Impacts of andrographolide on sciatic nerve function damage in diabetic peripheral neuropathy rats by regulating HMGB1/RAGE signaling pathway / 中国药房

OBJECTIVE To investigate the impacts of andrographolide on sciatic nerve function injury in diabetic peripheral neuropathy （DPN） rats by regulating high-mobility group protein box 1 （HMGB1）/receptor for advanced glycation end products （RAGE） signal pathway. METHODS A total of 84 rats were randomly divided into the control group （normal saline）， DPN group （normal saline）， low-dose andrographolide group （0.833 mg/kg）， high-dose andrographolide group （3.332 mg/kg）， lipoic acid group （positive control， 0.1 g/kg）， recombinant rat HMGB1 protein （rHMGB1） group （8 μg/kg）， and high-dose andrographolide+ rHMGB1 group， with 12 rats in each group. All rats except those in the control group were fed with high glucose and high fat diet combined with intraperitoneal injections of streptozotocin to establish the DPN rat model. After 24 hours of successful modeling， medication was administered daily for 8 weeks. The changes in fasting blood glucose， mechanical pain threshold， heat pain threshold and sciatic nerve conduction velocity were detected. Pathological changes in the sciatic nerve of rats and the activity of superoxide dismutase （SOD） and the content of malondialdehyde （MDA） in the sciatic nerve of rats were also detected. Besides， the expressions of HMGB1， RAGE proteins and phosphorylation level of nuclear factor κB p65（NF-κB p65） protein in rat sciatic nerves were found. RESULTS Compared with the control group， the pathological damage of the sciatic nerve of rats in the DPN group was strengthened， the fasting blood glucose， heat pain threshold， MDA content and the 诊治。E-mail：dqiaur@163.com expressions of HMGB1， RAGE proteins and phosphorylation level of NF-κB p65 protein were increased （P＜0.05）， while the mechanical pain threshold， sensory nerve conduction velocity， motor nerve conduction velocity， and SOD activity were decreased/slowed down （P＜0.05）. Compared with the DPN group， the above indexes were significantly potentiated in the andrographolide low- and high-dose groups and lipoic acid group （P＜0.05）， and the corresponding trends in the rHMGB1 group were opposite to those in the above three administration groups （P＜0.05）. Moreover， rHMGB1 attenuated the hypoglycemic effect of high-dose andrographolide on blood glucose and the improvement of oxidative stress injury in the sciatic nerve of DPN rats （P＜0.05）. CONCLUSIONS Andrographolide may reduce blood glucose by inhibiting the HMGB1/RAGE pathway and oxidative stress， thus ameliorating sciatic nerve injury in DPN rats.

Improvement effect and mechanism of “Layers adjusting external application”paste on synovial fibrosis in rats with knee osteoarthritis / 中国药房

OBJECTIVE To investigate the improvement effect and potential mechanism of “Layers adjusting external application” paste on synovial fibrosis （SF） in rats with knee osteoarthritis （KOA）. METHODS Male SD rats were randomly divided into sham operation group， KOA group and Layers adjusting external application group， with 8 rats in each group. KOA model was induced by the anterior cruciate ligament disruption method in KOA group and Layers adjusting external application group. Fourteen days after modeling， the Layers adjusting external application group was given “Layers adjusting external application” paste [Sanse powder （8 g for every 100 cm2）， Compound sanhuang ointment （5 g for every 100 cm2）] on the knee joint， 8 h every day， for 28 d in total. After the last administration， the degree of synovitis and fibrosis in rats was observed， and Krenn scoring was performed in each group. The expressions of collagen Ⅰ， high mobility group protein B1 （HMGB1） and phosphorylated nuclear factor-κB p65 （p-NF-κB p65） were detected in the synovial membrane； the contents of interleukin-1β （IL- 1β）， IL-6 and tumor necrosis factor-α （TNF-α） in serum as well as the expressions of fibrosis-related and HMGB1/Toll-like receptor 4 （TLR4）/NF-κB signaling pathway-related proteins and mRNA were detected in synovial tissue. RESULTS Compared with the sham operation group， the synovial lining cells in the KOA group showed significant proliferation and disordered arrangement， the inflammatory cell infiltration and collagen fiber deposition were obvious； the positive expressing cells of collagen Ⅰ， HMGB1 and p-NF-κB p65 were increased significantly； the contents of IL-1β， IL-6 and TNF-α in serum， the expressions of fibrosis-related protein （transforming growth factor-β， collagen Ⅰ， tissue inhibitor of metalloproteinase 1， α-smooth muscle actin） and their mRNA as well as theexpressions of HMGB1， TLR4 protein and their mRNA， the expressions of p-NF-κB p65 protein and NF-κB p65 mRNA were all increased significantly in synovial tissues of rats （P＜0.01）. Compared with the KOA group， the pathological changes in the synovial tissue of rats in Layers adjusting external application group were significantly improved， and the above quantitative indicators were significantly reversed （P＜0.05 or P＜0.01）. CONCLUSIONS “Layers adjusting external application” paste could significantly improve SF in KOA rats， the mechanism of which may be associated with the inhibition of the activation of HMGB1/ TLR4/NF-κB signaling pathway.

Mechanism of resveratrol regulating ovarian cancer cell apoptosis by inhibiting HMGB1 / 中华内分泌外科杂志

Objective:To investigate the effect of resveratrol on apoptosis of ovarian cancer cells and its molecular mechanism, and to find a potential new target for the treatment of ovarian cancer.Methods:Ovarian cancer SKOV-3 cells were divided into control group and resveratrol group.The survival rate of SKOV-3 cells treated with resveratrol was measured by MTT assay. 24 h after resveratrol intervention in SKOV-3 cells, Western blot was used to detect the expression of Bcl-2, Bax, Caspase-3, HMGB1, TLR4 and NF-?B. Ovarian cancer SKOV-3 cells were transfected with si-NC, si-HMGB1, Rb1+pcDNA3.1 or Rb1+pcDNA3.1-HMGB1, and the sensitivity of cells to resveratrol was detected by MTT assay. The transfected cells were treated with resveratrol and apoptosis was detected by flow cytometry.Results:Resveratrol could inhibit the growth of ovarian cancer SKOV-3 cells, and the higher the concentration of resveratrol, the more significant the inhibitory effect. The expression level of HMGB1 in control cells and resveratrol group was 1.24±0.15 and 0.86±0.11, respectively. 25μM resveratrol inhibited HMGB1 protein level ( P<0.01) . The sensitivity to resveratrol was increased after HMGB1 was downregulated, and the apoptosis of SKOV-3 cells was promoted. HMGB1 overexpression increased the resistance to resveratrol and inhibited the apoptosis of SKOV-3 cells. TLR4 expression quantity control and resveratrol cells were 0.98±0.12 and 0.63±0.08, amount of NF-?B expression were 1.21±0.14 and 0.45±0.07, 25 μM resveratrol could cut TLR4 and NF-?B protein levels. Conclusions:Resveratrol can promote apoptosis of ovarian cancer SKOV-3 cells. This mechanism may be related to the down-regulation of HMGB1/TLR4/NF-?B, which provides a basis for resveratrol treatment of ovarian cancer.

MiR-340 mediates the involvement of high mobility group box 1 in the pathogenesis of liver fibrosis / 中华肝脏病杂志

Objective: To explore the pathogenic mechanism of the miR-340/high mobility group box 1 (HMGB1) axis in the formation of liver fibrosis. Methods: A rat liver fibrosis model was established by injecting CCl(4) intraperitoneally. miRNAs targeting and validating HMGB1 were selected with gene microarrays after screening the differentially expressed miRNAs in rats with normal and hepatic fibrosis. The effect of miRNA expressional changes on HMGB1 levels was detected by qPCR. Dual luciferase gene reporter assays (LUC) was used to verify the targeting relationship between miR-340 and HMGB1. The proliferative activity of the hepatic stellate cell line HSC-T6 was detected by thiazolyl blue tetrazolium bromide (MTT) assay after co-transfection of miRNA mimics and HMGB1 overexpression vector, and the expression of extracellular matrix (ECM) proteins type I collagen and α-smooth muscle actin (SMA) was detected by western blot. Statistical analysis was performed by analysis of variance and the LSD-t test. Results: Hematoxylin-eosin and Masson staining results showed that the rat model of liver fibrosis was successfully established. Gene microarray analysis and bioinformatics prediction had detected eight miRNAs possibly targeting HMGB1, and animal model validation had detected miR-340. qPCR detection results showed that miR-340 had inhibited the expression of HMGB1, and a luciferase complementation assay suggested that miR-340 had targeted HMGB1. Functional experiments results showed that HMGB1 overexpression had enhanced cell proliferation activity and the expression of type I collagen and α-SMA, while miR-340 mimics had not only inhibited cell proliferation activity and the expression of HMGB1, type I collagen, and α-SMA, but also partially reversed the promoting effect of HMGB1 on cell proliferation and ECM synthesis. Conclusion: miR-340 targets HMGB1 to inhibit the proliferation and ECM deposition in hepatic stellate cells and plays a protective role during the process of liver fibrosis.

Mechanism of high mobility group protein B1 in lipopolysaccharide-induced acute lung injury/acute respiratory distress syndrome / 中华危重病急救医学

Objective:To investigate the role and possible pathogenesis of high mobility group protein B1 (HMGB1) in lipopolysaccharide (LPS)-induced acute lung injury/acute respiratory distress syndrome (ALI/ARDS).Methods:① In vivo, 24 SPFC57BL/6 male mice were randomly divided into normal control group, ALI/ARDS model group, ethyl pyruvate (EP) treatment group and EP control group, with 6 mice in each group. The ALI/ARDS model was established by intraperitoneal injection of 20 mg/kg LPS. Mice in normal control group and EP control group were intraperitoneally injected with the same amount of sterile normal saline. Then, mice in the EP treatment group and EP control group were intraperitoneally injected with 40 mg/kg HMGB1 inhibitor EP. After 6 hours, the mice were sacrificed and lung tissues were collected. The expressions of heparan sulfate (HS), syndecans-1 (SDC-1), heparanase (HPA) and matrix metalloproteinases-9 (MMP-9) in lung tissues were detected by immunofluorescence technique. Orbital blood of mice was collected and serum was extracted to detect the content of HMGB1 by enzyme linked immunosorbent assay (ELISA). ② In vitro, human umbilical vein endothelial cells (HUVECs) were randomly divided into 6 groups: normal control group, HUVECs damage group (treated with 1 mg/L LPS for 6 hours), HMGB1 group (treated with 1 μmol/L recombinant HMGB1 for 6 hours), HMGB1+EP group (treated with recombinant HMGB1 for 1 hour and then added 1 μmol/L EP for 6 hours), LPS+EP group (treated with LPS for 1 hour and then added 1 μmol/L EP for 6 hours), EP group (treated with 1 μmol/L EP for 6 hours). The expressions of HS, SDC-1, HPA and MMP-9 in endothelial cells were detected by immunofluorescence technique. Results:① In vivo, light microscopy showed that the alveolar space was thickened after LPS stimulation, and there were a large number of inflammatory cells infiltrating in the alveolar space. Compared with ALI/ARDS model group, the expressions of HS and SDC-1 in lung tissue of EP treatment group were significantly increased [HS (fluorescence intensity): 0.80±0.20 vs. 0.53±0.02, SDC-1 (fluorescence intensity): 0.72±0.02 vs. 0.51±0.01, both P < 0.05], and the expressions of HPA and MMP-9 were significantly decreased [HPA (fluorescence intensity): 2.36±0.05 vs. 3.00±0.04, MMP-9 (fluorescence intensity): 2.55±0.13 vs. 3.26±0.05, both P < 0.05]; there were no significant changes of the above indexes in EP control group. Compared with ALI/ARDS model group, the content of serum HMGB1 in EP treatment group decreased significantly (μg/L: 131.88±16.67 vs. 341.13±22.47, P < 0.05); there was no significant change in the EP control group. ② In vitro, compared with HMGB1 group, the expressions of HS and SDC-1 in HMGB1+EP group were significantly higher [HS (fluorescence intensity): 0.83±0.07 vs. 0.56±0.03, SDC-1 (fluorescence intensity): 0.80±0.01 vs. 0.61±0.01, both P < 0.05], and the expressions of HPA and MMP-9 were significantly lower [HPA (fluorescence intensity): 1.30±0.02 vs. 2.29±0.05, MMP-9 (fluorescence intensity): 1.55±0.04 vs. 2.50±0.06, both P < 0.05]; the expression of HS, SDC-1, HPA and MMP-9 had no significant changes in EP group. Conclusion:HMGB1 participates in LPS-induced injury of endothelial cell glycocalyx, leading to increased lung permeability, and inhibition of HMGB1 can alleviate lung injury.

The Value of Serum HMGB1 and sRAGE in the Diagnosis, Efficacy Monitoring and Prognosis of Newly Diagnosed Multiple Myeloma / 中国实验血液学杂志

OBJECTIVE@#To evaluate the value of high mobility group protein B1 (HMGB1) and soluble receptor for advanced glycation end products (sRAGE) in the diagnosis, efficacy monitoring and prognosis of newly diagnosed multiple myeloma (MM) patients.@*METHODS@#Fifty newly diagnosed MM patients before and after chemotherapy and 50 hematological outpatients from October 2018 to May 2020 were selected. Enzyme linked immunosorbent assay (ELISA) was used to detect the serum HMGB1 and sRAGE levels of the patients. ROC was used to further analyze the efficacy of serum HMGB1 and sRAGE levels on the diagnosis of MM. At the same time, the serum levels of HMGB1 and sRAGE before and after chemotherapy were compared, and their values in the evaluation of curative effect of MM patients were analyzed. According to the mean values of serum HMGB1 and sRAGE, all the patients were divided into different groups, the clinical characteristics and survival status of the patients were compared.@*RESULTS@#Before treatment the serum HMGB1 level of the patients in MM group was higher than that in control group, while sRAGE level was lower (t=11.363,6.127, P<0.001). The AUC of serum HMGB1 and sRAGE in the MM patients was 0.955 and 0.811, respectively. After 3 courses of chemotherapy, HMGB1 level of the patients in CR group was lower than before chemotherapy, while in PD group was higher, as well as sRAGE level of the patients in PR group (P<0.05). There were significant differences in R-ISS stage, HGB, CRP, ESR, CD56, CD117, D13S319 deletion between HMGB1 high expression group and HMGB1 low expression group (χ2=3.920, 6.522, 6.65, 4.16, 3.945, 6.65, 4.16, P<0.05), while there were significant differences in ISS stage, CRP and CD56 between sRAGE low expression group (28 cases) and sRAGE high expression group (22 cases) (χ2=4.565, 4.711, 5.547, P<0.05). Kaplan-Meier survival analysis showed that the patients in HMGB1 low expression group had better survival condition, for PFS Tlow>Thigh (χ2=9.470, P<0.05), and for OS Tlow>Thigh (χ2=7.808, P<0.05); there was no difference in the survival of sRAGE high expression group and low expression group, for PFS Tlow<Thigh (χ2=1.661, P>0.05), and for OS Tlow<Thigh (χ2=2.048, P>0.05). Cox analysis showed that LDH and HMGB1 were the factors affecting the prognosis of the patients, and both of them affected PFS (HR=2.771, 95% CI: 1.002-7.662, P=0.049; HR=6.022, 95% CI: 1.689-21.470, P=0.006), while HMGB1 also affected OS (HR=4.275, 95% CI: 1.183-15.451, P=0.027).@*CONCLUSION@#The serum HMGB1 and sRAGE have certain auxiliary value for the diagnosis and curative effect monitoring of newly diagnosed MM patients, and serum HMGB1 is expected to be an auxiliary detection index for the prognosis of MM.

Dihydromyricetin improves cardiac insufficiency by inhibiting HMGB1 in diabetic rats / 南方医科大学学报

OBJECTIVE@#To investigate the effect of dihydromyricetin (DHM) on cardiac insufficiency in diabetic rats and explore the underlying mechanism.@*METHOD@#Twenty-four male SD rats were randomized equally into normal control group, type 2 diabetes (T2DM) group fed on a high-glucose and high-fat diet for 6 weeks with low-dose streptozotocin (STZ) injection, metformin (MET) group with daily intragastric administration of MET (150 mg/kg) for 8 weeks after T2DM modeling, and dihydromyricetin (DHM) group with daily intragastric administration of DHM (250 mg/kg) for 8 weeks after modeling. The levels of fasting blood glucose, low density lipoprotein (LDL-C), triglyceride (TG), total cholesterol (TC), high density lipoprotein (HDL-C) and glycosylated hemoglobin (HbA1c) of the rats were measured, and plasma levels of insulin and high mobility group protein-1 (HMGB1) were detected with ELISA. The cardiac function of the rats was assessed using color echocardiography, ECG was measured using a biological signal acquisition system, and myocardial pathology was observed with HE staining. The protein expressions of HMGB1, nuclear factor-κB (NF-κB) p65 and phospho-NF-κB p65 (p-NF-κB p65) in the myocardial tissue were detected using Western blotting.@*RESULTS@#Compared with the control group, the rats in T2DM group showed significant anomalies in cardiac function after modeling with significantly increased plasma HMGB1 level and expressions of HMGB1, NF-κB p65 and p-NF-κB p65 proteins in the myocardial tissue (P < 0.05 or 0.01). Treatment with DHM significantly improved the indexes of cardiac function of the diabetic rats (P < 0.05 or 0.01), decreased plasma HMGB1 level and down-regulated the protein expressions of HMGB1 and p-NF-κB p65 in the myocardial tissue (P < 0.05 or 0.01).@*CONCLUSION@#DHM treatment can improve cardiac function in diabetic rats possibly by down-regulation of HMGB1 and phospho-NF-κB p65 expressions in the myocardium.

Effect of Compound Phylanthus urinaria Ⅱ in Regulating Expression of lncRNA CCAT1 in Hepatocellular Carcinoma HepG2 Cells / 中国实验方剂学杂志

Objective:To explore the anti-hepatoma effect of compound <italic>Phylanthus urinaria</italic> Ⅱ （ CPU Ⅱ） by inhibiting the expression of the long non-coding RNA （lncRNA） colon cancer associated transcript-1 （CCAT1） and restoring the expression of microRNA let-7a. Method:Real-time fluorescence quantitative polymerase chain reaction （PCR） was used to detect the expression of lncRNA CCAT1 in normal liver cells （LO2 cells） and hepatocellular carcinoma HepG2 cells， and the differences in expression between these two types of cells were compared. The methylthiazolyl tetrazolium（MTT） assay was used to detect the proliferation of HepG2 cells after treatment with different concentrations of CPU Ⅱ and 5-fluorouracil（5-FU） for 24， 48 and 72 h. Hepatocellular carcinoma HepG2 cells were cultured <italic>in vitro </italic>and set into three gropes： cell control group， CPU Ⅱ low-dose group （0.8 g·L^-1） and high-dose group （1.6 g·L^-1）. Real-time PCR was used to detect the mRNA expression of lncRNA CCAT1， microRNA let-7a and its target genes high mobility group protein A2（HMGA2）， and N-RAS in each grope. Western blot was used to detect the protein expression of HMGA2， and Cyclin D₁ in each grope. Result:As compared with LO2 cells， expression of lncRNA CCAT1 in HepG2 cells was significantly up-regulated （<italic>P</italic><0.05）. Results of MTT assay showed that the 50% inhibiting concentration（IC₅₀）of CPU Ⅱ and 5-FU on hepatocellular carcinoma HepG2 cells was 1.649， 0.044 648 g·L^-1respectively. As compared with the control group， CPU Ⅱ high-and low-dose groups （1.6， 0.8 g·L^-1） significantly inhibited the proliferation of HepG2 cells （<italic>P</italic><0.05）， and the effect was most remarkable in CPU Ⅱ high-dose group （<italic>P</italic><0.05）. The results of Real-time PCR showed that as compared with control group， the expression of lncRNA CCAT1 mRNA was significantly inhibited in CPU Ⅱ high-and low-dose groups （<italic>P</italic><0.05）， and the expression of microRNA let-7a mRNA was obviously up-regulated in high-dose group （<italic>P</italic><0.05）， but the expression of HMGA2 mRNA in CPU Ⅱ high-and low-dose groups as well as the expression of N-RAS mRNA in CPU Ⅱ low-dose group were down-regulated （<italic>P</italic><0.05）. Western blot results showed that as compared with the cell control group， the protein expression of HMGA2 and Cyclin D₁ in CPU Ⅱ high-and low-dose groups （1.6， 0.8 g·L^-1） was significantly down-regulated （<italic>P</italic><0.05）. Conclusion:CPU Ⅱ can inhibit the expression of lncRNA CCAT1， recover the expression of microRNA let-7a， and suppress the mRNA and protein expression of related downstream target genes in hepatoma cells line HepG2， thereby inhibiting the proliferation of hepatocellular carcinoma cells and exerting anti-hepatocellular carcinoma effect.

Research progress on release mechanism of high mobility group protein B1 in acute respiratory distress syndrome / 中华危重病急救医学

High mobility group protein B1 (HMGB1), a highly conversed non-histone nucleoproteins with strong pro-inflammatory property, is one of the inflammatory mediator of the acute respiratory distress syndrome (ARDS). Numerous studies have confirmed that HMGB1 regulates ARDS by binding to receptor for advanced glycation end product (RAGE), Toll-like receptor (TLR) and etc. And it can significantly increase the mortality of ARDS. But the mechanism of HMGB1 release is still unclear. This study focuses on the HMGB1 release progress, which connected with Janus kinases/signal transducer and activator of transcription (JAK/STAT), nuclear factor-κB (NF-κB), Notch, inflammasome, tumor necrosis factor (TNF), mitogen-activated protein kinase (MAPK), reactive oxygen species (ROS), peroxisome proliferator-activated receptor (PPAR) and other signaling or dependent pathways in ARDS.

Effects of silencing HMGB1 combined with docetaxel chemotherapy on the proliferation and apoptosis of prostate cancer cells and its action mechanism / 中华男科学杂志

Objective@#To investigate the effect of silencing the high-mobility group box-1 protein (HMGB1) combined with docetaxel (DTX) on the proliferation and apoptosis of PCa cells and its possible action mechanism.@*METHODS@#The expression of HMGB1 mRNA in different PCa cell lines and normal prostatic epithelial cells was detected by RT-qPCR. The PC-3 cells were transfected with different HMGB1 small interfering RNAs (si-HMGB1, si-HMGB1-2 and si-HMGB1-3), and the silencing effect was detected. The effects of different concentrations of DTX on the proliferation of the PC-3 cells was determined by MTT. Then the PC-3 cells were randomly divided into five groups: control (conventional culture), si-HMGB1-NC (si-HMGB1-NC transfection), si-HMGB1 (si-HMGB1-3 transfection), DTX (20 nmol/L DTX), and si-HMGB1+DTX (si-HMGB1-3+20 nmol/L DTX transfection), followed by measurement of the survival rate of the cells by MTT, their apoptosis rate by flow cytometry, and the expressions of HMGB1, B-cell lymphoma-2 (Bcl-2) and Bcl-2-associated X (Bax) proteins in different groups by Western blot.@*RESULTS@#The expression of HMGB1 mRNA in the PC-3 cells was the highest and the lowest after transfection with si-HMGB1-3. DTX inhibited the proliferation of the PC-3 cells at various concentrations. Compared with the control group, the si-HMGB1 and DTX groups showed significantly decreased A values, cell survival rates and HMGB1 and Bcl-2 expressions, but increased cell apoptosis rates and Bax expressions (P < 0.05). In comparison with the si-HMGB1 and DTX groups, the si-HMGB1+DTX group exhibited a remarkably decreased A value, cell survival rate and Bcl-2 expression, but increased cell apoptosis and Bax expression. The expression of the HMGB1 protein was markedly lower in the si-HMGB1+DTX than in the DTX group (P < 0.05).@*CONCLUSIONS@#Silencing HMGB1 combined with DTX chemotherapy can inhibit the proliferation and promote the apoptosis of PCa cells, which may be attributed to its regulatory effect on the expressions of the Bcl-2 family-related proteins.、.

Inflammatory mechanism of chlorpyrifos-induced lung injury in rats / 中国职业医学

OBJECTIVE: To investigate the inflammatory mechanism of chlorpyrifos-induced lung injury in rats. METHODS: Specific pathogen free SD rats were randomly divided into control and low-, medium-and high-dose groups, 8 rats in each group. Rats were exposed to 48% chlorpyrifos by continuous oral administration for 28 consecutive days, once a day, with the doses of 0.0, 4.1, 8.2 and 16.3 mg/kg body mass. After the exposure, the enzyme-linked immunosorbent assay was used to detect the level of tumor necrosis factor-α(TNF-α) and interleukin-1(IL-1) in rat lung tissue. The expression of high mobility group protein-1(HMGB1) and nuclear transcription factor-κB(NF-κB) was detected by immunohistochemistry and Western blotting, respectively. RESULTS: The activity of the rats decreased after exposure in the low-, medium-and high-dose groups compared with the control group. The rats in the medium and high dose groups increased salivation, nasal secretions, and difficulty breathing. The rats in the high dose group also developed diarrhea, muscle tremor, unstable gait, as well as muscarinic and nicotinic symptoms. After the exposure, the lung tissues of rats in the low-, medium-and high-dose groups showed different degrees of inflammation, which increased in a dose-dependent manner; the body mass of rats in the 3 dose groups was lower than that in the control group(all P values were <0.05), and the body mass of rats decreased with the increase of chlorpyrifos dose(all P values were <0.05). The levels of TNF-α, IL-1 and the relative expression of HMGB1, NF-κB were higher in the 3 dose groups than those in the control group(all P values were <0.05), and the levels of TNF-α, IL-1 and the relative expression of HMGB1 increased with the increasing exposure dose(all P values were <0.05). CONCLUSION: Chlorpyrifos can activate the NF-κB signaling pathway and eventually induce the macrophages to secrete large amount of TNF-α and IL-1, resulting in lung inflammation and injury in rats. The effect of chlorpyrifos has a dose-effect relationship.

The role of high mobility group protein 1 in toluene diisocyanate-induced NLPR3 inflammasome activation in human bronchial epithelial cells / 中国职业医学

OBJECTIVE: To investigate the role of high mobility group protein 1(HMGB1) in toluene diisocyanate(TDI) induced nucleotide-binding oligomerization domain like receptor family pyrin domain-containing 3(NLRP3) inflammasome activation in human bronchial epithelial cells(HBECs). METHODS: i) The TDI-human serum albumin(HSA) stimulation experiment: the HBECs in logarithmic growth phase were randomly divided into control group, low-, medium-and high-dose groups that were pretreated with TDI-HSA with the final concentration of 0.00, 40.00, 80.00 and 120.00 mg/L for 12 hours. ii) The HMGB1 expression inhibition experiment: the HBECs in logarithmic growth phase were divided into control group, TDI-HSA group, TDI-HAS+negative-siRNA group, and TDI-HAS+HMGB1-siRNA group. The cells in TDI-HAS+negative-siRNA group and TDI-HAS+HMGB1-siRNA group were infected with HBECs with negative-siRNA lentivirus and HMGB1-siRNA lentivirus, respectively. Cells in these two groups and the TDI-HSA group were treated with 120.00 mg/L of TDI-HSA for 12 hours. The cells in the control group were not treated with TDI-HAS. iii) The expression of HMGB1, NLRP3, apoptosis-associated speck-like protein containing CARD(ASC), pro-caspase-1 and caspase-1 p20 proteins in all groups were detected by Western blot. The number of NLRP3 and caspase-1 inflammasome in TDI-HSA stimulation experiment was observed by immunofluorescence method. RESULTS: i) TDI-HSA stimulation experiment: the relative protein expression of HMGB1 and ASC was higher in HBECs of medium-and high-dose groups than that of control group(all P values were <0.01). The relative protein expression of NLRP3 and casepase-1 p20 and the number of NLRP3-caspase-1 inflammasome were higher in HBECs of 3 dose groups than that of control group(all P values were <0.01). The number of NLRP3-caspase-1 inflammasome in HBECs increased obviously in low-, medium-and high-dose groups as compared to the control group(all P values were <0.05). The number of NLRP3-caspase-1 inflammasome in HBECs increased with the increase of TDI-HSA dose(all P values were <0.01). ii) The HMGB1 expression inhibition experiment: the relative protein expression of HMGB1, NLRP3, ASC, pro caspase-1 and caspase-1 p20 in HBECs were higher in the TDI-HSA group and TDI-HSA + negative-siRNA group than those of the control group(all P values were <0.01). The above indexes of HBECs were lower in the TDI-HAS + HMGB1-siRNA group than those in the TDI-HSA group and TDI-HSA + negative-siRNA group(all P values were <0.01).CONCLUSION: TDI treatment in HBECS can induce the increase of HMGB1 protein expression and activate NLPR3 inflammasome. Inhibition of HMGB1 expression can down-regulate the expression of NLPR3 and its related proteins.

Study on the correlation of HMGB1 and coagulation disorder in severe heatstroke rats / 解放军医学杂志

Objective To explore the association between HMGB1 and coagulation disorder in severe heat stroke rats. Methods A total of 48 rats were randomized equally into 8 groups: Room temperature group (Sham), severe heatstroke (HS) re-cooling 0 h (HS-0 h), 3 h (HS-3 h), 6 h (HS-6 h), 9 h (HS-9 h), 12 h (HS-12 h), 18 h (HS-18 h), 24 h (HS-24 h) groups. Sham group rats were housed at room temperature of (25.0±0.5) ℃ and humidity of (50.0%±5.0%), while severe heatstroke group rats were kept in an incubator at a temperature of (39.5±0.2) ℃ and humidity of 60.0%±5.0%. Rats with the rectal temperature (Tr) reached 43 ℃ were defined as the onset of severe heatstroke, followed by transferring to the room temperature for natural cooling. The blood samples were collected at 0, 3, 6, 9, 12, 18 and 24 h after natural cooling. Prothrombin time (PT), activated partial thromboplastin time (APTT), thrombin time (TT), fibrinogen (Fib) were evaluated by clotting methods. HMGB1and thrombin-antithrombin (TAT) were detected by ELISA. Results A linear association between HMGB1 and PT was found (P0.05); a nonlinear association between HMGB1 and PLT was found (P0.05). Conclusions HMGB1 has a significant association with coagulation disorder in severe heat stroke rats. The mechanism needs to be further studied.

The effect of losartan on the inflammatory damage through HMGB1 in the hepatocytes stimulated by heat stress / 解放军医学杂志

Objective To explore the function of losartan on heat-stress induced high-mobility group protein B1 (HMGB1) mediated inflammatory damage in the hepatocytes. Methods Rats were randomized into three groups: sham group (without heat stress), heatstroke group (heatstroke induction followed by i.p. injection of normal saline) and heatstroke+losartan group (heatstroke induction followed by i.p. injection of 50 mg/kg losartan). The serum and liver tissue were harvested nine hours after heatstroke to invest the serum alanine aminotransferase (ALT) levels, liver myeloperoxidase (MPO) levels, liver pathological morphology, serum HMGB1 levels as well as the expression of interleukin(IL)-1β and IL-18 in the liver. In vitro, the HBL3A hepatocyte cell lines were divided into the sham group (without heat stress), heat stress group and heat stress +losartan group (10 μmol/L losartan added into the supernatant after heat stress). Nine hours after heat stress, the cell viability and the levels of supernatant lactate dehydrogenase, supernatant HMGB, cytoplasm and total HMGB1 were all examined. Besides, the level of activated caspase-1 in hepatocytes, supernatant IL-1β and IL-18, as well as the cellular level of reactive oxygen species (ROS), were detected. The effects of recombinant HMGB1 with concentration gradients and 0.2 mmol/L hydrogen peroxide on the levels of IL-1β, IL-18 and HMGB1 in the heat stress hepatocytes treated by losartan were observed. Results In vivo, liver damage occurred in the rats of the heatstroke group. Compared with the heatstroke group, the levels of serum ALT, liver MPO, serum HMGB1, liver tissue IL-1β and IL-18 decreased in the heatstroke+losartan group (P<0.05). In vitro, hepatocytes in the heat stress group were apparently damaged. Compared with the heat stress, the levels of cytoplasmic HMGB1, supernatant HMGB1, IL-1β and IL-18 decreased in the heat stress+losartan group, and the cell survival rate increased (P<0.05). In addition, the HMGB1 inhibitor also reduced the levels of IL-1β and IL-18 in heat stress group hepatocytes. And the supplementation with HMGB1 increased the levels of IL-1β and IL-18 in heat stress hepatocytes treated by losartan (P<0.05). Losartan reduced the level of reactive oxygen species in heat stress hepatocytes, and the supplementation with hydrogen peroxide increased the level of HMGB1 in heat stress hepatocytes treated by losartan (P<0.05). Conclusion Losartan decreased the heat-stress induced HMGB1 mediated inflammatory damage in the hepatocytes.

Effect of bone morphogenetic protein-7 combined with porous tantalum on chondrogenic differentiation and function of bone marrow mesenchymal stem cells / 中国组织工程研究

BACKGROUND: The establishment of coculture system combined with physical factors and scaffold materials and the Induction of cytokines have become the focus of chondrogenlc differentiation of bone marrow mesenchymal stem cells. OBJECTIVE: To observe the effect of bone morphogenetic protein 7 combined with porous tantalum on chondrogenlc differentiation of bone marrow mesenchymal stem cells. METHODS: Bone marrow mesenchymal stem cells of Sprague-Dawley rats (provided by Beijing Huafukang Biology) were Isolated and cultured. Group Intervention: (1) in the experimental group, porous tantalum tablet was added, while In the control group, porous tantalum tablet was not added. At 5 days after culture, cell growth on the surface of porous tantalum tablet was observed by phalloidin staining. At 1, 3,5, and 7 days after culture, CCK-8 method was used to detect cell proliferation. (2) Group A was added with chondrocyte Inducer; group B with chondrocyte Inducer and bone morphogenetic protein 7; group C with domestic porous tantalum material and chondrocyte Inducer; group D with domestic porous tantalum material and chondrocyte Inducer and bone morphogenetic protein 7. At 7,14 and 21 days after culture, the levels of type II collagen, SRY type high mobility group protein and matrix metalloproteinase-13 secreted by cells In each group was detected by ELISA. Western blot assay was used to detect the expression of type II collagen, SRY type high mobility group protein and matrix metalloproteinase-13. This study was approved by the Animal Experimental Ethics Committee of North China University of Science and Technology. RESULTS AND CONCLUSION: (1) The phalloidin staining results showed that bone marrow mesenchymal stem cells grew well on and around the porous tantalum surface. (2) At 3 and 5 days after culture, the proliferation of bone marrow mesenchymal stem cells was slower In the experimental group than in the control group (P 0.05). (3) At 7,14 and 21 days, the expression of type II collagen and SRY high mobility group protein Increased gradually among groups A, B, C and D (P 0.05). At 21 days, there was no significant difference among groups A, B, C and D (P > 0.05). (4) Western blot assay showed that at 7,14 and 21 days after culture, the expression level of type II collagen and SRY high mobility group protein Increased gradually In groups A, B, C and D (P 0.05). (5) The results showed that bone morphogenetic proteln-7 combined with domestic porous tantalum could Induce cartilage differentiation of bone marrow mesenchymal stem cells, facilitate the expression of type II collagen and SRY high mobility group protein, and Inhibit the expression of matrix metalloproteinase-13.

Electroacupuncture at "Ganshu" (BL18) and "Yanglingquan" (GB34) alleviates hepatic ischemia-reperfusion injury by inhibiting translocation and release of high mobility group protein 1 in rats / 针刺研究

OBJECTIVE: To explore the protective effect of electroacupuncture (EA) on hepatic ischemia-reperfusion injury (HIRI) and the expression of high mobility group protein 1 (HMGB1) in liver tissues in rats. METHODS: A total of 40 male SD rats were randomly divided into 4 groups, namely sham control, HIRI model, "Ganshu"(BL18) -"Yanglingquan"(GB34) and non-acupoint group, with 10 rats in each group. The HIRI model was induced by blocking the arteries, veins and bile ducts supplying the middle and left lobes of the liver for 1 h, and reperfusion for 4 h to induce an area of about 70% HIRI. EA was applied to bila-teral BL18 and GB34, or non-acupoints about 6-8 mm to the bilateral BL18 for 30 min before modeling. Serum alanine transaminase (ALT) and aspartate aminotransferase (AST) levels were measured by using an automatic biochemical analyzer. Serum tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and HMGB1 levels were assayed by ELISA. Hematoxylin - eosin (H.E.) staining was used to observe histopathological changes of the liver tissue by using tissue injury scaling (0-3 scores). The expression of HMGB1 protein in liver tissues was detected by immunohistochemical staining, Western blot and PCR, separately. RESULTS: Following modeling and compared with the sham group, the levels of serum ALT, AST, TNF-α, IL-6, and HMGB1 contents, the number of HMGB1 immunoreaction (IR)-positive cells, and HMGB1 protein and mRNA were significantly increased (P＜0.01). After the treatment, the contents of serum ALT, AST, TNF-α, IL-6, and HMGB1, liver HMGB1 IR-positive cells, protein and mRNA were considerably down-regulated in the BL18-GB34 group (P0.05) in contrast to the model group. H.E. stain showed a higher liver injury score in the model group than in the sham group (P<0.01), and a lower liver injury score in the BL18-GB34 group (not the non-acupoint group) relevant to the model group (P<0.05). CONCLUSION: EA of BL18 and GB34 points has a protective effect on ischemic liver injury in rats with HIRI, which may be associated with its functions in inhibiting the migration and release of HMGB1 from the nucleus to the cytoplasm and in down-regulating the expression of inflammatory factors.

Apatinib enhances the anti-tumor effect of cisplatin on gastric cancer by inhibiting HMGA2 / 中国肿瘤生物治疗杂志

@#[Abstract] Objective: To investigate the effect of apatinib (APA) combined with cisplatin (DDP) on the proliferation, invasion and migration capacity of gastric carcinoma (GC) cells and its molecular mechanism. Methods: Cancer and para-cancerous tissue samples resected from 50 GC patients, who were surgically treated in Wuwei People's Hospital from January 2016 to June 2019, were collected for this study; in addition, GC cell lines MGC803 and SGC7901 were also collected. qPCR was used to detect the HMGA2 expression in tissues and mRNA expressions of molecules related to cell proliferation, migration and invasion in GC cell lines. MGC803 and SGC7901 cells were transfected with pcHMGA2 by liposome transfection technology. After treatment with DDP and APA at different concentrations, the cells were divided into NC, pcHMGA2, pcHMGA2+DDP and pcHMGA2+DDP+APA groups. Protein expression of HMGA2 in GC cells was detected by Western blotting, and proliferation, migration and invasion of the cells were detected by MTT and Transwell assay, respectively. Results: The mRNA expression of HMGA2 in GC tissues was higher than that in para-cancerous tissues (P<0.05), and the survival rate of GC patients in the high expression group was significantly reduced (P<0.01). DDP significantly inhibited the proliferation, invasion and migration of MGC803 and SGC7901 cells (all P<0.01); the proliferation, invasion and migration of MGC803 and SGC7901 cells in DDP+APA group significantly decreased (all P<0.01) as compared with DDP group; APA significantly enhanced the inhibitory effect of DDP on HMGA2 expression in GC cells (P<0.01); APA enhanced the anticancer activity of DDP against GC by down-regulating HMGA2 expression. Conclusion: APA promotes the anticancer activity of DDP against GC, and its molecular mechanism is the promotion of the inhibitory effect of DDP on HMGA2 expression.

Study on Mechanism of Taoren Chengqitang in Regulating Intestinal Myoelectrical Activity and Microenvironment Homeostasis in Intestinal Sepsis Rats Based on HMGB1/TLR4/NF-κB Pathway / 中国实验方剂学杂志

Objective:To study the mechanism of Taoren Chengqitang in regulating intestinal myoelectric activity and microenvironment homeostasis in intestinal sepsis rats based on high mobility group protein 1(HMGB1)/Toll-like receptor 4(TLR4)/nuclear factor -κB(NF-κB) pathway. Method:The 60 SD rats were randomly divided into sham operation group, model group, glycyrrhizic acid (HMGB1 inhibitor, 0.03 g·kg-1) group, Taoren Chengqitang group (10 g·kg-1), glycyrrhizic acid+Taoren Chengqitang group (0.03 g·kg-1+10 g·kg-1), with 12 rats in each group. Except the sham operation group, the other groups established intestinal sepsis rat models, each group was treated with medicine, hematoxylin-eosin (HE) staining was used to detect the histopathological changes of small intestinal mucosa in rats of each group, the changes of mucosal thickness and villus height were compared, the levels of secretory immunoglobulin A (sIgA), diamine oxidase (DAO) and D-lactic acid in intestinal mucosa of rats were detected by kit, the intestinal myoelectrical activity of rats in each group was measured, the slow wave frequency and amplitude of small intestinal smooth muscle were compared, the intestinal flora of rats in each group was detected, the contents of E. coli, Bifidobacterium and Lactobacillus were compared, and the expressions of HMGB1/TLR4/NF-κB pathway proteins HMGB1, TLR4, MyD88 and NF-κB p65 in small intestinal tissues were detected by Western blot. Result:Compared with sham operated group, the villus height, mucosal thickness, sIgA content, slow wave frequency and amplitude of smooth muscle, Bifidobacterium and Lactobacillus contents in intestinal mucosa of model group rats were significantly decreased, and serum DAO and D-lactic acid levels, intestinal E. coli content, intestinal tissue HMGB1, TLR4, MyD88 and NF-κB p65 proteins were significantly increased (P<0.05). Compared with the model group, the villus height, mucosal thickness, sIgA content, slow wave frequency and amplitude of smooth muscle, Bifidobacterium and Lactobacillus contents in intestinal mucosa of the Taoren Chengqitang group, glycyrrhizic acid group, and glycyrrhizic acid + Taoren Chengqitang group were significantly increased, and serum DAO and D-lactic acid levels, intestinal E. coli content, intestinal tissue HMGB1, TLR4, MyD88 and NF-κB p65 proteins were significantly decreased (P<0.05). Compared with the Taoren Chengqitang group and the glycyrrhizic acid group, the villus height, mucosal thickness, sIgA content, slow wave frequency and amplitude of smooth muscle, Bifidobacterium and Lactobacillus contents in intestinal mucosa of glycyrrhizic acid+Taoren Chengqitang group were significantly increased, and serum DAO and D-lactic acid levels, intestinal E. coli content, intestinal tissue HMGB1, TLR4, MyD88 and NF-κB p65 proteins were significantly decreased, the differences were statistically significant (P<0.05). Conclusions:Taoren Chengqitang can alleviate intestinal mucosal injury, regulate intestinal myoelectrical activity and microenvironment homeostasis, restore intestinal function and maintain flora balance in intestinal sepsis rats, which may be achieved by down-regulating HMGB1/TLR4/NF-κB pathway.

Effect of Whenshen Prescription on Hepatic Encephalopathy in Patients with HBV-related Hepatic Cirrhosis and Content of HMGB1/TLR4 / 中国实验方剂学杂志

Objective:To investigate the effect of Whenshen prescription on hepatic encephalopathy in patients with HBV-related hepatic cirrhosis and kidney Yang deficiency syndrome and the content of high-mobility group protein B1 (HMGB1) and Toll-like receptor 4 (TLR4). Method:The 66 patients treated in the Hospital of Chengdu University of Traditional Chinese Medicine from August 2013 to January 2015 were included. A prospective, parallel controlled design was used. The 66 cases were randomly divided into treatment group and control group according to 1:1 ratio, with 33 cases in each group.The control group was treated with comprehensive therapy, colon dialysis and placebo enema, while treatment group was treated with comprehensive therapy, colon dialysis and Whenshen prescription enema for 10 days. Finally, liver function, number connect test (NCT), digit-symbol test (SDT), awake time, the total effective rate and content of HMGB1 and TLR4 were analyzed. Result:There was no significant difference between two groups at baseline. The total effective rate in treatment group was 93.9%, which was higher than 72.7% in control group (PP1 and TLR4 in treatment group were lower than those in the control group. Conclusion:Whenshen prescription can significantly improve the clinical efficacy by inhibiting the contents of HMGB1 and TLR4.

Dual roles of immune cells and cytokines in hepatic ischemia reperfusion injury / 中华肝胆外科杂志

Ischemia-reperfusion injury is one of the main causes of complications related to liver transplantation, hepatectomy, trauma and hemorrhagic shock. The cells of ischemia and hypoxic injury release of injury-related molecular patterns, lead to the activation of immune cells and cytokine, which further aggravates the inflammatory response and enlarges the injury. It’s indicated that injury-related molecular patterns, liver resident immune cells and cytokines play a key role in promoting inflammation and liver ischemia-reperfusion injury. However, recent studies suggested that the ischemia cells and cytokines played acomplex role in this process. Relevant progresses were reviewed in this article.