OBJECTIVES: To investigate the role of the calcium/calmodulin (CaM)-mediated activation of calcineurin (CN)-nuclear factor of activated T cells (NFAT) signaling pathway in mediating the regulatory effect of hyperforin (HY) on stress-induced depression-like disorder (DP) in mice. METHODS: C57BL/6J mice were randomly divided into control group, DP model group, and hyperforin treatment group (n=15). Behavioral changes of the mice were assessed using open field test (OFT), sucrose preference test (SPT), tail suspension test (TST), light/dark box test (LDB), and novel object suppression test (NSFT). Immunohistochemistry was used to detect tyrosine hydroxylase (TH) expression in the CA1 region of the hippocampus, and serum serotonin (5-HT) and norepinephrine (NA) levels were detected with ELISA. Western blotting was used to analyze the expressions of TNF-α, IL-1β, IL-2, and CN-NFAT pathway proteins. In cultured BV-2 microglial cells with lipopolysaccharide (LPS) stimulation, the effects of hyperforin and CN inhibitor (CNIS) on expressions of ionized calcium-binding adapter molecule 1 (IBA-1), 5-HT, NA, inflammatory cytokines and CN-NFAT pathway proteins were examined using immunofluorescence assay, ELISA or Western blotting. RESULTS: Compared with the control mice, the mice in DP group showed significantly reduced activity in OFT, decreased sucrose consumption in SPT, reduced shuttle crossing in LDB, and lowered food intake in NSFT with significantly increased immobility in TST. The mice with DP showed significantly decreased TH-positive neurons, lowered 5-HT and NA levels, and increased expressions of TNF-α, IL-1β, IL-2 and CaM-CN-NFAT pathway proteins. In cultured BV-2 cells, LPS stimulation strongly increased cellular IBA-1 expression, decreased the levels of neurotransmitters (5-HT and NA), and increased the levels of inflammatory cytokines and CN-NFAT signaling, and these changes were effectively reversed by treatment with hyperforin or CNIS. CONCLUSIONS Hyperforin improves stress-induced depression-like behaviors in mice and activated BV-2 cells by targeting the CN-NFAT signaling pathway.
Animals ; Mice, Inbred C57BL ; Mice ; Microglia/drug effects* ; Depression/etiology* ; Perylene/pharmacology* ; Calcineurin/metabolism* ; NFATC Transcription Factors/metabolism* ; Calcium Signaling/drug effects* ; Stress, Psychological ; Phloroglucinol/pharmacology* ; Signal Transduction ; Male ; Behavior, Animal/drug effects* ; Terpenes

Objective To investigate the effect and mechanism of Congrong Shujing Granules on promoting microglial polarization and inhibiting neuroinflammation through the nucleotide-binding oligomeric domain-like receptor protein 3(NLRP3)/Caspase-1 signaling pathway.Methods Twenty Sprague-Dawley rats were assigned to the blank serum and Congrong Shujing Granules containing serum groups using random number table method,with 10 rats in each group.Rats in the Congrong Shujing Granules containing serum group received intragastric administration of Congrong Shujing Granules(2.57 g/kg)and the rats in the blank serum group received intragastric administration of physiological saline of equal volume.Blank serum and Congrong Shujing Granules containing serum were prepared separately.Mouse microglia cells BV-2 were cultured in vitro,and the optimal concentration of 1-methyl-4-phenylpyridine(MPP+)and optimal volume fraction of Congrong Shujing Granules containing serum were selected by the CCK-8 assay and immunofluorescence staining.And the NLRP3 inhibitor MCC950 was used as a postive control.Cells were divided into the blank serum group(10%blank serum),model group(10%blank serum+500 μmol/L MPP+),Congrong Shujing Granules containing serum group(10%Congrong Shujing Granules containing serum+500 μmol/L MPP+),and MCC950 group(10%blank serum+10 μmol/L MCC950+500 μmol/L MPP+),and intervened separately.After 14 h of intervention,morphological changes in BV-2 cells were observed.The contents of interleukin(IL)-1β,tumor necrosis factor-α(TNF-α),IL-6,and IL-4 were detected by an enzyme-linked immunosorbent assay.The mRNA expressions of differentiation cluster 86(CD86),inducible nitric oxide synthase(iNOS),CD206,and arginase 1(Arg1)were detected by real-time fluorescence PCR.The expressions of CD86,Arg1,Ionized calcium binding adaptor molecule 1(Iba1),and NLRP3 were detected by immunofluorescence staining.The expression of iNOS,Arg1,TNF-α,IL-6,IL-4,NLRP3,pro-cysteinyl aspartate specific proteinase 1(pro-Caspase-1),and Caspase-1 proteins was detected by Western blotting.Results Iba1 activation and expression increased under the MPP+(12 h,500 μmol/L)intervention(P<0.05),and cell viability was not affected.There was no statistically significant effect on cell viability after treatment with 10%Congrong Shujing Granules containing serum alone or in combination with MPP+(P>0.05).Compared to the blank serum group,BV-2 cells in the model group showed multiple branches and protruded in the shape of an amoeba.The contents of IL-1β,TNF-α,and IL-6 increased,while the contents of IL-4 decreased.The mRNA expressions of CD86 and iNOS increased,while mRNA expressions of CD206 and Arg1 decreased.The mean fluorescence intensity of CD86,Iba1,and NLRP3 increased,while the mean fluorescence intensity of Arg1 decreased.The protein expressions of iNOS,TNF-α,IL-6,NLRP3,pro-Caspase-1,and Caspase-1 increased,while the protein expressions of Arg1,IL-4 decreased,P<0.05.Compared to the model group,the Congrong Shujing Granules containing serum group and MCC950 group showed a decrease in the branch of cell protrusions,reduced cell activation,decreased levels of IL-1β,TNF-α,and IL-6,increased levels of IL-4,decreased expression of CD86 and iNOS mRNA,increased expression of CD206 mRNA,the decreased mean fluorescence intensity of CD86,Iba1,and NLRP3,the increased mean fluorescence intensity of Arg1,decreased expression of iNOS,TNF-α,IL-6,NLRP3,pro-Caspase-1,and Caspase-1 proteins,and increased expression of Arg1 and IL-4 proteins,P<0.05.Conclusion Congrong Shujing Granules containing serum may alleviate the MPP+-induced neuroinflammatory response by inhibiting the NLRP3/Caspase-1 signaling pathway to regulate M1/M2 phenotype polarization of microglia.

Objective To investigate the effect and mechanism of Congrong Shujing Granules on promoting microglial polarization and inhibiting neuroinflammation through the nucleotide-binding oligomeric domain-like receptor protein 3(NLRP3)/Caspase-1 signaling pathway.Methods Twenty Sprague-Dawley rats were assigned to the blank serum and Congrong Shujing Granules containing serum groups using random number table method,with 10 rats in each group.Rats in the Congrong Shujing Granules containing serum group received intragastric administration of Congrong Shujing Granules(2.57 g/kg)and the rats in the blank serum group received intragastric administration of physiological saline of equal volume.Blank serum and Congrong Shujing Granules containing serum were prepared separately.Mouse microglia cells BV-2 were cultured in vitro,and the optimal concentration of 1-methyl-4-phenylpyridine(MPP+)and optimal volume fraction of Congrong Shujing Granules containing serum were selected by the CCK-8 assay and immunofluorescence staining.And the NLRP3 inhibitor MCC950 was used as a postive control.Cells were divided into the blank serum group(10%blank serum),model group(10%blank serum+500 μmol/L MPP+),Congrong Shujing Granules containing serum group(10%Congrong Shujing Granules containing serum+500 μmol/L MPP+),and MCC950 group(10%blank serum+10 μmol/L MCC950+500 μmol/L MPP+),and intervened separately.After 14 h of intervention,morphological changes in BV-2 cells were observed.The contents of interleukin(IL)-1β,tumor necrosis factor-α(TNF-α),IL-6,and IL-4 were detected by an enzyme-linked immunosorbent assay.The mRNA expressions of differentiation cluster 86(CD86),inducible nitric oxide synthase(iNOS),CD206,and arginase 1(Arg1)were detected by real-time fluorescence PCR.The expressions of CD86,Arg1,Ionized calcium binding adaptor molecule 1(Iba1),and NLRP3 were detected by immunofluorescence staining.The expression of iNOS,Arg1,TNF-α,IL-6,IL-4,NLRP3,pro-cysteinyl aspartate specific proteinase 1(pro-Caspase-1),and Caspase-1 proteins was detected by Western blotting.Results Iba1 activation and expression increased under the MPP+(12 h,500 μmol/L)intervention(P<0.05),and cell viability was not affected.There was no statistically significant effect on cell viability after treatment with 10%Congrong Shujing Granules containing serum alone or in combination with MPP+(P>0.05).Compared to the blank serum group,BV-2 cells in the model group showed multiple branches and protruded in the shape of an amoeba.The contents of IL-1β,TNF-α,and IL-6 increased,while the contents of IL-4 decreased.The mRNA expressions of CD86 and iNOS increased,while mRNA expressions of CD206 and Arg1 decreased.The mean fluorescence intensity of CD86,Iba1,and NLRP3 increased,while the mean fluorescence intensity of Arg1 decreased.The protein expressions of iNOS,TNF-α,IL-6,NLRP3,pro-Caspase-1,and Caspase-1 increased,while the protein expressions of Arg1,IL-4 decreased,P<0.05.Compared to the model group,the Congrong Shujing Granules containing serum group and MCC950 group showed a decrease in the branch of cell protrusions,reduced cell activation,decreased levels of IL-1β,TNF-α,and IL-6,increased levels of IL-4,decreased expression of CD86 and iNOS mRNA,increased expression of CD206 mRNA,the decreased mean fluorescence intensity of CD86,Iba1,and NLRP3,the increased mean fluorescence intensity of Arg1,decreased expression of iNOS,TNF-α,IL-6,NLRP3,pro-Caspase-1,and Caspase-1 proteins,and increased expression of Arg1 and IL-4 proteins,P<0.05.Conclusion Congrong Shujing Granules containing serum may alleviate the MPP+-induced neuroinflammatory response by inhibiting the NLRP3/Caspase-1 signaling pathway to regulate M1/M2 phenotype polarization of microglia.

Biocompatibility has always been the focal point in the study of biomaterials applied to medical apparatus. But at present, the standard system hasn't formed completely to evaluate the biomaterials. In recent years scientists tend to use general standard of evaluation. In this article, based on the recalcification time, the adhesion of platelets as well as the total quantity of plasma protein and some other evidences, combining with the Analytic hierarchy process (AHP), the method of general evaluation on the biocompatibility of anticoagulant biomaterials was discussed.
Anticoagulants ; analysis ; Biocompatible Materials ; standards ; Blood Proteins ; analysis ; Humans ; Materials Testing ; methods ; standards ; Platelet Adhesiveness ; drug effects

Good anticoagulant biomaterials need good surface chemical properties, good mechanics performances and particularly good characteristics of biocompatibility, including tissue compatibility and hemocompatibility. In order to understand with greater clearness the anticoagulant biomaterial, we have to characterize them by different methods. In this paper, the approaches to assessing and displaying the characteristics of anticoagulant biomaterial are reviewed in three aspects, namely the surface chemical properties and structure, the mechanics performances the and the biocompatibility of anticoagulant biomaterial.
Anticoagulants ; Biocompatible Materials ; chemistry ; Blood Coagulation ; drug effects ; Humans ; Materials Testing ; Prostheses and Implants ; adverse effects ; Prosthesis Design ; Surface Properties