Objective:Mesenchymal stem cells( MSCs) have self-renewal capacity and potential to differentiate into the cells.It was reported that the expression of miR-30a changed in some immune diseases.But it remains unclear the effect of miR-30a on the im-munoregulatory functions of MSCs.Here we studied the impact of miR-30a on the phenotype,cell viability,apoptosis,cell cycle and im-munoregulatory functions of MSCs.Methods: The mixed enzyme methods were used for the isolation of human umbilical cord MSCs.Flow cytometry(FCM)was used to investigate the effect of overexpressed miR-30a on the phenotype of MSCs.CCK-8 was used to examine the cell viability of miR-30a-overexpressed MSCs.Annexin V/PI was used for the detection of apoptosis of MSCs.Q-PCR and Western blot were used to investigate the effect of miR-30a on the expression of Cyclin E2( CCNE2).CCNE2 was one putative target of miR-30a predicted by Targetscan database.The effects of miR-30a-overexpressed MSCs on the maturation of dendritic cells(DCs)were determined.Results:Overexpression of miR-30a blocked the cell cycle of MSCs in the G0/G1 phase by inhibiting the expression of CCNE2,but did not affect the phenotype, cell viability and apoptosis of MSCs.When co-cultured with DCs, although MSCs down-regulated the expression of CD40 and CD86 on DCs,overexpression of miR-30a more significantly enhanced the suppressive impact of MSCs on the maturation of DCs.Conclusion: miR-30a affects the cell cycle of MSCs and enhances its immunosuppressive effect on DCs.

Objective: To investigate the antitumor actions of polysaccharide extracts from the fruiting body of coriolus versicolor (CVE). Methods:Hepatoma HepA cells were injected into mice subcutaneously. Different doses of CVE were given by gavage. On the 7 th and 14 th day, tumor inhibitive rates were calculated. ELISA was performed to measure the serum IgG level; MTT was used to examine CVE′s effects on the proliferation of T lymphocytes of thymus. Immunohistochemistry was used to determine CVE′s influence on the expression of tumor related genes P53 and VEGF in liver. Results: CVE may evidently inhibit the growth of the transplanted HepA tumors. Its effects on the serum IgG level and on the proliferation of T lymphocytes of thymus were also significantly. Also, CVE markedly decreased the expression of P53, VEGF genes in liver. Conclustion: CVE had significant antitumor effects in vivo . The mechanisms may involve immune modulation effects and antimetastasis actions.

BACKGROUND: Although shape memory alloy has been extensively used in modem medicine, including orthopaedics and dental surgery, the body fluid could influence the stability of biomaterial and some ions released by materials may cause toxic and side effect to human body. The technology for the modification of shape-memory materials is of crucial importance for clinical use of shape memory alloy.OBJECTIVE: To investigate the biocompatibility of diamond-like carbon (DLC) coated Nickel-Titanium shape memory alloy and uncoated shape memory alloy with osteoblasts cultured in vitro. DESIGN, TIME AND SETTING: A comparative observation. The study was performed at the Institute of Othopaedics of Lanzhou University from March to June 2005.MATERIALS: A total of 30 DLC-coated Nickel-Titanium shape memory alloy disks, 6 minx7 mmx0.5 mm, and the same amount of uncoated ones of equal size were provided by College of Materials, Lanzhou University. METHODS: Rabbit osteoblast suspension of the third passage at density of 10 × 108/L were incubated with DLC-coated Nickel-Titanium shape memory alloy disks and uncoated shape memory alloy in 12-well culture plate in 5% CO2 at 37 ℃.MAIN OUTCOME MEASURES: Culture media were counted at 1, 2, 3, 4 and 5 days of culture to determine alkaline phosphatase (AKP) activity and nickel (Ni2+) concentration.RESULTS: The proliferation of osteoblasts and the concentration of AKP in DLC- coated group were greater than uncoated group; while the uncoated group released more Ni2. into the cells culture media than coated group (P < 0.05). CONCLUSION: DLC-ceated Nickel-Titanium shape memory alloy appears to have better biocompatibility with osteoblast cultured in vitro compared to uncoated ones.

BACKGROUND:Several researches have highlighted the selective dissolution of Ni ion from the nickel-titanium (NiTi) alloy during the corrosion process,which can lead to potential damage to human body.Different surface treatments will improve the corrosion resistance of NiTi implants.In modern medicines,it is necessary to analyze the characteristics of surface modified NiTi implants.OBJECTIVE:To study the effects of coated and uncoated materials made by elastic nickel-titanium alloy internal fixator on fracture healing and to compare the effects of continuous compressive stress after internal fixator of different types on fracture healing by setting up control group of bone nail internal fixation.DESIGN,TIME AND SETTING:A randomized,controlled,animal experiment was performed at the Laboratory of Tissue Engineering,Institute of Orthopedics,Second Hospital Affiliated to Lanzhou University between September 2004 and March 2005.MATERIALS:Diamond-like carbon coated and nickel-titanium alloy and uncoated nickel-titanium alloy embracing fixator (type 4H8-40) were provided by Lanzhou Ximai Memory Co.,Ltd.,China.Intramedullary nails (type ZQY-01) were purchased from Tianjin Jinxingda Industries Co.,Ltd.,China.METHODS:Thirty Chinchilla rabbits of 4-6 months old were randomly divided into 3 groups (n = 10):diamond-like carbon coated nickel-titanium alloy embracing fixator (group A),uncoated nickel-titanium alloy embracing fixator (group B),and intramedullary fixator (group C).Following anesthesia by injection of 1% sodium pentobarbital (25 mg/kg),transverse fracture models in middle part of the femur were made through a lateral femoral incision and fixed with diamond-like carbon coated nickel-titanium alloy embracing fixator,uncoated nickel-titanium alloy embracing fixator,and intramedullary fixator respectively.MAIN OUTCOME MEASURES:The inorganic substance level,osteocalcin,alkaline phosphatase (ALP) and tumor necrosis factor (TNF) expression in callus surrounding fracture site were detected 4 weeks postoperatively.Ni ion level in muscles surrounding fracture site,live tissue,and brain tissue were also detected.RESULTS:Inorganic substance level and ALP,osteocalcin,and TNF expression were significantly higher in the groups A and B than in group C (P＜0.05).Ni ion level in the liver tissue,brain tissue,and muscles surrounding the fracture were significantly lower in the groups A and C than in group B (P＜0.05).CONCLUSION:Elastic fixation promotes fracture healing.Diamond-like carbon coated nickel-titanium alloy embracing fixator has a better histocompatibility than uncoated group.

Objective To investigate the role of astroglial glutamate-glutamine shuttle in the development of neuropathic pain (NP) in rats. Methods Forty-eight adult male SD rats weighing 200-230 g were randomly divided into 8 groups (n =6 each): Ⅰ control group (group C);Ⅱ sham operation group (group S);group ⅢNP;Ⅳ-Ⅶ 0.01, 0.03, 0.05 and 0.10 mmol/L methionine sulfoximine (MSO, an inhibitor of glutamine synthetase (GS)) group (group M1-4 );Ⅷ MSO + glutaminate group (group MG). In group C no operation was performed. In group S the sciatic nerve was only exposed but not ligated. NP was induced by ligation of the tibial nerve and commom peroneal nerve according to the technique described by Dixon. After the establishment of the model, intrathecal PBS 50 μl was injected in group NP, IT 0.01, 0.03, 0.05 and 0.10 mmol/L MSO 50 μl was injected intrathecally in group M1-4, and 0.05 mmol/L MSO 50 μl was injected intrathecally and then 0.25 mmol/L glutamine 50 μl was injected intrathecally 15 min later in group MG. Mechanical pain threshold was measured 1 week before ligation (T0 , baseline), 1 week after ligation (T1) and 15, 30, 45 and 60 min after injection of MSO (T2-5). Then rats were killed and the lumbar segment of the spinal cord was removed for determination of the expression of glial fibrillary acidic protein (GFAP) and GS and the co-expression (GFAP/GS) in the dorsal horn.Results Mechanical pain threshold was significantly lower at T1-5 in group MG and NP and at T2-4 in group M3.4 ,and the expression of GFAP, GS and GFAP/GS was significantly higher in group MG,NP and M3 than in group S and C ( P ＜ 0.05) .Conclusion Astroglial glutamate-glutamine shuttle in the spinal cord is involved in the development of neuropathic pain in rats.

Objective:To explore the effect of histone demethylase JMJD3 on B cell activation and apoptosis.Methods:B cells were sorted and purified from the peripheral blood of healthy people and SLE patients by using magnetic bead.After B cells were treated with IFN-αor R848 or IFN-α+R848,the percentages of CD86+B cells,CD69+B cells,CD86+Annexin V+B cells and CD69+Annexin V+B cells were detected by flow cytometry.The expression of JMJD3 was detected by Real Time PCR and Western blot.Results:The purity of sorted B cells was up to 95%.IFN-αenhanced both the activation and apoptosis and the JMJD3 expression of TLR7-activated B cells.The expression of JMJD3 was dependent on MAPK signal pathway,but not the NF-κB signaling pathway.Moreover,JMJD3 was highly expressed in B cells of peripheral blood from SLE patients compared to those from healthy people.Furthermore,JMJD3 inhibitors could inhibit the activation and apoptosis of IFN-αand R848 activated B cells.Conclusion:JMJD3 participated in the activation and apoptosis of IFN-αand TLR7-induced B cells, suggesting JMJD3 inhibitors may possess therapeutic effect for alleviating symptom of SLE.

Objective:To explore the change of B cell numbers in active MRL/lpr lupus mice , and their regulation mechanisms.Methods:B cell cycle and the percent of B cells in spleen lymphocytes of active MRL /lpr lupus mice and normal C 57/B6 mice were analyzed by using flow cytometry .The apoptotic B cells and their subclass were analyzed by Annexin V and PI staining.Further more ,B cells were purified by magnetic sorting , and real-time quantitative PCR was carried out to detect apoptosis-related gene.Results:Compared with the C57/B6 mice,the percent of B cells in active MRL/lpr lupus mice were significantly reduced (P<0.01),while the percent of apoptotic cells were significantly increased (P<0.01).The percent of early apoptotic B cells were sig-nificantly increased ( P <0.01 ) which including the immature and mature B cells , while the late apoptotic B cells were unchanged.Further more,we found that the anti-apoptotic protein BIRC3 was significantly reduced in active lupus B cells (P<0.01), while the pro-apoptotic protein BCL2L1 and BBC3(PUMA) were significantly increased(P<0.01).Conclusion: B cells in active lupus mice were significantly reduced while early apoptotic B cells were increased , which may be attributed to the changed balance between the anti-apoptotic and pro-apoptotic proteins , suggesting the reduction of B cells in SLE patients may be related to their increased early apoptosis .

Objective:To investigate the effect of HSM on the immunoreaction and renal fibrosis in mice with cecal ligation and puncture (CLP).Methods: Renal fibrosis was induced by CLP;mice were divided into three groups,including sham group (sham,n =5),model group (CLP,n =11),therapy group (HSM,n =11).HSM extract [200 mg/(kg?d)] was orally administered to HSM group2 hour before surgery and repeated everyday throughout 10 days,while sham group and model group were given the same dose of normalsaline.FACS assay was used to analyze the amount of macrophages ,neutrophils and Treg in PBMC,as well as macrophages in peritonealfluid;we used Q-PCR assay to analyze the expression of inflammatory molecules (IL-1βand TNF-α) and fibrogenic cytokines (TGF-β,MMP9 and TIMP1) from renal sections.Besides,renal sections were subjected to HE stain and immunohistochemical staining with α-SMA and fibronectin.Results: The amount of model group′s macrophages and neutrophils in PBMC,as well as macrophages in theperitoneal fluid were significantly higher than sham group ′s,whereas HSM succeeded in lowering them;contrast to sham group,Tregs′amount of CLP group and HSM group in PBMC had no significant changes .The expression of inflammatory molecules (IL-1βand TNF-α) and fibrogenic cytokines (TGF-β,MMP9 and TIMP1) from CLP group′s renal sections were remarkably improved ,whereas HSM inhibitedthat.The CLP group′s HE results showed obvious renal inflammatory damage ,whereas HSM reduced the histopathologicalterations;contrast to sham group,model group′s expression of α-SMA and fibronectin was remarkably improved,while HSM groupshowed lower expression.Conclusion: HSM extract could regulate immunity response and had effect in improving renal fibrosis .

Objective:To investigate the effect of HSM on the inflammation and pulmonary fibrosis in mice with cecal ligation and puncture ( CLP) . Methods: Both 5 day and 10 day timepoint pulmonary fibrosis were induced by CLP;mice were randomly divided into three groups,including sham group (sham,n=5),model group (CLP,n=11),therapy group (HSM,n=11). HSM extract (200 mg/kg,less than human dose of 0. 27 g/kg) was orally administered to HSM group 2 hour before surgery and repeated everyday, while sham group and model group were given the same dose of normal saline. We used Q-PCR assay to analyze the expression of in-flammatory molecules ( IL-1β and TNF-α) and fibrogenic cytokines ( TGF-β, MMP9 and TIMP1 ) from lung tissues , we used FACS assay to analyze the amount of Th1 cells in PBMC and spleen sections,as well as Treg in spleen sections. Besides,lung sections were subjected to HE stain and immunohistochemical staining withα-SMA and fibronectin. Results:The amount of model group's Th1 cells in PBMC and spleen sections,as well as Treg in spleen sections were significantly lower than sham group's,whereas HSM succeeded in raising them. By day 5, the expression of inflammatory molecules IL-1β and fibrogenic cytokines (TGF-β,MMP9 and TIMP1) from CLP group's lung sections were remarkably improved,by day 10,the expression had decreased but was higher than sham group's;HSM inhibited these cytokines' expression. The expression of TNF-α of CLP group and HSM group had no significant changes. The CLP group's HE results showed obvious pulmonary inflammatory damage,by day 10,the situation had improved,whereas HSM reduced the histopathologic alterations;contrast to sham group,model group's expression of α-SMA and fibronectin was remarkably improved,while HSM group showed lower expression. Conclusion: HSM extract can suppress inflammation, balances immune system and relieves pulmonary fibrosis.

Objective To explore the effects of HAVCR2 siRNA on apoptosis and cell cycle in the radiation-caused genomic instable liver cells.Methods RNAi was used to inhibit HAVCR2 gene transcription.Cellular apoptosis and cell cycle distribution were detected by flow cytometry (FCM).Expression of p53 gene was assayed by real time fluorescence quantitative PCR.Results HAVCR2 gene was effectively inhibited by RNAi (t =19.21,P ＜ 0.05).After siRNA transferring,cell apoptosis (t =3.65,P ＜ 0.05) and p53 gene expression (t =4.82,P ＜ 0.05) were decreased,and G2-phase arrest was induced(t =-3.41,P ＜ 0.05).Conclusion HAVCR2 siRNA can decrease the generation of apoptosis in the genomic instable liver cells and blocks the cells at G2 phase.