BACKGROUND:Bone marrow mesenchymal stem cels have low immunogenicity and can induce immune tolerance. At present, the mechanism of immune regulation of bone marrow mesenchymal stem cels is not completely understood. It has been rarely reported whether the bone marrow mesenchymal stem cels can migrate to the thymus after transplantation.
OBJECTIVE:To observe the distribution and survival of bone marrow mesenchymal stem cels in the thymus of aging rats after transplantation.
METHODS: Bone marrow mesenchymal stem cels cultured in vitrowere transfected by adenovirus vectors expressing green fluorescent protein. Transfected bone marrow mesenchymal stem cels were injected into the portal vein of aging rats. At days 3, 7, 14, 21 after transplantation, the survival of bone marrow mesenchymal stem cels homing to the thymus was observed under fluorescence microscope. At day 3 after transplantation, thymus tissues were taken and stained with hematoxylin-eosin for pathological observation.
RESULTS AND CONCLUSION:Green fluorescent protein-labeled bone marrow mesenchymal stem cels had a strong green fluorescence at days 3 and 7 after transplantation, and the cel contour was clear. There was no significant difference in the mean absorbance values at days 3 and 7 (P> 0.05). Expression of green fluorescent protein was weakened significantly at days 14 and 21 compared with that at day 3 (P < 0.05). At 3 days after transplantation, the transplanted bone marrow mesenchymal stem cels were clearly visible in the thymus, and acute rejection was not observed. The results show that bone marrow mesenchymal stem cels can migrate to the damaged thymus tissue through the blood circulation, and can survive at least 1 week.

Objective To investigate the photodynamic therapy(PDT)of human erythroleukemia cell line K562 and drug-resistance cell line K562/ADM using hexyl 5-aminolevulinate(He-ALA)in vitro.Methods Four groups were set:PDT group(photosensitizer with light irradiation),laser group(light irradiation only),dark-cytoxocity group(photosensitizer only)and normal control group(neither photosensitizer nor light irradiation).K562 cells and K562/ADM cells cultured were incubated with various concentrations of He-ALA(0.025,0.1,0.4 and 1.6mmol/L)for 4 hours,then illuminated with different light doses(4.5,9,18 and 36 J/cm2)using a laser with a wave length of 630nm.After 12 hours incubation,Wright's staining method was used to observe the cell's morphological changes,the survival rates of cells were analyzed by CCK8 assay,and the changes in colony-forming abilities of cells were analyzed by methylcellulose colony-forming assay.Results He-ALA or light irradiation alone had no evident cytotoxicity,while the application of He-ALA plus light irradiation effectively killed the leukemia cells.With the increase of the concentrations of He-ALA and the dosages of irradiation,the viability of cells decreased.Under the same conditions,the survival rate of K562 was significantly lower than that of K562/ADM(P

Objective:To investigate the effects of TJU103 on T lymphocytes proliferation and cytokine production in vitro.Methods:MTT and ELISA assays were used respectively to detect the proliferation and the cytokine production of donor T lymphocytes blocked by TJU103.Results:There are significant difference in the inhibition of T-cell proliferation and the cytokine production between groups of with TJU103 or without.Conclusion:TJU103 could significantly inhibit the T-cell proliferation and the cytokine production.

Objective To observe the effect of combination of Tju103 and CTLA4-Ig on engraftment, graft versus host disease (GVHD), graft versus leukemia (GVL) and anti-infection post major histocompatibility complex (MHC) haploidentical bone marrow transplantation in mice in order to seek an effective access to transplantation with less GVHD and more potential GVL and anti-infection.Methods In the presence of the recipient's antigen (normal CB6F1, H-2 bd) as a stimulus for induction of specific immune tolerance, T cells from the MHC haploidentical donors (C57BL/6, H-2 b) were first in vitro cultured with Tju103 and CTLA4-Ig, then were transfused with the donors' bone marrow cells into the preconditioned recipients. At last, the effect of combination of Tju103 and CTLA4-Ig on hematopoietic rebuilding, GVHD, GVL and anti-infection was observed in compared with CsA, Tju103 and CTLA4-Ig as controls.Results The only irradiated group (group A): All the mice (10 mice) died of failure of hematopoiesis within 11 days post irradiation, of which most (8 mice) died within 4～7 days post transplantation. The CTX-treated leukemia group (group B): All the mice (10 mice) died of leukemia within 16～23 days post leukemia cells infusion (11～18 days post BMT). CTX treatment prolonged the survival time. The only transplanted group (group C): The mice began to die from day 16 post transplantation, and all (10 mice) died of GVHD within 3 weeks. The CsA prophylaxis group (group D): 4 mice died within 8～22 days after transplantation, of which one died of leukemia, two died of infection and one died of GVHD, and the remaining 6 survived over 30 days post transplantation. The Tju103 treated group (group E): 4 mice died within 9～26 days post transplantation, of which one died of leukemia, one died of infection and two died of GVHD, and the remaining 6 mice survived over 30 days post transplantation. The CTLA4-Ig treated group (group F): 3 mice died within 14～23 days after transplantation, of which one died of infection and two died of GVHD, and the remaining 7 survived over 30 days post transplantation. The Tju103/CTLA4-Ig treated group (group G): one died of GVHD on day 19 after transplantation, and the remaining 9 mice survived over 30 days post transplantation. Conclusions CsA, Tju103 or CTLA4-Ig alone could prolong survival time and reduce incidence and degree of GVHD severity. But CTLA4-Ig could spare much ability of GVL and anti-infection while Tju103, just like CsA, couldn't. Combination of both was the most favorable way for transplantation with the most remarkable efficiency on prolongation of survival time and reduction of GVHD.

Objective To investigate the effect of donors' activating killer cell immunoglobulin-like receptor (aKIR) on recipients prognosis post haemopoietic stem cell transplantation (HSCT).Methods Fifty-nine cases of related HLA full matched HSCT were studied. Sequence specific primer polymerase chain reaction (SSP-PCR) was used to type donors' KIR. Virus,bacterium,and fungi infection as well as bleeding,relapse,survival were observed in the recipients post transplantation. Relationship between aKIR and clinical indexes mentioned above were analyzed.Results Donors' aKIR showed no beneficial effects on recipients' virus and bacterium infection. Furthermore,the incidence of fungi infection was increased in HSCT if donors expressed KIR3DS1 (?2= 4.804 ,P= 0.028 ). No significant difference was found in survival curve between donor aKIR positive HSCT or negative one (KIR2DS1+/-: P= 0.651 ; KIR2DS2+/-: P= 0.847 ; KIR3DS1+/-: P= 0.341 ). No effect of donor's aKIR on relapse was observed,too.Conclusion In Guangdong Han,so far as related HLA full matched HSCT is concerned,no improvement in recipient prognosis due to donor's aKIR is verified.

PC84045 human lung carcinoma cells (PC84045 HLCCs) were mixed with the BM cells activated by anti-CD3 McAb and IL-2 and then CD3-ALTO4 bi-specific McAbs was put into the cell suspension. The mixed cells were cultured for 3 days in presence of IL-2 and IL-3. Tumor cell colony formation method in half-solid culture system was used to assess the purging efficiency (PE) of the activated T cells to PC84045 HLCCs. The ratio of T cells to PC84045 HLCCs was 8 to 1, the PE was 4 Logs. The rario of T cells to PC84045 HLCCs was 16 tol, the PE was 5 Logs. When the purged BM cells were injected into the NC nude mice, none of subcutaneous node was found. The number of the CFU-GM, BFU-E of the purged BM cells was more than 85 % compared with that of the fresh BM cells (P

0.05). The survival time of the second control group and experimental groups was much longer than that of the first control group (P

Objective To study the feasibility of HLA non-identical allogeneic peripheral blood hematopoietic stem cell transplantation (allo-PBHSCT) for the treatment of patients with severe aplastic anemia (SAA). Methods Four patients with SAA received non-identical allo-PBHSCT from HLA one or three loci mismatched related donors. Donors were given G-CSF 5?g/(kg?d) for 5 days, then PBSC were harvested on days 4 and 5. Prime regimen consisted of fludarbine, CTX, ATG and TBI. Combination of CsA, MTX, MMF and Simulect (CD25 monoclonal antibody) were used for GVHD prophylaxis. Results Engraft was achieved in all patients. The median days of neutrophil exceeding 0.5?109/L and platelet exceeding 20?109/L were 15.3 days (range 14~16 days) and 17.5 days (range 16~19 days), respectively. In the 14-month median follow-up duration, only one of four patients developed grade Ⅱ aGVHD, and one patient died of systemic candidiasis, the other three were alive in disease-free condition. Conclusion HLA non-identical allo-PBHSCT was well tolerated by patients with SAA, and it provided rapid and sustained engraftment without increase in incidence of GVHD.

Objective: To clone the 5＇-flanking region of the human CD34 gene containing transcriptional regulatory sequence (TRS). Methods: According to the registered 5＇-flanking region of CD34 gene, two pairs of primers were designed and net-PCR was used to amplify 661 bp long TRS of CD34 gene. The CD34 TRS fragment was cloned into reported plasmid pEGFP-1. The role of the regulating the specific expression of recombinant plasmid pCD34 EGFP in hematopoietic and non-hematopoietic cells was observed. Results: Restrictive endonuclease identification and DNA sequencing provedthat the CD34 promoter cloned was consistent with the sequence reported to a large extent. It could induce the EGFP gene to express in hematopoietic cell line K562 specifically, while has no effect on hepatocellular carcinoma cell HepG-2. Conclusion: The cloned CD34 gene TRS has the effect of regulating gene expression specifically. The study established the fundament for the construction of specific gene expression vector used in hematopoietic system cells.

Objectives To study the effect of mesenchymal stem cells on the aging rat kidney and to explore the underlying mechanism. Methods Rat models of senile kidney were built with hypodermic injection of D-galactose daily. Injections of MSCs of 3 × 10<'6>were given to each rat through vena caudalis and CFSE was used as a tracing label to detect the distribution of MSCs in vivo. After 24 h, rats were dissected and their kidneys were frozen for section. MSCs were observed with Fluophot and quantitative analysis of the various parameters of kidney was performed under a light microscope with B12000 image analysis system.The contents of superoxide dismutase (SOD) and malondialdehyde (MDA) in serum and kidney were measured wtih hydroxylamine and chromatometry. The expression of VEGF and P16 mRNA in kidney tissue was detected with real-time PCR and Western blotting. Results MSCs was found homing in the rat kidney,and the glomerular size, sklerosis-rate and the average cell count of glomerulus in the treated group were different from those of the model group (P<0.05). In the treated group, the activity of SOD was significantly higher and the content of MDA was lower in serum and kidney than that in the model control group (P<0.05). The expression of VEGF mRNA and protein in the kidneys of MSCs group increased significantly as compared with the model group (P<0.05). The expression of P16 mRNA and protein in the kidney of MSCs group decreased significantly compared with the model group (P<0.05). Conclusion MSCs can increase the expression of VEGF while decrease the expression of P16, so as to play a key role in the anti-aging on rat kidney.