Improvement effects of glycyrrhizin on Helicobacter pylori-associated gastritis in rats and its mechanism / 中国药房

OBJECTIVE To investigate the improvement effects of glycyrrhizin （GL） on Helicobacter pylori （HP）-associated gastritis in rats and its mechanism. METHODS HP-associated gastritis rat model was induced by inoculating with 1×109 cfu/mL HP. The model rats were randomly divided into model group， positive control group （HP standard quadruple group）， GL low-dose， medium-dose and high-dose groups （5， 20， 50 mg/kg）， with 12 rats in each group. Another 12 healthy rats were selected as normal control group. Except the normal control group and model group were given constant volume of normal saline intragastrically， the other groups were given corresponding drugs intragastrically， once a day， for 30 consecutive days. After administration， rats received 13C urea breath test， and delta-over-baseline （DOB） was recorded； the pathological and cellular morphological changes of gastric mucosa in rats were observed， and pathological scoring was performed； the levels of interleukin-8 （IL-8）， IL-1β， tumor necrosis factor-α （TNF-α）， reactive oxygen species （ROS） and malondialdehyde （MDA） were detected in gastric mucosa of rats； mRNA expressions of high mobility group box-1 protein （HMGB1） and nuclear factor-κ-B （NF-κB）， relative expressions of nitric oxide synthases （iNOS） and HMGB1， the phosphorylation level of NF- κBp65 were also detected in rats. RESULTS Compared with normal control group， the DOB value， histopathological score of gastric mucosa， the levels of IL-8， IL-1β， TNF-α， ROS and MDA， relative expressions of HMGB1 and NF- κB mRNA， relative expressions of iNOS and HMGB1 protein and the phosphorylation level of NF-κB p65 were all increased significantly in model group （P＜0.05）； the epithelial cells of gastric mucosa in rats were incomplete in structure and decreased in the number， with an increase in cell fragments and vacuoles， and significant cell pyknosis. Compared with model group， the changes of the above indexes in GL groups and positive control group were significantly reversed （P＜0.05）； the changes in the above indicators in the GL high-dose group were more significant than GL low-dose and medium-dose groups （P＜0.05）； the pathological changes of gastric mucosal cells in rats had all improved. CONCLUSIONS GL may inhibit inflammation and oxidative stress by inhibiting the activation of HMGB1/NF-κB pathway， thus relieving HP-induced gastric mucosal injury.

Mechanism of HMGB1 in scarring after glaucoma drainage valve implantation / 国际眼科杂志(Guoji Yanke Zazhi)

AIM: To explore the dynamic expression of high mobility group box 1(HMGB1)in scar tissues after glaucoma drainage valve implantation, and to further reveal the role and possible mechanism of HMGB1 in scarring after glaucoma surgery.METHODS: A total of 60 New Zealand white rabbits were randomly divided into control group(n=20), model group(n=20, silicone implantation under conjunctival sac)and model with drug administration group(n=20, silicone implantation under conjunctival sac combined with 5-fluorouracil injection). The conjunctival tissues were collected at 4 and 8 wk after surgery. HE staining and Masson staining were used to detect the proliferation and distribution of fibroblasts and collagen fibers in conjunctival tissues. Immunohistochemistry was utilized to detect the distribution and changes of HMGB1, transforming growth factor(TGF)-β1, Smad3 and α-smooth muscle actin(SMA)in conjunctival tissues. RT-PCR and Western blot were adopted to detect the mRNA and protein expression of HMGB1, TGF-β1, Smad3 and α-SMA in conjunctival tissues.RESULTS: HE staining and Masson staining showed that the proliferation of inflammatory cells, fibroblasts and collagen fibers in the model group was significantly higher than that in the control group at both 4 and 8 wk. Meanwhile, the proliferation of fibroblasts and collagen fibers in the model with drug administration group was significantly lower than that in the model group. Immunohistochemical staining showed that the expression of HMGB1, TGF-β1, Smad3 and α-SMA protein was observed in the conjunctival tissues of the model group both 4 and 8 wk, with brown and significantly deeper staining of the model group at 8 wk. Meanwhile, the positive staining in the model with drug administration group at both 4 and 8 wk was significantly lower than that in the model group. There was positive correlations between the number of fibroblasts stained with HE and the expression of HMGB1 in the conjunctival tissue of the model group at both 4 and 8 wk(r=0.602, 0.703, all P<0.05). RT-PCR and Western blot revealed that the mRNA and protein expression levels of HMGB1, TGF-β1, Smad3 and α-SMA in the model group were significantly higher than those in the control group at both 4 and 8 wk(all P<0.05). Meanwhile, the mRNA and protein expression levels of HMGB1, TGF-β1, Smad3 and α-SMA in the model with drug administration group were significantly lower than those in the model group(all P<0.05). There was positive correlations between mRNA expressions of HMGB1 and TGF-β1, Smad3 in the model group and the model with drug administration group(all P<0.05).CONCLUSION: The expression of HMGB1 increased at a time-dependent manner after glaucoma valve implantation. HMGB1 acts an indispensable role in the initiation and progression of scar formation after glaucoma surgery, which may be involved in the regulation of TGF-β/Smad signaling pathway.

Dahuang Mudantang Alleviates Intestinal Injury in Rat Model of Acute Pancreatitis by Regulating HMGB1/RAGE/NF-κB Signaling Pathway / 中国实验方剂学杂志

ObjectiveTo explore the mechanism of Dahuang Mudantang in alleviating the intestinal injury in the rat model of acute pancreatitis via the high-mobility group box 1 （HMGB1）/receptor for advanced glycation endproduct （RAGE）/nuclear factor-κB （NF-κB） signaling pathway. MethodOne hundred and twenty SPF-grade Wistar rats received retrograde injection of 5% sodium taurocholate into the biliopancreatic duct for the modeling of intestinal injury in acute pancreatitis. The rats were randomized into blank， model， low-， medium-， and high-dose （3.5， 7， 14 g·kg-1， administrated by gavage） Dahuang Mudantang， and octreotide （1×10-5 g·kg-1， subcutaneous injection） groups （n=20）. The rats in blank and model groups received equal volume of distilled water by gavage. Drugs were administered 1 h before and every 12 h after modeling， and samples were collected 24 h after modeling. The general status of the rats was observed. The biochemical methods were employed to measure the levels of amylase （AMS） and C-reactive protein （CRP） in the serum. The enzyme-linked immunosorbent assay （ELISA） was employed to measure the levels of tumor necrosis factor （TNF）-α， interleukin （IL）-1β， and IL-6 in the colon tissue. The morphological changes of pancreatic and colon tissues were observed by hematoxylin-eosin （HE） staining. Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR） and Western blot were employed to measure the expression levels of HMGB1， RAGE， inhibitor of NF-κB kinase （IKK）， and NF-κB suppressor protein α（IκBα）in the colon tissue. ResultThe rats in the model group showed poor general survival， writhing response， reduced frequency of defecation， and dry stool. The symptoms of rats in the model group were mitigated in each treatment group， and the high-dose Dahuang Mudantang showed the most significant effect. Compared with the normal group， the model group had elevated AMS and CRP levels （P<0.05）， which were lowered by Dahuang Mudantang （P<0.05）， especially that at the high dose （P<0.05）. Compared with the normal group， the modeling elevated that levels of TNF-α， IL-1β， and IL-6 （P<0.05）. Such elevations were lowered by Dahuang Mudantang （P<0.05）， and the high-dose group and the octreotide group showed better performance （P<0.05）. The modeling caused necrotic， congested， and destructed pancreatic and colonic tissues， which were ameliorated by the drugs， especially high-dose Dahuang Mudantang. Compared with the normal group， the modeling up-regulated the mRNA levels of HMGB1， RAGE， IKK， IκBα， and NF-κB （P<0.05）. Compared with the model group， Dahuang Mudantang and octreotide down-regulated the mRNA levels of HMGB1， RAGE， IKK， IκBα， and NF-κB （P<0.05）， and the high-dose Dahuang Mudantang demonstrated the best performance （P<0.05）. Western blot results showed a trend consistent with the results of Real-time PCR. ConclusionDahuang Mudantang can improved the general status， reduce inflammation， and alleviate histopathological changes in the pancreatic and colon tissues in the rat model of acute pancreatitis by inhibiting the HMGB1/RAGE/NF-κB signaling pathway.

High mobility group box-1 inhibition reduces expression of matrix metalloproteinase-9 in astrocytes in rats after spinal cord injury / 中华创伤骨科杂志

Objective:To investigate the role and underlying mechanisms of inhibiting high mobility group box-1 (HMGB1) in the expression of matrix metalloproteinase-9 (MMP-9) in spinal cord astrocytes (AS) in rats after spinal cord injury (SCI).Methods:After an SCI model was established in Sprague-Dawley (SD) rats using a modified Allen's Weight-Dropping method and ethyl pyruvate (EP) or glycyrrhizin (GL) was used to inhibit the effect of HMGB1, the rats were divided into a sham group, an SCI group, an SCI+EP (50 mg/kg) group, and an SCI+GL (100 mg/kg) group. The expression levels of glial fibrillary acid protein (GFAP) and MMP-9 in spinal cord AS were observed. After the spinal cord AS in SD rats was cultured and incubated by the oxygen-glucose deprivation/reoxygenation (OGD/R) procedure, the expression of MMP-9 protein was detected at 6 h/R 6 h, 12 h, 24 h, and 48 h after OGD. The time point with the highest expression was chosen in the subsequent experiments as an OGD/R group. HMGB1 was inhibited by HMGB1 shRNA or EP to observe the effect of HMGB1 on the expression of MMP-9 protein in AS treated with OGD/R. Then, toll-like receptor 4 (TLR4) inhibitor, TIR-domain-containing adaptor inducing interferon- β (TRIF) inhibitor, and nuclear factor-kappa B (NF- κB) inhibitor were used to investigate the effects of TLR4/TRIF/NF- κB signaling pathway during the regulation of HMGB1 on MMP-9 in vitro. Results:Western blot showed that the expression of MMP-9 protein in the spinal cord was significantly increased in rats at 1 d after SCI, and the expression of MMP-9 protein in the SCI+EP group and the SCI+GL group was significantly lower than that in the SCI group ( P<0.001). Immunofluorescence showed that GFAP and MMP-9 proteins were co-localized in the spinal cord after SCI, and the expression of GFAP and MMP-9 proteins in the SCI+EP and SCI+GL groups was significantly lower than that in the SCI group ( P<0.05). Since the expression of MMP-9 protein in the spinal cord AS cultured in vitro was significantly higher in the OGD 6h/R 12h group than that in the normal group and the OGD 6h/R 6h, 24, and 48 h groups, the OGD 6h/R 12h was taken as the OGD/R group. The MMP-9 protein expression in AS in the OGD/R+HMGB1 shRNA group and the OGD/R+EP group was significantly lower than that in the OGD/R group ( P<0.001). In the cultured AS, moreover, inhibiting TLR4, TRIF, and NF- κB reduced MMP-9 protein expression after OGD 6 h/R 12 h when compared with that in the OGD/R group ( P<0.001). Conclusions:HMGB1 inhibition may result in a reduction in MMP-9 expression both in the spinal cord AS in SCI rats and in AS after OGD/R treatment in vitro. HMGB1 may regulate MMP-9 protein expression in AS after OGD/R treatment via the TLR4/TRIF/NF- κB signal pathway.

Mechanism of regulatory effect of recombinant human HMGB1 on endothelial cell angiogenesis / 器官移植

Objective To unravel the possible mechanism of the role of recombinant human high mobility group box 1 (rhHMGB1) protein in regulating the angiogenesis of endothelial cells. Methods Endothelial cells were divided into the control group, bone marrow mesenchymal stem cells (MSC) supernatant group and rhHMGB1 group. The proliferation and survival of endothelial cells were detected by cell counting kit(CCK)-8 assay. The relative expression levels of vascular endothelial growth factor (VEGF), Yes-associated protein (YAP), CD31 and hypoxia inducible factor (HIF)-1α proteins were determined by Western blot. The relative expression levels of VEGF, YAP, CD31 and HIF-1α messenger RNA (mRNA) were detected by real-time fluorescent quantitative polymerase chain reaction (RT-qPCR). The migration ability of endothelial cells was assessed by Transwell chamber test. The localization of YAP was detected by immunofluorescence staining. Results Compared with the control group, the migration rate of endothelial cells was increased in the rhHMGB1 group (P < 0.05), and the cell migration rate was enhanced over time. Compared with the control group, the relative expression levels of VEGF and p-YAP proteins were up-regulated in the MSC supernatant group, and the differences were statistically significant (both P < 0.05). Compared with the control group, the relative expression levels of VEGF and HIF-1α proteins, VEGF and CD31 mRNA and YAP and p-YAP proteins were up-regulated, and YAP/p-YAP ratio was increased in the rhHMGB1 group, and the differences were statistically significant (all P < 0.05). Compared with the MSC supernatant group, the relative expression levels of CD31 mRNA and YAP protein were up-regulated, and the YAP/p-YAP ratio was increased in the rhHMGB1 group, and the differences were statistically significant (all P < 0.05). Conclusions Exogenous high-concentration rhHMGB1 may promote the migration ability of endothelial cells and up-regulate the expression levels of angiogenesis-related proteins by regulating the recruitment of YAP to the nucleus.

Effect of Modified Erchentang on Bronchioles of Rats with Chronic Obstructive Pulmonary Disease by HMGB1/RAGE/NF-κB Signaling Pathway / 中国实验方剂学杂志

ObjectiveTo study the effect of modified Erchentang on the expression of key molecules in the high mobility group Box 1 protein （HMGB1）/receptor for advanced glycation endproduct （RAGE）/nuclear factor-κB （NF-κB） signaling pathway in bronchioles of rats with chronic obstructive pulmonary disease （COPD）， to explore the mechanism of modified Erchentang against bronchiolar inflammation of COPD rats via HMGB1/RAGE/NF-κB signaling pathway. MethodSixty SD rats were randomly divided into normal group， model group， modified Erchentang low-， medium- and high-dose groups （5， 10， 20 g·kg-1·d-1） and ethyl pyruvate （HMGB1 inhibitor） group， with 10 in each group. The COPD rat model was prepared by cigarette smoke combined with tracheal injection of lipopolysaccharide （LPS）. After modeling， the modified Erchentang groups were given corresponding drugs （ig） and Ringer's solution （4 mL， ip）， while the EP group was treated with equal volume of normal saline （ig） and EP （0.04 g·kg-1·d-1， ip）. The normal group and the model group received equal volume of normal saline （ig） and Ringer's solution （ip） for 21 consecutive days. The contents of HMGB1， chemokine （C-X-C motif） ligand 1 （CXCL1）， CXCL2 and monocyte chemotactic protein-1 （MCP-1） in bronchoalveolar lavage fluid （BALF） were detected by enzyme-linked immunosorbent assay （ELISA）. The mRNA expressions of HMGB1， RAGE and NF-κB p65 were determined by Real-time polymerase chain reaction （Real-time PCR）， and the protein expressions of HMGB1， RAGE， p-NF-κB p65， and alpha-smooth muscle actin （α-SMA） in bronchioles tissue of rats were determined by immunohistochemistry （IHC）. ResultCompared with the conditions in the normal group， the forced vital capacity （FVC）， forced expiratory volume in the first second （FEV1） and FEV1/FVC in the model group were decreased （P<0.01） while the contents of HMGB1， CXCL1， CXCL2 and MCP-1 in BALF were increased （P<0.01）. And the model group presented higher mRNA expressions of HMGB1， RAGE and NF-κB p65 （P<0.01） and protein expressions of HMGB1， RAGE， p-NF-κB p65 and α-SMA （P<0.05， P<0.01） than the normal group. Compared with the model group， the modified Erchentang medium- and high-dose groups had increased FEV1/FVC （P<0.05， P<0.01）， lowered contents of HMGB1， CXCL1， CXCL2 and MCP-1 in BALF （P<0.05， P<0.05）， and reduced mRNA expressions of HMGB1， RAGE and NF-κB p65 （P<0.05， P<0.01） and protein expressions of HMGB1， RAGE， p-NF-κB p65 and α-SMA （P<0.05， P<0.01）. ConclusionModified Erchentang can resist bronchiolar inflammation of COPD rats. The mechanism may be related to down-regulating the mRNA expressiona of HMGB1 and RAGE， inhibiting the activity of NF-κB， and reducing the release of HMGB1， CXCL1， CXCL2 and MCP-1， thus suppressing the inflammatory injury and abnormal repair of bronchioles.

Compound Tinglizi Decoction intervenes COPD-associated pulmonary hypertension through regulation of HMGB1-mediated pyroptosis and immune imbalance / 中国中药杂志

This paper aimed to investigate the effects of high mobility group box 1(HMGB1)-mediated pulmonary artery smooth muscle cell pyroptosis and immune imbalance on chronic obstructive pulmonary disease-associated pulmonary hypertension(COPD-PH) in rats and the intervening mechanism of Compound Tinglizi Decoction. Ninety rats were randomly divided into a normal group, a model group, low-dose, medium-dose, and high-dose Compound Tinglizi Decoction groups, and a simvastatin group. The rat model of COPD-PH was established by fumigation combined with lipopolysaccharide(LPS) intravascular infusion, which lasted 60 days. Rats in the low, medium, and high-dose Compound Tinglizi Decoction groups were given 4.93, 9.87, and 19.74 g·kg~(-1) Compound Tinglizi Decoction by gavage, respectively. Rats in the simvastatin group were given 1.50 mg·kg~(-1) simvastatin by gavage. After 14 days, the lung function, mean pulmonary artery pressure, and arterial blood gas of rats were analyzed. Lung tissues of rats were collected for hematoxylin-eosin(HE) staining to observe the pathological changes. Real-time fluorescent quantitative polymerase chain reaction(qRT-PCR) was used to determine the expression of related mRNA in lung tissues, Western blot(WB) was used to determine the expression of related proteins in lung tissues, and enzyme linked immunosorbent assay(ELISA) was used to determine the levels of inflammatory factors in the lung tissues of rats. The ultrastructure of lung cells was observed by transmission electron microscope. The forced vital capacity(FVC), forced expiratory volume in 0.3 second(FEV_(0.3)), FEV_(0.3)/FVC, peek expiratory flow(PEF), respiratory dynamic compliance(Cdyn), arterial partial pressure of oxygen(PaO_2), and arterial oxygen saturation(SaO_2) were increased, and resistance of expiration(Re), mean pulmonary arterial pressure(mPAP), right ventricular hypertrophy index(RVHI), and arterial partial pressure of carbon dioxide(PaCO_2) were decreased by Compound Tinglizi Decoction in rats with COPD-PH. Compound Tinglizi Decoction inhibited the protein expression of HMGB1, receptor for advanced glycation end products(RAGE), pro caspase-8, cleaved caspase-8, and gasdermin D(GSDMD) in lung tissues of rats with COPD-PH, as well as the mRNA expression of HMGB1, RAGE, and caspase-8. Pulmonary artery smooth muscle cell pyroptosis was inhibited by Compound Tinglizi Decoction. Interferon-γ(IFN-γ) and interleukin-17(IL-17) were reduced, and interleukin-4(IL-4) and interleukin-10(IL-10) were incresead by Compound Tinglizi Decoction in lung tissues of rats with COPD-PH. In addition, the lesion degree of trachea, alveoli, and pulmonary artery in lung tissues of rats with COPD-PH was improved by Compound Tinglizi Decoction. Compound Tinglizi Decoction had dose-dependent effects. The lung function, pulmonary artery pressure, arterial blood gas, inflammation, trachea, alveoli, and pulmonary artery disease have been improved by Compound Tinglizi Decoction, and its mechanism is related to HMGB1-mediated pulmonary artery smooth muscle cell pyroptosis and helper T cell 1(Th1)/helper T cell 2(Th2), helper T cell 17(Th17)/regulatory T cell(Treg) imbalance.

The mechanism of the receptors for advanced glycation end products affecting pericyte detachment from blood vessels after traumatic brain injury / 西安交通大学学报(医学版)

【Objective】 To observe the reactive change of cortical perivascular cells after craniocerebral injury and explore its mechanism. 【Methods】 The controllable cortical impact animal model was used to simulate craniocerebral injury, the expressions of cortical pericyte markers at different time points after trauma were studied by Western blotting, and the biological behavior of vascular pericytes after craniocerebral injury was determined by transmission electron microscopy. Post-traumatic high mobility group box 1 (HMGB1), receptor for advanced glycation end product (RAGE), and nuclear factor κB (NF-κB) were detected by Western blotting. The experimental animals were divided into FPS-ZM1 (a specific RAGE receptor blocker) injection group and wild-type group. Wet and dry brain weight and transmission electron microscopy were used to study the post-traumatic effects of HMGB1-RAGE on pericytes. The primary mouse brain microvascular pericytes were cultured and supplemented with HMGB1 recombinant protein; the cultured pericytes supplemented with FPS-ZM1 were used as the control to explore the effect of HMGB1-RAGE pathway on vascular pericytes in vitro. 【Results】 The expression levels of early post-traumatic cortical pericyte markers platelet-derived growth factor receptor beta (PDGFR-β) and NG2 proteoglycan (NG2) decreased (PDGFR-β, Control vs. CCI 3D P<0.05; NG2, Control vs. CCI 6H P<0.05; Control vs. CCI 1D P<0.05). We found that pericytes were detached from blood vessels, accompanied by local blood-brain barrier opening. The expression of HMGB1-RAGE-NF-κB signaling pathway was increased in the early cortex after trauma (HMGB1, Control vs. CCI 6H P<0.05, Control vs. CCI 1D P<0.05; RAGE, Control vs. CCI 6H P<0.05, Control vs. CCI 1D P<0.05, Control vs. CCI 3D P<0.05, Control vs. CCI 5D P<0.05, Control vs. CCI 7D P<0.05; NF-κB, Control vs. CCI 6H P<0.05, Control vs. CCI 1D P<0.05). After blocking the binding of RAGE with the ligand, cortical edema was reduced (CCI 6H P<0.05, CCI 1D P<0.05), and neurovascular unit damage was reduced. HMGB1 recombinant protein could increase the migration ability of cultured pericytes (Control vs. HMGB1 P<0.05, Control vs. HMGB1+FPS-ZM1 P<0.05), and could be reversed by FPS-ZM1 (HMGB1 vs. HMGB1+FPS-ZM1 P<0.05). 【Conclusion】 High-level HMGB1 after traumatic brain injury mediates pericytes’ detachment from blood vessels through RAGE on pericytes and leads to the occurrence of local cerebral edema.

miR-142-3p alleviates house dust mite induced airway inflammation among children / 中国学校卫生

Objective@#To investigate the role of miR-142-3p in alleviation of house dust mite induced allergic airway inflammation among children, so as to provide insights into unraveling the pathogenesis of allergic airway inflammation.@*Methods@#Serum samples were collected from 15 patients with house dust mite induced allergic asthma and 15 healthy children in Jiangnan University Medical Center from September to November 2022, and serum miR-142-3p expression was quantified using a fluorescent quantitative real time PCR (qPCR) assay. The levels of interleukin 6 (IL 6) and tumor necrosis factor α (TNF α) were measured in the cell culture supernatant using enzyme linked immunosorbent assay (ELISA), and the expression of high mobility group box 1 (HMGB1) was detected at transcriptional and translational lvels using qPCR and Western blotting assays. The negative regulation of the HMGB1 gene by miR 142 3p was identified using a dual luciferase gene reporter assay, and the expression of downstream regulatory proteins was determined in human normal lung epithelial cells (BEAS 2B) cells transfected with miR 142 3p using Western blotting. In addition, female C57BL/6 mice at ages of 6-8 weeks were randomly assigned to the phosphate buffer saline (PBS) group, house dust mite sensitized airway inflammation group and house dust mite sensitized airway inflammation + miR 142 3p intervention group. Mouse airway inflammation was evaluated using hematoxylin eosin staining, and the expression of inflammatory cells and inflammatory cytokines were detected in mouse bronchoalveolar lavage fluid (BALF) using Giemsa staining and ELISA.@*Results@#Lower serum miR-142-3p expression was quantified among children with house dust mite induced allergic asthma than among healthy controls (1.33±0.21 vs. 4.74±0.62, t=5.22, P <0.05). Stimulation with dermatophagoides farinae extract (DFE) resulted in a reduction in miR-142-3p expression in BEAS-2B cells (0.82±0.25), while transfection with miR-142-3p mimics resulted in a rise in miR-142-3p expression in BEAS-2B cells (0.55±0.14)( t=3.31, 3.94, P <0.05). Pre treatment with miR-142-3p reduced the expression of IL 6(2.25±0.46)and TNF α(6.58±1.95) ( t=4.86, 3.38, P <0.05) in BEAS 2B cells stimulated with DFE, and treatment with miR-142-3p mimics resulted in a reduction in TLR4 and NF-κB expression in BEAS-2B cells via negative regulation of the HMGB1 expression. In addition, treatment with miR-142-3p was found to alleviate inflammatory cell infiltration in lung tissues of house dust mite sensitized mice, and results in a reduction in interleukin 4 (IL-4)[(107.60±10.43)pg/mL], interleukin 5 (IL 5)[(95.78±13.14)pg/mL] and HMGB1[(2.52±0.87)pg/mL] expression in BALF ( t=10.32, 7.29, 2.90, P <0.05).@*Conclusion@#miR-142-3p alleviates house dust mite induced allergic airway inflammation among children via negative regulation of the HMGB1/TLR4/NF-κB pathway.

Effect of HMGB1/Caspase-1/GSDMD signaling axis-mediated hepatocyte pyroptosis on liver ischemia-reperfusion injury / 器官移植

Objective To evaluate the effect of high mobility group box 1 (HMGB1)/ cysteinyl aspartate specific proteinase (Caspase)-1/Gasdermin D (GSDMD) signaling axis-mediated hepatocyte pyroptosis on liver ischemia-reperfusion injury (IRI). Methods C57BL/6 mice were randomly divided into the sham operation group (Sham group), IRI 2 h group, IRI 6 h group, IRI 12 h group, glycyrrhizic acid (GA)+Sham group and GA+IRI 12 h group (n=8 in each group). AML12 cells were evenly divided into the Sham group, IRI 12 h group, GA+Sham group and GA+IRI 12 h group. The serum levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), interleukin (IL)-1β and IL-6 in each group were detected by enzyme-linked immune absorbent assay(ELISA). The messenger ribonucleic acid (mRNA) levels of IL-1β and IL-6 were detected by reverse transcription polymerase chain reaction(RT-PCR). The pathological score of liver ischemia and cell apoptosis were compared among all groups. The expression level of HMGB1 in the liver tissues of each group was determined by immunohistochemistry. The expression levels of HMGB1, Caspase-1 and GSDMD proteins in the mouse liver tissues and AML12 cells were measured by Western blot. Results Compared with the Sham group, the serum levels of ALT, AST, IL-1β and IL-6 and the relative expression levels of IL-1β and IL-6 mRNA in the liver tissues were all significantly up-regulated after IRI in each group (all P < 0.05), and showed significant time-dependent pattern along with the prolongation of reperfusion time. Compared with the Sham group, the pathological score of hepatic ischemia and the apoptosis rate of hepatocytes were significantly increased after IRI in each group (all P < 0.05). Immunohistochemical results showed that the expression level of HMGB1 in the liver tissues was significantly up-regulated after IRI, which showed an increasing trend along with the prolongation within the period of 2-12 h. Western blot showed that compared with the Sham group, the relative expression levels of HMGB1, Caspase-1 and GSDMD proteins in vivo and in vitro were up-regulated in the IRI 12 h group. The relative expression level of HMGB1 protein was significantly up-regulated, whereas those of Caspase-1 and GSDMD proteins were significantly down-regulated in the GA+IRI 12 h group compared with those in the IRI 12 h group (all P < 0.05). Conclusions Hepatocytes probably activate the Caspase-1/GSDMD signaling pathway by releasing HMGB1, thereby triggering hepatocyte pyroptosis and leading to liver IRI. Inhibition of extracellular release of HMGB1 by GA may mitigate liver IRI.

Role of high-mobility group box 1 in cancer / 中南大学学报(医学版)

High-mobility group box 1 (HMGB1) is a non-histone nuclear protein in most eukaryocytes. Inside the nucleus, HMGB1 plays an important role in several DNA events such as DNA repair, transcription, telomere maintenance, and genome stability. While outside the nucleus, it fulfils more complicated functions, including promoting cell proliferation, inflammation, angiogenesis, immune tolerance and immune escape, which may play a pro-tumoral role.Meanwhile, HMGB1 acts as an anti-tumoral protein by regulating immune cell recruitment and inducing immunogenic cell death (ICD) during the carcinogenesis process. Therefore, abnormal expression of HMGB1 is associated with oncogenesis, development, and metastasis of cancer, which may play a dual role of pro-tumor and anti-tumor.

Metformin improves implant osseointegration in type 2 diabetic rats through inhibition high mobility group box 1 and receptor for advanced glycation end product / 中华内分泌代谢杂志

Objective:To investigate the effects of metformin on the expression of high mobility group box 1 (HMGB1) and receptor for advanced glycation end product(RAGE) around implants in type 2 diabetic rats and underlying mechanism on bone bonding. To investigate the effect of metformin on osseointegration in non-diabetic rats.Methods:Forty male SD rats aged 6 to 8 weeks were randomly divided into 4 groups by random number table with 10 rats each: normal group (T0 group), normal+ metformin group (T1 group), diabetic group (T2 group), and diabetic+ metformin group (T3 group). After type 2 diabetes model was established in T2 and T3 groups, pure titanium implants were implanted in bilateral tibial epiphyseal of all rats. On the same day, T1 and T3 groups were given 300 mg·kg -1·d -1 metformin, and other groups were gavaged with the same amount of normal saline. At the 4th and 8th week after surgery, 5 rats in each group were randomly sacrificed, and bone structure was examined using HE staining and micro-computed tomography (micro-CT). The expression of related factors was detected with immunohistochemistry and enzyme linked immunosorbent assay. Results:At the 4th and 8th week after surgery, the trabecular bone structure, new bone formation quality, and bone microparameters in T3 group were better than those in T2 group. Compared with T2 group, the expression of HMGB1 and RAGE was decreased, the content of osteocalcin was increased, and the expression of tumor necrosis factor-α was decreased (all P<0.01). 4 weeks after operation, bone volume/tissue volume and trabecular number in T1 group was higher than that in T0 group [(0.569±0.013)% vs (0.523±0.030)%, P=0.014; (1.695±0.059)/mm vs (1.569±0.050)/mm, P=0.007]. 8 weeks after operation, trabecular number in T1 group was higher than that in T0 group [(2.324±0.337)/mm vs (1.882±0.057)/mm, P=0.042]. Compared with T0 group, the content of RAGE in T1 group was significantly decreased [(35.49±2.77)ng/L vs (44.92±7.99)ng/L, P=0.005]. The osteocalcin content in T1 group was significantly higher than that in T0 group [(1.32±0.19)ng/L vs (0.89±0.26)ng/L, P=0.001]. Conclusions:Metformin can reduce the expression of HMGB1 and RAGE in type 2 diabetic rats, which may be one of the mechanisms promoting implant bone binding. Metformin has bone protective effect on non-diabetic rats.

Suppression of HMGB1 inhibits neuronal autophagy and apoptosis to improve neurological deficits in rats following intracerebral hemorrhage / 南方医科大学学报

OBJECTIVE@#To investigate the effect of suppressing high-mobility group box 1 (HMGB1) on neuronal autophagy and apoptosis in rats after intracerebral hemorrhage (ICH) in rats.@*METHODS@#Rat models of ICH induced by intracerebral striatum injection of 0.2 U/mL collagenase Ⅳ were treated with 1 mg/kg anti-HMGB1 mAb or a control anti-IgG mAb injected via the tail immediately and at 6 h after the operation (n=5). The rats in the sham-operated group (with intracranial injection of 2 μL normal saline) and ICH model group (n=5) were treated with PBS in the same manner after the operation. The neurological deficits of the rats were evaluated using modified neurological severity score (mNSS). TUNEL staining was used to detect apoptosis of the striatal neurons, and the expressions of HMGB1, autophagy-related proteins (Beclin-1, LC3-Ⅱ and LC3-Ⅰ) and apoptosis-related proteins (Bcl-2, Bax and cleaved caspase-3) in the brain tissues surrounding the hematoma were detected using Western blotting. The expression of HMGB1 in the striatum was detected by immunohistochemistry, and serum level of HMGB1 was detected with ELISA.@*RESULTS@#The rat models of ICH showed significantly increased mNSS (P < 0.05), which was markedly lowered after treatment with anti- HMGB1 mAb (P < 0.05). ICH caused a significant increase of apoptosis of the striatal neurons (P < 0.05), enhanced the expressions of beclin-1, LC3-Ⅱ, Bax and cleaved caspase-3 (P < 0.05), lowered the expressions of LC3-Ⅰ and Bcl-2 (P < 0.05), and increased the content of HMGB1 (P < 0.05). Treatment with anti-HMGB1 mAb obviously lowered the apoptosis rate of the striatal neurons (P < 0.05), decreased the expressions of Beclin-1, LC3-Ⅱ, Bax and cleaved caspase-3 (P < 0.05), increased the expressions of LC3-Ⅰ and Bcl-2 (P < 0.05), and reduced the content of HMGB1 in ICH rats (P < 0.05).@*CONCLUSION@#Down- regulation of HMGB1 by anti-HMGB1 improves neurological functions of rats after ICH possibly by inhibiting autophagy and apoptosis of the neurons.

Associations of high-mobility group box 1 and receptor for advanced glycation end products with acute lung injury in patient with acute aortic dissection

SUMMARY OBJECTIVE: To investigate the associations of high-mobility group box 1 and its specific receptor, receptor for advanced glycation end products with acute lung injury in patients with acute aortic dissection. METHODS: A total of 96 acute aortic dissection patients were divided into acute aortic dissection with acute lung injury group (38 cases) and acute aortic dissection without acute lung injury group (58 cases), according to partial pressure of oxygen/fraction of inspired oxygen. In addition, 44 healthy individuals were selected for the control group. The blood samples were taken. The serum high-mobility group box 1 and receptor for advanced glycation end products levels were detected by enzyme-linked immunosorbent assay, and the partial pressure of oxygen/fraction of inspired oxygen was measured. RESULTS: 24 h after admission, the high-mobility group box 1 and receptor for advanced glycation end products levels in acute aortic dissection with acute lung injury and acute aortic dissection without acute lung injury groups were significantly higher than those in the control group, respectively (p<0.05), and each index in acute aortic dissection with acute lung injury group was significantly higher than that in acute aortic dissection without acute lung injury group (p<0.05). At each time point within 96 h after admission, compared with acute aortic dissection without acute lung injury group, in acute aortic dissection with acute lung injury group, the high-mobility group box 1 and receptor for advanced glycation end products levels were increased, respectively, and the partial pressure of oxygen/fraction of inspired oxygen was decreased. The correlation analysis showed that, in acute aortic dissection patients, the high-mobility group box 1 and receptor for advanced glycation end products levels were negatively correlated with partial pressure of oxygen/fraction of inspired oxygen, respectively (p<0.05). CONCLUSIONS: The serum high-mobility group box 1 and receptor for advanced glycation end products levels may be associated with the occurrence of acute lung injury in acute aortic dissection patients. Monitoring the high-mobility group box 1 and receptor for advanced glycation end products levels can evaluate the risk of acute aortic dissection with acute lung injury.

Effects of oleanolic acid on early brain injury after subarachnoid hemorrhage through SIRT1 / 国际生物医学工程杂志

Objective:To investigate the effect of oleanolic acid (OA) on Sirtuin1 (SIRT1)-mediated high-mobility group box 1(HMGB1) deacetylation in the early brain injury after subarachnoid hemorrhage (SAH).Methods:A total of 176 male Sprague-Dawley rats were randomly divided into Sham operation (Sham group) ( n=48), SAH group ( n=48), OA group ( n=48) and Sirtinol group ( n=32). Rats in the SAH group, OA group and the Sirtinol group all adopted internal carotid artery puncture to construct SAH model, while rats in the sham group did not adopt puncture. One hour after modeling, the rats in the OA group were given intraperitoneal injection of OA (20 mg/kg), and the rats in the Sirtinol group were given intracerebroventricular injection of Sirtinol (2 mmol/L, 30 μL/kg). The rats in the sham group and SAH group were injected with equal volumes of sodium chloride injection. The SAH score and neurological score were performed 24 h after SAH, and the water content in the brain tissue and Evans blue exudation rate were measured. The expressions of HMGB1, SIRT1 and acetylated HMGB1 proteins in the brain tissue of rats were detected by Western Blot. The expression of HMGB1 mRNA in the brain of the rats was detected by quantitative real-time PCR. The distribution of HMGB1 protein in the brain of the rats was observed by immunofluorescence staining. TUNEL staining was used to observe the neuronal apoptosis in the brain tissue of the rats. Results:Compared with the SAH group, the SAH score of the OA group was significantly reduced ( P<0.001), the Garcia score was increased ( P<0.01), and the brain water content and Evans blue exudation rate were both reduced (all P<0.01). Compared with the OA group, the SAH score of the Sirtinol group was increased ( P<0.01), the Garcia score was significantly decreased ( P<0.001), and the brain water content and Evans blue exudation rate were both increased (all P<0.01). The results of Western Blot and real-time fluorescent quantitative PCR showed that, compared with the SAH group, the protein level ( P<0.01) and mRNA level ( P<0.05) of HMGB1 in the OA group were decreased, the expression of SIRT1 protein was significantly increased ( P<0.001), and the expression of acetylated HMGB1 protein was decreased ( P<0.01). Immunofluorescence staining showed that OA inhibited the migration of HMGB1 protein from the nucleus to the cytoplasm. TUNEL staining showed that OA could effectively reduce the number of TUNEL-positive cells. Compared with the OA group, Sirtinol significantly increased the number of TUNEL-positive cells. Conclusions:OA can reduce the release of HMGB1 through the SIRT1/HMGB1 pathway, thereby protecting the early brain injury after SAH.

The mechanism of high mobility group box 1 in coronary artery lesions of Kawasaki disease / 国际儿科学杂志

Kawasaki disease(KD)is a common acute febrile vasculitis in children, but the etiology and pathogenesis of KD are unknown.The most serious complication is coronary artery lesions.The damage of vascular endothelial cell may be the key mechanism of coronary artery lesions in KD.It is found that serum levels of high mobility group box 1(HMGB1) increase in the acute phase of KD, especially in children with coronary artery lesions, and that the gene of HMGB1 is associated with coronary artery lesions in Korean children with KD.HMGB1 is a non-histone DNA binding protein related to inflammation, which plays an important role in promoting the apoptosis of coronary artery endothelial cells in KD.The article reviews the mechanism of HMGB1 in coronary artery lesions of KD.

Recent progress on the roles of DAMP and NET in organ ischemia-reperfusion injury / 器官移植

Ischemia-reperfusion injury (IRI) is a common pathophysiological phenomenon, secondary to multiple pathological processes, such as organ transplantation, acute kidney injury and myocardial infarction. IRI may significantly aggravate the severity of diseases and increase the fatality of patients. Aseptic inflammation is one of the critical mechanisms of IRI. Damage-associated molecular pattern (DAMP) is a pivotal substance, which mediates aseptic inflammation. After released into extracellular space, it could effectively activate the immune system, and initiate and maintain the inflammatory responses by binding with pattern recognition receptor (PRR). Neutrophil extracellular trap (NET) is a DNA-based network structure released by neutrophils during the process of inflammatory responses, which contains histones and multiple granular proteins. Recent studies have demonstrated that DAMP and NET may aggravate IRI via aseptic inflammation. In this article, relevant studies of DAMP, NET and their relationship in IRI were reviewed, which was of great significance for understanding the pathophysiological mechanism of IRI and studying the corresponding prevention and treatment strategies.

Endogenous HMGB1 Modulates Fatty Acid Metabolism of Hypoxic and Nutrient-poor Pancreatic Cancer Cells and Related Mechanism / 肿瘤防治研究

Objective To explore the role and mechanism of HMGB1 in the fatty acid metabolism reprogramming and mitochondrial fusion/fission of hypoxic and nutrient-poor pancreatic cancer cells. Methods The correlation between the expression level of HMGB1 in pancreatic cancer tissue and the survival rate of pancreatic cancer patients were analyzed by GEPIA database. CCK-8 assay was used to measure cell proliferation rate, and scratch test and Transwell chamber method were carried out to detect the effects of endogenous HMGB1 on the invasion and migration abilities of human pancreatic cancer cell line Patu8988 after hypoxic and nutrient-poor treatment. Laser confocal microscope was used to observe the changes of mitochondrial morphology of Patu8988 cells. Western blot was used to detect the expression levels of mitochondrial fusion/fission and de novo fatty acid synthesis-related proteins. Results GEPIA database analysis results showed that HMGB1 was highly expressed in pancreatic cancer tissues (P < 0.01), and the expression level was negatively correlated with the survival time of pancreatic cancer patients (P=0.00097). Knockdown of HMGB1 expression could inhibit the proliferation, invasion and migration abilities of Patu8988 cells under hypoxic and nutrient-poor conditions. However, mitochondrial fission in patu8988 cells was increased. Knockdown of HMGB1 in Patu8988 cells increased the expression of fission-related protein FIS1 while decreased the expression of p-DRP1(Ser637) and fusion-related protein MFN1 and MFN2 in hypoxic and nutrient-poor environment; ACLY, p-ACLY and FASN protein expression levels were down-regulated. Conclusion Endogenous HMGB1 can promote the fusion and inhibit the fission of mitochondria in hypoxic and nutrient-poor Patu8988 cells, maintain mitochondrial morphology and function, and thereby up-regulate ACLY protein expression and phosphorylation level, promote FA synthesis, and maintain the proliferation, invasion and migration abilities of pancreatic cancer cells.

Glaucocalyxin A attenuates allergic responses by inhibiting mast cell degranulation through HMGB1/TLR4/NF-κB signaling pathways / 药学学报

The study is to investigate the effect of glaucocalyxin A (GLA) on mast cell-mediated anaphylaxis. The animal welfare and experimental process of this experiment followed the regulations of the Animal Ethics Committee of Yanbian University. BALB/c mice were used in the animal experiment and randomly divided into five groups, control group, model group, and GLA low, medium, and high dose groups (10, 20, and 40 mg·kg-1). Mice were sensitized by intradermal injection of anti-dinitrophenyl-immunoglobulin E (DNP-IgE) into the ears and challenged with a mixture of DNP-human serum albumin (HSA) and 4% evans blue into the tail veins to prepare an animal skin passive cutaneous anaphylaxis (PCA) model, which was collected from both ears for measurement of dye staining and histology. Rat peritoneal mast cells (RPMCs) were used in the cell experiment and divided into control, IgE + antigen (Ag), and IgE + Ag + GLA groups to determine histamine release as well as calcium influx levels. High-affinity IgE receptor (FcεRI)-mediated signaling pathway proteins and HMGB1/TLR4/NF-κB (high mobility group box 1/toll like receptor 4/nuclear transcription factor kappa B) signaling proteins were detected by Western blot. The results of animal experiments suggest that GLA inhibits PCA, reduces evans blue dye exudation, and reduces ear inflammation and ear thickness in mice. The results of cellular experiments suggested that GLA could reduce histamine release and calcium influx, and inhibit tumor necrosis factor-α (TNF-α), interleukin (IL)-4, IL-13, and IL-1β production; Western blot results showed that GLA inhibited FcεRI-mediated phosphorylation levels of spleen tyrosine kinase (Syk), Lck/Yes novel tyrosine kinase (Lyn), tyrosine kinase Fyn (Fyn), growth-factor receptor-bound protein 2 (Gab2), and phospholipase C (PLC) γ1, while GLA inhibited HMGB1/TLR4 signaling pathway to limit NF-κB p65 nuclear metastasis. The results indicate that GLA inhibits mast cell degranulation and attenuates allergic inflammation through the HMGB1/TLR4/NF-κB signaling pathway.

Effect and mechanism of glycyrrhizin on glial scar formation after spinal cord injury in rats / 中国修复重建外科杂志

Objective: To explore the effect and potential mechanism of glycyrrhizin (GL) by inhibiting high mobility group box 1 (HMGB1) on glial scar formation after spinal cord injury (SCI) in rats. Methods: Seventy-two female Sprague Dawley rats were randomly divided into sham group ( n=12), SCI model group (SCI group, n=36), GL intervention group (SCI+GL group, n=12), and nuclear factor κB (NF-κB) inhibitor [pynolidine dithiocarbamate (PDTC)] intervention group (SCI+PDTC group, n=12). The SCI models of SCI group, SCI+GL group, and SCI+PDTC group were made by modified Allen's method, the sham group was only exposed the spinal cord without any injury. First of all, Basso-Beattie-Bresnahan (BBB) score of hind limbs and slope test were performed in SCI group at 1, 2, and 3 weeks after operation; Western blot was used to detect the expressions of glial fibrillary acidic protein (GFAP) and HMGB1 proteins. Compared with the sham group, the most significant time point in the SCI group was selected for subsequent experiment, in which the most significant glial scar was formed. Then, behavioral tests (BBB score of hind limbs and slope test), histological observation of spinal cord tissue structure, Western blot detection of HMGB1, GFAP, and NF-κB proteins, and immunohistochemical staining observation of GFAP and chondroitin sulfate proteoglycan (CSPG) were used to explore the effect of GL on the formation of glial scar after SCI and its potential mechanism. Results: The BBB score and slope angle of the SCI group increased gradually with time, which were significantly lower than those of the sham group at each time point ( P<0.05). Western blot detection showed that the relative expressions of HMGB1 and GFAP proteins in the SCI group at 1, 2, and 3 weeks after operation were significantly higher than those in sham group ( P<0.05). The change was most obvious at 3 weeks after SCI, therefore the spinal cord tissue was selected for subsequent experiments at this time point. At 3 weeks after operation, compared with the SCI group, BBB score and slope angle of SCI+GL group significantly increased ( P<0.05); the relative expressions of HMGB1, GFAP, and NF-κB proteins detected by Western blot and the expressions of GFAP and CSPG proteins detected by immunohistochemical staining significantly decreased ( P<0.05); the disorder of spinal cord tissue by HE staining improved, inflammatory cell infiltration reduced, and glial scar formation decreased. At 3 weeks after operation, the expressions of NF-κB, GFAP, and CSPG proteins of the SCI+PDTC group significantly reduced when compared with the SCI group ( P<0.05); and the expression of NF-κB protein significantly decreased and the expressions of GFAP and CSPG proteins significantly increased when compared with the SCI+GL group ( P<0.05). Conclusion: After SCI in rats, the application of GL to inhibit the expression of HMGB1 can reduce the expression of GFAP and CSPG in the injured spinal cord, then reduce the formation of glial scars and promote the recovery of motor function of the hind limbs, and GL may play a role in inhibiting glial scar through HMGB1/NF-κB pathway.