.其虽可起到一定的治疗效果,但去除牙髓后,牙体组织会变脆、变薄,长时间后会出现患牙折断现象,患者甚至面临拔牙风险.因此,寻求一种既能有效去除牙髓感染,又能保留或重建牙髓功能的方法具有重要意义.近年来,生物组织工程牙髓血管再生、牙髓再血管化在牙髓再生中具有广泛的应用前景,为牙髓再生提供了更多的可能性.本文就以上两种方法作一概述,以期为临床牙髓再生方法的选择提供些许参考.

再生医学在缺损或受损组织器官的修复中具有重要作用.组织工程是再生医学的重要手段,其中干细胞发挥着重要的作用.干细胞根据其来源可分为胚胎干细胞(ESCs)、成体干细胞(ASCs)和诱导多能干细胞(iPSCs).目前,研究发现外胚层间充质干细胞(EMSCs)是一种有良好自我更新和多向分化潜能的干细胞群,并且近年来越来越受到临床关注.多项研究已经证明,EMSCs在组织再生中起到关键作用,尤其体现在颅颌面复杂的发生发育过程中.本文就EMSCs的起源、生物学特性、多向分化潜能及在牙组织再生中的应用做一总结与探讨.

OBJECTIVE:To detect the binding of Mage-D1 with activated p75NTR and explore their role in regulating mineralization of ectomesenchymal stem cells (EMSCs).METHODS:EMSCs were isolated from the tooth germs of embryonic SD rats (19.5 days of gestation) by tissue explant culture and were identified for surface markers using flow cytometry. The cultured cells were divided into blank control group, 100 ng/mL nerve growth factor (NGF) stimulation group, and lentivirus-mediated Mage-D1 interference (SH-Mage-D1) group. Proximity ligation assay was used to detect the binding of Mage-D1 with activated p75NTR in the EMSCs, and the binding strength was compared among the 3 groups. Alizarin red staining and ALP staining were used to observe mineralization of the induced cells. The expressions of ALP, Runx2, OCN, BSP, OPN, Msx1 and Dlx1 at both the mRNA and protein levels were detected using RT-PCR and Western blotting.RESULTS:The isolated EMSCs expressed high levels of cell surface markers CD44, CD90, CD29, CD146, and CD105 with a low expression of CD45. The results of proximity ligation assay showed that the binding of Mage-D1 with activated p75NTR in the cells increased over time, and the binding strength was significantly greater in NFG-treated cells than in the cells in the other two groups (P < 0.05). Alizarin red staining and ALP staining of the induced cells showed that the changes in the mineralization nodules were consistent with those of ALP activity. The cells treated with 100 ng/mL NGF exhibited significantly increased expressions of ALP, Runx2, OCN, BSP, OPN, Col1, Msx1 and Dlx1 as compared with the cells in the other two groups (P < 0.05).CONCLUSION:Mage-D1 directly binds to activated p75NTR in embryonic rat EMSCs to positively regulate the mineralization of the EMSCs.

Human periodontal ligament stem cells (hPDLSCs) are a promising source in regenerative medicine. Due to the complexity and heterogeneity of hPDLSCs, it is critical to isolate homogeneous hPDLSCs with high regenerative potential. In this study, p75 neurotrophin receptor (p75NTR) was used to isolate p75NTR(+) and p75NTR(-) hPDLSCs by fluorescence-activated cell sorting. Differences in osteogenic differentiation among p75NTR(+), p75NTR(-) and unsorted hPDLSCs were observed. Differential gene expression profiles between p75NTR(+) and p75NTR(-) hPDLSCs were analysed by RNA sequencing. alpha 1 Integrin (ITGA1) small interfering RNA and ITGA1-overexpressing adenovirus were used to transfect p75NTR(+) and p75NTR(-) hPDLSCs. The results showed that p75NTR(+) hPDLSCs demonstrated superior osteogenic capacity than p75NTR(-) and unsorted hPDLSCs. Differentially expressed genes between p75NTR(+) and p75NTR(-) hPDLSCs were highly involved in the extracellular matrix-receptor interaction signalling pathway, and p75NTR(+) hPDLSCs expressed higher ITGA1 levels than p75NTR(-) hPDLSCs. ITGA1 silencing inhibited the osteogenic differentiation of p75NTR(+) hPDLSCs, while ITGA1 overexpression enhanced the osteogenic differentiation of p75NTR(-) hPDLSCs. These findings indicate that p75NTR optimizes the osteogenic potential of hPDLSCs by up-regulating ITGA1 expression, suggesting that p75NTR can be used as a novel cell surface marker to identify and purify hPDLSCs to promote their applications in regenerative medicine.

Augmenter of liver regeneration (ALR) is a thermostable cytokine that was originally identified to promote the growth of hepatocytes. This study was conducted to explore the expression and function of ALR in multiple myeloma (MM), a common hematologic malignancy. Real-time PCR and western blot analysis were performed to detect the expression of ALR in U266 human MM cells and healthy peripheral blood mononuclear cells (PBMCs). U266 MM cells were exposed to 20 or 40 μg/mL of recombinant ALR and tested for cell proliferation. Small interfering RNA-mediated silencing of ALR was done to investigate the role of ALR in cell proliferation, apoptosis, and cytokine production. Compared to PBMCs, U266 MM cells exhibited significantly higher levels of ALR at both the mRNA and protein levels. The addition of recombinant ALR protein significantly promoted the proliferation of U266 cells. In contrast, knockdown of ALR led to a significant decline in the viability and proliferation of U266 cells. Annexin-V/PI staining analysis demonstrated that ALR downregulation increased apoptosis in U266 MM cells, compared to control cells (20.1±1.1 vs 9.1±0.3%, P<0.05). Moreover, ALR depletion reduced the Bcl-2 mRNA level by 40% and raised the Bax mRNA level by 2-fold. Additionally, conditioned medium from ALR-depleted U266 cells had significantly lower concentrations of interleukin-6 than control cells (P<0.05). Taken together, ALR contributed to the proliferation and survival of U266 MM cells, and targeting ALR may have therapeutic potential in the treatment of MM.