As activated hepatic stellate cells (aHSCs) play a central role in fibrogenesis, they have become key target cells for anti-fibrotic treatment. Nevertheless, the therapeutic efficiency is constrained by the exosomes they secrete, which are linked to energy metabolism and continuously stimulate the activation of neighboring quiescent hepatic stellate cells (qHSCs). Herein, an intercellular communication interference strategy is designed utilizing paeoniflorin (PF) loaded and hyaluronic acid (HA) coated copper-doped ZIF-8 (PF@HA-Cu/ZIF-8, PF@HCZ) to reduce energy-related exosome secretion from aHSCs, thus preserving neighboring qHSCs in a quiescent state. Simultaneously, the released copper and zinc ions disrupt key enzymes involved in glycolysis to reduce bioenergy synthesis in aHSCs, thereby promoting the reversion of aHSCs to a quiescent state and further decreasing exosome secretion. Therefore, PF@HCZ can effectively sustain both aHSCs and qHSCs in a metabolically dormant state to ultimately alleviate liver fibrosis. The study provides an enlightening strategy for interrupting exosome-mediated intercellular communication and remodeling the energy metabolic status of HSCs with boosted antifibrogenic activity.

Objective To investigate the influence mechanism of Cordyceps sinensis (CS) on β-glycerophosphate-induced vascular smooth muscle cell (VSMC) calcification.Methods The effect of CS on VSMC cell viability was detected by CCK-8.The cellular models of rat VSMC calcification were established by treating with β-glycerophosphate (β-GP,10 mmol/L);then CS (10 mg/L),autophagy inhibitor 3-methyladenine (3-MA,5 mmol/L),and AMPK inhibitor compound C (CC,10 μmol/L) were added to the cell cultures.There were a total of 5 experiment groups:VSMC cultured in normal medium (Control),VSMC treated with β-GP,VSMC treated with β-GP and CS,VSMC treated with 3-MA,β-GP and CS,and VSMC treated with CC,β-GP and CS.The calcium nodules and calcium content were examined with alizarin red S staining and the O-cresolphthaleincomplexone method,respectively.The autophagosomes within the VSMC were observed using transmission electron microscope (TEM).Immunofluorescence showed the accumulation of microtubule-associated protein 1 light chain 3 (LC3) puncta.In addition,levels of osteogenic related proteins,autophagy related proteins,and AMPK/mTOR pathway related proteins were evaluated by Western blotting.Results CS increased the number of autophagosomes and the accumulation of LC3 puncta within VSMC.It also upregulated the protein levels of LC3 Ⅱ/LC3 Ⅰ,beclin1,α-SMA,and p-AMPK;whereas,the protein levels of Runx2 and p-mTOR,as well as calcium nodules and calcium content were reduced (all P ＜ 0.01).When the cells were pretreated with 3-MA before treating with β-GP and CS,the autophagosomes,accumulation of LC3 puncta,and protein levels of LC3 Ⅱ/LC3 Ⅰ,beclinl,and α-SMA were decreased (all P ＜ 0.01);however,the protein level of Runx2,and the calcium nodules and calcium content were increased (all P ＜ 0.01).Nevertheless,when the cells were pretreated with CC before giving β-GP and CS,the autophagosomes,the accumulation of LC3 puncta,and the expression levels of p-AMPK,LC3 Ⅱ/LC3 Ⅰ,beclin1,and α-SMA were significantly down-regulated (all P ＜ 0.01);whereas,the expression levels of Runx2 and p-mTOR,as well as calcium nodules and calcium content were increased (all P ＜ 0.01).Conclusions CS can effectively alleviate β-GP-induced VSMC calcification,which may be due to the activation of autophagy by AMPK/mTOR signaling pathway.

Objective: To investigate the role and mechanism of Hydroxysafflor yellow A (HSYA) in the calcification of vascular smooth muscle cells (VSMC) induced by β-glycerol phosphate (β-GP). Methods: VSMC were cultured with 10% fetal bovine serum+1% double anti-high glucose DMEM medium at 37℃ and 5%CO₂ incubator, and were subcultured according to cell growth density at 1∶4 ratio. The cells were divided into three groups: control group (NC), high-phosphate-induced calcification (HP) group, and HSYA intervention (HSYA) group. The Calcium deposition amount was measured by alizarin red staining and calcium determination kit. The expressions of ALP, RUNX2, RANKL, α-SMA and inflammation indicators TLR4, TNF-α, IL-8 were detected by Western blotting method; Western blotting was also used to detect calcification index alkaline phosphatase (ALP) and Runt-related transcription factor 2 (RUNX2). Nuclear factor kappa B receptor activating factor ligand(RANKL), α-smooth muscle actin (α-SMA), and the expressions of TLR4/NF-κB pathway and inflammatory response-related indicators Toll-like receptor 4 (TLR4), interleukin-8 (IL-8) and tumor necrosis factor alpha (TNF-α). The nuclear protein and cytoplasmic proteins were respectively extracted. The expressions of p65 in nucleus and cytoplasm, as well as the expressions of p65 and phosphorylated p65 in total proteins were detected by Western blotting method. Superoxide dismutase (SOD) and malondialdehyde (MDA) kit were used to detect the content of antioxidant enzymes and oxidation end products in cells. Results: Western blotting showed that the expressions of ALP, RUNX2 and RANKL in HSYA group were significantly lower than that in HP group. The expression of α-SMA was increased than that of HP group (all P<0.01). The expression levels of TLR4, TNF-α, IL-8 and p-NF-κB/p65 in HSYA group were decreased compared with that in the HP group, and p65 was decreased in nucleus and increased in cytoplasm (all P<0.05). SOD content in HSYA group was significantly higher than that in HP group, and MDA content was significantly lower than that in HP group (all P<0.01). Conclusions HSYA can reduce calcification and calcium deposition of vascular smooth muscle cells induced by high phosphorus. The mechanism is related to the inhibition of the activation of TLR4/NF-κB pathway and the inhibition of oxidative stress.

Objective To investigate the possible mechanism of sclerostin/Lrp4 in calcification of VSMC induced by high phosphorus and the protective effect of Ginkgo biloba extract.Methods Aortic vascular smooth muscle cells (VSMCs) of SD rats were extracted and identified.VSMCs were divided into normal control group,high phosphorus induced calcification group (10 mmol/L β-glycerophosphate+50 μg/ml ascorbic acid),and high phosphorus induced calcification+Ginkgo biloba extract intervention group (10 mmol/L β-glycerophosphate+50 μg/ml ascorbic acid+0.5 mg/ml GBE),cultured in different mediums for 14 days.Vonkossa staining and alizarin red staining were used to detect the calcification of VSMCs.The mRNA level of BGP was detected by real time PCR,and the protein expressions of sclerostin and Lrp4 were detected by Western blot.Results Compared with normal control group,vonkossa staining and alizarin red staining showed significant calcium deposition in calcification group.Compared with calcification group,calcium salt deposition was significantly reduced in GBE treatment group.Real time PCR results showed β-catenin and BGP mRNA expressions in VSMC calcification group were higher than those in normal control group (P＜ 0.05).mRNA expressions of β-catenin and BGP in GBE treatment group were lower than those in calcification group (all P ＜ 0.05).Compared with normal control group,the protein expression of sclerostin was increased,but the protein expression of Lrp4 was decreased in calcified group (all P ＜ 0.05).Compared with calcification group,the protein expression of sclerostin decreased and the protein expression of Lrp4 increased in GBE treatment group (all P ＜ 0.05).Conclusions High phosphorus can induce VSMC calcification by activating Wn/β-catenin signaling pathway.Sclerostin/Lrp4 is involved in hyperphosphine-induced VSMC calcification.GBE can reduce the high phosphorus induced VSMC calcification by regulating the Wnt/β-catenin signaling pathway.

Objective To observe the protection and distribution of bone marrow mesenchymal stem cells (MSCs) by distinct intravenous infusion time on renal ischemia reperfusion injury (IRI) in rats.Methods We used unilateral nephrectomy and contralateral vascular occlusion method to establish renal IRI model in rats.The experimental groups which received 2 × 106 MSCs infusion through the tail vein,were subsequently divided into 3 subgroups:2 h pre-reperfusion (PreOp,n =16),immediately after reperfusion (Op,n =16),6 h post-reperfusion (PostOp,n - 16).The control groups included sham operation group (n =16) and ischemia group (n =16).Chemotaxis of DAPI-labeled MSCs was detected 6 h after administration in the IR kidney.Renal function was detected at 6,24,and 48 h respectively after operation. Forty eight h after operation,the renal tissues were harvested to observe the pathological changes by HE staining and the tubular epithelial cell apoptosis via TUNEL assay.Results MSCs were found in the experimental groups after IR in the kidney,most in PostOp group.Twenty-four and 48 h after reperfusion,there was no significant difference in Cr and BUN between the experimental groups and sham operation group (P＞0.05),but the levels of Cr and BUN in the experimental groups were significantly lower than in the IR group (P＜ 0.05). As compared with IR group,the renal pathological injury was alleviated,the number of apoptotic cells was decreased in the experimental group,most significantly in PostOp group (P＜0.05).Conclusion MSCs can reduce the inflammatory response and inhibit renal tubular cell apoptosis in rat renal IRI.Post-reperfusion administration of MSCs leads to the best chemotaxis efficiency and protection.

Objective To study the expression of EI-1 in myocardium during cerebral ischemia and reper-fusion, and to investigate the mechanism of cerebral cardiac syndrome. Method Two hundred and eight SD rats weighting 220～250 gram, were divided into three groups: sham control group (n=48), cerebral ischemia group (n=80), cerebral ischemia/reperfusion group (n=80). The area of cerebral ischemia, and the concentration of sennn ET-1 and CK-MB, and the content of myocardial ET-1 were determined in 0,6, 12,24,48,72 h after cerebral ischemia and reperfusion, and were analyzed by t-test or F-test. Results Cerebral necrosis area was ob-served at 6 h after cerebral ischemia in cerebral ischemia group, and peaked at 12 h (P>0.05). The concentra-tion of CK-MB increased gradually after cerebral ischmia, peaked at 12 h (P<0.05), and then gradually de-creased. The serum concentration of ET-1 peaked at 6 h and then gradually decreased. The content of ET-1 in my-ocardium began to increase at 6 h after cerebral ischemia, and peaked at 12 h (P<0.05). In cerebral ischemia/reperfusion group, all of cerebral necrosis size, CK-MB concentration and myocardial ET-1 concent paelced at 12 h and then gradually decreased (P<0.05). Change of ET-1 concentration in blood was similar to that in cerebral ischemia group. Compared with cerebral ischemia group, the size of cerebral necrosis reduced obviously at 24 h,48 h,72 h in cerebral ischemia/reperfusion group (P<0.05). The concentration of CK-MB in cerebral ischemia/ reperfusion group was higher than that in cerebral isehemia group (P<0.05). The peak time of myocardial ET-1 was shatened in cerebral ischemia/reperfusion group. The change of serum ET-1 was not different between two groups. Conclusions Large area of cerebral ischemia, might cause myocardial injury. ET-1 is involved in the course of myocardial injury following cerebral ischemia. Though cerebral reperfusion can protect brain,but it make myocardial injury more serious,and ET-1 might participate in this course.

Objective　To study β-AR density relativity between porcine myocardial tissue and circulating lymphocytes and clinical significance ．Methods　 48 porcines(seven months old），were　taken a few of myocardial tissue and peripheral blood．Assay　β-AR density with radioactive isotope to study the　relativity．Results　Scatter plot of β-AR　density　 circulating lymphocytes and myocardial tissue showed　 linear　Correlation（r＝0.845，P＜0.001　Y＝0.002527X＋0.726）．Conclusion　There　was　obvious　 correlation on β-AR　density in circulating lymphocytes and myocardial membrane　of pigs．Determination of β-AR　density　in　circulating lymphocytes can reflect alteration of that in myocardial tissue．Therefore，we can use it to monitor and investigate　self-induced and drug-induced β-AR changes．