Traumatic brain injury (TBI) is a significant global public health challenge, with an estimated mortality rate of 13 per 100,000 people in China. Bedsides the immediate neurological consequences,[1] TBI is associated with long-term sequelae and an elevated risk of neurodegenerative conditions such as dementia, Parkinson’s disease, and epilepsy. Moreover, TBI can induce systemic effects, including acute gastrointestinal injury (AGI), contributing to poor patient outcomes.[2]

Objective To investigate the effect of Angelica polysaccharide(APS)on the differentiation and function of M2 macrophages and underlying molecular mechanism.Methods Mouse bone marrow derived macrophages(BMDM)and M2 macrophages were induced and treated with APS(0,80,160,320 μg/mL);Mouse peritoneal macrophages were isolated and treated with APS(0,160 μg/mL).Flow cytometry(FCM)was used to detect mannose receptor(MR),CD11b,F4/80,CD163,and ARG-1 expression levels,apoptosis,and phagocytic ability of M2 macrophages and peritoneal macrophages.Mice were randomly divided into APS gavage group and control group,APS was intragastrically administered to mice,and macrophage MR expression level in blood and spleen were detected by FCM.Fluorescence microscopy was used to observe the morphology of BMDM-differentiated M2 macrophages.RT-qPCR was employed to detect the mRNA expression levels of MR and ARG-1 in M2 macrophages.Immunofluorescence assay was performed to detect the expression of the proteins related to molecular mechanism of differentiation and function of M2 macrophages.Results Compared with the 0 μg/mL APS group,the MR expression level in the M2 macrophages was decreased with the increase of APS concentration within a certain concentration range(80～320 μg/mL),and the MR expression level in peritoneal macrophages was also decreased in the 160 μg/mL APS treatment group(P＜0.01).The expression level of macrophage MR was also significantly decreased in peripheral blood and spleen in the APS gavage mice than the control group(P＜0.05).Compared with the 0 μg/mL APS group,the expression levels of CD11b,F4/80,and CD163 in the macrophages were increased in the 80～320 μg/mL APS treatment groups(P＜0.01).The morphology of macrophage had changed,from mostly spindle-shaped and pseudopodia to mostly round or irregular,and even a few cells with pseudopodia.APS induced apoptosis in M2 macrophages(P＜0.05).Compared with the 0 μg/mL APS group,M2 macrophages treated with 160 μg/mL APS had an increased ability to phagocytose fluorescent microspheres(P＜0.01),but the expression level of ARG-1 was decreased(P＜0.01).The mRNA expression of MR and ARG-1 in M2 macrophages was decreased(P＜0.05).The mean fluorescence intensity of phosphate acidified-signal transducers and activators of transcription 6(p-STAT6)-positive signals in M2 macrophages was significantly reduced in the 160 μg/mL APS-treated group(P＜0.05).Conclusion APS has bidirectional regulation on the differentiation and function of M2 macrophages,which may be associated with its downregulation of signal transducers and activators of transcription 6(STAT6)signaling pathway.

Objective To investigate the action of chondrogenesis differentiation of bone marrow stromal cells (BMSCs) transfected with adeno-hTGF-β1. Methods In the experiment group, replication-deficient a denoviruses carrying human hTGF-β1 complementary DNA (adeno-hTGF-β1 was constructed and applied to transfect to the first generation BMSCs. As a control, each BMSCs was transduced with 200 pfu of adeno-LacZ gene. One day after transfer, BMSCs were trypsinized, counted, and 5×105 cells aliuots were spun down at 500 rpm per minute in 15 ml polypropylene conical tubes and then cultured in a defined medium in an incubator at 37℃ for 21 days. The aggregates were harvested at time points to 21 days and assessed by gross observation, histological analyses and immunohistochemical localization of type Ⅱ collagen. Results When harvested at 21 days, each pellet shrinked to spheroid tissue with apearly opalescence in gross morphology and found to be relatively firm. H.E staining showed elongate dlining cells appeared as perichon drium-like cells at the surface. Some nests of cartilage were observed at the substrate of the tissue. Mature chon drocytes were embeded in the lacuna in the experiment group. In addition, Safranin'O staining confirmed the presence of sulfated proteoglycans in the ECM of chondrogenesis region. Immunohistochemical staining revealed the presence of type Ⅱ collagen in chondrogenesis region. By contrast, HE staining showed no evidence of cartilage formation in the control group. They were fibrous tissue with no architectural feature. Safranin'O staining and Immunohistochemical staining showed no evidence of sulfated proteoglycans or typeⅡ collagen expression. Conclusion BMSCs transfected with adeno-hTGF-β1 could induce its chondro-genesis when aggregate cultured in a defined medium in vitro, laying a foundation for the application of hTGFβ1 gene-transfected BMSCs in cartilage tissue engineering.