Enhancing Effects of Ligustrazine on Expression of CD31 and Hematopoietic Reconstitution in Syngenic Bone Marrow Transplantation of Mice

Summary： The effect of ligustrazine on the expression of CD31 in syngenic bone marrow transplantation （BMT） mice was studied. Fifty-six Balb/c mice were divided into 3 groups： normal control group. BMT control group, and ligustrazine treated group. Syngenic BMT mouse models were established according to the literatures. In BMT control group and the ligustrazine treated group, the mice were given respecxively orally 0.2 mL saline and 2 mg ligustrazine twice a day. On the 7th, 14th, and 21st day after BMT, the mice were killed. The expression of CD31 on the surface of bone marrow nuclear cells （BMNC） was detected by flow cytometry. Peripheral blood leukocytes, platelets and BMNC were counted. Histological observation of bone marrow was made. The results showed thai in ligustrazine treated group the peripheral blood leukocylcs, platelets and BMNC counts, and the expression of CD31 on the day 7, 14, 21 after BMT were higher than in BMTcontrol group （P〈0.01 or P〈0.05）. In conclusion, ligustrazine could obviously enhance the CD31 expression on the surface of BMNC after syngcnic BMT in mice, which may be one of the mecha- nisms underlying the ligustrazine accelerating hematopoietic reconstitution in syngenic BMT.

Study on the Effect of Ligustrazine on Hematopoietic Reconstitution in Bone Marrow Transplantation Mice

To explore tile effects of ligustrazine on hematopoietic reconstitution and its mechanism after bone marrow transplantation (BMT), the allogenic BMT mice were given intra-abdominal injection of 2,mg ligustrazine twice a day. On the 1st, 7th, 14th, and 28th day after BMT, peripheral blood cells and bone marrow nuclear cells (BMNC) were counted, and the histological features were evaluated. On the 7th, 14th, 21st day after BMT, CXCR4 expression on the BMNC was assayed. The results showed that peripheral blood cell counts and BMNC counts in ligustrazine-treated group on the 7th, 14th, 28th day were higher than those in BMT group (P<0. 01 or P<O. 05). The percentage of hematopoietic tissue volume, fat tissue hyperplasia and congestion and dilation degree of microvessel in ligustrazine-treated group on the 7th, 14th, 21st, 28th day was higher than those in BMT group. The CXCR4 expression levels in ligustrazine-treated group were higher than in BMT group (P<0.01 or P<0. 05) on the 7th and 14th day, and were lower than in BMT group on the 21st day (P<0. 01 ). It is concluded that the ligustrazine can accelerate hematopoietic reconstruction, enhance growth of hematopoietic tissues and promote the repair of microvessels. The CXCR4 expres- sion levels on BMNC may be responsible for the effect of ligustrazine.

How mesenchymal stem cell cotransplantation with hematopoietic stem cells can improve engraftment in animal models

BACKGROUND Bone marrow transplantation(BMT)can be applied to both hematopoietic and nonhematopoietic diseases;nonetheless,it still comes with a number of challenges and limitations that contribute to treatment failure.Bearing this in mind,a possible way to increase the success rate of BMT would be cotransplantation of mesenchymal stem cells(MSCs)and hematopoietic stem cells(HSCs)to improve the bone marrow niche and secrete molecules that enhance the hematopoietic engraftment.AIM To analyze HSC and MSC characteristics and their interactions through cotransplantation in murine models.METHODS We searched for original articles indexed in PubMed and Scopus during the last decade that used HSC and MSC cotransplantation and in vivo BMT in animal models while evaluating cell engraftment.We excluded in vitro studies or studies that involved graft versus host disease or other hematological diseases and publications in languages other than English.In PubMed,we initially identified 555 articles and after selection,only 12 were chosen.In Scopus,2010 were identified,and six were left after the screening and eligibility process.RESULTS Of the 2565 articles found in the databases,only 18 original studies met the eligibility criteria.HSC distribution by source showed similar ratios,with human umbilical cord blood or animal bone marrow being administered mainly with a dose of 1×10^(7) cells by intravenous or intrabone routes.However,MSCs had a high prevalence of human donors with a variety of sources(umbilical cord blood,bone marrow,tonsil,adipose tissue or fetal lung),using a lower dose,mainly 106 cells and ranging 104 to 1.5×107 cells,utilizing the same routes.MSCs were characterized prior to administration in almost every experiment.The recipient used was mostly immunodeficient mice submitted to low-dose irradiation or chemotherapy.The main technique of engraftment for HSC and MSC cotransplantation evaluation was chimerism,followed by hematopoietic reconstitution and survival analysis.Besides the engraftment,homing and cellularity were also evaluated in some studies.CONCLUSION The preclinical findings validate the potential of MSCs to enable HSC engraftment in vivo in both xenogeneic and allogeneic hematopoietic cell transplantation animal models,in the absence of toxicity.

Hepatocyte growth factor gene transfer effects on the femoral and intramuscular nerve in a canine model of lower limb ischemia

BACKGROUND： Recent advancements in gene therapy have provided new methodology for treating ischemia in lower extremities. Gene transfer of angiogenic factors to ischemic tissues may promote local proliferation of new vessels and form collateral circulation. OBJECTIVE： To observe histopathological changes in the femoral and intramuscular nerve three months after intramuscular injection of hepatocyte growth factor （HGF） into the peripheral skeletal muscle in a canine model of lower limb ischemia. DESIGN： Randomized occlusion modelled and verification animal study. SETTING： Experimental Center, Lanzhou General Hospital of Lanzhou Military Area Command of Chinese PLA. MATERIALS： This study was performed at Animal Experimental Center, Lanzhou General Hospital of Lanzhou Military Area Command of Chinese PLA from September to November 2006. A total of eight male mongrel dogs, weighing 12–15 kg and 1.5–3 years of age, were selected for this study. This experimental study was in accordance with local ethics standards. Recombinant plasmid carrying HGF （pUDKH） and occlusion model plasmid （pUDK） were provided by the Third Laboratory of Radiation Medical Institute, Academy of Military Medical Sciences of PLA. METHODS： Grouping and model establishment： under anesthesia, complete vascular occlusion models were established on the left lower extremities. The experimental dogs were randomly divided into a model group and a pUDKH treatment group, with four dogs in each group. Dogs in the pUDKH group were injected with 0.15 mg/kg pUDKH. Ten minutes later, intramuscular injections were performed at three spots into the peripheral skeletal muscle of the left hind limb, as well as lateral injections at two spots. The injection volume at each spot was 0.2 mL. Dogs in the model group were injected with pUDK, and dosage and injection method were identical to the treatment group. MAIN OUTCOME MEASURES： Histopathological changes in the femoral nerve, as well as internal and external intramuscular nerve tissues in the hind limb of dogs three months after plasmid injection under optic microscope. RESULTS： （1） Histopathological changes in the femoral nerve： tiny nerves from the femoral nerve to the intramuscular nerve exhibited marked degeneration in the model group. The degenerating features included neurites, myelin sheaths, and Schwann cell nuclei. Neuropathy in the pUDKH treatment group was not detected. （2） Histopathological changes of the intramuscular nerve： large and irregular vacuoles were present on several longitudinal sections of intramuscular nerve fibers in the model group, as well as annular-shaped blank regions on transverse sections of peripheral neurites. In the pUDKH treatment group, large, blank regions were present in several segments of partial nerve fibers of the longitudinal intramuscular nerve region, but only a few nerve fibers exhibited annular-shaped blank regions on the transverse section of peripheral neurites. CONCLUSION： Local pUDKH injection may relieve or block femoral and intramuscular nerve tissue injury in a canine mocel of lower limb ischemia.

Effects of Ligustrazine on Hematopoiesis in the Early Phase of Bone Marrow Trans plantation Mice

To investigate the effects of Ligustrazine on histogenesis of bone marrow in the early phase of hematopoietic reconstruction in bone marrow transplantation (BMT ) mice. The syngeneic BMT mice model was established. The syngeneic BMT mice were orally given 2 mg Ligustrazine twice a day. 1, 3, 5, 7, 10, 15 and 21 day(s) after BMT, peripheral blood granulocytes and bone marrow nucleated cells (BMNC) were counted and the diameter of central vein and the area of micro vessel in femur were measured. The effect of Ligustrazine on hematopoietic stem cells was observed by colony forming unit of spleen (CFU S). The effect of Ligustrazine on hemopoietic progenitors was studied by observing the number of progenitors of Granulocytes/Macrophage on day 10 and day 20 after BMT. In Ligustrazine treated group, the diameter of center veins and the area of micro vessel of femur were all significantly less than the control group 7, 10, 15, 21 days after BMT ( P <0.01). In addition, Ligustrazine significantly increased the number of CFU S on day 10 and the number of CFU GM on day 10, 20 after BMT. These results indicate that Ligustrazine can accelerate the histogenesis of hemopoietic bone marrow, which may be one mechanism by which Ligustrazine promotes hematopoietic reconstitution after BMT.

Effects of Ligustrazine on Expression of Bone Marrow Heparan Sulfates in Syngeneic Bone Marrow Transplantation Mice

To explore the effects of ligustrazine on bone marrow heparan sulfates (HS) expression in bone marrow transplantation (BMT) mice, the syngeneic BMT mice were orally given 2 mg ligustrazine twice a day. On the 7th, 10th, 14th, 18th day after BMT, peripheral blood cells and bone marrow nuclear cells (BMNC) were counted, and the expression levels of HS in bone marrow and on the stromal cell surfaces were detected by immunohistochemistry and flow cytometry assay respectively. In ligustrazine-treated group, the white blood cells (WBC) and BMNC on the 7th, 10th, 14th, 18th day and platelets (PLT) on the 7th, 10th day were all significantly more than those in control group (P<0.05). The bone marrow HS expression levels in ligustrazine-treated group were higher than those in control group (P<0.05) on the 7th, 10th, 14th, 18th day. However, the HS expression levels on the stromal cell surfaces showed no significant difference between the two groups on the 18th day (P>0.05). It was concluded that ligustrazine could up-regulate HS expression in bone marrow, which might be one of the mechanisms contributing to ligustrazine promoting hematopoietic reconstitution after BMT.