Objective　To investigate the relationship between the tumor angiogenesis and the clinicopathological variables of hepatocellular carcinoma (HCC) and determine whether tumor angiogenesis can be used as an indicator for predicting invasion, metastasis and prognosis of HCC. Methods　Using the immunohistochemical S-P method, CD34 expression was detected in liver tissue from 20 normal individuals, 20 patients with liver cirrhosis and 85 patients with HCC and juxtacancerous tissues and the microvascular density (MVD) determined in those from 85 patients with HCC. Results　CD34 was mainly located in the cell cytoplasm of endothelial cells. The CD34 staining was confined to vessels in the portal triad in the above-mentioned various tissues. In HCC tissues, however, sinusoid-like vessels reacted intensively with anti-CD34 and the MVD was 156.5±62.4 (per 0.74 mm2). MVD was correlated to tumor size, number, differentiation, emboli in the portal vein and capsule (P<0.05 or 0.01) but not to HBsAg and AFP (P>0.05). It was also correlated with prognosis of HCC. The mean tumor-free survival of MVD ≥156 was 14 months while that of MVD <156 was 53 months (P<0.01). Conclusion　The expression of CD34 in sinusoid-like vessels in HCC tissues represents the neovascularization. Angiogenesis in HCC is associated with the progression of HCC. MVD can be used as an indicator to judge the invasion, metastasis and prognosis of HCC.

Objective To investigate the alteration of lymphocyte P-gp expression level and function in liver transplant recipient and its effect on immunosuppressive therapy.Methods Using flow cytometry,the level of lymphocyte P-gp and analyzed the P-gp function in 53 cases of liver transplant recipieints was observed.Results Among 53 cases,the preoperative mean level of lymphocyte P-gp was(10.34?4.0)%,P-gp level began to increase in the 1st month postoperatively(17.8?8.0)%,peaked in the 3rd month(27.5?13.3)%,and then was stable at this level thereafter.There were no significantly difference between different sex of age group.But the P-gp level in CD+4T cells was significantly higher than that in CD+8T cells and B cells,RH123test showed that high P-gp expression of lymphocyte enhanced its transport activity of medical agents,and decreased the intracellular accumulation of drug more significantly than that of low P-gp expression(P

Objective To investigate the effects of probucol and losartan on cell proliferation after balloon angioplasty in hypercholesterolaemic rabbits.Methods Forty male New Zealand rabbits were randomly divided into high cholesterol diet group,probucol group,losartan group and combined drugs group.The iliac arteries of the rabbits were balloon-injured and the insulin-like growth factor-I receptor(IGF-IR),vascular endothelial growth factor(VEGF),and proliferating cell nuclear antigen(PCNA) were detected by using immunohistochemical method.Results Compared with high cholesterol diet group,the lumen areas were enlarged and the intimal areas were decreased in the probucol group,losartan group and combined drugs group(all P

Objective To investigate the effect of mild hypothermia on the expression of hypoxia-inducible factor-1α (HIF-1α) and glucose transporter-1 (GLUT-1) in a rat model of chronic cerebral ischemia-reperfusion.Methods Thirty-six female SD rats weighing 170-210 g were randomly divided into 3 groups (n = 12 each):sham operation group (group S), normal body temperature group (group NT ) and mild hypothermia group (group MH). Arterio-venous fistula was established by end-to-end anastomosis between the right common carotid artery and right external jugular vein for 6 weeks followed by reperfusion. In group MH, mild hypothermia was induced at the initiation of reperfusion and the rectal temperature was reduced to 31.5-32.5 ℃. In group S and NT, the rectal temperature was maintained at 37-38 ℃. Six rats in each group were sacrificed at 3 and 48 h of reperfusion. The brains were immediately removed for determination of the expression of HIF-1α, GLUT-1, HIF-1α mRNA and GLUT-1 mRNA and microscopic examination. Results Compared with group S, the expression of HIF-1α and HIF-1α mRNA at 3 and 48 h of reperfusion and GLUT-1 mRNA at 3 h of reperfusion was up-regulated, while the expression of GLUT-1 and GLUT-1 mRNA at 48 h of reperfusion was down-regulated in group NT (P ＜ 0.05).Compared with group NT, the expression of HIF-1α and HIF-1α mRNA at 48 h of reperfusion and HIF-1α mRNA and GLUT-1 mRNA at 3 h of reperfusion was down-regulated, while the expression of GLUT-1 and GLUT-1 mRNA at 48 h of reperfusion was up-regulated in group MH (P ＜ 0.05).Microscopic examination showed that the injury to the ultrastructure of blood-brain barrier was significantly attenuated in group MH compared with group NT. Conclusion Mild hypothermia can attenuate chronic ischemia-reperfusion injury by down-regulating the expression of HIF-1α and up-regulating the expression of GLUT-1.

Objective To investigate the influence and the clinical significance of interleukin-2 (IL-2) on T lymphocyte subgroup after chemotherapy in acute myeloid leukemia patients. Methods Fiftyfour acute myeloid leukemia patients were divided into treatment group (chemotherapy combined with IL-2,28 cases) and control group (pure chemotherapy,26 cases) by radom digits table. In treatment group, IL-2400 000 U was dripped after chemotherapy for 14 days. Then the T lymphocyte subgroup change before and after treatment was examined. Results After 14 days' treatment, the levels of CD3 (0.6026±0.2275 ),CD4(0.4972±0.1224),CD56(0.3016±0.1053 ) in treatment group were significantly higher than those before treatment(0.3926±0.2010,0.2264±0.1190,0.1729±0.1226) and in control group (0.4352±0.1930,0.2738±0.1362,0.1937±0.1268)(P< 0.05). Conclusion IL-2 treatment can obviously raise the levels of T lymphocyte subgroup and can raise the immunologic function.

Objective To choose one protocol that can quickly ,safely and effectively provide amount enough of bone marrow derived mesenchymal stem cells(MSCs) for use of clinical or experimental test through comparison of their growth characteristics and growth factors levels in culture solution .Methods Cells extracted from bone marrow of C57BL/C mice respectively underwent two different isolation protocols :whole bone marrow adherent culture(WBMAC) or gradient density separation(GDS);characteris‐tic surface antigens of MSCs were identified by flow cytometry on cells isolated in different ways ;the distinct growth curve of pri‐mary stem cells cultured in vitro described their different proliferation rate;levels of vascular endothelial growth factor(VEGF) and stromal cell‐derived factor‐1α(SDF‐1α) in culture medium were detected by ELISA .Results Primary MSCs obtained by WBMAC proliferated at higher speed and exhibited shorter growth cycle than those separated by GDS ;on MSCs from both groups ,surface antigens CD29 were detected positively ,and antigens including CD31 ,CD34 and CD45 were assayed negatively ;concentration of VEGF and SDF‐1αin both two nutrient solution primarily keep at low levels ,comparatively ,level of VEGF and SDF‐1αin the di‐shes which contain MSCs by WBMAC was higher than the one in the dishes which contain MSCs by GDS .Conclusion MSCs ex‐tracted by WBMAC shows unimpaired cell function ,can build automatically more suitable microenviroment for their growth;this classic method was qualified for clinical and experimental use in a safe ,rapid ,effective way .

Objective To discuss whether mild hyperuricemia can lead to kidney damage and the protection of decreased uric acid,through observing that hyperuricemia did damage to glomerulus endothelial function and cell proliferation of vascular smooth muscle in rats. Methods Fifty-four male SD rats were divided into four groups,the control group,model group (Oxonate),allopurinol group and Oxonate+allopurinol group.Rats were administered on a low sodium diet and their systolic blood pressure (SBP) were measured each 10 days.ELISA was used to detect rat plasma markers of endothelial function damage [nitric oxide (NO),type-1 plasminogen activator inhibitor (PAI-1),endothelin 1 (ET-1)] and cell proliferation of vascular smooth muscle[plateletderived growth factor (PDGF),cycloxygenase 2 (COX2),monocyte chemotactic protein-1 (MCP-1)],and the markers of inflammatory reaction[interleukin-18 (IL-18),tumor necrosis factor α(TNF-α)].PDGF and nitric oxide synthase (NOS) levels of rats were detected by immunohistochemical method.Renal tissue pathology of rats was observed. Results Compared to the control group,the plasmic concentration of COX2,ET-1,IL-18,PAI-1,PDGF,TNF-o,MCP-1 increased,and NO decreased significantly in rats of model group (all P＜0.05),expression of NOS significantly reduced and PDGF increased (all P＜0.05).Under light microscope,vascular wall thickening,intimal proliferation and lumen slight stricture without uric acid crystals in renal tissue were found in model group,which were obviously improved by using allopurinol. Conclusion Mild hyperuricemia can do damage to endothelial function of glomerulus and lead to vascular cell proliferation,which can be improved through decreasing uric acid.

BACKGROUND: Bushen Huoxue Tongluo Capsule, a well-known traditional prescription, exhibits remarkable treatment outcomes in knee osteoarthritis, but the underlying mechanism is not fully understood. OBJECTIVE: To explore the regulatory effect of Bushen Huoxue Tongluo Capsule on miR-27a of synovial cells in Sprague-Dawley rats with knee osteoarthritis. METHODS: The model of knee osteoarthritis was established in 120 Sprague-Dawley rats using Hulth method, and then these rat models were randomized into four groups, followed by given75 mg/kg normal saline (blank control group), 34 (low-dose group), 75 (medium-dose group) and 140 (high-dose group) mg/kg Bushen Huoxue Tongluo Capsule aqueous solvent via gavage according to body mass, respectively, twice daily, for consecutive 2 courses (15 days as a course) with 2 days in between. The synovium and cartilages were removed after each course, and the expression of miR-27a in the knee joint synovium and chondrocytes was determined by real-time quantitative PCR. RESULTS AND CONCLUSION: Gross observation found that the degeneration of synovium and cartilage in the Bushen Huoxue Tongluo Capsule groups was milder than that in the blank control group. Real-time PCR revealed that there was an increasing trend in the expression of miRNA-27a in the synovium and cartilage in each group, especially in the high-dose group, and there was a significant difference between groups (P < 0.05). Our findings show that Bushen Huoxue Tongluo Capsule can suppress the reduced expression of miRNA-27a in the synovium and chondrocytes in rats with knee osteoarthritis, further alleviating knee degeneration and protecting the knee from damage.

BACKGROUND:Different anesthesia methods significantly impact mean arterial pressure, systolic and diastolic blood pressure, arterial blood pH and blood viscosity, but what kind of anesthesia method is more suitable for preparation of animal model does not reach an agreement. OBJECTIVE: To compare the effectiveness of local anesthesia and general anesthesia for preparation of rabbit intervertebral disc degeneration model in procession of percutaneous acupuncture rotary cut. METHODS:A total of 48 New Zealand white rabbits were randomly divided into the local anesthesia group and the general anesthesia group. The local anesthesia group was anesthetized with 0.5% lidocaine. The general anesthesia group was intraperitoneally injected with 3% sodium pentobarbital (30 mg/kg). In the two groups, models of intervertebral disc degeneration were established by minimally invasive percutaneous acupuncture rotary cut at L4/5, L5/6 levels. Modeling time was observed in both groups. Disc degeneration was evaluated by the gross observation, MRI detection, and histopathological examination at 4, 8, 12, and 16 weeks after modeling. RESULTS AND CONCLUSION: (1) General observation: the nucleus pulposus gradually with darker color and elasticity reduced in two groups after modeling. (2) MRI T2-weighted image results showed no significant changes in disc signal intensity at early stage, but the signal strength showed a decreasing trend with time. (3) Disc degeneration was evaluated according to Pfirrmann grading. Disc degeneration was gradually increased with time in two groups (P< 0.05), but there was no significant difference at postoperative time points in both groups (P > 0.05). (4) Masson staining showed that different degrees of annulus irregular arrangement appear in the two groups 8 and 12 weeks after modeling, but the structure was still intact. 16 weeks later, annulus disorganized, or even breakage in the two groups; no significant difference was detected between the two groups. (5) Safranin O staining showed that nucleus pulposus cells were not significantly reduced in the two groups at 4 weeks, but were significantly reduced at 16 weeks. (6) The average modeling time of local anesthesia group [(15.24±2.67) minutes] was significantly shorter than that in the general anesthesia group [(25.64±6.85) minutes] (P < 0.05). These results indicate that intervertebral disc degeneration model can be successfully established by minimally invasive percutaneous acupuncture rotary cut by using local anesthesia and general anesthesia; however, local anesthesia has a shorter operating time and simple anesthetic procedure. The efficacy was identical between the two methods.

BACKGROUND:The Herbal Compound formula Yaotu Granules for lumbar disc degeneration has achieved satisfactory efficacy, but the underlying mechanism remains unclear. OBJECTIVE: To explore the possible molecular mechanisms ofYaotu Granules for lumbar disc degeneration based on transcriptome sequencing (RNA-seq). METHODS: Ten New Zealand white rabbits were randomly divided into control and experimental groups (n=5 per group), followed by intragastric injection of 20 mL of normal saline and 20 mL of water decoction of Yaotu Granules, respectively, twice daily for consecutive 1 week. The different drug serums were prepared using serum samples obtained from the common carotid artery at 2 hours after the last injection. Human nucleus pulposus cells were cultured in DMEM containing 10% different drug serums to observe the cellular morphology in each group. The cell viability was detected by trypan blue staining. The passage 3 human nucleus pulposus cells were pretreated for 48 hours, the mRNA level was detected using RNA-seq, and the differentially expressed mRNA was screened by RNA-seq. Subsequently, the differential gene expression, Gene Ontology (GO) enrichment analysis and Pathway enrichment analysis were performed. RESULTS AND CONCLUSION: In the experimental group, there were round or fusiform cells with abundant cytoplasm and clear nuclei that showed smooth surface and complete nuclear membrane; elongated rough endoplasmic reticulum arranged regularly, and less mitochondrion, scattered lysosomes and filaments were visible in the cytoplasm. In the control group, spindle and polygonal cells were found, and the large nuclei with 2 or 3 nucleoli and slightly rough surface were observed; there were mildly dilated endoplasmic reticulum, and few mitochondria with incomplete membrane. The adherent rate, generation time and cell viability of passage 1, 2 and 3 cells in the experimental group were significantly better than those in the control group (P < 0.05). Sequencing results found that there were 464 differentially expressed genes including 143 upregulated, and 321 downregulated genes. GO analysis revealed that the differentiated genes were mainly concentrated on cell regulation and metabolism. Pathway analysis found that mainly differentiated genes focused on the metabolism related pathways. These findings suggest that the differentially expressed gene profile of Yaotu Granules for lumbar disc degeneration is obtained by RNA-seq technology.Yaotu Granules mainly upregulate extracellular matrix metabolism-related genes and downregulate polysaccharide synthesis related genes in the prevention and treatment of lumbar disc degeneration, which provides basis for further research.