OBJECTIVETo investigate the expression profiles of myocardin gene during the differentiation of bone marrow-derived mesenchymal stem cell to smooth muscle cells in the conditional medium combined with a high concentration of fetal bovine serum (FBS).

METHODSMarrow-derived mesenchymal stem cells were isolated and purified from mouse femoral bone and shinbones using differential adherent methods. Cells at the third passage were induced by 20% FBS in conditioned medium, conditioned medium alone, 20% FBS or 10% FBS alone respectively. Mouse aortic smooth muscle cells were cultured as the positive control. Levels of mRNA and protein expression of myocardin and several smooth muscle cells marker genes were determined by immunofluorescence, RT-PCR and Western blot before and 3, 7, 10, 14 d after the induction. The presence of smooth muscle myofilaments was detected by using transmission electron microscope.

RESULTSNaive bone marrow-derived mesenchymal stem cells displayed multiple morphological forms including fusiform, polygon, oval, and micro-spherical, as compared to the single macro-spindle form after the induction. Typical appearance of peak valley was displayed on the 21st day after induction. At the same time, the expression of smooth muscle marker genes was reinforced along with an up-regulation of myocardin expression. Immunofluorescence study showed that the cells expressing myocardin and smooth muscle marker genes such as alpha-SMA and SM-MHC increased. Fluorescence domain of myocardin translocated from cytoplasm to nucleus and the amounts of double positive cells for myocardin with alpha-SMA or SM-MHC also increased. RT-PCR confirmed that the mRNA expression of myocardin increased gradually and remained stabilized after achieving its peak on the 7th day after induction. The expression of smooth muscle marker genes, alpha-SMA and SM22alpha, remained stable on the 10th day of induction. It was also confirmed by Western blot that the protein expression of both myocardin and alpha-SMA were markedly increased during the induction. Finally, transmission electron microscopy revealed the presence of myofilament on the 21st day after induction.

CONCLUSIONSBone marrow-derived mesenchymal stem cells can be effectively induced into smooth muscle-like cells by conditioned medium combined with 20% FBS. Myocardin plays an important role in the differentiation process of bone marrow-derived mesenchymal stem cells to the peripheral smooth muscle cells.

Animals ; Bone Marrow Cells ; cytology ; physiology ; Cattle ; Cell Differentiation ; physiology ; Male ; Mesenchymal Stromal Cells ; cytology ; metabolism ; Mice ; Muscle, Smooth, Vascular ; cytology ; Myocytes, Smooth Muscle ; physiology ; Nuclear Proteins ; genetics ; metabolism ; physiology ; Reverse Transcriptase Polymerase Chain Reaction ; Trans-Activators ; genetics ; metabolism ; physiology ; Up-Regulation

OBJECTIVEConstruction of a small interfering RNA (siRNA) eukaryotic expression vector specific to mouse myocardin gene and study on the role of myocardin-siRNA on differentiation of mouse bone mesenchymal stem cells (MSCs) into smooth muscle-like cells induced by PDGF-BB in vitro.

METHODSMouse MSCs were isolated from bone marrow and cultured with 50 mg/L PDGF-BB and fetal bovine serum (20%). Specific myocardin-siRNA sequence was cloned into a plasmid pGenesil-1.0 vector, which contained U6 promoter. The recombinant plasmid and control plasmid were transfected into MSCs which had been cultured with PDGF-BB for 6 days beforehand. The expression of myocardin mRNA was detected by RT-PCR 48 hours after the transfection. Immunohistochemistry was used to detect the SM-MHC and to identify the smooth muscle-like cells.

RESULTSThe recombinant plasmids carrying myocardin-siRNA sequences were constructed successfully and the myocardin mRNA was reduced 42.86% by pGen-myo-shRNA in comparing with that of the controls (P<0.01); and the expression of SM-MHC protein was down-regulated (P<0.01).

CONCLUSIONSubset of mouse MSCs have the potential to differentiate into smooth muscle-like cells, a possible cell source responsible for atherosclerotic plaque formation, and myocardin expression may play an important role during this process.

Animals ; Bone Marrow Cells ; cytology ; metabolism ; Cell Differentiation ; Cells, Cultured ; Down-Regulation ; Gene Silencing ; Genetic Vectors ; Male ; Mesenchymal Stromal Cells ; cytology ; metabolism ; Mice ; Muscle, Smooth, Vascular ; cytology ; Myocytes, Smooth Muscle ; cytology ; metabolism ; Myosin Heavy Chains ; metabolism ; Nuclear Proteins ; biosynthesis ; genetics ; physiology ; Plaque, Atherosclerotic ; pathology ; Plasmids ; Platelet-Derived Growth Factor ; pharmacology ; Proto-Oncogene Proteins c-sis ; RNA, Messenger ; metabolism ; RNA, Small Interfering ; Trans-Activators ; biosynthesis ; genetics ; physiology ; Transfection

OBJECTIVETo determine the distribution of HLA-B alleles in the Chinese Yi ethnic group and its association with HIV infection.

METHODSOne hundred and six unrelated healthy HIV negative and 73 HIV positive Chinese Yi ethnic individuals were typed by PCR-SSP.

RESULTSThe frequency of alleles B*07, Bx 35, and B*46 were increased in HIV-1-positive subjects, whereas the alleles B*55, B*44 and B*78 were absent in the HIV-infected persons studied. The B*46 allele was present in a significantly higher gene frequency among HIV-1-positive individuals (P=0.02, OR=3.32, 95% CI=1.13-9.78) compared with control subjects.

CONCLUSIONHLA-B*46 may be associated with its susceptibility to HIV-1 infections.

Case-Control Studies ; China ; ethnology ; DNA ; analysis ; Ethnic Groups ; Gene Frequency ; Genetic Predisposition to Disease ; Genotype ; HIV Infections ; blood ; genetics ; HIV Seropositivity ; blood ; HIV-1 ; pathogenicity ; HLA-B Antigens ; genetics ; Humans ; Polymerase Chain Reaction ; methods ; Surveys and Questionnaires

Objective To explore the effect of specific antagonist of estrogen-related receptor alpha——XCT790 on tumor growth, weight, liver index(LI), spleen index(SI) and kidney index (KI) in the diffe-rent models of tumor -bearing mice.Methods The H22 ascitic and solid tumor-bearing mice models were established , then mice were ran-domized into five groups , including model group (20%DMSO), control group(cyclophosphamide:CTX 30 mg· kg -1), experimental -L,-M,-H groups(XCT790:2,4,6 mg· kg -1).The samples were obtained in 24 h after continuous intraperitoneal administration of drug or solvent to mice for 7 d.The ascitic volume and tumor weight were measured .The ratios of LI,SI,KI were calculated.Results The ascitic volume of mice in model group, control group,and experimental -L,-M,-H groups were (6.17 ±3.04),(3.28 ±1.62),(3.60 ±1.67),(4.67 ±2.57), (4.73 ±2.66 ) mL; comparing between control group , experimental -L group and model group,the differences were significant(all P<0.01).In H22 solid tumor -bearing mice, the tumor weight of mice in model group, control group, experimental -L,-M,-H groups were (2.53 ±0.39),(1.25 ±0.45),(1.27 ±0.61),(1.14 ±0.56),(1.24 ±0.39) g with significant difference com-pared with model group ( all P<0.05 ) .LI,SI and KI had no statistically significant differences in ascitic or solid tumor-bearing groups(all P>0.05 ) .Conclusion XCT790 had anti -tumor effect on H22 tumor-bearing mice without influences on ratios of liver ,spleen and kidney.

Objective To explore the effect of kaempferol on tumor growth,weight,liver index (LI) and spleen index (SI) in different models of H22 tumor-bearing mice.Methods The H22 ascitic and solid tumor-bearing mice model were established,then mice were randomized into five groups,including model group,control group (cyclophosphamide 30 mg · kg-1),experimental-L,-M,-H groups (kaempferol 150,300,600 mg · kg-1).The samples were obtained in 24 h after continuous intragastrical administration of drug or solvent to mice for 8 d.The ascitic volume and tumor weight were measured.The liver index(LI),spleen index (SI) were calculated.Results In H22 ascitic tumor-bearing mice,the ascitic volume of mice in model group,control group,experimental-L,-M,-H groups were (9.85 ± 1.99),(2.28 ±2.74),(8.44 ±2.51),(5.91 ±2.29),(4.98 ±4.25) mL;compared with model group,the difference in control group,experimental-M,-H groups was significantly (all P ＜ 0.05).LI and SI in above five groups were 4.72 ± 2.22,5.42 ±0.77,4.69 ± 0.94,5.22 ± 0.60,5.27±0.61;0.49 ±0.33,0.62 ±0.19,0.51 ±0.13,0.49 ±0.14,0.41 ±0.14 with not significantly between five groups(all P ＞0.05).In H22 solid tumor-beating mice,the tumor weight of mice in model group,control group,experimental-L,-M,-H groups were (1.70 ± 0.80),(0.73 ±0.51),(1.14 ±0.91),(1.07 ±0.34),(0.95 ±0.60) g with significantly compared with model group(all P ＜ 0.05).Conclusion The kaempferol had anti-tumor effects toward H22 tumor-bearing mice with less influence on liver and spleen.

OBJECTIVETo study the quantity and function of bone marrow (BM) T follicular helper (Tfh) cells of the cytopenia patients with positive bone marrow mononuclear cells (BMMNC)- Coombs test (also known as immuno-related pancytopenia, IRP), and explore the role of Tfh cells in the pathogenesis of IRP.

METHODSForty- three untreated IRP patients, 47 recovered IRP patients and 25 healthy donors were enrolled in this study. The percentages of Tfh cells, Tfh-related molecules ICOS, CD40L, IL-21 and Bcl-6 in BM were investigated by flow cytometry and semiquantitive RT-PCR.

RESULTSThe ratio of CD4⁺CXCR5⁺/CD4⁺ cells of untreated IRP patients [(28.79 ± 19.70)%] was significantly higher than that of recovered IRP patients [(21.15 ± 12.81)% ] and normal controls ([ 13.42 ± 6.72)% ](P<0.05). The ratio of CD4⁺CXCR5⁺ICOS⁺/CD4⁺CXCR5⁺ cells of untreated IRP patients [(5.05 ± 4.71)% ] was significantly higher than that of recovered IRP patients [(2.96 ± 2.89)% ] and normal controls [(2.99 ± 2.23)% ] (P<0.05). The ratio of CD4⁺CXCR5⁺CD40L⁺/CD4⁺CXCR5⁺ cells of untreated IRP patients [(5.87 ± 4.14)%] and recovered IRP patients [(6.52±5.47)%] were significantly higher than that of normal controls [(2.93 ± 2.92)%] (P<0.05). The ratio of intracytoplasmic CD4⁺CXCR5⁺IL-21⁺/CD4⁺CXCR5⁺ cells of untreated IRP patients [(8.20 ± 7.41)% ] and recovered IRP patients [(6.30 ± 6.03)% ] were significantly higher than that of normal controls [(3.43 ± 3.40)%] (P<0.05). The relative expressions of Bcl-6 mRNA in BMMNC were 0.625 ± 0.248, 0.485 ± 0.253, 0.306 ± 0.210 in three groups, respectively. The differences between untreated IRP patients, recovered IRP patients and normal controls were significant (P<0.05).

CONCLUSIONThere exists increased quantity and hyperfunction of Tfh cells in the IRP patients, they may play important role in the pathogenesis of IRP. Tfh cells and their related effector molecules could be a potential therapeutic target for the disease.

Adolescent ; Adult ; Aged ; Aged, 80 and over ; Case-Control Studies ; Child ; Child, Preschool ; Coombs Test ; Female ; Flow Cytometry ; Humans ; Interleukins ; metabolism ; Lymphocyte Count ; Male ; Middle Aged ; Pancytopenia ; blood ; diagnosis ; etiology ; T-Lymphocytes, Helper-Inducer ; cytology ; metabolism ; Young Adult

OBJECTIVETo observe the efficacy and safety of linezolid for the treatment of gram positive coccus infections in hematological disease patients.

METHODSOne hundred and two hematological disease patients with suspected or proven gram positive coccus bacteria infection were enrolled in this study. Linezolid was given at a dosage of 600 mg, iv, q12h. The mean treatment period was (10.82 ± 5.12) days (1 to 51 days) with 74.5% over 7 d and 51.0% over 10 d.

RESULTSAmong 102 patients, 57 were male, 45 female aged 11 to 81 years, with a mean of (45.26 ± 19.15) years. Ninety four cases were nosocomial infection (92.2%) and 8 community infection (7.8%); There were pneumonia in 80 (78.4%), septicemia in 11 (10.8%), and infection of other organsin 11 (10.8%); Forty five cases were proven gram positive coccus bacteria infection, and 57 were suspected infection; Fifty one bacteria strains were isolated from cultivated samples of proven patients, in which 22 were staphylococcus aureus with 19 methicillin resistant 13 hemolytic streptococcus, 9 staphylococcus epidermidis with 7 methicillin resistant 6 enterococcus faecom, and 1 enterococcus hirae. Seven cases were mixed with one kind gram negative bacillus infection, 4 mixed with two kinds of gram negative bacillus infection, and 12 mixed with fungal infection; Total clinical response rates by ITT (intention to treatment) analysis was 69.6%, in which 40 (39.2%) were curative and 31 (30.4%) obviously effective; PP (per-protocol) analysis was 70.9%, in which 39 (41.9%) were curative and 27 (29.0%) obviously effective. Bacteria clearance rate was 70.6%, and in this group the clinical effective rate was 88.9%; Adverse effect rate was 2.9%, being transient thrombocytopenia and increased transaminase.

CONCLUSIONLinezolid is a safe and effective antibiotic used in hematological disease patients complicated with infections of gram positive coccus.

Acetamides ; Anti-Bacterial Agents ; therapeutic use ; Cross Infection ; microbiology ; Gram-Positive Bacterial Infections ; drug therapy ; Gram-Positive Cocci ; Hematologic Diseases ; drug therapy ; Humans ; Linezolid ; Oxazolidinones ; Staphylococcus aureus

Adult ; Aged ; Aged, 80 and over ; Bone Marrow Cells ; physiology ; Female ; Humans ; Immune Evasion ; Male ; Middle Aged ; Myelodysplastic Syndromes ; etiology ; immunology

OBJECTIVETo investigate the function of dendritic cells (DC) of patients with immune related pancytopenia (IRP) and explore the role of DC in IRP.

METHODSThe expression of CD80 and CD86 on myeloid DC (mDC, Lin-HLA-DR(+) CD11c(+) cells) and plasmacytoid DC (pDC, Lin-HLA-DR(+) CD123(+) cells) of 65 IRP (37 untreated and 28 remitted) patients and 17 healthy controls were analyzed by flow cytometry.

RESULTSThe expression of CD86 on pDC was (82.47 ± 13.17)% in untreated group and (60.08 ± 14.29)% in remission group, which were significantly higher than that of controls (47.95 ± 18.59)% (P < 0.05), while the expression in untreated group was higher than that of remission group (P < 0.05). The expression of CD80 on pDC was (6.31 ± 4.49)% in untreated group, which was significantly higher than that of remitted patients (3.09 ± 2.93)% and controls (2.33 ± 2.25)% (P < 0.05). The expression of CD86 on mDC was (97.06 ± 4.82)% in untreated group and (91.35 ± 12.20)% in control group, while the expression in untreated group was higher than that of control group (P < 0.05). The expression of CD80 on mDC was (6.20 ± 5.44)% in untreated group and (3.97 ± 3.24)% in remission group, which were significantly higher than that of controls (1.86 ± 1.73)% (P < 0.05). The expression of CD86 on pDC was negatively correlated to Th1/Th2 (r = -0.733, P < 0.05), it was positively correlated to the antibody on membrane of BMMNC (r = 0.283, P < 0.05) and the quantity of CD5(+)B cells (r = 0.436, P < 0.05), while it was negatively correlated to the level of hemoglobin, platelets and white blood cells (r = -0.539, P < 0.05; r = -0.519, P < 0.05; r = -0.567, P < 0.05, respectively). The expression of CD80 on pDC was negatively correlated to the level of hemoglobin and platelets (r = -0.431, P < 0.05; r = -0.464, P < 0.05).

CONCLUSIONThe function of pDC in PB of IRP were strengthened, which was relevant to the immunopathogenesis of IRP.

Adolescent ; Adult ; Autoimmune Diseases ; complications ; B7-1 Antigen ; metabolism ; B7-2 Antigen ; metabolism ; Case-Control Studies ; Child ; Child, Preschool ; Dendritic Cells ; metabolism ; Female ; Flow Cytometry ; Humans ; Male ; Middle Aged ; Pancytopenia ; blood ; etiology ; pathology ; Young Adult

This study was purposed to investigate the expressions of miR-21, miR-155 and miR-210 in plasma of patients with lymphoma, and explore their role played in diagnosis, evaluation of chemotherapy effect and prognosis of lymphoma. The expressions of miR-21, miR-155 and miR-210 were assayed by RT-PCR in plasma of 54 cases of lymphoma, 10 cases of lymphonode inflammation and 27 cases of normal controls. The results indicated that the expressions of miR-21, miR-155 and miR-210 in plasma of lymphoma patients were higher than those of control group and lymphonode inflammation group (P < 0.05). The expressions of miR-21 and miR-210 in plasma of control group and lymphonode inflammation group had no significant differences (P > 0.05). The expression of miR-21 in plasma of lymphoma patient group significantly correlated with their serum LDH level. The expressions of miR-21 and miR-210 in plasma of previously untreated lymphoma patient group were higher than those of the patients treated for 6 or more courses (P < 0.05). The diagnostic accuracy of miR-21, miR-155 and miR-210 used for lymphoma patients was 56, 65, 48 respectively, and reached to 83 when combined three of them. It is concluded that the expressions of miR-21, miR-155 and miR-210 in plasma of lymphoma patients were significantly higher. Detection of these 3 miRNA in plasma of patients can contribute to the clinical diagnosis, treatment and prognosis evaluation of lymphoma.
Adult ; Aged ; Aged, 80 and over ; Case-Control Studies ; Female ; Humans ; Lymphoma ; blood ; diagnosis ; Male ; MicroRNAs ; blood ; Middle Aged ; Plasma ; metabolism ; Prognosis ; Young Adult