Objective To explore the differences of Th17 population and serum transforming growth factor (TGF)-β1 levels between early-and late-stage primary biliary cirrhosis (PBC) and their roles in pathogenesis.Methods Peripheral Th17 counts were analyzed by flow cytometry.The expression of IL-17A in peripheral blood mononuclear cells and TGF-β1 were measured by real-time quantitative polymerase chain reaction.Serum concentration of TGF-β1 was measured by enzyme-linked immunosorbent assay.Liver biopsies were stained with hematoxylin-eosin to determine the pathological stage.Results were evaluated using KrustalWallis test followed by Mann-Whitney U tests for comparisons of Th17 population between patients with early and late PBC,patients with chronic hepatitis B (CHB) and health controls (HCs).ANOVA followed by LSD t-tests were used for comparing IL-17 mRNA,TGF-β1 mRNA and TGF-β1 serum concentration between groups.The correlations between Mayo risk score and peripheral Th17 of PBC patients,Mayo risk score and serum concentration of TGF-β1 was analyzed by Pearson correlation analysis separately.Results The peripheral Th17 population increased in patients with early PBC (1.03±0.33)％,compared to those with late PBC [(0.48± 0.13％,U=14.0,P＜0.01],CHB [(0.56±0.35)％,U=104.5,P＜0.01],and HCs [(0.36±0.17)％,U=8.0,P＜0.01],while TGF-β1 changed in the opposite direction.Serum concentration of TGF-β1 elevated in late PBC (43.0± 18.7) ng/ml compared with early PBC (29.5±12.2) ng/ml,t=2.85,P=0.006.Conclusion The opposite changes of Th17 population and TGF-β1 level in early and late PBC indicated their different roles in different stages.Th17 may contribute to the autoimmune response in early PBC,participate in the occurrence of autoimmune inflammation,while TGF-β1 to fibrogenesis in late stage.In addition,the possible regulation mechanisms of differentiation of Th17 by TGF-β1 cannot be ignored.

The origin and development of round magnetic needle was explored, and the structure of round magnetic needle was introduced in detail, including the handle, the body and the tip of the needle. The clinical opera tion of round magnetic needle were standardized from the aspects of the methods of holding needle, manipulation skill, tapping position, strength of manipulation, application scope and matters needing attention, which laid foundation for the popularization and application of round magnetic needle.
Acupuncture Therapy ; history ; instrumentation ; methods ; standards ; China ; History, Ancient ; Humans ; Medicine in Literature ; Needles ; history ; standards

Aging can cause degenerative changes in the function of multiple tissues and organs in the body. Gastrointestinal diseases and intestinal dysfunction are very common in the elderly people. The purpose of this study is to explore the effect of the total extract of Astragalus membranaceus (Fisch.) Bge. on intestinal function and gut microbiota homeostasis in natural aging mice, which will provide clues for further mechanism study. The natural aging mice model is established and animal experiments follow the regulations of the Animal Ethics Committee of Nanjing University of Traditional Chinese Medicine. The overall health of the mice was evaluated by the "frailty index" scoring method. The intestinal absorption and transport function were measured by detecting intestinal glucose absorption capacity, transport time, lipase and amylase activities of aging mice. Intestinal inflammation was assessed by detecting inflammatory cytokines by enzyme-linked immunosorbent assay (ELISA). The pathological changes in the intestines of aging mice were tested by hematoxylin-eosin (H&E) staining and alizarin blue (AB) staining. The qRT-PCR method was used to explore the gene transcription level related with the proliferation and differentiation of intestinal stem cells. Microbiota analysis based on 16S rDNA were used to evaluate the composition of gut microbiota. The results showed that Astragalus had a tendency to reduce the "frailty index" of aging mice, but did not show a significant difference. In some indicators of aging phenotype, Astragalus has the most significant effect on hair loss and physical fitness. In terms of intestinal function, Astragalus could increase intestinal glucose absorption capacity, shorten intestinal transportation time and promote lipase secretion in aging mice. The levels of inflammatory cytokines such as tumor necrosis factor-α (TNF-‍α) in the aging intestinal tissue were reduced after Astragalus administration. Astragalus also ameliorated the pathological degeneration of the intestinal tissue of aging mice by increasing the length of small intestinal villi, the thickness of colonic mucosa and goblet cell number. In addition, Astragalus elevated the expression of genes associated with the proliferation and differentiation in jejunum and modulated gut microbiota, especially restoring the abundance of Lachnospiraceae. Taken together, the above research results demonstrate the total extract of Astragalus as a key factor improving the intestinal function and gut microbiota homeostasis of aging mice.

To investigate the immune regulatory effects of human bone marrow mesenchymal stem cells on alloantigen T lymphocyte in vitro, human MSCs were isolated and expanded from bone marrow cells, and identified with cell morphology, and the phenotypes were assessed by immunohistochemistry and flow cytometry. As the stimulation factor of T lymphocytes proliferation, either PHA or dendritic cells isolated from cord blood were cocultured with CD2(+) T lymphocytes from peripheral blood mononuclear cells by magnetic beads with or without MSC in 96-well plats for seven days. T cell proliferation was assessed by [(3)H]-thymidine incorporation using a liquid scintillation counter. T cell subsets, Th1, Th2, Tc1 and Tc2 were analyzed by flow cytometry after co-culture of CD2(+) T cells with MSCs for 10 days. The results showed that a significant decrease of CD2(+) T cell proliferation was evident when MSC were added back to T cells stimulated by DC or PHA, and an increase of Th2 and Tc2 subsets were observed after co-culture of MSC with T lymphocytes. It is suggested that allogeneic MSC can suppress T cell proliferation in vitro and the cause of that was partly depend on interaction of cells and the alteration of T cell subsets.
Bone Marrow Cells ; cytology ; immunology ; CD2 Antigens ; immunology ; Cell Communication ; immunology ; Cell Proliferation ; Cells, Cultured ; Coculture Techniques ; Flow Cytometry ; Humans ; Immunohistochemistry ; Mesenchymal Stromal Cells ; cytology ; immunology ; T-Lymphocyte Subsets ; cytology ; immunology ; T-Lymphocytes ; cytology ; immunology

To synthesize salbutamol immunogen and develop an enzyme immunoassay (ELISA), a new salbutamol immunogen was synthesized using 4-aminobenzoic acid as a linker to connect hapten with carrier protein. An enzyme immunoassay based on the antibody prepared was developed and applied to detect salbutamol residue spiked in swine liver. An unusual coating antigen, clenbuterol-ovalbumin (OVA) conjugate instead of salbutamol-OVA conjugate, was used in the immunoassay and the results were discussed based on the structures of related compounds. The antibodies showed high sensitivity in the heterologous assay when using clenbuterol-OVA as a coating antigen, with an IC50 value of 8.97 ng mL(-1) toward salbutamol. The antibodies prepared showed high cross-reactivity with clenbuterol (107%) and were promising for the simultaneous determination of salbutamol and clenbuterol residues in food and food products. Recovery rates from the salbutamol-spiked swine liver samples were in the range of 70%-99%, while the intra-assay and inter-assay coefficients of variation were <13.3% and <14.3%, respectively. In summary, the antibodies of salbutamol have been successfully prepared. Sensitive and stable analysis for the detection of salbutamol residues in swine liver was obtained based on the competitive ELISA methods developed in this study.
4-Aminobenzoic Acid ; chemistry ; Adrenergic beta-Agonists ; analysis ; immunology ; Albuterol ; analysis ; immunology ; Animals ; Antibodies ; immunology ; Antibody Specificity ; Clenbuterol ; analysis ; immunology ; Drug Residues ; analysis ; Enzyme-Linked Immunosorbent Assay ; methods ; Food Contamination ; Haptens ; immunology ; Immunization ; Liver ; chemistry ; Male ; Ovalbumin ; chemistry ; immunology ; Rabbits ; Serum Albumin, Bovine ; chemistry ; immunology ; Swine

The purpose of this study was to investigate whether pretransplant infusion of reactive natural killer cells (NK cells) from donor or recipient can reduce graft-versus-host disease (GVHD) and enhance engraftment in bone marrow transplantation (BMT). Recipient BALB/c mice were divided into 4 groups after received 6.5 Gy total-body irradiation (TBI): control group 1 was treated with nothing, control group 2 received BMT alone, experiment group 1 received BMT and autoreactive NK cells, experiment group 2 received BMT and alloreactive NK cells. Life span, clinical and pathologic changes of GVHD and chimerism rate of each group were evaluated. The results showed that all mice were survival in control group 1. The life span was shorter in experiment group 1 than that in control group 2 (P < 0.05) and longer in experiment group 2 than that in control group 2 (P < 0.01). GVHD was higher in score of experiment group 1 than in control group 2 (P < 0.05) but lower in experiment group 2 than that in control group 2 (P < 0.01). The donor chimerism rate in both two experiment groups were higher than that in control group 2 (P < 0.05), however, the donor chimerism rate was higher in experiment group 2 than that in experiment group 1 (P < 0.01). It is concluded that pretransplant infusion of alloreactive donor NK cells can prolong life span, reduce the degree of GVHD and enhance engraftment. But autoreactive recipient NK cells can shorten life span, aggravate the degree of GVHD and also enhance engraftment, which is weaker than that using alloreactive donor NK cells.
Animals ; Bone Marrow Transplantation ; adverse effects ; Graft Survival ; immunology ; Graft vs Host Disease ; prevention & control ; Killer Cells, Natural ; transplantation ; Male ; Mice ; Mice, Inbred BALB C ; Mice, Inbred C57BL ; Whole-Body Irradiation

The purpose of this research is to study the effect of small molecule compound piceatannol (PIC) on host inflammation in adenine induced chronic kidney disease (CKD) mice, and then to explore its mechanism based on the regulation of gut microbiota. All procedures were approved by the Institutional Animal Care and Use Committee of the Nanjing University of Chinese Medicine. The level of interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) was detected by enzyme linked immunosorbent assay (ELISA); UPLC-TQ/MS technology was used to monitor the level of proinflammatory uremic toxin indoxyl sulfate (IS) and p-cresol sulfate (PCS); the expression of occludin was tested by Western blot; in vitro anaerobic culture of gut bacteria was used to produce indole; the abundance of gut microbiota was evaluated by 16S rDNA sequencing. The results showed that PIC had no effect on inflammatory infiltration in kidney tissue of CKD mice, but could decrease IL-6 level in blood and IL-6/TNF-α level in colon tissue. PIC did not improve intestinal occludin protein expression in CKD mice; while it could significantly reduce the levels of IS and PCS in blood and liver of CKD mice. Further mechanism studies showed that PIC could inhibit the synthesis of IS precursor indole in gut bacteria. Moreover, PIC could decrease the abundance of gut bacteria which producing uremic toxin, such as reducing the abundance of indole and p-cresol producing gut bacteria. In conclusion, PIC could regulate gut microbiota and inhibit the synthesis of uremic toxin precursor, thereafter reducing the accumulation of IS and PCS in vivo, ultimately relieving the inflammation of CKD mice.

OBJECTIVETo investigate the clinicopathologic and prognostic implications of phosphoinositide 3 kinase (PI3K)/AKT pathway alterations in endometrial cancers of Chinese women.

METHODSThe expression of PTEN, p-AKT, and ER/PR was assessed in 71 cases of endometrial carcinoma by immunohistochemistry (EnVision method). The PIK3CA mutation at exon 9 and exon 20 was analyzed by PCR and direct sequencing in 34 tumors.

RESULTS(1) Of the 71 cases of endometrial carcinoma, 65 cases were endometrioid adenocarcinoma (EEC) and 6 cases were nonendometrioid adenocarcinoma (NEEC). PTEN loss of expression was found in 63.4% (45/71) of tumors, and more commonly occurred in EEC (66.2%, 43/65) than that in NEEC (2/6, P = 0.18). Patients with PTEN loss in their tumors (45 cases) had a better survival than those without (26 cases, P = 0.07). In ER negative subgroup, the patients with PTEN loss of expression (12 cases) had longer survival than those with normal PTEN expression (7 cases; P = 0.04). (2) The frequency of PIK3CA mutation was 41.2% (14/34) with a hot mutation spot at T544 in exon 9. PIK3CA mutations more commonly occurred in EEC (44.8%, 13/29) than in NEEC (1/5, P > 0.05). The mutations at exon 9 more commonly occurred in EEC, well- and moderately-differentiated EEC, and tumors at early stage (P > 0.05). On the contrary, in tumors at early stages, the frequency of mutations in exon 20 (14.3%, 4/28) was significantly lower than that at late stages (4/6, P = 0.01). (3) p-AKT was positive in 59.2% (42/71) of tumors that were more frequently found in EEC (60.0%, 39/65) than that in NEEC (3/6, P = 0.68). However, the significant difference of p-AKT expression was found between well- and moderately-differentiated EEC (75.0%, 21/28; 53.6%, 15/28) and poorly-differentiated EEC (3/9, P = 0.02). Moreover, p-AKT expression was significantly correlated with positive ER (r = 0.339, P = 0.00).

CONCLUSIONSEndometrial carcinoma patients with loss of PTEN and p-AKT positivity have a favorable prognosis. PIK3CA mutations at exon 9 or 20 may have different impact on the prognosis. The function of PTEN loss and p-AKT expression may vary according to different hormone status.

Adult ; Aged ; Carcinoma, Endometrioid ; genetics ; metabolism ; pathology ; surgery ; Class I Phosphatidylinositol 3-Kinases ; Endometrial Neoplasms ; genetics ; metabolism ; pathology ; surgery ; Exons ; Female ; Follow-Up Studies ; Humans ; Hysterectomy ; Middle Aged ; Mutation ; Neoplasm Invasiveness ; Neoplasm Staging ; PTEN Phosphohydrolase ; metabolism ; Phosphatidylinositol 3-Kinases ; genetics ; metabolism ; Phosphorylation ; Proto-Oncogene Proteins c-akt ; metabolism ; Receptors, Estrogen ; metabolism ; Receptors, Progesterone ; metabolism ; Signal Transduction ; Survival Rate

OBJECTIVETo explore the effects of adenovirus-delivered tissue inhibitor of metalloprotein- ases-3 (Ad-TIMP-3) on the irradiation sensitivity of human papillomavirus (HPV)-positive cervical cancer cells.

METHODSAn adenovirus expressing TIMP-3 (Ad-TIMP-3), alone or in combination with irradiation,was used to treat HPV-positive cervical cancer cells HeLa-Luc and CaSki. The effects of Ad-TIMP-3 on the proliferation of HeLa-Luc and CaSki cells were detected with MTT assay. The effect of the combination of Ad-TIMP-3 and X-ray on the proliferation of cells were determined by clone formation assay. Twenty nude mice were equally randomly divided into four groups: normal control group,Ad-TIMP-3 group,X-ray group,and combination group. The size of tumor was measured separately,and tumor growth curves were drawn.

RESULTSAd-TIMP-3 significantly inhibited the proliferation of HPV-positive cervical cancer cells in a dose-dependent manner. Combination of Ad-TIMP-3 and X-ray significantly decreased the clones of HeLa-Luc and CaSki than Ad-TIMP-3 or X-ray alone (P<0.05). The tumor weights were (0.216±0.098), (0.276±0.073), and (0.044±0.043) g, respectively, in Ad-TIMP-3 group, X-ray group,and combination group, which were all significantly lower than that in normal control group [(0.534±0.218) g] (all P<0.05). In addition,the tumor weight in the combination group was significantly lower than that in Ad-TIMP-3 group and X-ray group (both P<0.05). The tumor inhibition rate was 59.60%, 48.30%, and 91.80% in X-ray group, Ad-TIMP-3 group and combination group, respectively.

CONCLUSIONSAd-TIMP-3 can effectively inhibit the proliferation of cervical cancer cells. When combined with X-ray,it can remarkably increase the irradiation sensitivity of HPV-positive cervical cancer cells,and thus suppress the tumorigenesis capability of these cells in vivo.

Adenoviridae ; genetics ; Animals ; Cell Line, Tumor ; Cell Proliferation ; Female ; Genetic Vectors ; Humans ; Mice ; Mice, Nude ; Papillomaviridae ; genetics ; Radiation Tolerance ; Tissue Inhibitor of Metalloproteinase-3 ; genetics ; Transfection ; Uterine Cervical Neoplasms ; pathology ; radiotherapy ; virology

OBJECTIVETo study the relationship of Chinese familial Parkinson disease with alpha-synuclein gene.

METHODSPolymerase chain reaction-single strand conformational polymorphism (PCR-SSCP) and polymerase chain reaction-heteroduplex analysis(PCR-HA) were employed to detect the abnormal mobilization in the familial Parkinson disease and sporadic Parkinson disease patients, then it was verified by gene sequencing.

RESULTSNo mutation was found in alpha-synuclein gene exons 3 and 4 by PCR-SSCP together with PCR-HA. An inserted c and an inserted t were found in intron 4, position 23 and position 67 respectively.

CONCLUSION(1) Exons 3 and 4 of alpha-synuclein gene are not the mutational hot spots of Chinese familial Parkinson disease. (2) Two polymorphisms were found in intron 4 of alpha-synuclein gene. They are 23 ins c and 67 ins t.

Adult ; Aged ; Exons ; Female ; Heteroduplex Analysis ; Humans ; Male ; Middle Aged ; Mutation ; Nerve Tissue Proteins ; genetics ; Parkinson Disease ; genetics ; Polymerase Chain Reaction ; Polymorphism, Single-Stranded Conformational ; Synucleins ; alpha-Synuclein