Da Chengqitang Protects Intestinal Barrier in Septic Mice via Endogenous Antimicrobial Peptide mCRAMP / 中国实验方剂学杂志

ObjectiveMolecular docking and animal experiments were employed to explore the protective effect and mechanism of Da Chengqitang （DCQD） on intestinal barrier in septic mice. MethodText mining method was used to screen the active ingredients in DCQD. AutoDock Tools and Discovery Studio were used to study the interactions of active components with the core target proteins ［claudin-1， tumor necrosis factor （TNF）-α， interleukin （IL）-6， endogenous antimicrobial peptide mCRAMP， Toll-like receptor 4 （TLR4）， and myeloid differentiation primary response gene 88 （MyD88）］ in sepsis. Fifty C57BL/6 mice were randomized into sham， model， low- and high-dose （4 g∙kg-1 and 8 g∙kg-1） DCQD， and ulinastatin groups （n=10）. Before， during， and after the day of modeling surgery， each group was administrated with corresponding drugs. The mice in other groups except the model group were subjected to modeling by cecal ligation and puncture. Enzyme-linked immunosorbent assay （ELISA） was used measure the serum level of D-lactic acid to assess intestinal mucosa permeability. Hematoxylin-eosin staining was employed to observe the histopathological changes in the ileum and assess the intestinal mucosal damage and inflammatory infiltration. Western blotting was employed to determine the expression levels of tight junction proteins claudin-1 and occludin in the ileal tissue， which were indicative of the bowel barrier function. The TNF-α and IL-6 levels were measured by ELISA to assess the intestinal inflammation. The expression of mCRAMP in the ileal tissue was observed by immunohistochemistry. The mRNA levels of mCRAMP， TLR4， and MyD88 in mouse ileal tissue were determined by Real-time polymerase chain reaction， on the basis of which the mechanism of DCQD in protecting the intestinal barrier of septic mice was explored. ResultMolecular docking results showed that most of the 10 active ingredients of DCQD that were screened out by text mining could bind to sepsis targets by van der Waals force， hydrogen bonding， and other conjugated systems. The results of animal experiments showed that compared with the model group， low- or high-dose DCQD lowered the D-lactic acid level in the serum （P<0.01）， alleviated damage to the ileal tissue and mucosal edema， protected the small intestine villus integrity， reduced inflammatory cell infiltration， promoted the expression of claudin-1 （P<0.01）， lowered the IL-6 level （P<0.01）， up-regulated the mRNA and protein levels of mCRAMP （P<0.01）， and down-regulated the mRNA and protein levels of TLR4 and MyD88 （P<0.01） in the ileal tissue. In addition， high-dose DCQD lowered the TNF-α level and promoted the expression of occludin in the ileum tissue （P<0.01）， and low-dose DCQD up-regulated the protein level of occludin in the ileum tissue （P<0.05）. ConclusionDCQD has a protective effect on intestinal barrier in septic mice. It can reduce intestinal inflammation， repair intestinal mucosal damage， improve the tight junction protein level， and reduce intestinal mucosal permeability by up-regulating the mRNA and protein levels of mCRAMP and the down-regulating the expression of genes in the TLR4/MyD88 pathway.

Protective effect of Qingjie Huagong decoction on pancreatic tissue of mice with severe acute pancreatitis by regulating the NOD-like receptor protein 3/Toll-like receptor 4/nuclear factor-kappa B signaling pathway / 临床肝胆病杂志

ObjectiveTo investigate the therapeutic effect of Qingjie Huagong decoction （QJHGD） on a mouse model of severe acute pancreatitis （SAP） and the mechanism of action of QJHGD against inflammatory response. MethodsA total of 36 male C57BL/6J mice were randomly divided into blank group， model group， Western medicine group （ulinastatin）， and low-， middle-， and high-dose QJHGD groups， with 6 mice in each group. All mice except those in the blank group were given 5% sodium taurocholate by retrograde pancreaticobiliary injection to establish a model of SAP. After modeling， the mice in the low-， middle-， and high-dose groups were given QJHGD （1， 2， and 4 g/kg， respectively） by gavage， and those in the Western medicine group were given intraperitoneal injection of ulinastatin （5×104 U/kg）， for 7 days in total. HE staining was used to observe the histopathological changes of the pancreas； ELISA was used to measure the levels of α-amylase， lipase， interleukin-1β （IL-1β）， interleukin-6 （IL-6）， interleukin-8 （IL-8）， interleukin-18 （IL-18）， and tumor necrosis factor-α （TNF-α） in mice； RT-qPCR was used to measure the mRNA expression levels of NOD-like receptor protein3 （NLRP3）， Toll-like receptor 4 （TLR4）， and nuclear factor-kappa B （NF-κB） in pancreatic tissue； immunohistochemistry was used to measure the positive expression rates of NLRP3， TLR4， and NF-κB in pancreatic tissue； Western blot was used to measure the protein expression levels of NLRP3， TLR4， NF-κB， IL-1β， and IL-6. An analysis of variance was used for comparison of continuous data between multiple groups， and the least significant difference t-test was used for further comparison between two groups. ResultsCompared with the blank group， the model group had diffuse destruction of pancreatic tissue structure， focal dilatation of pancreatic lobular septum， pancreatic acinar atrophy， and massive inflammatory cell infiltration， as well as significant increases in the content of α-amylase， lipase， IL-1β， IL-6， IL-8， IL-18， and TNF-α （all P<0.05）， the mRNA expression levels and positive expression rates of NLRP3， TLR4， and NF-κB （all P<0.05）， and the protein expression levels of NLRP3， TLR4， NF-κB， IL-1β， and IL-6 （all P<0.05）. Compared with the model group， the low-， middle-， and high-dose QJHGD groups and the Western medicine group had slightly tighter and more intact structure of pancreatic tissue， ordered arrangement of pancreatic acinar cells， a small amount of inflammatory cell infiltration， and hemorrhagic foci of pancreatic lobules， as well as significant reductions in the content of α-amylase， lipase， IL-1β， IL-6， IL-8， IL-18， and TNF-α （all P<0.05）， the mRNA expression levels and positive expression rates of NLRP3， TLR4， and NF-κB （all P<0.05）， and the protein expression levels of NLRP3， TLR4， NF-κB， IL-1β， and IL-6 （all P<0.05）. ConclusionQJHGD may exert a protective effect on the pancreatic tissue of SAP mice by inhibiting the activation of NLRP3/TLR4/NF-κB signaling pathway-related proteins， reducing the release of inflammatory mediators， and preventing the enhancement of inflammatory cascade response.

Screening of adjuvant for PD-L1 vaccine based on nitrated T cell epitope / 中国药科大学学报

@#Abstract: To enhance the anti-tumor activity of tumor vaccine targeting PD-L1 based on the nitrated T-epitope (PD-L1-NitraTh), this research compared several adjuvants with different mechanisms to screen out the adjuvant most suitable for PD-L1-NitraTh. The results showed that Poly(I:C), CPG1018, swollen knotted polysaccharide SGP2 and GM-CSF could enhance the immunogenicity of PD-L1-NitraTh when used as adjuvants, with the Poly(I:C) group inducing the highest antibody titer. The results of qPCR for T cell differentiation-related cytokines showed that Poly(I:C) reduced the expression of GATA3 and FoxP3, indicating a strong effect on CD4+ T cell differentiation. Besides, compared with other adjuvants, Poly(I:C) could assist PD-L1-NitraTh to increase the infiltration of T cells as well as CD11b+ cells within tumor, suggesting that Poly(I:C) may be the suitable adjuvant for tumor vaccines based on the nitrated T epitopes.

MiR-145-5p targeting TLR4 participates in inflammation and oxidative stress of atherosclerosis foam cells / 中华老年心脑血管病杂志

Objective To explore the role of microRNA-145-5p(miR-145-5p)in the regulation of inflammatory response and oxidative stress process in cellular model of atherosclerosis.Methods Human monocytic leukemia THP-1 cells-derived foam cells were constructed in vitro.Then,the relative expression of miR-145-5p in the model and control groups of cells were detected by qRT-PCR.TargetScan database was used to predict the targeting relationship between miR-145-5p and Toll-like receptor 4(TLR4).The 293T cells were divided into wild-type+mimic group,wild-type+mimic negative control(NC)group,and mutant+mimic group,mutant+mimic NC group,and dual luciferase assay was employed to verify the targeting relationship of miR-145-5p and TLR4.Foam cells were cultured in vitro and divided into miR-145-5p mimic group,mimic NC group,miR-145-5p inhibitor group,and inhibitor NC group according to the corresponding treat-ments.The expression of TLR4 at mRNA and protein levels was detected by qRT-PCR and Wes-tern blotting.The contents of TNF-α,IL-1β and IL-6 were detected by ELISA.Biochemical reagent kits were applied for generation of reactive oxygen species(ROS),content of MDA and activity of SOD.Results The expression of miR-145-5p was significantly reduced in the model group than the control group(0.29±0.01 vs 1.00±0.08,t=11.180,P＜0.01).Dual luciferase assay showed that luciferase activity was significantly lower in the miR-145-5p mimic group than the mimic NC group(t=8.612,P＜0.01).Compared with the mimic NC group,the mimic group had obviously lower mRNA and protein levels of TLR4 and contents of TNF-α,IL-1β,lL-6,ROS and MDA,and higher miR-145-5p expression level and SOD activity(P＜0.05,P＜0.01).The treatment of inhibi-tor resulted in increased TLR4 mRNA and protein levels and TNF-α,IL-1β,IL-6,ROS and MDA contents,and decreased miR-145-5p expression and SOD activity when compared with the above levels in the inhibitor NC group(P＜0.05,P＜0.01).Conclusion MiR-145-5p inhibits inflamma-tion and oxidative stress in cellular model of atherosclerosis by targeting TLR4.

Role of PPARγ in Ulcerative Colitis and Traditional Chinese Medicine Intervention： A Review / 中国实验方剂学杂志

Ulcerative colitis （UC） is a chronic inflammatory bowel disease primarily affecting the colon and rectum， with the typical symptoms such as abdominal pain， bloody diarrhea， and tenesmus. The pathogenesis of UC remains to be fully elucidated. The disease is prone to recurrence， seriously affecting the patients' quality of life. Conventional therapies for UC have limitations， including unsatisfactory clinical efficacy， lengthy courses， and adverse reactions. Therefore， there is an urgent need to explore new therapeutic agents. Peroxisome proliferator-activated receptor gamma （PPARγ）， a ligand-dependent nuclear receptor protein that plays a crucial role in maintaining intestinal homeostasis， is closely associated with the onset and development of UC. Traditional Chinese medicine （TCM） has advantages such as multi-targeting and mild side effects in the treatment of UC. Recent studies have shown that TCM can exert the therapeutic effects on UC by modulating PPARγ. The TCM methods for regulating PPARγ include clearing heat， drying dampness， moving Qi， activating blood， resolving stasis， invigorating the spleen， warming the kidney， and treating with both tonification and elimination. On one hand， TCM directly activates PPARγ or mediates signaling pathways such as nuclear factor-κB （NF-κB）， mitogen-activated protein kinase （MAPK）， Toll-like receptor 4 （TLR4）， and regulates helper T cell 17 （Th17）/regulatory T cell （Treg） balance to promote macrophage polarization from the pro-inflammatory M1 phenotype to the anti-inflammatory M2 phenotype， thereby inhibiting intestinal inflammation. On the other hand， TCM regulates the intestinal metabolism to activate PPARγ， lower the nitrate level， and maintain local hypoxia. In this way， it can restore the balance between specialized anaerobes and facultative anaerobes， thereby improving the gut microbiota and treating UC. This article summarizes the role of PPARγ in UC and reviews the research progress of TCM in treating UC by intervening in PPARγ in the last five years， aiming to give insights into the treatment and new drug development for UC.

Traditional Chinese Medicine Intervention in Acute Pancreatitis Based on TLR4 Signaling Pathway： A Review / 中国实验方剂学杂志

Acute pancreatitis （AP） is one of the most clinically common acute digestive disorders characterized by quick onset，rapid progression，severe condition，and high mortality. If the disease is not timely intervened in the early stage，it can develop into severe AP in the later stage，which damages the long-term quality of life and brings serious economic burden to patients and their families. However， the pathogenesis of this disease is complex and has not been fully explained. The generation and development of AP is closely related to many signaling pathways. Among them，Toll-like receptor 4（TLR4），as a transmembrane signal transduction receptor，can mediate immune response and inflammatory response，and play a key role in the occurrence and development of AP. Traditional Chinese medicine（TCM）can regulate the TLR4 signaling pathway with multiple targets，multiple effects，and multiple administration methods to inhibit inflammatory response，and effectively intervene in the progression of AP， which has gradually become a new craze for preventing and treating AP. Many studies have shown that TCM has obvious advantages in the prevention and treatment of AP. It can effectively treat AP by regulating TLR4 signaling pathway，strengthening immune resistance and defense，and inhibiting inflammatory response. Despite of the research progress，there is still a lack of comprehensive review on TCM regulation of TLR4 signaling pathway in the treatment of AP. Therefore，the literature on TCM regulation of TLR4 signaling pathway published in recent years was systematically reviewed and elaborated，aiming to provide new ideas for the treatment of AP and further drug development.

Effect of Tongluo Juanbi Granules on Inflammatory Injury and Apoptosis of Osteoarthritis Based on TLR4/MyD88/NF-κB Signaling Pathway / 中国实验方剂学杂志

ObjectiveTo investigate the effects of Tongluo Juanbi granules on chondrocyte apoptosis and Toll-like receptor 4 （TLR4）/myeloid differentiation factor 88 （MyD88）/nuclear factor-κB （NF-κB） signaling pathway of rabbits with knee osteoarthritis （KOA） and study the mechanism of Tongluo Juanbi granules in the prevention and treatment of KOA. MethodThirty New Zealand rabbits were randomly assigned to the following five groups （n=6）： sham group， model group， low-dose and high-dose groups of Tongluo Juanbi granules （4.1 and 8.2 g·kg-1·d-1）， and celecoxib group （10.9 mg·kg-1·d-1）. The KOA model was established by destabilization of the medial meniscus （DMM） for six weeks. Six weeks after the modeling， the drug was given once a day for eight weeks. The pathological changes of cartilago articularis were observed by hematoxylin-eosin （HE） staining and Safranin O-Fast Green staining. Terminal deoxynucleotidyl transferase dUTP nick end labeling （TUNEL） staining was performed to detect chondrocyte apoptosis. Enzyme-linked immunosorbent assay （ELISA） was used to detect the contents of interleukin-1β （IL-1β） and tumor necrosis factor-α （TNF-α） in synovial fluid. The mRNA and protein expression levels of genes related to the TLR4/MyD88/NF-κB signaling pathway were detected by real-time fluorescence quantitative polymerase chain reaction （Real-time PCR） and Western blot， respectively. ResultCompared with the sham group， the cartilago articularis of the model group significantly degenerated. Mankin's score was increased （P<0.01）， and the contents of IL-1β and TNF-α in synovial fluid were increased （P<0.01）. The number of apoptosis of chondrocytes was increased （P<0.01）. The mRNA and protein expressions of TLR4， MyD88， and NF-κB p65 in cartilage tissue were up-regulated （P<0.01）， while the mRNA and protein expressions of Bcl-2 were down-regulated （P<0.01）. Compared with the model group， chondrocyte degeneration in both low-dose and high-dose groups of Tongluo Juanbi granules was improved， and Mankin's score was decreased （P<0.01）. The contents of IL-1β and TNF-α were decreased （P<0.01）， and the number of apoptosis of chondrocytes was decreased （P<0.01）. The mRNA and protein expressions of TLR4， MyD88， and NF-κB p65 in cartilage tissue were down-regulated （P<0.01）， while the mRNA and protein expressions of Bcl-2 were up-regulated （P<0.01）. In addition， in the above observation indicators， the high-dose group of Tongluo Juanbi granules was significantly superior to the low-dose group of Tongluo Juanbi granules. ConclusionTongluo Juanbi granules could inhibit chondrocyte apoptosis in rabbits with KOA and improve cartilage degeneration， which may be related to inhibiting inflammatory responses mediated by TLR4/MyD88/NF-κB signaling pathway.

Traditional Chinese Medicine in Treatment of Parkinson's Disease by Regulating TLR4/NF-κB Signaling Pathway： A Review / 中国实验方剂学杂志

Parkinson's disease （PD） is a common neurological degenerative disease in the middle-aged and elderly， characterized by pathological changes of progressive degeneration of dopaminergic neurons in the substantia nigra and Lewy body formation， with high prevalence and long course of disease. The drug is mainly used to treat PD in western medicine， and the early curative effect is remarkable. However， with the progression of the disease and the long-term use of the drug， the efficacy will be significantly reduced， or there may be sports complications， and the long-term efficacy is not good. As a traditional medical system， traditional Chinese medicine has a unique understanding of PD. Traditional Chinese medicine plays an important role in the treatment of PD， which is natural， mild， safe， and effective， and it can cooperate with western medicine to enhance its efficacy and reduce the adverse reactions of western medicine. The pathogenesis of PD is complex， involving multiple levels such as mitochondrial dysfunction and apoptosis. Neuroinflammation is also involved in the progressive degeneration of dopaminergic neurons in PD. The Toll-like receptor 4 （TLR4）/nuclear factor-κB （NF-κB） signaling pathway is a classic inflammatory pathway， and its expression changes play an important role in the occurrence and development of inflammatory response in the body. In recent years， the research on this pathway in TCM is increasing. This paper summarized the literature of traditional Chinese and western medicine in the past 10 years and reviewed the relevant mechanism of TCM regulation of TLR4/NF-κB pathway in the treatment of PD from the aspects of TCM monomer， compound， and other TCM therapies， so as to provide some references for the search for new targets of drug therapy and gene therapy and the in-depth study of TCM prevention and treatment of PD.

Regulation of Toll-like receptors on function of osteoblasts and osteoclasts / 中国组织工程研究

BACKGROUND:Toll-like receptors are an important class of pattern recognition receptors that have important functions in pathogen immunity and cytokine synthesis by recognizing specific molecular patterns.Previous studies have found that different types of bone tissue cells also express Toll-like receptors.Activation or inhibition of Toll-like receptors can have significant effects on osteoblast and osteoclast function through multiple pathways. OBJECTIVE:To summarize the expression and action pathways of Toll-like receptors in osteoblasts and osteoclasts,in order to further elucidate the biological mechanisms involved in the regulation of Toll-like receptors under physiological and pathological conditions. METHODS:Relevant literature was retrieved from databases such as PubMed and CNKI as of December 2022.The Chinese and English search terms included"Toll-like receptor,osteoblast,osteoclast,mesenchymal stem cells,macrophage,cytokine,signaling pathway".According to the research needs,the corresponding criteria were established to screen the final literature. RESULTS AND CONCLUSION:(1)Toll-like receptors could directly regulate osteoblast and osteoclast differentiation through the activation of related signaling pathways.(2)Toll-like receptor activation induces cytokine production and exerts regulatory effects.(3)Toll-like receptor activation can affect the survival and migration ability of osteoblasts and osteoclasts.(4)Toll-like receptors in osteoblasts and osteoclasts are activated in certain diseases and pathological settings and are involved in intercellular interactions.

Electroacupuncture reduces inflammatory factor expression by suppressing Toll-like receptor 4/nuclear factor-kappa B signaling in rats with cerebral ischemia-reperfusion injury / 中国组织工程研究

BACKGROUND:Inflammation is one of the important factors that induce cerebral ischemia-reperfusion injury.Studies have shown that electroacupuncture can effectively reduce inflammation after ischemic stroke and improve the symptoms of neurological deficits,but the mechanism is not clear. OBJECTIVE:To observe the effect of electroacupuncture on Toll-like receptor 4/nuclear factor-κB in rats with cerebral ischemia-reperfusion injury. METHODS:Forty-eight male Sprague-Dawley rats were randomly divided into sham operation group,model group and electroacupuncture group,with 16 rats in each group.The rat model of cerebral ischemia-reperfusion injury was prepared by middle cerebral artery occlusion.At 24 hours after modeling,the rats in the electroacupuncture group were treated with electroacupuncture,once a day,20 minutes each time,for a total of 5 days.The sham operation group and the model group did not do any intervention.After 5 days of intervention,Longa method was used to evaluate the degree of neurological injury in rats.Triphenyl tetrazolium chloride staining and hematoxylin-eosin staining were used to measure the volume of cerebral infarction and the pathological changes of brain tissue in rats.Serum interleukin-6,interleukin-18 and tumor necrosis factor-α were detected by ELISA.Expressions of Toll-like receptor 4 and nuclear factor-κB in the cerebral cortex at mRNA and protein levels were detected by fluorescence quantitative PCR and western blot,respectively. RESULTS AND CONCLUSION:Compared with the sham operation group,the neurological function scores,serum interleukin-6,interleukin-18,and tumor necrosis factor-α levels,Toll-like receptor 4 and nuclear factor-κB mRNA and protein expression levels were significantly higher in the model group(P＜0.01).Compared with the model group,electroacupuncture significantly reduced the neurological function scores,serum interleukin-6,interleukin-18,and tumor necrosis factor-α levels,Toll-like receptor 4 and nuclear factor-κB mRNA and protein expression levels(P＜0.05,P＜0.01).Compared with the sham operation group,the volume of cerebral infarction in the model group increased significantly(P＜0.01).Compared with the model group,the volume of cerebral infarction in the electroacupuncture group decreased(P＜0.05).In the model group,the arrangement of neurons was disordered,some nerve cells disappeared,nuclei presented with pyknosis and incomplete structure.After electroacupuncture intervention,the degree of neuronal degeneration and neuronal loss in the cerebral cortex of rats were reduced compared with those in the model group.To conclude,electroacupuncture can significantly improve the neurobehavior of rats with cerebral ischemia-reperfusion injury,reduce brain tissue injury,and effectively reduce the level of serum inflammatory factors.The mechanism may be related to the inhibition of Toll-like receptor 4/nuclear factor-κB signaling pathway.

Clinical effect of procaterol hydrochloride combined with Xiaokechuan capsule in the treatment of cough variant asthma and its impact on serological indicators and airway function of children / 新乡医学院学报

Objective To explore the clinical effect of procaterol hydrochloride combined with Xiaokechuan capsule in the treatment of cough variant asthma(CVA)and its impact on serological indicators,airway function of children.Methods A total of 124 children with CVA admitted to the Zigong First People's Hospital from March 2019 to April 2022 were selected as the research subjects.The children were divided into control group and observation group according to random number table method,with 62 cases in each group.The children in the control group were treated with procaterol hydrochloride,and the children in the observation group were treated with procaterol hydrochloride and Xiaokechuan capsule for two weeks.The clinical efficacy of children was compared between the two groups after treatment.The cough scores during the day and night of children were evaluated in the two groups before and 2 weeks after treatment.The serum high mobility group protein B1(HMGB1),Toll like receptor 4(TLR4),nuclear factor-κB(NF-κB),interleukin-4(IL-4),interferon-γ(INF-γ)levels of children in the two groups were measured by enzyme linked immunosorbent assay before and 2 weeks after treatment,and the ratio of INF-γ/IL-was calculated.The 25％maximal expiratory flow-volume(MEF25),50％maximal expiratory flow-volume(MEF50),75％maximal expiratory flow-volume(MEF75)of children in the two groups were measured by lung function detector before and 2 weeks after treatment.The adverse reactions of children in the two groups were recorded during treatment.Results The total effective rate of children in the control group and observation group was 82.26％(51/62)and 95.16％(59/62),respectively;the total effective rate of children in the observation group was significantly higher than that in the control group(P＜0.05).There was no significant difference in cough scores during the day and night of children between the two groups before treatment(P＞0.05);the cough scores during the day and night of children after treatment were significantly lower than those before treatment in the two groups(P＜0.05);after treatment,the cough scores during the day and night of children in the observation group were significantly lower than those in the control group(P＜0.05).There was no significant difference in serum HMGB1,TLR4,NF-κB levels of children between the two groups before treatment(P＞0.05);the serum HMGB1,TLR4,NF-κB levels of children after treatment were significantly lower than those before treatment in the two groups(P＜0.05);after treatment,the serum levels of HMGB1,TLR4,and NF-κB of children in the observation group were significantly lower than those in the control group(P＜0.05).There was no significant difference in MEF25,MEF50,and MEF75 of children between the two groups before treatment(P＞0.05);the MEF25,MEF50,and MEF75 of children after treatment were significantly higher than those before treatment in the two groups(P＜0.05);after treatment,the MEF25,MEF50,and MEF75 of children in the observation group were significantly higher than those in the control group(P＜0.05).There was no significant difference in serum IL-4,INF-γ levels and the ratio of INF-γ/IL-4 of children between the two groups before treatment(P＞0.05);the serum IL-4 level of children after treatment were significantly lower than those before treatment,the INF-γ level and the ratio of INF-γ/IL-4 were significantly higher than those before treatment in the two groups(P＜0.05);after treatment,the serum IL-4 level of children in the observation group was significantly lower than that in the control group,the INF-γ level and the ratio of INF-γ/IL-4 were significantly higher than those in the control group(P＜0.05).All children had good drug tolerance during the treatment period,and no significant adverse drug reactions were observed.Conclusion The combination of Xiaokechuan capsules and procaterol hydrochloride has a significant therapeutic effect for pediatric CVA,and its mechanism of action may be related to the regulation of HMGB1-TLR4-NF-κB signal pathway.

Mechanism of electroacupuncture on improving corneal inflammation in dia-betic dry eye rats by regulating Toll-like receptor 4-mediated inflammatory signaling pathway / 眼科新进展

Objective To observe the effect of electroacupuncture on corneal Toll-like receptor 4(TLR4)-mediated inflammatory signaling pathway in diabetic dry eye rats and to explore the mechanism of electroacupuncture in the treat-ment of diabetic dry eye.Methods A type 2 diabetic rat model was established in 32 healthy male Sprague-Dawley(SD)rats by intraperitoneal injection of streptozotocin(30 mg·kg-1)for 12 weeks after feeding with high-sugar and high-fat di-et for 4 weeks.Twenty-five successfully modeled diabetic dry eye rats were randomly divided into the model group(non-in-tervention),electroacupuncture group(the"Jingming","Cuanzhu","Sizhukong","Taiyang"and"Tongziliao"acupoints were treated with acupuncture,and then"Cuanzhu"and"Tongzilao"acupoints were treated with electroacupuncture,15 min for each time,once a day),sham acupuncture group(blunt-tip needle pricking was performed at the same acupoints as the electroacupuncture group),and fluorometholone group(1 g·L-1fluorometholone eye drops were used in both eyes at 8 o'clock,13 o'clock,and 18 o'clock,1 drop each time),with 6 rats in each group,lasting for 2 weeks.Another 6 healthy male SD rats were selected as a blank group.Random blood glucose,tear film breakup time(BUT),tear secre-tion,corneal fluorescein staining(FL)score,and corneal touch threshold(CTT)of rats in each group were detected be-fore modeling,after modeling,and after the intervention.Hematoxylin and Eosin(HE)staining was performed to observe the corneal morphologic changes in each group.Immunofluorescence histochemical staining was adopted to detect the cor-neal TLR4-positive expression in each group.The TLR4,phosphorylated nuclear factor-kappa P65(P-NF-KB P65),inter-leukin(IL)-1β,and IL-18 expression levels in the cornea were detected by Western blot.Results After modeling,com-pared with the blank group,BUT,tear secretion and CTT decreased and FL increased in all experimental groups,and the differences were statistically significant(all P＜0.01).After the intervention,compared with the model group,FL de-creased,and BUT,tear secretion and CTT increased in the electroacupuncture group,and the differences were statistically significant(all P＜0.05).Corneal HE staining showed that after the intervention,the corneal surface of rats in the model group and sham acupuncture group was not smooth,and the corneal epithelial cells were thickened and disorganized;the corneal surface of rats in the electroacupuncture group and fluorometholone group was smooth,and the corneal epithelial cells were arranged neatly.After the intervention,compared with the blank group,the corneal TLR4 expression of rats in all other groups was elevated,and the differences were statistically significant(all P＜0.05);compared with the model group,the corneal TLR4 expression of rats in the electroacupuncture group and fluorometholone group was reduced(both P＜0.01).Compared with the blank group,corneal TLR4,P-NF-κBP65,IL-1β and IL-18 protein expressions increased in the model group and sham acupuncture group(all P＜0.05);compared with the model group,these expressions decreased in the electroacupuncture group and the fluorometholone group(all P＜0.05).Conclusion Electroacupuncture can im-prove the ocular surface signs and inhibit the expressions of TLR4,P-NF-κB P65,IL-1 β,and IL-18 in the cornea of type 2 diabetic dry eye rats,and its mechanism of action may be related to the regulation of TLR4-mediated inflammatory signaling pathway,which inhibits ocular surface inflammation in diabetic dry eye rats.

The role of innate immunity in diabetic nephropathy and their therapeutic consequences / 药物分析学报

Diabetic nephropathy(DN)is an enduring condition that leads to inflammation and affects a substantial number of individuals with diabetes worldwide.A gradual reduction in glomerular filtration and emergence of proteins in the urine are typical aspects of DN,ultimately resulting in renal failure.Mounting evidence suggests that immunological and inflammatory factors are crucial for the develop-ment of DN.Therefore,the activation of innate immunity by resident renal and immune cells is critical for initiating and perpetuating inflammation.Toll-like receptors(TLRs)are an important group of re-ceptors that identify patterns and activate immune responses and inflammation.Meanwhile,inflam-matory responses in the liver,pancreatic islets,and kidneys involve inflammasomes and chemokines that generate pro-inflammatory cytokines.Moreover,the activation of the complement cascade can be triggered by glycated proteins.This review highlights recent findings elucidating how the innate immune system contributes to tissue fibrosis and organ dysfunction,ultimately leading to renal failure.This re-view also discusses innovative approaches that can be utilized to modulate the innate immune responses in DN for therapeutic purposes.

Tectorigenin improves cognitive deficits in rats with vascular dementia by regulating TLR4/MyD88/NF-κB signaling pathway / 中国免疫学杂志

Objective:To analyze effects of tectorigenin on improving cognitive deficits in rats with vascular dementia(VD)by regulating Toll-like receptor 4(TLR4)/myeloid differentiation factor 88(MyD88)/nuclear factor-κB(NF-κB)signaling pathway.Methods:A total of 72 rats were randomly divided into sham operation group,model group,low,medium and high doses[25,50,100 mg/(kg·d)]tectorigenin groups and positive control group[piracetam 324 mg/(kg·d)],with 12 rats in each group.Except for sham operation group,VD models were replicated in other groups.After successful modeling,different doses tectorigenin groups and positive control group were administered intragastrically with different doses of tectorigenin and piracetam,while other groups were administered intragastrically with same volume of normal saline for 28 d.Spatial learning and memory ability were detected by Morris water maze.Neurotransmitter levels in hippocampus interstitial fluid were detected by high performance liquid chromatography-electro-chemical.Brain-derived neurotrophic factor(BDNF)and tyrosine kinase receptor b(TrkB)expressions in hippocampus were detected by RT-qPCR and Western blot.TLR4/MyD88/NF-κB pathway-related proteins in hippocampus were detected by Western blot.Results:Compared with sham operation group,escape latency was longer,while stay time in target area and times of crossing platform were lower in model group(P<0.05).Compared with model group,escape latency was shorter,while stay time in target area and times of crossing platform were higher in medium and high doses tectorigenin groups(P<0.05).NE,DA,5-HT and 5-HIAA levels in model group were lower than those in sham operation group(P<0.05),which were higher in medium and high doses tectorigenin groups than model group(P<0.05).Compared with sham operation group,BDNF and TrkB mRNA and proteins levels were lower,while TLR4,MyD88 and p-NF-κB p65/NF-κB p65 proteins levels were higher in model group(P<0.05).Compared with model group,BDNF and TrkB mRNA and proteins levels were higher,while TLR4,MyD88 and p-NF-κB p65/NF-κB p65 proteins levels were lower in medium and high doses tectorigenin groups(P<0.05).Conclusion:Tectorigenin can improve cognitive deficits in VD rats,which may be related to regulating TLR4/MyD88/NF-κB signaling pathway.

The influence of Liuwei Dihuang Wan on the behavior and TLR4/NF-κB signaling pathway of Alzheimer's disease model mice / 中华行为医学与脑科学杂志

Objective:To explore the effects of Liuwei Dihuang Wan on the behaviors and Toll-like receptor 4/nuclear factor kappa-B(TLR4/NF-κB) signal transduction pathway of amyloid β-precursor protein/presenilin-1(APP/PS1) double transgenic mice.Methods:Forty 3-month-old female APP/PS1 mice were randomly divided into model group, low-dose group(0.59 g/kg), medium-dose group(1.18 g/kg), high-dose group(2.36 g/kg)of Liuwei Dihuang Wan(gavaged according to grouped doses), and ibuprofen group(0.04 g/kg, gavage) using a random number table method, with 8 mice in each group.Eight 3-month-old wild-type female C57BL/6 mice with matched body weight were used as the control group.The mice in control group and model group were given an equal volume of 0.9% sodium chloride solution by gavage.The gavage administration was twice a day for a continuous period of 3 months.Morris water maze test was used to detect the learning and memory abilities of mice. ELISA was used to detect the serum levels of tumor necrosis factor-α (TNF-α), interleukin-1β(IL-1β) and immunohistochemistry was used to detect the levels of amyloid β-protein (Aβ), glial fibrillary acidic protein(GFAP) and NF-κB in hippocampal tissue.Western blot was used to detect the expression levels of TLR4, myeloid differentiation primary response gene 88(MyD88), and phosphorylated NF-κB(p-NF-κB) proteins in hippocampal tissue.The SPSS 20.0 software was used for data analysis. Multiple group comparisons were conducted by repeated measure ANOVA or one-way ANOVA.Results:The results of repeated measure ANOVA showed that as for the escape latency of the 6 groups of mice, the interaction effect between time and group was significant ( Finteraction=117.219, P<0.001). The escape latencies of mice in the 6 groups on the 5th day were all lower than those on the 1st day (all P<0.05). The escape latencies of mice in the ibuprofen group and the medium-dose and high-dose groups of Liuwei Dihuang Wan were lower than that in the model group from 1st day to 5th day(all P<0.05). On the 3rd to 5th day, the escape latencies of mice in the medium-dose and high-dose groups of Liuwei Dihuang Wan were lower than those in the low-dose group of Liuwei Dihuang Wan (all P<0.05). There were statistically significant differences in the percentage of residence time in the platform quadrant and the numbers of crossing platform among the 6 groups of mice ( F=5.451, 4.824, both P<0.05). The percentage of residence time in the platform quadrant (50.77±5.49)%, (54.39±5.71)%, (51.98±6.12)%), and the numbers of crossing platform((5.9±1.1) times, (6.0±1.3) times, (5.1±0.8) times) in the high-dose and medium-dose groups of Liuwei Dihuang Wan and the ibuprofen group were all higher than those in the model group ((27.32±3.22)%, (2.2±1.0) times )(all P<0.05). The immunohistochemical results showed that there were statistically significant differences in the integrated optical density values of Aβ, GFAP and NF-κB in the hippocampal tissues of 6 groups of mice ( F=57.52, 45.37, 79.10, all P<0.05). The integrated optical density values of Aβ, GFAP and NF-κB in the high-dose and medium-dose groups of Liuwei Dihuang Wan and the ibuprofen group were all lower than those in the model group (all P<0.05). And the integrated optical density values of Aβ, GFAP, and NF-κB in the high-dose and medium-dose groups of Liuwei Dihuang Wan were all lower than those in the low-dose group of Liuwei Dihuang Wan (all P<0.05). There were statistically significant differences in the levels of serum TNF-α and IL-1β detected by ELISA ( F=3.996, 6.395, both P<0.05) and the proteins levels of TLR4, MyD88, and p-NF-κB in hippocampal tissue detected by Western blot among the 6 groups( F=15.710, 3.522, 4.119, all P<0.05). The serum TNF-α and IL-1β levels in the high-dose and medium-dose groups of Liuwei Dihuang Wan and ibuprofen group were all lower than those in the model group (all P<0.05). The serum TNF-α ((18.90±2.33) ng/L, (21.56±2.49) ng/L) and IL-1β ((5.88±0.80) ng/L, (6.75±0.83) ng/L) levels in the high-dose and medium-dose groups of Liuwei Dihuang Wan were lower than those in the low-dose group ((30.77±2.89) ng/L, (9.11±1.27) ng/L) (all P<0.05). The protein expression levels of TLR4, MyD88, and p-NF-κB in the hippocampus of the high-dose and medium-dose groups of Liuwei Dihuang Wan and ibuprofen group were lower than those of the model group (all P<0.05). The protein expression levels of TLR4 ((0.254±0.091), (0.318±0.122)), MyD88 ((0.229±0.077), (0.386±0.119)), and p-NF-κB ((0.412±0.188), (0.358±0.119)) in the hippocampus of the high-dose and medium-dose groups of Liuwei Dihuang Wan were lower than those of the low-dose group ((0.617±0.172), (0.672±0.166), (0.799±0.227)) (all P<0.05). Conclusion:Liuwei Dihuang Wan can significantly alleviate learning and memory impairment in Alzheimer's disease model mice, possibly by inhibiting TLR4/NF-κB signal pathway, reducing TNF-α and IL-1β expression, thereby alleviate central immune inflammatory response and exert anti Alzheimer's disease effects.

Analysis of drug resistance of pathogens of puerperal infection in pregnant women with diabetes and its influence on toll like receptor 4 inflammatory pathway of peripheral blood mononuclear cells / 中国医师杂志

Objective:To explore the drug resistance of pathogens in puerperal infection of pregnant women with diabetes mellitus (GDM), and analyze the influence of puerperal infection on the expression of toll like receptor 4 (TLR4) inflammatory pathway in peripheral blood monocytes.Methods:A retrospective selection was conducted on 120 GDM postpartum women who underwent regular prenatal check ups and delivery at the 903th Hospital of the PLA (People′s Liberation Army) Joint Logistic Support Force from January 2020 to October 2022. The postpartum infection status, pathogenic characteristics of the infected pathogens, and drug resistance of the mothers were analyzed; According to the postpartum infection situation, the parturients were divided into an infected group and an uninfected group. Logistic regression analysis was used to analyze the relevant factors affecting postpartum infection, and the TLR4 protein and mRNA expression levels of peripheral blood mononuclear cells in the two groups were compared.Results:Among 120 GDM pregnant women, 21 cases (17.50%) developed post infection, including 8 cases (38.10%) of incision infection, 6 cases (28.57%) of uterine cavity infection, 4 cases (19.05%) of urinary system infection, and 3 cases (14.28%) of blood infection; A total of 43 pathogenic bacteria were detected, including 26 Gram negative bacteria (60.46%), 14 Gram positive bacteria (32.56%), and 3 fungi (6.98%). Among the main Gram negative bacteria, escherichia coli had the highest resistance rate to ceftazidime and tetracycline, and had not developed resistance to meropenem; Pseudomonas aeruginosa had the highest resistance rate to ceftazidime and gentamicin. Among the main Gram positive bacteria, staphylococcus aureus had the highest resistance rate to penicillin G and ceftazidime, and had not developed resistance to vancomycin; Enterococcus faecalis had the highest resistance rate to clindamycin. The results of multivariate logistic regression analysis showed that postpartum hemorrhage, premature rupture of membranes, and poor control of prenatal blood sugar were independent risk factors for postpartum infection in GDM mothers (all P<0.05). The expression rate of TLR4 protein, relative expression level of TLR4 mRNA, and levels of tumor necrosis factor-α (TNF-α), interleukin (IL)-1, and IL-10 in the infected group were significantly higher than those in the non infected group (all P<0.05). Conclusions:The distribution and drug resistance of pathogenic bacteria in postpartum infections of GDM mothers have certain characteristics. Postpartum hemorrhage, premature rupture of membranes, and poor control of prenatal blood sugar are independent risk factors affecting postpartum infections in GDM mothers; The TLR4 inflammatory pathway in peripheral blood mononuclear cells may be involved in the occurrence and development of postpartum infection in GDM mothers.

Effect of oxymatrine on Cryptosporidium parvum infection in mice based on the HMGB1-TLR2/TLR4-NF-κB pathway / 中国血吸虫病防治杂志

Objective To investigate the involvement of the high mobility group box protein B1 (HMGB1)-Toll-like receptor 2 (TLR2)/TLR4-nuclear factor κB (NF-κB) pathway in the intestinal mucosal injury induced by Cryptosporidium parvum infection, and to examine the effect of oxymatrine (OMT) on C. parvum infection in mice. Methods Forty SPF 4-week-old BALB/c mice were randomly divided into four groups, including the control group, infection group, glycyrrhizin (GA) group and OMT group. Each mouse was orally administered with 1 × 105 C. parvum oocysts one week in the infection, GA and OMT groups following dexamethasone-induced immunosuppression to model C. parvum intestinal infections in mice. Upon successful modeling, mice in the GA group were intraperitoneally injected with GA at a daily dose of 25.9 mL/kg for successive two weeks, and animals in the OMT group were orally administered OMT at a daily dose of 50 mg/kg for successive two weeks, while mice in the control group were given normal food and water. All mice were sacrificed two weeks post-treatment, and proximal jejunal tissues were sampled. The pathological changes of mouse intestinal mucosal specimens were observed using hematoxylin-eosin (HE) staining, and the mouse intestinal villous height, intestinal crypt depth and the ratio of intestinal villous height to intestinal crypt depth were measured. The occludin and zonula occludens protein 1 (ZO1) expression was determined in mouse intestinal epithelial cells using immunohistochemistry, and the relative expression of HMGB1, TLR2, TLR4, myeloid differentiation primary response gene 88 (MyD88) and NF-κB p65 mRNA was quantified in mouse jejunal tissues using quantitative real-time PCR (qPCR) assay. Results HE staining showed that the mouse intestinal villi were obviously atrophic, shortened, and detached, and the submucosal layer of the mouse intestine was edematous in the infection group as compared with the control group, while the mouse intestinal villi tended to be structurally intact and neatly arranged in the GA and OMT groups. There were significant differences among the four groups in terms of the mouse intestinal villous height (F = 6.207, P = 0.000 5), intestinal crypt depth (F = 6.903, P = 0.000 3) and the ratio of intestinal villous height to intestinal crypt depth (F = 37.190, P < 0.000 1). The mouse intestinal villous height was lower in the infection group than in the control group [(321.9 ± 41.1) μm vs. (399.5 ± 30.9) μm; t = 4.178, P < 0.01] and the GA group [(321.9 ± 41.1) μm vs. (383.7 ± 42.7) μm; t = 3.130, P < 0.01], and the mouse intestinal crypt depth was greater in the infection group [(185.0 ± 35.9) μm] than in the control group [(128.4 ± 23.6) μm] (t = 3.877, P < 0.01) and GA group [(143.3 ± 24.7) μm] (t = 2.710, P < 0.05). The mouse intestinal villous height was greater in the OMT group [(375.3 ± 22.9) μm] than in the infection group (t = 3.888, P < 0.01), and there was no significant difference in mouse intestinal villous height between the OMT group and the control group (t = 1.989, P > 0.05). The mouse intestinal crypt depth was significantly lower in the OMT group [(121.5 ± 27.3) μm] than in the infection group (t = 4.133, P < 0.01), and there was no significant difference in mouse intestinal crypt depth between the OMT group and the control group (t = 0.575, P > 0.05). The ratio of the mouse intestinal villous height to intestinal crypt depth was significantly lower in the infection group (1.8 ± 0.2) than in the control group (3.1 ± 0.3) (t = 10.540, P < 0.01) and the GA group (2.7 ± 0.3) (t = 7.370, P < 0.01), and the ratio of the mouse intestinal villous height to intestinal crypt depth was significantly higher in the OMT group (3.1 ± 0.2) than in the infection group (t = 15.020, P < 0.01); however, there was no significant difference in the ratio of the mouse intestinal villous height to intestinal crypt depth between the OMT group and the control group (t = 0.404, P > 0.05). Immunohistochemical staining showed significant differences among the four groups in terms of occludin (F = 28.031, P < 0.000 1) and ZO1 expression (F = 14.122, P < 0.000 1) in mouse intestinal epithelial cells. The proportion of positive occluding expression was significantly lower in mouse intestinal epithelial cells in the infection group than in the control group [(14.3 ± 4.5)% vs. (28.3 ± 0.5)%; t = 3.810, P < 0.01], and the proportions of positive occluding expression were significantly higher in mouse intestinal epithelial cells in the GA group [(30.3 ± 1.3)%] and OMT group [(25.8 ± 1.5)%] than in the infection group (t = 7.620 and 5.391, both P values < 0.01); however, there was no significant differences in the proportion of positive occluding expression in mouse intestinal epithelial cells between the GA or OMT groups and the control group (t = 1.791 and 2.033, both P values > 0.05). The proportion of positive ZO1 expression was significantly lower in mouse intestinal epithelial cells in the infection group than in the control group [(14.4 ± 1.8)% vs. (24.2 ± 2.8)%; t = 4.485, P < 0.01], and the proportions of positive ZO1 expression were significantly higher in mouse intestinal epithelial cells in the GA group [(24.1 ± 2.3)%] (t = 5.159, P < 0.01) and OMT group than in the infection group [(22.5 ± 1.9)%] (t = 4.441, P < 0.05); however, there were no significant differences in the proportion of positive ZO1 expression in mouse intestinal epithelial cells between the GA or OMT groups and the control group (t = 0.037 and 0.742, both P values > 0.05). qPCR assay showed significant differences among the four groups in terms of HMGB1 (F = 21.980, P < 0.000 1), TLR2 (F = 20.630, P < 0.000 1), TLR4 (F = 17.000, P = 0.000 6), MyD88 (F = 8.907, P = 0.000 5) and NF-κB p65 mRNA expression in mouse jejunal tissues (F = 8.889, P = 0.000 7). The relative expression of HMGB1 [(5.97 ± 1.07) vs. (1.05 ± 0.07); t = 6.482, P < 0.05] 、TLR2 [(5.92 ± 1.29) vs. (1.10 ± 0.14); t = 5.272, P < 0.05] 、TLR4 [(5.96 ± 1.50) vs. (1.02 ± 0.03); t = 4.644, P < 0.05] 、MyD88 [(3.00 ± 1.26) vs. (1.02 ± 0.05); t = 2.734, P < 0.05] and NF-κB p65 mRNA [(2.33 ± 0.72) vs. (1.04 ± 0.06); t = 2.665, P < 0.05] was all significantly higher in mouse jejunal tissues in the infection group than in the control group. A significant reduction was detected in the relative expression of HMGB1 (0.63 ± 0.01), TLR2 (0.42 ± 0.10), TLR4 (0.35 ± 0.07), MyD88 (0.70 ± 0.11) and NF-κB p65 mRNA (0.75 ± 0.01) in mouse jejunal tissues in the GA group relative to the control group (t = 8.629, 5.830, 11.500, 4.729 and 6.898, all P values < 0.05), and the relative expression of HMGB1, TLR2, TLR4, MyD88 and NF-κB p65 mRNA significantly reduced in mouse jejunal tissues in the GA group as compared to the infection group (t = 7.052, 6.035, 4.084, 3.165 and 3.274, all P values < 0.05). In addition, the relative expression of HMGB1 (1.14 ± 0.60), TLR2 (1.00 ± 0.24), TLR4 (1.14 ± 0.07), MyD88 (0.96 ± 0.25) and NF-κ B p65 mRNA (1.12 ± 0.17) was significantly lower in mouse jejunal tissues in the OMT group than in the infection group (t = 7.059, 5.320, 3.510, 3.466 and 3.273, all P values < 0.05); however, there were no significant differences between the OMT and control groups in terms of relative expression of HMGB1, TLR2, TLR4, MyD88 or NF-κB p65 mRNA in mouse jejunal tissues (t = 0.239, 0.518, 1.887, 0.427 and 0.641, all P values > 0.05). Conclusions C. parvum infection causes intestinal inflammatory responses and destruction of intestinal mucosal barrier through up-regulating of the HMGB1-TLR2/TLR4-NF-κB pathway. OMT may suppress the intestinal inflammation and repair the intestinal mucosal barrier through inhibiting the activity of the HMGB1-TLR2/TLR4-NF-κB pathway.

Zuoguiwan Prevent Decreased Ovarian Reserve in Prenatal Stressed Offspring Rats via TLR4/NF-κB p65 Signaling Pathway / 中国实验方剂学杂志

ObjectiveTo observe the effect of Zuoguiwan on ovarian reserve in the female offspring rat model of prenatal stress （PS） and explore the mechanism based on Toll-like receptor 4/nuclear factor-κB p65 （TLR4/NF-κB p65） signaling pathway. MethodThirty-two pregnant rats were prepared and randomized into four groups （n=8）： control， model， Zuoguiwan （18.9 mg·kg-1）， and vitamin E （1.44 mg·kg-1）. Except the control group， the other three groups were subjected to chronic unpredictable mild stress （CUMS） from day 11 of pregnancy， and the modeling was accompanied by gavage with corresponding drugs until delivery. The PS model was evaluated by the sucrose preference test， open field test， and serum corticosterone （CORT） level. The estrous cycle was monitored and the morphological changes in the ovarian tissue were observed. The serum levels of estradiol （E2）， luteinizing hormone （LH）， follicle-stimulating hormone （FSH）， and anti-Mullerian hormone （AMH） in the 75-day-old offspring rats were measured by enzyme-linked immunosorbent assay （ELISA） to evaluate the ovarian reserve. The ovary and uterus indices were calculated. The serum levels of interleukin-1β （IL-1β） and tumor necrosis factor-α （TNF-α） were measured by enzyme-linked immunosorbent assay （ELISA）. The morphology of the ovarian tissue in the offspring on the day of birth and day 75 after birth was observed by hematoxylin-eosin staining. The transport of NF-κB p65 to the nucleus in the ovaries of the 75-day-old offspring was detected by the immunofluorescence （IF） assay. The expression of TLR4， NF-κB p65 and other related proteins in the ovarian tissue was determined by Western blot. ResultCompared with the control group， the model group showed reduced primordial follicles in the offspring on the day of birth （P<0.01） as well as disturbed estrous cycle， decreased ovary index and uterus index （P<0.01）， reduced corpus luteum， increased atretic follicles （P<0.01）， lowered serum levels of AMH and E2 （P<0.01）， elevated serum levels of LH， FSH， IL-1β， and TNF-α （P<0.05， P<0.01）， and up-regulated protein levels of TLR4， NF-κB p65， recombinant myeloid differentiation factor 88 （MyD88）， and phosphorylated NF-κB inhibitor （p-IκBα） （P<0.01） in the 75-day-old offspring rats. Compared with the model group， Zuoguiwan and vitamin E increased the primordial follicles in the offspring on the day of birth （P<0.01）. Moreover， they resumed the estrous cycle， increased the ovary and uterine indices （P<0.05， P<0.01） and corpus luteum （P<0.01）， reduced atretic follicles （P<0.01）， elevated the serum levels of AMH and E2 （P<0.05， P<0.01）， lowered the serum levels of LH， FSH， IL-1β， and TNF-α （P<0.05， P<0.01）， and down-regulated the expression of TLR4， NF-κB p65， MyD88， and p-IκB-α （P<0.05， P<0.01） in the 75-day-old offspring. ConclusionZuoguiwan can improve the ovarian reserve in the offspring rat model of congenital kidney deficiency by regulating the TLR4/NF-κB p65 signaling pathway.

Protective Effect and Mechanism of Mori Folium Extract on Kidney of db/db Diabetic Mice / 中国实验方剂学杂志

ObjectiveTo investigate the protective effects of Mori Folium extract （MLE） on the kidney of db/db diabetic mice and its mechanism. MethodTwenty-four male C57BLKS/JGpt-Leprdb/Leprdb （db/db） mice were randomly divided into model group， metformin group， low-dose group of MLE （MLE-L）， and high-dose group of MLE （MLE-H） according to their fasting blood glucose （FBG）， with six mice in each group， and other six C57BLKS/JGpt wild littermate （m/m） mice were selected as normal group. The mice in the drug administration groups were given corresponding drugs by gavage， and the mice in the normal group and model group were given the same dose of deionized water by gavage once a day for continuous eight weeks. Body weight， bilateral kidney weight， and FBG were measured， and an oral glucose tolerance test （OGTT） was performed. The pathological changes in the kidney tissue of mice were observed by hematoxylin-eosin （HE） and periodic acid-silver （PAS） staining， and serum creatinine （SCr） and blood urea nitrogen （BUN） levels were detected. Enzyme-linked immunosorbent assay （ELISA） was used to detect the levels of tumor necrosis factor-alpha （TNF-α） and interleukin-6 （IL-6） in serum and urinary microalbumin （U-mAlb） of mice. The expression levels of toll-like receptor 4 （TLR4）， myeloid differentiation factor 88 （MyD88）， and nuclear factor-kappa B p65 （NF-κB p65） protein in kidney tissue of mice were tested by Western blot. ResultCompared with the normal group， the body weight， absolute renal weight， FBG， and the area under the curve （AUC） of OGTT of mice in the model group were significantly increased （P<0.01）， and the levels of SCr， BUN， and U-mAlb， as well as TNF-α and IL-6 in serum were significantly increased （P<0.01）. The glomerular basement membrane in the kidney tissue of mice was thicker， with obvious inflammatory cell infiltration. The protein expression levels of TLR4， MyD88， and NF-κB p65 in the kidney tissue of mice were increased significantly （P<0.01）. Compared with the model group， there was no statistical difference in the body weight of mice in each drug administration group. The absolute renal weight of mice in the MLE-H and metformin groups was significantly reduced （P<0.05， P<0.01）. The FBG levels of mice in the metformin， MLE-L， and MLE-H groups started to decrease after treatment for four to eight weeks （P<0.05， P<0.01）. The AUC of mice in the MLE-H and metformin groups was significantly decreased （P<0.01）. The levels of SCr， BUN， and U-mAlb of mice in the MLE-H and metformin groups were significantly decreased （P<0.01）， and those of SCr and U-mAlb of mice in the MLE-L group were significantly decreased （P<0.01）. The levels of TNF-α and IL-6 in the serum of mice in the MLE-H and metformin groups were significantly decreased （P<0.01）. The renal tissue pathology of mice in each drug administration group was improved to varying degrees， and the protein expression levels of TLR4， MyD88， and NF-κB p65 in the MLE-H group were decreased significantly （P<0.05， P<0.01）. ConclusionMLE can improve the renal structure and function of db/db diabetic mice， and its mechanism may be related to the inhibition of the TLR4/MyD88/NF-κB signaling pathway.

HMGB1 gene knockout alleviates acute lung injury of sepsis mice via inhibiting TLR4/NF-κB pathway / 安徽医科大学学报

Objective To study the effect of high mobility group box B1(HMGB1)gene knockout on alleviating a-cute lung injury and inhibiting toll-like receptor 4(TLR4)/nuclear factor-KB(NF-κB)pathway of sepsis mice.Methods Wild-type(WT)mice were divided into WT-Sham group and WT-model group,and HMGB1 knockout(KO)mice were divided into KO-sham group and KO-model group.Sepsis ALI model was established by cecal ligation and perforation in WT-model group and KO-model group.Sham operation was performed in WT-Sham group and KO-Sham group.24 h after modeling,the partial pressure of arterial oxygen(PaO2)was detected,oxy-genation index(OI)was calculated,pathological changes of lung tissue were detected and lung injury score was calculated,the concentrations of tumor necrosis factor-α(TNF-α),interleukin-1 β(IL-1 β),interleukin-6(IL-6),reactive oxygen species(ROS),malondialdehyde(MDA),superoxide dismutase(SOD),in serum and lung tissues and the expression of HMGB1,TLR4 and nuclear NF-κB in lung tissues were detected.Results The PaO2,OI and the concentration of SOD in serum and lung tissue of WT-model group were lower than those of WT-Sham group,the lung injury scores,the concentrations of TNF-α,IL-1 β,IL-6,ROS and MDA in serum and lung tissue,and the expression levels of HMGB1,TLR4 and nuclear NF-κB in lung tissue were higher than those in WT-Sham group(P＜0.05).HMGB1 was not expressed in lung tissue of KO-model group,and the concentrations of PaO2,OI and the concentration of SOD in serum and lung tissue of KO-model group were higher than those of WT-model group,the lung injury scores,the concentrations of TNF-α,IL-1β,IL-6,ROS and MDA in serum and lung tissue,and the expression levels of TLR4 and nuclear NF-κB in lung tissue were lower than those of the WT-model group(P＜0.05).Conclusion HMGB1 gene knockout alleviates acute lung injury of sepsis mice,the re-lated molecular mechanism may be the inhibition of TLR4/NF-κB pathway mediated inflammation and oxidative stress.