Gut microbiota is one of the complicated eco-systems in human body.A large amount of bacteria colonize in the healthy human intestine, which not only play a variety of biological functions, but also are associated with various diseases.By adding microecological agents (probiotics, prebiotics, and synbiotics), we are able to improve the gut microbiota structure and reduce the related carcinogenic metabolites, and also to improve the clinical manifestations of certain diseases.Therefore, it is meaningful to apply microecological agents (probiotics, prebiotics, and synbiotics) both in healthy population and patients.We reviewed the researches on microecological aspect of gut microbiota-related diseases, aiming to shed some light on fully understanding and popularizing of the application of microecological agents among different populations.

Objective To investigate the expression of JMJD3 and its functions for cell growth in human gastric carcinoma.Methods The expression of JMJD3 was detected by immunohistochemistry.Recombinant JMJD3 plasmids were transfected into MGC-803 cells.Cell proliferation was determined by CCK-8 assay and cell apoptosis was detected by flow cytometry.Results The expression of JMJD3 was down-regulated in gastric carcinoma tissues (P ＜ 0.05).Low expression of JMJD3 was associated with advanced TNM stage (Ⅲ+Ⅳ,P ＜ 0.05).Overexpression of JMJD3 could inhibit cell proliferation and induce cell apoptosis (P ＜ 0.05).Conclusion Low expression of JMJD3 was commonly existed in gastric cancer.JMJD3 could inhibit cell proliferation and induce cell apoptosis.

Objective To investigate the effects of chondroitin sulfate proteoglycans (CSPGs) degradation induced by chondroitinase ABC (ChABC) on the rat flash visual evoked potential (F-VEP) around the end of the critical period of visual development.Methods A total of 30 Long Evans rats were randomly divided into 3 groups according to their age of 14,21 and 35 d.Then in each group,5 animals served as CSPGs degradation model induced by ChABC microinjection into 5 sites of the rat cortex [750 nL (48 U/ml) for 1 site,for 2 times at a 3-day interval],and the other 5 rats served as normal control and received normal saline microinjection.Immunofluorescence microscopy and laser scanning confocal microscopy were used to determine the establishment of animal model.F-VEPs were evaluated in 7 d after first microinjection.Results In the normal rats,the latency of F-VEP main wave became shorter with the increase of their age,the number was [(63.5?10.1),(50.8?6.4),(44.9?6.3) ms,P

Objective To measure the number of peripheral blood CD34+ hematopoietic stem/progenitor cells (HSC/HPCs) and membrane expression of CD34 on these cells in patients with SLE. Methods Lymphocytes were isolated from peripheral blood of 30 patients with SLE and 14 normal human controls. Flow cytometry using FITC-labeled antibodies was performed to determine the percentage of CD34+ HSC/HPCs and mean fluorescence intensity (MFI) of CD34 on these cells. Their correlation with clinical data was analyzed.Results The percentage of CD34+ HSC/HPCs in peripheral lymphocytes was (0.15 ± 0.10)% and (0.09 ±0.07)% in active and stable SLE patients, respectively, significantly lower than that in normal controls [(0.37 +0.17)%, F = 17.18, P ＜ 0.01], however, there was no significant difference between active and stable SLE patients (t = 1.51, P＞ 0.05). The MFI of CD34 was higher in active SLE patients than in the normal controls (41.35 ± 19.24 vs. 27.43 ± 7.57, F= 3.13, P ＜ 0.05), but no difference was observed between stable SLE patients and normal controls (F= 3.13, P ＞ 0.05). In patients with SLE, the percentage of CD34+ HSC/HPCs was negatively correlated with serum IgG levels (r = -0.588, P ＜ 0.01 ), but uncorrelated with SLE disease activity index (SLEDAI) or serum levels of complement, anti-dsDNA antibodies, anti-C1q antibodies, antinucleosome antibodies, etc. Conclusions The count of CD34+ HSC/HPCs is reduced while the MFI of CD34 antigen is elevated in SLE patients, hinting that there is a functional abnormality of HSC/HPCs in SLE patients, which may be involved in the pathogenesis of SLE.

Objective To measure the number of peripheral blood CD34+ hematopoietic stem/progenitor cells (HSC/HPC) expression of CD34 in the peripheral blood of patients with rheumatoid arthritis and exploreits relationship with clinical manifestations. Methods CD34+ HSC/HPCs in the peripheral blood of RA patients (n=32) and healthy controls (n=16) were detected using flow cytometry. The relationship between the frequency of HSC/HPCs, mean fluorescence intensities (MFI) of CD34 and clinical manifestations and rheumatoid factor (RF), anti-cyclic citrullinated peptide (CCP) antibodies, disease activity score (DAS) 28,X rays stages and healthy assessment questionnaire (HAQ) were analyzed. Student's t-test and pearont test were used for statistical analysis. Results Frequency of CD34+ HSC/HPC in the peripheral blood of RA patients was decreased compared with normal controls [ (0.13±0.09)% vs (0.38±0.21)%, P＜0.05 ], CD34 MFIwere higher in RA patients than those in the normal controls (57±33 vs 3111, P＜0.05). The frequency was positively correlated with the number of (RBC red blood cell), (Hb hemoglobin), and was negatively correlated with C-reactive protein (CRP), and the MFI of RA patients was positively correlated with healthy assessment questionnaire (HAQ) and X ray stages, but negatively correlated with the number of platelets.Conclusion CD34+ HSC/HPC of the peripheral blood of RA patients are significantly abnormal, which is characterized by decreased CD34+ hematopoietic stem cell, and the decrease is positively correlated with RBC and Hb, but negatively correlated with CRP. CD34+ hematopoietic stem cell may play an important role in the pathogenesis of RA.

Oocyte-like cells (OLC) can be generated by stem cells after the induction and differentiation in vitro, and maturated when transplanted in vivo to improve the development potential. Human amniotic fluid stem cells (hAFSC) were cultured for 10 days in porcine follicle fluid (pFF) that was extracted from the medium follicle with high levels of hormones and Bmp 15 protein. After the induction, the cell aggregates showed the germ cell-like cells and produced the germ cell marker oct4, and triggered epigenetic changes with high expression of methylation transferase gene dnmt3b. The cell aggregates were packaged into porcine theca folliculi to form grafts, which were then transplanted into mouse renal capsule. After one month of transplantation, the morphology of OLC from a graft was not only similar to oocytes, but also expressed the germ cells markers (oct4, nanog, stella, ifitm3, dazl, nanos3, bmp15, and gd9). The results demonstrate that the in vivo differentiation model was useful for OLC development.
Amniotic Fluid ; cytology ; Animals ; Biomarkers ; Cell Differentiation ; Female ; Humans ; Mice ; Oocytes ; cytology ; Ovarian Follicle ; Stem Cells ; cytology ; Swine

The aim of this study is to identify the express specificity of bone morphogenetic protein 15 (Bmp15) in porcine. The pBMP15-EGFP reporter vector was constructed from the 2.2 kb fragment of porcine bmp15 promoter to trace the differentiation process of stem cells into oocyte-like cells. We used porcine ovary and Chinese Hamster Ovary cell line (CHO), mouse myoblast cell line (C2C12) and porcine amniotic fluid stem cell (pAFSC) to investigate the expression and regulation of this gene via RT-PCR, immunofluorescence, cell transfection, and microinjection methods. We also used single layer cell differentiation to detect the application potential of bmp15. The results show that bmp15 gene was specifically expressed in the porcine ovary and CHO rather than in C2C12 and pAFSC. In addition, the characteristic of tissue-specific of Bmp15 was detected on CHO instead of other cell lines by transient transfection. We also detected the expression of Bmp15 in oocyte at different development stages by immunofluorescence of fixed paraffin-embedded ovary sections. Furthermore, microinjection results show that bmp15 expressed in oocytes at 18 h of maturation in vitro, and continued up to 4-cell stage embryos. Most importantly, we found that the expression of Bmp15 started at day 12 after inducing pAFSC into oocyte-like cells by transfection; green fluorescent was visible in round cell masses. It indicated that bmp15 has the expression specificity and the pBMP15-EGFP reporter vector can be used to trace Bmp15 action in the differentiation of stem cells into germ cells.
Animals ; Bone Morphogenetic Protein 15 ; genetics ; CHO Cells ; Cell Differentiation ; Cricetinae ; Cricetulus ; Female ; Genes, Reporter ; Genetic Vectors ; Mice ; Microinjections ; Myoblasts ; cytology ; Oocytes ; metabolism ; Ovary ; metabolism ; Stem Cells ; cytology ; Swine

Objective To determine the inhibitory effect and mechanism of exogenous carbon monoxide against excessive neutrophil infiltration in liver and lung tissues during sepsis.Methods Thirty-two mice were subjected to sham operation (sham group),cecal ligation and perforation (CLP) group,CLP with 8 mg/kg of exogenous carbon monoxide releasing molecule Ⅱ (CORM-2) (CORM-2 group),and CLP with 8 mg/kg of inactive variants of CORM-2 (iCORM-2) (iCORM-2 group) according to the random number table,with 8 mice per group.Liver and lung tissues were collected at 24 hours after surgery to examine the pathologic changes,myeloperoxidase (MPO) activity and malonaldehyde (MDA) content.Another 60 mice were enrolled into the same 4 groups with 15 mice per group and were tested for 72-hour survival rate.Bone marrow neutrophils were isolated and divided into normal control group,1 μg/ml lipopolysaccharide (LPS) group,1 μg/ml LPS plus 10 μmol/L CORM-2 group (low dose group),1 μg/ml LPS plus 50 μmol/L CORM-2 group (high dose group),1 μg/ml LPS plus 50 μmol/L iCORM-2 group (iCORM-2 group).Under the agarose chemotaxis,qPCR and immunofluorescence detection of formyl peptide receptor 1 (FPR1) were performed.Results CLP group presented enhanced activity of MPO [liver:(9.1 ± 1.1) U/g,lung:(16.3 ± 2.8) U/g],increased MDA content [liver:(76.5 ±11.3) nmol/mg,lung:(32.4 ± 10.3) nmol/mg] and 72-hour survival rate of 20％ as compared with the sham group (all P ＜ 0.05).CORM-2 group showed inhibited activity of MPO [liver:(5.2 ± 0.8) U/g,lung:(7.5 ± 2.4) U/g],increased MDA content [liver:(46.7 ± 6.1) nmol/mg,lung:(23.8 ±7.3) nmol/mg] and 72-hour survival rate of 67％ as compared with the sham group (all P ＜ 0.05).LPS enhanced neutrophil migration (61.3 ± 7.1) (P ＜ 0.05) and expression of FPR1 which was enriched in the membrane.Meanwhile,neutrophil migration was significantly inhibited in a dose-dependent of CORM-2 (low dose group:43.3 ±6.1,high-dose group:23.3 ±5.9) (P＜0.05).Conclusions Exogenous carbon monoxide is effective to inhibit the excessive neutrophil infiltration,attenuate oxidative stress or pathological injury,and improve the survival from sepsis.The mechanism is associated with the down-regulation of FPR1,inhibition of FPR1 enrichment in the membrane,and decreased neutrophil migration.

Objective To investigate the suppressive effect of exogenous carbon monoxide (CO) on abnormal platelet exocytosis and its possible molecular mechanism. Methods Venous blood was collected from healthy volunteers. Platelet-rich plasma (PRP) was isolated from the blood by differential centrifugation. The PRP was randomly divided into five groups by random number table, namely normal control group, lipopolysaccharide (LPS) group (challenged with 10 mg/L LPS), inactively exogenous carbon monoxide releasing molecule 2 (iCORM-2) group (given 10 mg/L LPS + 50 μmol/L iCORM-2 for intervention), exogenous carbon monoxide releasing molecule 2 (CORM-2) 10 μmol/L and 50 μmol/L groups (given 10 mg/L LPS + CORM-2 10 μmol/L or 50 μmol/L for intervention). After 30 minutes, enzyme linked immunosorbent assay (ELISA) was used to determine the platelet-derived growth factor BB (PDGF-BB) and matrix metalloproteinase 2 (MMP-2). Chemical fluorescein method was used to determine the platelet adenosine triphosphate (ATP). Flow cytometer was used to determine the expression of P-selectin. The expressions of Toll-like receptor 4 (TLR4), phosphorylation of protein kinase Cθ (PKCθ) and syntaxin binding protein 1 (STXBP-1) were determined by Western Bolt. The soluble N-ethylmaleimide-sensitive factor-attachment protein receptors (SNAREs) complex formation [syntaxin 2-synaptosomal-associated protein 23-vesicle associated membrane protein 8 (STX2-SNAP23-VAMP8)] mediated by STXBP-1 was determined by immunoprecipitation. Results ① Compared with normal control group, the platelet release of PDGF-BB, MMP-2 and ATP was significantly increased after LPS challenge, and the P-selectin expression of platelet was also obviously up-regulated [PDGF-BB (μg/L): 127.53±1.78 vs. 94.35±5.84, MMP-2 (ng/L): 51.87±9.20 vs. 35.83±3.17, ATP (μmol/L): 1.288±0.056 vs. 0.975±0.010, P-selectin: (3.93±0.19)% vs. (0.44±0.10)%, all P < 0.05]. The increases in platelet release of PDGF-BB, MMP-2 and ATP were suppressed by 10 μmol/L or 50 μmol/L CORM-2 administration, as well as high-expression of P-selectin in a dose-dependent manner [PDGF-BB (μg/L): 114.68±1.35, 97.08±6.14 vs. 127.53±1.78, MMP-2 (ng/L): 32.67±8.00, 24.63±1.63 vs. 51.87±9.20, ATP (μmol/L): 0.999±0.015, 0.965±0.008 vs. 1.288±0.056, P-selectin: (1.95±0.27)%, (0.94±0.11)% vs. (3.93±0.19)%, all P < 0.05]. ② Compared with normal control group, LPS challenge resulted in a significant increase in the expression of TLR4 and the phosphorylation of PKCθ and STXBP-1 [TLR4 (gray value): 1.21±0.38 vs. 0.67±0.06, p-PKCθ (gray value): 1.36±0.20 vs. 0.44±0.03, p-STXBP-1 (gray value): 1.13±0.06 vs. 0.59±0.04, all P < 0.05]. The increases in above parameters were suppressed by 10 μmol/L or 50 μmol/L CORM-2 administration in a dose-dependent manner [TLR4 (gray value): 0.76±0.05, 0.65±0.04 vs. 1.21±0.38; p-PKCθ (gray value): 0.71±0.07, 0.47±0.10 vs. 1.36±0.20; p-STXBP-1 (gray value): 0.56±0.02, 0.48±0.01 vs. 1.13±0.06, all P < 0.05]. ③ Compared with normal control group, the SNAREs proteins in platelet that combined with STXBP-1, including STX2, SNAP23 and VAMP8, were obviously increased after LPS challenge [STX2 (gray value): 1.35±0.06 vs. 0.57±0.04, SNAP23 (gray value): 0.97±0.04 vs. 0.30±0.12, VAMP8 (gray value): 1.37±0.12 vs. 0.77±0.10, all P < 0.05]. The increases in SNAREs complex formation were suppressed by 10 μmol/L or 50 μmol/L CORM-2 administration in a dose-dependent manner [STX2 (gray value): 0.77±0.02, 0.39±0.03 vs. 1.35±0.06, SNAP23 (gray value): 0.41±0.03, 0.22±0.08 vs. 0.97±0.04, VAMP8 (gray value): 0.85±0.07, 0.66±0.07 vs. 1.37±0.12, all P < 0.05]. There was no significant difference in the above mentioned parameters between iCORM-2 group and LPS group. Conclusions LPS-induced abnormal secretion of platelet was suppressed by CORM-2 administration. The mechanism may involve the TLR4/PKCθ/STXBP-1 signaling pathway activation and the SNAREs complex formation.

Objective To observe the effects of nourishing lung and kidney formulas on inflammatory response of alveolar epithelial cells stimulated by monocytes conditioned medium and study the anti-inflammatory mechanism of the formulas for the treatment of chronic obstructive pulmonary diseases (COPD).Methods The reproduction of inflammation models of A549 cells were stimulated by monocyte THP-1 cell strain conditioned medium.A549 cells were randomly divided into blank control group (20％ blank rabbits serum),model group (20％ blank rabbits serum+25.0％ THP-1 cell conditioned medium),nuclear transcription factor activator protein-1 (AP-1) pathway inhibitor T-5224group (20％ blank rabbits serum+25.0％ THP-1 cells conditioned medium+100 μmol/L T-5224),nourishing lung and kidney group (20％ rabbits serum with nourishing lung and kidney formulas+25.0％ THP-1 cells conditioned medium).The contents of interleukins (IL-6,IL-8),tumor necrosis factor-α (TNF-αα),matrix metalloprotein-9 (MMP-9) in cell culture supernatant were detected with enzyme linked immunosorbent assay (ELISA),the supernatant content of malondialdehyde (MDA) was detected with thibabituric acid (TBA) method,the total activity of superoxide dismutase (T-SOD) was detected with hydroxylamine method,and the activity of AP-1 pathway was detected with electrophoretic mobility shift assay (EMSA) method.Results Compared with the blank control group,the A549 cell proliferation were significantly increased at 24 hours,48 hours stimulation by 25.0％ cell conditioned medium (A value:24 hours was 0.41 ± 0.02 vs.0.37 ± 0.04,48 hours was 1.30 ± 0.09 vs.1.15 ± 0.19).Compared with the blank control group,the contents of IL-6,IL-8,TNF-αα,MMP-9,MDA,AP-1 expression were significantly increased in model group [IL-6 (ng/L):35.00±3.63 vs.23.15±1.72,IL-8 (ng/L):273.09± 164.36 vs.231.45±33.90,TNF-α(ng/L):51.61 ± 9.51 vs.28.87 ± 3.34,MMP-9 (ng/L):442.85 ± 78.86 vs.235.60 ± 14.62,MDA (μmol/L):6.90 ± 0.11 vs.6.01 ± 0.12,AP-1 expression (A value):2.260 ± 0.062 vs.1.000 ± 0.000],MDA/T-SOD ratio was increased (4.43 ± 0.05vs.3.96 ± 0.06).Compared with model group,the levels of IL-8 (ng/L:100.29 ± 17.03),TNF-α (ng/L:25.13 ± 0.46),AP-1 expression (A value:1.38 ± 0.02),and the MDA/T-SOD ratio (4.23 ± 0.23) in T-5224 group,and MMP-9 (ng/L:195.44±9.80),MDA (μmol/L:5.86±0.30),MDA/T-SOD ratio (3.56±0.41),AP-1 expression (A value:0.76 ± 0.01) in nourishing lung and kidney group were all reduced significantly (all P ＜ 0.05).Conclusion Nourishing lung and kidney formulas can suppress the inflammatory response through regulating the alveolar epithelial cells AP-1 signaling pathways.