Mechanism of dexmedetomidine preconditioning on spinal cord analgesia in rats with functional chronic visceral pain

Purpose: To analyze the effect and mechanism of dexmedetomidine (DEX) analgesia pretreatment on functional chronic visceral pain in rats. Methods: Rats were divided into six groups: W1, W2, W3, W4, W5, and W6. The behavioral changes and electrophysiological indexes of rats in each group before and after DEX treatment were detected. Results: The levels of abdominal withdrawal reflex (AWR) in W5 and W6 groups were significantly lower than those in group W3, while the levels of thermal withdrawal latency (TWL) and mechanical withdrawal threshold (MWT) were significantly higher than those in group W3 (p < 0.05). The electromyographic signals of W1, W5, and W6 groups showed little fluctuation, while those of groups W2, W3, and W4 showed obvious fluctuation. TLR4 mRNA expression, IRF3, P65, and phosphorylation levels in W4, W5, and W6 groups were significantly lower than those in group W2 (p < 0.05). Conclusions: Dexmedetomidine epidural anesthesia pretreatment could significantly inhibit visceral pain response in rats with functional chronic visceral pain, and its mechanism was related to the activation of TLR4 in spinal dorsal horn tissue of rats and the activation inhibition of IRF3 and P65 in the downstream key signals.

Effect of salvianolic acid on behaviors and brain inflammatory factors in depression model rats / 中华行为医学与脑科学杂志

Objective:To investigate the effect of salvianolic acid on depressive behavior in depression model rats induced by chronic mild stress (CMS) and its mechanism.Methods:Fifty healthy male clean grade Sprague-Dawley(SD) rats were divided into five groups according to a random number table with 10 in each group: control group (nCMS+ Nal group), CMS+ normal saline group (CMS+ Nal group), CMS+ fluoxetine group (CMS+ Flu group), CMS+ salvia acid group (CMS+ Sal group), CMS+ fluoxetine+ Salvia acid group (CMS+ Flu+ Sal group). Except the control group, the rats in the other four groups were all received CMS modeling for 21 days. Twenty-one days after CMS modeling, rats were intraperitoneally injected with 0.9% normal saline (10 mg·kg -1·d -1), fluoxetine (20 mg·kg -1·d -1), salvia acid(40 mg·kg -1·d -1), fluoxetine(20 mg·kg -1·d -1)+ salvia acid(40 mg·kg -1·d -1)for 21 days. During the administration period, rats in the other four groups continued to receive CMS intervention for 21 days. Forced swimming test and sucrose preference test were conducted at baseline (day 0), after modeling (day 21) and after intervention (day 42) so as to evaluate depression like behavior. Then the rats were sacrificed and the hippocampus and prefrontal cortex were taken. The mRNA levels of Toll like receptor 4 (TLR4) and myeloid differentiation primary response 88 (MyD88) were detected by RT-qPCR. The cytokines including interleukin-1β(IL-1β), interleukin-2(IL-2), interferon-γ (IFN-γ) and tumor necrosis factor-α (TNF-α) were detected by Luminex technique.SPSS 21.0 was used for statistical analysis.Repeated measurement ANOVA was used for behavioral data analysis, one-way ANOVA was used for molecular index data analysis, and Spearman was used for correlation analysis. Results:The results of repeated measurement ANOVA showed that the interaction effects between group and time of body mass, sucrose preference, forced swimming immobility time were significant at baseline, after modeling and after intervention ( F=18.238, 6.921, 7.591, all P<0.05). After modeling, compared with nCMS+ Nal group, the rats in CMS+ Flu group, CMS+ Sal group, CMS+ Flu+ Sal group and CMS+ Nal group had lower body weight, lower sucrose preference rate and longer forced swimming immobility time (all P<0.05). After intervention, compared with CMS+ Nal group(body weight (350.15±41.65)g, sucrose preference(52.95±11.13)%, static time(91.40±15.22)s), the body weight((378.21±30.78)g, (385.12±43.19)g, (391.41±31.21)g, (402.33±18.67)g, all P<0.05) and sucrose preference((69.30±15.56)%, (68.12±10.99)%, (71.18±9.51)%, (75.47±11.55)%, all P<0.05) of CMS+ Flu group, CMS+ Sal group, CMS+ Flu+ Sal group and nCMS+ Nal group were all increased, while the forced swimming immobility time ((68.81±21.74)s, (66.10±25.51)s, (63.53±22.32)s, (71.21±21.41)s, all P<0.05) were shorter (all P<0.05). After intervention, among the body weight, sucrose preference and the immobility time of CMS+ Flu group、CMS+ Sal group and CMS+ Flu+ Sal group, there were no differences between each two groups(all P>0.05). After intervention, the levels of TLR4 mRNA and MyD88 mRNA in prefrontal cortex and hippocampus of CMS+ Flu group, CMS+ Sal group, CMS+ Flu+ Sal group and nCMS+ Nal group were all lower than those in CMS+ Nal group (all P<0.05). In prefrontal cortex, the levels of TLR4 mRNA (0.715±0.358) and MyD88 mRNA (0.739±0.233) in CMS+ Flu+ Sal group were lower than those in CMS+ Sal group (1.943±0.606, 1.815±0.897) (both P<0.05). The level of TLR4 mRNA in prefrontal cortex and hippocampus of rats were positively correlated with the level of MyD88 mRNA and TNF-α level and forced swimming immobility time and negatively correlated with sucrose preference rate (prefrontal cortex r=0.915, 0.041, 0.027, -0.178, all P<0.05; hippocampus r=0.810, 0.070, 0.011, -0.153, all P<0.05). Conclusion:The antidepressant effect of salvianolic acid is presumedly achieved by inhibiting the immunoinflammatory response mediated by the TLR4/Myd88 signaling pathway in CMS rats.

Investigating the effect of the Toll-like receptor-4 inhibitor TAK-242 on alveolar bone resorption in severe periodontitis in rats / 口腔疾病防治

Objective@# The goal of this study was to investigate the effect of the Toll-like receptor-4 (TLR-4) inhibitor TAK-242 on bone resorption in severe periodontitis in rats in order to provide an experimental basis for finding new adjuvant treatments for severe periodontitis.@*Methods @# Eighteen three-week-old male Wistar rats were randomly divided into three groups (n=6). One group was the control group, and the other two groups were modelled with severe periodontitis in bilateral maxillary molars ligated with 5-0 silk thread containing P. gingivalis ATCC33277 (periodontitis and TAK-242 groups, respectively). The TAK-242 group was injected with TAK-242 (2 mg/kg) in DMSO every other day through the tail vein from the first day of silk ligation, and the other two groups were injected with DMSO solvent at the same proportion of body weight for 8 consecutive weeks. At the end of the 8th week, rats in the 3 groups were sacrificed, and maxillary specimens were taken. Three-dimensional reconstruction was achieved after micro-CT scanning to measure the distance between the enamel cementum boundary and alveolar crest at specific sites to assess the amount of bone loss. The parameters related to alveolar bone and bone microstructure were analyzed, and the pathological changes of periodontal tissue were observed by hematoxylin-eosin (HE) staining. Alveolar bone resorption was observed by Methyl Green staining, and the distribution of osteoclasts was observed by anti-tartrate acid phosphatase double staining (TRAP).@*Results@#Micro-CT quantitative analysis showed that alveolar bone resorption in the periodontitis group and TAK-242 group was significantly higher than that in the control group. Compared with the periodontitis group, bone loss of the mesial and distal root resorption sites of maxillary first molars was significantly reduced in the TAK-242 group (P < 0.001), the bone mineral density (P < 0.05) and bone volume/total volume fraction (P < 0.01) were significantly increased, and the number of trabeculae and the trabeculae thickness (P < 0.01) were relatively increased. The trabecular separation (P < 0.01) and trabecular structure model index were significantly decreased. In the periodontitis group, the bone exhibited a sparse and porous honeycomb structure, with deterioration of the trabecular structure and a shift towards a rod-strength structure. In the TAK-242 group, the bone microstructure was improved, the bone volume was enriched, the distribution of trabeculae was relatively denser and the trabecular structure shared more similarities with the control group. HE staining displayed that the attachment loss and bone absorption were significantly higher in the periodontitis group and TAK-242 group than in the control group. Compared with the periodontitis group, MG staining displayed significantly alleviated bone absorption in the TAK-242 group. TRAP staining showed that osteoclast infiltration decreased in the TAK-242 group compared with the periodontitis group (P < 0.001)@*Conclusions @#The TLR-4 inhibitor TAK-242 can alleviate bone resorption in severe periodontitis and improve the porous, sparse and disorganized inflammatory bone trabecular structure in rats.

Effect of Chuanshanlong Granule on TLR4/MyD88/NF-κB Signaling Pathway in Rats with Autoimmune Thyroiditis / 中国实验方剂学杂志

ObjectiveTo investigate the effect of Chuanshanlong granule on Toll-like receptor 4 （TLR4）/myeloid differentiation protein 88 （MyD88）/nuclear transcription factor -κB （NF-κB） signaling pathway， and to explore the mechanism of its treatment of autoimmune thyroiditis （AIT） in rats. MethodForty AIT models were established following excess iodine and injection of porcine thyroglobulin and Freund's adjuvant into Lewis rats for six weeks. Then the rats were randomly divided into the model group， Chuanshanlong granule low-， medium- and high-dose group （0.52， 1.03， 2.06 g·kg-1·d-1）， with ten in each group. Rats in the Chuanshanlong granule low-， medium- and high-dose groups were separately given 0.01 mL·g-1·d-1 Chuanshanlong granule， and those in the normal group and the model group were given the same volume of deionized water for eight weeks. Serum of rats was taken to measure thyroglobulin antibody （TgAb） and thyroid peroxidase antibody （TPOAb） by enzyme-linked immunosorbent assay （ELISA）， and the concentrations of free triiodothyronine （FT3）， free thyroxine （FT4） and thyroid stimulating hormone （TSH） were detected. The rat thyroid lobes were stained with hematoxylin and eosin （HE）， and the pathological changes were observed under light microscope. In addition， the relative expression of TLR4， MyD88， NF-κB protein and mRNA was determined by immunohistochemistry and real-time polymerase chain reaction （Real-time PCR）. ResultCompared with the conditions in the normal group， the serum concentrations of TPOAb and TgAb （P<0.01） and FT3 and FT4 （P<0.01） increased and TSH decreased （P<0.01） in the model group. Compared with the conditions in the model group， the concentrations of TPOAb and TgAb in the Chuanshanlong granule treatment groups reduced （P<0.01）， and the concentrations of FT3 and FT4 were lowered （P<0.01） while TSH increased （P<0.01） in the Chuanshanlong granule high-dose group. HE staining showed that there was lymphocyte infiltration in the thyroid follicular space， a large number of destroyed or diminished follicular cavities， decreased colloid content， and thinned or destroyed follicular wall in the model group， while the thyroid lymphocyte infiltration in the Chuanshanlong granule treatment groups was significantly less and the structure of thyroid follicles was more complete than those in the model group. Compared with the normal group， the model group had up-regulated relative expression of TLR4， MyD88 and NF-κB protein （P<0.01） and mRNA （P<0.01）. Compared with the model group， the Chuanshanlong granule high-dose group had down-regulated relative expression of TLR4 protein and mRNA （P<0.05）， MyD88 protein （P<0.01） and mRNA （P<0.05）， and NF-κB protein and mRNA （P<0.01）. ConclusionChuanshanlong granule may play a therapeutic role in AIT by inhibiting the activation of TLR4/MyD88/NF-κB signaling pathway.

Qiling Tongluo Prescription Improves Renal Injury in Rats with Idiopathic Membranous Nephropathy by Inhibiting TLR4/MyD88/NF-κB Signaling Pathway / 中国实验方剂学杂志

ObjectiveTo explore the therapeutic effect and mechanism of Qiling Tongluo prescription against idiopathic membranous nephropathy （IMN） in rats based on Toll-like receptor 4/myeloid differentiation factor 88/nuclear transcription factor-κB （TLR4/MyD88/NF-κB） signaling pathway. MethodSixty male SD rats were randomly divided into the normal group， model group， benazepril hydrochloride （10 mg·kg-1） group， and low-，medium-， and high-dose （6.48， 12.95， and 25.9 g·kg-1） Qiling Tongluo prescription groups. The IMN rat model was established by injection of cationized bovine serum albumin （C-BSA） into the tail vein. After the model was successfully prepared， the rats were gavaged with the corresponding drugs， once a day， for four consecutive weeks. After the treatment， the pathological changes in rat kidneys were observed by hematoxylin-eosin （HE） staining， Masson staining， and periodic acid-silver metheramine （PASM） staining， followed by the detection of 24 h urinary total protein （24 h UTP）， plasma albumin （ALB）， total serum protein （TP）， serum creatinine （SCr）， urea nitrogen （BUN）， and uric acid （UA） levels. The levels of interleukin-1β （IL-1β） and interleukin-6 （IL-6） were determined by enzyme-linked immunosorbent assay （ELISA）， and the mRNA and protein expression levels of TLR4， MyD88， and NF-κB in the kidney tissue were assayed by real-time fluorescent quantitative polymerase chain reaction （Real-time PCR）， immunohistochemistry （IHC）， and Western blot. ResultCompared with the normal group， the model group exhibited elevated 24 h UTP and serum SCr， BUN， UA， IL-1β， and IL-6 （P<0.05， P<0.01）， decreased ALB and TP （P<0.01）， up-regulated TLR4， MyD88， and NF-κB p65 mRNA and protein expression in kidney tissue （P<0.05， P<0.01）， obvious inflammation， disordered glomerular structure with enlarged volume， irregularly thickened basement membrane， inflammatory cell infiltration in the renal interstitium， reduced renal tubular epithelial cells due to shedding and apoptosis， and some vacuolar degeneration. Compared with the model group， benazepril hydrochloride and Qiling Tongluo prescription at the high dose remarkably lowered the serum SCr and UA （P<0.05） and increased ALB and TP （P<0.05）. Benazepril hydrochloride and Qiling Tongluo prescription at the low， medium， and high doses down-regulated the 24 h UTP， serum IL-1β and IL-6 levels， and renal TLR4， MyD88， and NF-κB p65 mRNA and protein expression to varying degrees （P<0.05， P<0.01）， alleviated IMN inflammatory reaction， glomerular swelling， and volume increase， slightly dilated glomerular capillaries， proliferated mesangial matrix， and relieved pathological and morphological damages in rat kidney， with inflammatory cell infiltration occasionally observed. ConclusionQiling Tongluo prescription may reduce the release and expression of inflammatory factors by regulating the TLR4/MyD88/NF-κB signaling pathway to inhibit the inflammatory response in IMN rats， ameliorate proteinuria and kidney damage， and protect kidney function.

Effect of Gegen Qinliantang on TLR4/IKKβ/NF-κB Pathway in KKAy Mice with Diabetes / 中国实验方剂学杂志

ObjectiveTo observe the effect of classical prescription Gegen Qinliantang（GGQLT） on inflammatory factors and key targets in the inflammatory pathways mediated by lipopolysaccharide in KKAy mice and explore its mechanism in improving spontaneous type 2 diabetes mellitus （T2DM）. MethodSixty-five SPF KKAy mice with spontaneous T2DM and 13 C57BL/6J mice （control） were selected in the barrier system and fed on a high-fat diet. The model was properly induced in 44 mice in the context of random blood glucose exceeding or equal to 13.9 mmol·L-1. Then the mice were assigned into a normal group （20 mL∙kg-1 normal saline）， a model group （20 mL∙kg-1 normal saline）， an acarbose group （3.9 mg∙kg-1）， and high- and low-dose GGQLT groups （1.82 and 0.45 g∙kg-1）， with 11 mice in each group. The mice in each group were treated correspondingly by gavage for eight weeks， once per day. Blood glucose and body weight were systematically evaluated. Twelve hours after the last administration， blood samples were collected from the eyes， and the serum and muscle and liver tissues were extracted. The levels of tumor necrosis factor-α （TNF-α）， interleukin-6 （IL-6）， and glucose transporter type 4 （GluT4） were detected by semi-quantitative enzyme-linked immunosorbent assay （ELISA）. The protein expression of IκB kinase β （IKKβ） and nuclear factor-κB （NF-κB） in muscle tissues and Toll-like receptor 4 （TLR4） in liver tissues was detected by Western blot. ResultCompared with the normal group， the model group showed increased body weight and blood glucose （P<0.01）. Compared with the model group， the acarbose group and the GGQLT groups showed reduced body weight and blood glucose （P<0.05， P<0.01）. As revealed by ELISA results， compared with the normal group， the model group showed increased levels of TNF-α and IL-6 （P<0.01） and deceased GluT4 level （P<0.05）. Compared with the model group， the groups with drug treatment showed reduced levels of TNF-α and IL-6 （P<0.05， P<0.01）， and the acarbose group and the high-dose GGQLT group showed increased GluT4 level （P<0.05， P<0.01）. As displayed by Western blot results， compared with the normal group， the model group showed increased protein expression of IKKβ， NF-κB， and TLR4 （P<0.01）. Compared with the model group， the acarbose group and the GGQLT groups showed reduced protein expression of IKKβ， NF-κB， and TLR4 （P<0.05， P<0.01）. ConclusionGGQLT can inhibit the inflammatory cascade effect and improve T2DM by down-regulating the levels of key inflammatory factors in the TLR4 pathway， inhibiting their activation， and increasing the translocation and activity of GluT4 on the basis of the regulation of intestinal flora.

Mechanism of Chaishao Liujunzi Tang in Treatment of Ulcerative Colitis Through TLR4/MyD88/NF-κB Signaling Pathway / 中国实验方剂学杂志

ObjectiveTo investigate the preventive and curative effect of Chaishao Liujunzi Tang （CSLJZT） on colonic mucosal injury induced by dextran sulfate sodium （DSS） in mice with ulcerative colitis （UC） and its mechanism. MethodFifty Balb/c male mice were randomly divided into normal group， model group， CSLJZT low-dose group， CSLJZT high-dose group， and sulfasalazine group. Except for the normal group， other groups were given 2.5% DSS freely for 7 d， and were given drug intervention after successful modeling for 7 d. Bodyweight， feces， and other general physiological statuses of mice were recorded every day， and disease activity index （DAI） scores were calculated.The colon length was measured， and stained by hematoxylin-eosin （HE） staining to observe the morphological changes of the colon.The enzyme-linked immunosorbent assay （ELISA） was used to determine the content of interleukin-1β （IL-1β）， myeloperoxidase （MPO）， and superoxide dismutase （SOD） in the serum. Western blot was used to determine the protein expression levels of Toll-like receptor 4 （TLR4）， myeloid differentiation factor 88 （MyD88）， nuclear factor kappa B （NF-κB）， inhibitor-kappa binding protein （IκB）， Caspase-1， and nucleotide-binding oligomerization domain-like receptor 3 （NLRP3） in the colon tissues. ResultAs compared with the normal group， mice in the model group had significantly decreased body weight （P<0.01）， severe diarrhea and hematochezia， and significantly increased DAI score （P<0.01）. As compared with the model group， the decreasing trend of body weight was significantly alleviated in the CSLJZT groups （P<0.01）， diarrhea and hematochezia were significantly improved， DAI score was significantly decreased （P<0.01）， and colon length increased （P<0.05）. HE staining showed that the pathological damage of colon tissue was significantly improved and the inflammatory cell infiltration was reduced in the CSLJZT groups as compared with the model group. As compared with the normal group， the serum levels of IL-1β and MPO were significantly higher （P<0.01） and SOD levels were significantly lower （P<0.01） of mice in the model group.Compared with the model group， the treated group reduced the serum IL-1β and MPO levels （P<0.01）， and raised the SOD level （P<0.01）. The results of Western blot showed that as compared with the normal group， the expression levels of TLR4， MyD88， NF-κB， Ccaspase-1， and NLRP3 proteins were significantly increased （P<0.01）， whereas the expression level of IκB protein was significantly decreased （P<0.01） in the colonic tissue of mice in the model group. As compared with the model group， the expression levels of TLR4， MyD88， NF-κB， Caspase-1， and NLRP3 proteins were decreased （P<0.01）， whereas the expression level of IκB protein was increased （P<0.01） in the colonic tissue of mice in the CSLJZT groups. ConclusionCSLJZT improves the inflammatory injury of the colon tissue in DSS-induced UC mice through TLR4/MyD88/NF-κB signaling pathway.

Mechanism of Chaishao Liujunzi Tang in Treatment of Ulcerative Colitis Through TLR4/MyD88/NF-κB Signaling Pathway / 中国实验方剂学杂志

ObjectiveTo investigate the preventive and curative effect of Chaishao Liujunzi Tang （CSLJZT） on colonic mucosal injury induced by dextran sulfate sodium （DSS） in mice with ulcerative colitis （UC） and its mechanism. MethodFifty Balb/c male mice were randomly divided into normal group， model group， CSLJZT low-dose group， CSLJZT high-dose group， and sulfasalazine group. Except for the normal group， other groups were given 2.5% DSS freely for 7 d， and were given drug intervention after successful modeling for 7 d. Bodyweight， feces， and other general physiological statuses of mice were recorded every day， and disease activity index （DAI） scores were calculated.The colon length was measured， and stained by hematoxylin-eosin （HE） staining to observe the morphological changes of the colon.The enzyme-linked immunosorbent assay （ELISA） was used to determine the content of interleukin-1β （IL-1β）， myeloperoxidase （MPO）， and superoxide dismutase （SOD） in the serum. Western blot was used to determine the protein expression levels of Toll-like receptor 4 （TLR4）， myeloid differentiation factor 88 （MyD88）， nuclear factor kappa B （NF-κB）， inhibitor-kappa binding protein （IκB）， Caspase-1， and nucleotide-binding oligomerization domain-like receptor 3 （NLRP3） in the colon tissues. ResultAs compared with the normal group， mice in the model group had significantly decreased body weight （P<0.01）， severe diarrhea and hematochezia， and significantly increased DAI score （P<0.01）. As compared with the model group， the decreasing trend of body weight was significantly alleviated in the CSLJZT groups （P<0.01）， diarrhea and hematochezia were significantly improved， DAI score was significantly decreased （P<0.01）， and colon length increased （P<0.05）. HE staining showed that the pathological damage of colon tissue was significantly improved and the inflammatory cell infiltration was reduced in the CSLJZT groups as compared with the model group. As compared with the normal group， the serum levels of IL-1β and MPO were significantly higher （P<0.01） and SOD levels were significantly lower （P<0.01） of mice in the model group.Compared with the model group， the treated group reduced the serum IL-1β and MPO levels （P<0.01）， and raised the SOD level （P<0.01）. The results of Western blot showed that as compared with the normal group， the expression levels of TLR4， MyD88， NF-κB， Ccaspase-1， and NLRP3 proteins were significantly increased （P<0.01）， whereas the expression level of IκB protein was significantly decreased （P<0.01） in the colonic tissue of mice in the model group. As compared with the model group， the expression levels of TLR4， MyD88， NF-κB， Caspase-1， and NLRP3 proteins were decreased （P<0.01）， whereas the expression level of IκB protein was increased （P<0.01） in the colonic tissue of mice in the CSLJZT groups. ConclusionCSLJZT improves the inflammatory injury of the colon tissue in DSS-induced UC mice through TLR4/MyD88/NF-κB signaling pathway.

Effects of butyphthalide on microglia polarization after intracerebral hemorrhage and the underlying mechanisms / 中南大学学报(医学版)

OBJECTIVES@#Because intracerebral hemorrhage (ICH) has high morbidity, disability and mortality, it is significant to find new and effective treatments for ICH. This study aims to explore the effect of butyphthalide (NBP) on neuroinflammation secondary to ICH and microglia polarization.@*METHODS@#A total of 48 healthy male SD rats were randomly divided into 6 groups: a sham 24 h group, a sham 72 h group, an ICH 24 h group, an ICH 72 h group, an ICH+NBP 24 h group, and an ICH+NBP 72 h group (8 rats per group). After operation, the neurological deficiencies were assessed based on improved Garcia scores and corner test. The expressions of Toll-like receptor 4 (TLR4)/nuclear factor-kappa B (NF-κB), nuclear factor erythroid 2-related factor 2 (Nrf2), aquaporin-4 (AQP4), zonula occludens-1 (ZO-1), occludin, CD68, CD86, and CD206 were observed by Western blotting. Inflammatory cytokines were detected by ELISA. The immunofluorescence was to detect the polarization of microglia.@*RESULTS@#1) Compared with the sham groups, the expression of TLR4 (24 h: P<0.05; 72 h: P<0.01), NF-κB (both P<0.01) and Nrf2 (both P<0.01) in the perihematoma of the ICH group was increased, leading to microglia activation (P<0.01). The expressions of IL-6 (24 h: P<0.05; 72 h: P<0.01) and TNF-α (both P<0.01), the pro-inflammatory cytokines were up-regulated, and the expression of anti-inflammatory cytokine IL-4 was down-regulated (both P<0.01). Besides, the expression of AQP4 was enhanced (both P<0.01). The protein level of tightly connected proteins (including ZO-1, occludin) was decreased (all P<0.01). The neurological function of the rats in the ICH group was impaired in the 2 time points (both P<0.01). 2) Compared with the sham group at 24 h and 72 h after the intervention of NBP, the expressions of TLR4 (both P<0.05) and NF-κB (both P<0.01) were significantly declined, and the expression of Nrf2 was further enhanced (both P<0.05) in the perihematoma of the ICH+NBP group. Furthermore, the expression of M1 microglia marker was inhibited (P<0.05), and the polarization of microglia to the M2 phenotype was promoted (P<0.01). 3) In terms of inflammation after ICH, the IL-4 expression in the ICH+NBP group was increased compared with the ICH group (24 h: P<0.05; 72 h: P<0.01); the expression of IL-6 was decreased significantly in the ICH+NBP 72 h group (P<0.01); the level of AQP4 was declined significantly in the ICH+NBP 24 h group (P<0.05), there was a downward trend in the 72-hour intervention group but without significant statistical difference. 4) Compared with the ICH group, the ZO-1 protein levels were increased (24 h: P<0.05; 72 h: P<0.01), and the symptoms of nerve defect were improved eventually (both P<0.05) in the ICH+NBP groups.@*CONCLUSIONS@#After ICH, the TLR4/NF-κB pathway is activated. The M1 microglia is up-regulated along with the release of detrimental cytokines, while the anti-inflammatory cytokines are down-regulated. The expression of AQP4 is increased, the tight junction proteins from the blood-brain barrier (BBB) is damaged, and the neurological function of rats is impaired. On the contrary, NBP may regulate microglia polarization to M2 phenotype and play a role in the neuroprotective effect mediated via inhibiting TLR4/NF-κB and enhancing Nrf2 pathways, which relieves the neuroinflammation, inhibits the expression of AQP4, repairs BBB, and improves neurological functional defects.

Target inhibition of Toll-like receptor 4 signaling pathway and necrotizing enterocolitis / 中华围产医学杂志

The pathogenesis of neonatal necrotizing enterocolitis (NEC) is still unclear. Toll-like receptor 4 (TLR4) signaling pathway, mediated by TLR4 in the intestinal epithelial cells, is considered to play an important role in activating inflammatory storm in NEC. This paper elaborates the association between NEC and the inhibition of TLR4 signal pathway and its upstream and downstream signal targets, critical pattern recognition receptors, and negative regulation of TLR4 by certain receptors to gain more insight into possible target interventions for NEC.

Research progress of NF-κB signaling pathway in kidney ischemia-reperfusion injury / 器官移植

Kidney ischemia-reperfusion injury (IRI) is the major cause of poor prognosis after kidney transplantation and partial nephrectomy. Besides, it is also a critical pathophysiological process of acute kidney injury. Consequently, the prevention and treatment of kidney IRI are of significance to improve clinical prognosis of recipients undergoing kidney transplantation. However, the mechanism underlying IRI is complex, and the exact mechanism remains elusive. Inflammation, as one of the main pathogenesis of IRI, plays a significant role in IRI-induced kidney injury. Nuclear factor (NF)-κB, as a rapid response transcription factor, has been proven to be involved in the regulation of inflammation during kidney IRI. Therefore, in this article, the structure of NF-κB, the activation pattern of NF-κB signaling pathway, the regulatory mechanisms of NF-κB upstream and downstream signaling pathways in kidney IRI were reviewed, and the role of NF-κB signaling pathway in kidney IRI was investigated, aiming to provide novel clinical ideas for the prevention and treatment of kidney IRI.

Electroacupuncture Attenuates Immune-Inflammatory Response in Hippocampus of Rats with Vascular Dementia by Inhibiting TLR4/MyD88 Signaling Pathway / 中国结合医学杂志

OBJECTIVE@#To investigate whether electroacupuncture (EA) alleviates cognitive impairment by suppressing the toll-like receptor 4 (TLR4)/myeloid differentiation factor 88 (MyD88) signaling pathway, which triggers immune-inflammatory responses in the hippocampus of rats with vascular dementia (VaD).@*METHODS@#The experiments were conducted in 3 parts and in total the Sprague-Dawley rats were randomly divided into 8 groups by a random number table, including sham, four-vessel occlusion (4-VO), 4-VO+EA, 4-VO+non-EA, sham+EA, 4-VO+lipopolysaccharide (LPS), 4-VO+LPS+EA, and 4-VO+TAK-242 groups. The VaD model was established by the 4-VO method. Seven days later, rats were treated with EA at 5 acupoints of Baihui (DV 20), Danzhong (RN 17), Geshu (BL 17), Qihai (RN 6) and Sanyinjiao (SP 6), once per day for 3 consecutive weeks. Lymphocyte subsets, lymphocyte transformation rates, and inflammatory cytokines interleukin-6 (IL-6) and tumor necrosis factor α(TNF-α) were measured to assess immune function and inflammation in VaD rats. Transmission electron microscopy was used to observe the ultrastructure of nerve cells in the hippocampus. The levels of TLR4, MyD88, IL-6, and TNF-α were detected after EA treatment. TLR4/MyD88 signaling and cognitive function were also assessed after intracerebroventricular injection of TLR4 antagonist TAK-242 or TLR4 agonist LPS with or without EA.@*RESULTS@#Compared with the 4-VO group, EA notably improved immune function of rats in the 4-VO+EA group, inhibited the protein and mRNA expressions of TLR4 and MyD88 in the hippocampus of rats, reduced the expressions of serum IL-6 and TNF-α (all P0.05).@*CONCLUSIONS@#EA attenuated cognitive impairment associated with immune inflammation by inhibition of the TLR4/MyD88 signaling pathway. Thus, EA may be a promising alternative therapy for the treatment of VaD.

Understanding the physiological functions of the host xenobiotic-sensing nuclear receptors PXR and CAR on the gut microbiome using genetically modified mice

Pharmacological activation of the xenobiotic-sensing nuclear receptors pregnane X receptor (PXR) and constitutive androstane receptor (CAR) is well-known to increase drug metabolism and reduce inflammation. Little is known regarding their physiological functions on the gut microbiome. In this study, we discovered bivalent hormetic functions of PXR/CAR modulating the richness of the gut microbiome using genetically engineered mice. The absence of PXR or CAR increased microbial richness, and absence of both receptors synergistically increased microbial richness. PXR and CAR deficiency increased the pro-inflammatory bacteria Helicobacteraceae and Helicobacter. Deficiency in both PXR and CAR increased the relative abundance of Lactobacillus, which has bile salt hydrolase activity, corresponding to decreased primary taurine-conjugated bile acids (BAs) in feces, which may lead to higher internal burden of taurine and unconjugated BAs, both of which are linked to inflammation, oxidative stress, and cytotoxicity. The basal effect of PXR/CAR on the gut microbiome was distinct from pharmacological and toxicological activation of these receptors. Common PXR/CAR-targeted bacteria were identified, the majority of which were suppressed by these receptors. hPXR-TG mice had a distinct microbial profile as compared to wild-type mice. This study is the first to unveil the basal functions of PXR and CAR on the gut microbiome.

Pure drug nano-assemblies: A facile carrier-free nanoplatform for efficient cancer therapy

Nanoparticulate drug delivery systems (Nano-DDSs) have emerged as possible solution to the obstacles of anticancer drug delivery. However, the clinical outcomes and translation are restricted by several drawbacks, such as low drug loading, premature drug leakage and carrier-related toxicity. Recently, pure drug nano-assemblies (PDNAs), fabricated by the self-assembly or co-assembly of pure drug molecules, have attracted considerable attention. Their facile and reproducible preparation technique helps to remove the bottleneck of nanomedicines including quality control, scale-up production and clinical translation. Acting as both carriers and cargos, the carrier-free PDNAs have an ultra-high or even 100% drug loading. In addition, combination therapies based on PDNAs could possibly address the most intractable problems in cancer treatment, such as tumor metastasis and drug resistance. In the present review, the latest development of PDNAs for cancer treatment is overviewed. First, PDNAs are classified according to the composition of drug molecules, and the assembly mechanisms are discussed. Furthermore, the co-delivery of PDNAs for combination therapies is summarized, with special focus on the improvement of therapeutic outcomes. Finally, future prospects and challenges of PDNAs for efficient cancer therapy are spotlighted.

Papel do imunometabolismo, receptores Toll-Like e ECA 2 na COVID-19 / Role of immunometabolism, Toll-Like receptors, and ACE-2 in COVID-19

No combate à infecção pelo coronavírus 2 da síndrome respiratória aguda grave (SARS-CoV-2), o organismo se utiliza de mecanismos da imunidade inata, dentre eles os receptores Toll- Like (TLR), responsáveis pela sinalização da inflamação através da liberação de mediadores químicos e recrutamento de células imunitárias. Na patologia causada pela doença do SARS-CoV-2 2019 (COVID-19), ganha especial importância o TLR-4, visto que a sua estimulação exacerbada vem sendo relacionada ao estado hiperinflamatório em fases avançadas da COVID-19. Outro receptor que desempenha um papel primordial na infecção pelo SARS-CoV-2, servindo como porta de entrada para o vírus e progressão da doença, é a enzima conversora de angiotensina 2 (ECA 2), cuja ligação com a proteína S viral causa desregulação de vários sistemas fundamentais para a homeostase, como o sistema renina-angiotensina-aldosterona. Pacientes com doenças cardiometabólicas como obesidade, diabetes, aterosclerose e hipertensão vêm sendo classificados como alto risco para desenvolver as formas graves da COVID-19, visto que o estado inflamatório, já existente nessas doenças, pode ser agravado pelo desequilíbrio metabólico causado pelo SARS-CoV-2. A elucidação desses e de outros mecanismos relacionados à fisiopatologia da COVID-19 é imprescindível para uma melhora na estratificação de risco, nas escolhas terapêuticas e no prognóstico desses pacientes. Desta forma, nesta revisão objetivamos discutir as relações entre TLR-4, ECA 2, doenças cardiometabólicas, infecção pelo SARS-CoV-2 e gravidade da COVID-19.

In the fight against the infection caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the body uses mechanisms from the innate immune system, such as Toll-Like receptors (TLR), responsible for inflammation signaling through release of chemical mediators and recruitment of immune cells. In the disease caused by SARS-CoV-2 (COVID-19), TLR-4 assumes special importance because its exacerbated stimulation has been related to a hyperinflammatory state in advanced stages of COVID-19. Another receptor that plays a major role in SARS-CoV-2 infection, serving as a gateway to the virus and impacting disease progression, is angiotensin-converting enzyme 2 (ACE-2), whose binding to the viral S protein causes dysregulation of several key systems for homeostasis, such as the renin-angiotensin-aldosterone system. The elucidation of these and other mechanisms related to the pathophysiology of COVID-19 is essential for an improvement in risk stratification, therapeutic choices, and prognosis for these patients. Thus, we aimed to discuss in this review the relationships between TLR-4, ACE-2, cardiometabolic diseases, SARS-CoV-2 infection, and severity of COVID-19.

Protective effect of TAK242 blocking Toll-like receptor 4 pathway on septic myocardial injury and cardiac dysfunction / 中华危重病急救医学

Objective:To investigate the role of Toll-like receptor 4 (TLR4) pathway on myocardial injury and cardiac dysfunction in septic rats.Methods:According to the random number table, 18 male Sprague-Dawley (SD) rats were divided into control group, lipopolysaccharide (LPS) group and TLR4 specific inhibitor TAK242 pretreatment group (TAK242+LPS group) with 6 rats in each group. The rat model of septic cardiac dysfunction was induced by intraperitoneal injection of LPS 15 mg/kg, and the control group was given the same amount of normal saline. The TAK242+LPS group was intraperitoneally given injection of TAK242 [it was injected intraperitoneally at a dose of 3 mg/kg and dissolved in 10% dimethyl sulfoxide (DMSO) and 90% corn oil according to the concentration of 0.2 g/L] 3 hours before LPS stimulation. The control group and LPS group were given the same amount of 10% DMSO and 90% corn oil. The cardiac function of rats in each group was examined by Doppler echocardiography 14 hours after injection of LPS. The blood of abdominal aorta was taken and the level of serum troponin (cTn) was measured by enzyme linked immunosorbent assay (ELISA). Myocardial tissue was harvested for hematoxylin-eosin (HE) staining, and the morphological changes of myocardial tissue were observed under light microscope. The mRNA expressions of interleukin-6 (IL-6) and tumor necrosis factor-α(TNF-α) in myocardial tissue were detected by real-time fluorescence quantitative polymerase chain reaction (qPCR). The protein expression of TLR4 in myocardial tissue was observed by immunohistochemical method. Western blotting was used to detect the levels of TLR4, nuclear factor-κB p65 (NF-κB p65) and its phosphorylation (p-NF-κB p65) in myocardial tissue.Results:① The cardiac function and myocardial injury: Doppler echocardiography showed that the levels of left ventricular end-diastolic volume (LVEDV) and left ventricular end-systolic volume (LVESV) in the LPS group were significantly higher than those in the control group, while left ventricular ejection fraction (LVEF) and left ventricular shortened fraction (LVFS) were significantly lower than those in the control group. The degeneration, necrosis and inflammatory cell infiltration of cardiomyocytes were found with light microscope in the LPS group, and the levels of serum cTn were significantly higher than those in the control group, indicating that LPS-induced sepsis could cause cardiac dysfunction and myocardial injury. TAK242 blocking TLR4 pathway had a protective effect on cardiac function and myocardium during sepsis. LVEDV and LVESV in the TAK242+LPS group were significantly lower than those in the LPS group [LVEDV (μL): 71.25±21.16 vs. 118.01±11.89, LVESV (μL): 9.57±5.75 vs. 32.70±9.22, both P < 0.01]. LVEF and LVFS were significantly higher than those in the LPS group [LVEF: 0.868±0.075 vs. 0.722±0.095, LVFS: (59.88±8.46)% vs. (42.37±8.71)%, both P < 0.05]. Myocardial tissue injury was significantly reduced, and the serum cTn level was significantly lower than that in the LPS group (μg/L: 107.85±21.38 vs. 152.25±27.46, P < 0.05). ② Inflammatory parameters: the results of qPCR, Western blotting and immunohistochemistry showed that the mRNA expressions of IL-6 and TNF-α, the expression of TLR4 protein and the p-NF-κB p65/NF-κB p65 ratio in the LPS group were significantly higher than those in the control group, indicating that LPS-induced sepsis could activate the inflammatory response mediated by TLR4/NF-κB pathway in the heart. However, blocking the TLR4 pathway by TAK242 could inhibit the TLR4/NF-κB pathway and reduce the myocardial inflammation in septic rats. The mRNA expressions of IL-6 and TNF-α, the expression of TLR4 protein and the p-NF-κB p65/NF-κB p65 ratio in the TAK242+LPS group were significantly lower than those in the LPS group [IL-6 mRNA (2 -ΔΔCT): 10.44±3.30 vs. 107.50±29.48, TNF-α mRNA (2 -ΔΔCT): 2.38±0.68 vs. 3.77±0.56, TLR4 protein (TLR4/GAPDH): 0.39±0.01 vs. 0.58±0.04, p-NF-κB p65/NF-κB p65 ratio: 1.21±0.11 vs. 2.10±0.18, all P < 0.05]. Conclusion:TAK242 can protect LPS-induced cardiac dysfunction and myocardial injury by blocking the TLR4 mediated inflammatory response.

Effect of acupuncture on intestinal flora and TLR4 in brain and intestinal tissues in rats with stress gastric ulcer / 中国针灸

OBJECTIVE@#To observe the effect of acupuncture at "Guanyuan" (CV 4) and "Xiajuxu" (ST 39) on intestinal flora and Toll-like receptors-4 (TLR4) in brain and intestinal tissue in rats with stress gastric ulcer (SGU), and to explore the possible mechanism of acupuncture for SGU.@*METHODS@#Thirty-one male SD rats were randomly divided into a blank group (@*RESULTS@#Compared with the blank group, the gastric mucosal damage index was significantly increased in the model group (@*CONCLUSION@#Acupuncture at "Guanyuan" (CV 4) and "Xiajuxu" (ST 39) could alleviate SGU in rats, and its mechanism may be related to increasing the diversity of intestinal flora, promoting the disorder of intestinal flora to normal, and reducing the overexpression of TLR4 in brain and intestinal tissues.

Opposite effects of miR-155 in the initial and later stages of lipopolysaccharide (LPS)-induced inflammatory response / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

Although microRNA-155 (miR-155) is considered a pro-inflammatory mediator, cumulative evidence indicates that it also has anti-inflammatory effects in macrophages and dendritic cells. In this study, we identified the dramatic expression changes of more than half of potential miR-155-targeted genes upon lipopolysaccharide (LPS) stimulation; 223 genes were down-regulated and 85 genes were up-regulated, including suppressor of cytokine signaling 1 (

Association between Toll-like receptor 4 and pancreatic cancer / 临床肝胆病杂志

Toll-like receptor 4 (TLR4) is a key regulator of innate and adaptive immune response. The role of TLR4 in pancreatic diseases is a research hotspot in recent years, and a large number of studies have shown that TLR4 is closely associated with pancreatic cancer. This article mainly discusses the abnormal expression and regulation mechanism of TLR4 in pancreatic cancer and its potential in cancer treatment, so as to provide new ideas for the pathogenesis and treatment of pancreatic cancer.

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.