Research on Runx2 gene induced differentiation of human amniotic mesenchymal stem cells into ligament fibroblasts in vitro and promotion of tendon-bone healing in rabbits / 中国修复重建外科杂志

OBJECTIVE@#To investigate whether the Runx2 gene can induce the differentiation of human amniotic mesenchymal stem cells (hAMSCs) to ligament fibroblasts in vitro and promote the tendon-bone healing in rabbits.@*METHODS@#hAMSCs were isolated from the placentas voluntarily donated from healthy parturients and passaged, and then identified by flow cytometric identification. Adenoviral vectors carrying Runx2 gene (Ad-Runx2) and empty vector adenovirus (Ad-NC) were constructed and viral titer assay; then, the 3rd generation hAMSCs were transfected with Ad-Runx2 (Ad-Runx2 group) or Ad-NC (Ad-NC group). The real-time fluorescence quantitative PCR and Western blot were used to detect Runx2 gene and protein expression to verify the effectiveness of Ad-Runx2 transfection of hAMSCs; and at 3 and 7 days after transfection, real-time fluorescence quantitative PCR was further used to detect the expressions of ligament fibroblast-related genes [vascular endothelial growth factor (VEGF), collagen type Ⅰ, Fibronectin, and Tenascin-C]. The hAMSCs were used as a blank control group. The hAMSCs, hAMSCs transfected with Ad-NC, and hAMSCs were mixed with Matrigel according to the ratio of 1 : 1 and 1 : 2 to construct the cell-scaffold compound. Cell proliferation was detected by cell counting kit 8 (CCK-8) assay, and the corresponding cell-scaffold compound with better proliferation were taken for subsequent animal experiments. Twelve New Zealand white rabbits were randomly divided into 4 groups of sham operation group (Sham group), anterior cruciate ligament reconstruction group (ACLR group), anterior cruciate ligament reconstruction+hAMSCs transfected with Ad-NC-scaffold compound group (Ad-NC group), and anterior cruciate ligament reconstruction+hAMSCs transfected with Ad-Runx2-scaffold compound group (Ad-Runx2 group), with 3 rabbits in each group. After preparing the ACL reconstruction model, the Ad-NC group and the Ad-Runx2 group injected the optimal hAMSCs-Matrigel compunds into the bone channel correspondingly. The samples were taken for gross, histological (HE staining and sirius red staining), and immunofluorescence staining observation at 1 month after operation to evaluate the inflammatory cell infiltration as well as collagen and Tenascin-C content in the ligament tissues.@*RESULTS@#Flow cytometric identification of the isolated cells conformed to the phenotypic characteristics of MSCs. The Runx2 gene was successfully transfected into hAMSCs. Compared with the Ad-NC group, the relative expressions of VEGF and collagen type Ⅰ genes in the Ad-Runx2 group significantly increased at 3 and 7 days after transfection ( P<0.05), Fibronectin significantly increased at 3 days ( P<0.05), and Tenascin-C significantly increased at 3 days and decreased at 7 days ( P<0.05). CCK-8 detection showed that there was no significant difference ( P>0.05) in the cell proliferation between groups and between different time points after mixed culture of two ratios. So the cell-scaffold compound constructed in the ratio of 1∶1 was selected for subsequent experiments. Animal experiments showed that at 1 month after operation, the continuity of the grafted tendon was complete in all groups; HE staining showed that the tissue repair in the Ad-Runx2 group was better and there were fewer inflammatory cells when compared with the ACLR group and the Ad-NC group; sirius red staining and immunofluorescence staining showed that the Ad-Runx2 group had more collagen typeⅠ and Ⅲ fibers, tending to form a normal ACL structure. However, the fluorescence intensity of Tenascin-C protein was weakening when compared to the ACLR and Ad-NC groups.@*CONCLUSION@#Runx2 gene transfection of hAMSCs induces directed differentiation to ligament fibroblasts and promotes tendon-bone healing in reconstructed anterior cruciate ligament in rabbits.

Diagnostic Efficiency of T2WI and DWI in LI-RADS Classification with Cirrhosis Caused by Hepatitis B / 中国医学影像学杂志

Purpose To investigate the value of T2WI mild-moderate signal and restricted diffusion in the context of liver imaging reporting and data system (LI-RADS) (2014 edition) in the diagnosis of hepatocellular carcinoma (HCC) with cirrhosis caused by hepatitis B virus.Materials and Methods A total of 77 lesions (LI-RADS 3-5,size of 1.1 cm×0.7 cm-12.7 cm×9.1 cm) of 69 HCC patients in Beijing Friendship Hospital from January 2012 to November 2016 were retrospectively analyzed.All these patients underwent MRI scan and multiphase dynamic enhanced scan.The images were analyzed by two radiologists.If a disagreement occurred,liver accelerated volume acquisition and multiphase dynamic enhanced scan were combined to reach a consensus.The contrast noise ratio (CNR) and apparent diffusion coefficient (ADC) of T2WI and diffusion weighted imaging (DWI) sequences were compared,as well as the identification of the two signs.Results There was no statistically significant difference between T2WI mild-moderate signal and restricted diffusion in the identification of lesions (LI-RADS 3-5) (P＞0.05),while the sensitivity with DWI b=0 (61.0％) was significantly lower than DWI b=600 s/mm2 (70.1％) (P＜0.05).The CNR of all DWI sequences (b=0,600 s/mm2) were larger than those of T2WI (P＜0.01).The ADC of small lesions (diameter ＜2 cm) were larger than those of larger lesions (diameter ＞2 cm) [(1.57+0.37)×10-3 mm2/s vs.(1.37+0.51)×10 3 mm2/s,P＜0.05].Conclusion There is no significant difference in sensitivity of lesions between T2WI mild-moderate signal and restricted diffusion.However,due to different CNRs,DWI with b=600 s/mm2 is more obvious for the lesions,and can be first investigated in practice.