Objective To observe the effect of methylprednisolone (MP) pre-intervention on ex-pressions of heat shock protein 27 (HSP27) and tumor necrosis factor alpha (TNF-α) in cells in rat spinal cord following ischemia-reperfusion injury. Methods One hundred and fifty male Sprague-Dawley (SD) rats were randomly divided into 3 equal groups: group A (control) in which the abdominal aorta was exposed without any treatment, group B in which the abdominal aorta was clipped for 30 minutes before reperfusion for 3 bours to establish a model of ischemia- reperfusion injury, and group C in which intravenous MP injection was conducted 30 minutes before the establishment of the ischemia-reperfusion injury model. Three hours later the spinal cords were harvested. Pathological changes of spinal cord cells were observed with HE staining and expressions of HSP27 and TNF-α in spinal cord cells were observed with immunohistochemical staining. The motor function of hind-limbs before was evaluated before sample harvest. The data were analyzed with SPSS software. Results There were significant differences between groups A and B in the expressions of TNF-α and HSP27. Compared with group B, the expression of TNF-α decreased and HSP27 increased in group C, with statistically significant differences between the 2 groups. The motor function score of hind-limbs decreased in group B but improved in group C. Conclusions Since MP can decrease the expression of TNF-α and up-regulate the expression of HSP27, it has a potency of neuro-protection. Spinal cord ischemia-reperfusion injury can be avoided or decreased after MP pre-intervention.

Objective:This study aimed to identify the morphology of mast cells by using a modified toluidine blue staining scheme,so as to provide a powerful reference for the experimental basis research of mast cells.Methods:Bone marrow-derived mast cells were induced in vitro.After 4 weeks,the cells were collected,fixed,and stained.Mast cells were fixed at different temperature during different time.The optimum condition was determined by comparing the effects of toluidine blue staining.Results:Bone marrow cells were induced to differentiate into mast cells by SCF and IL-3 in vitro.When mast cells were stained with modified toluidine blue staining,the staining effect was better.Mast cells were round or oval and the cell membrane was complete and the cytoplasm was filled with a large number of purple particles.Conclusion:In this study,we successfully applied a modified toluidine blue staining method to mast cells cultured in vitro.The results showed that the condition at 37 ℃ full fixation with staining could reduce the degeneration of mast cells.This method was easy to operate with good stability.It was suitable for the morphological observation of mast cells cultured in vitro.

Objective To evaluate the specificity,sensitivity and accuracy of miniature probe combined with radial scanning endoscopic ultrasonography(EUS)in preoperative TN staging of rectal cancer,and to assess its value in the choice of therapeutic strategy.Methods A total of 60 patients with rectal cancer received EUS assessment before surgery.Diagnosis was made according to TNM standard and compared with those of MRI and postoperative pathological examination.The reference value of EUS for therapy selection was studied.Results According to EUS staging,there were 4 cases of TI,18 T2,30 T3 and 8 T4,among which 7 cases were over-staged and 4 others were under-staged.MRI staging showed 1 case of T1,18 T2,30 T3 and 10 T4,among which 14 were over-staged and 3 others were under-staged.The total accuracy of EUS in T staging and N staging was 81.67%(49/60)and 78.33%,respectively,with the sensitivity and specificity at 71.43%and 91.03%,respectively.Accuracy of MRI for T staging and N staging were 71.67%(43/60)and 83.33%,respectively,with the sensitivity and specificity as 85.71%and 86.96%.Conclusion EUS with combination of miniature probe and radial scanning is effective in preoperative TN staging of rectal cancer with easy manipulation and less pain.

OBJECTIVETo explore the effect and safety of topical propranolol hydrochloride gel for treatment of infantile hemangioma. METHODS Thirty nude mice (BALA/c, nu/nu) were divided into three groups, experimental group, control group and normal group. Human hemangioma endothelial cells cultured in vitro were injected subcutaneously in experimental group and control group to establish infantile hemangioma model. Topical propranolol hydrochloride gel was applied on the surface of the hemangioman in experimental group and normal group. Tumor volumn change and the skin situations (edema, erythema, ulceration) were observed at different periods. 45 days after cell injection, the mice were killed and plasma concentration was detected in the experimental group and the control group by high performance liquid chromatography with evaporative light scattering detector, and tumors were subjected to histopathologic examination and immunohistochemistry for CD31 and CD34. The correlation between volumes and plasma concentration was statistically analyzed with SPSS 13.0 paired samples t test with α = 0.05 as statistical standard.

RESULTSAt 45 days, the volume of the tumor in control group was (366.57 ± 17.08) mm³, which has a significant difference as compared to the experimental group (13.36 ± 2.09) mm³ (P < 0.05); and the plasma concentration was (16.83 ± 1.53) ng/ml in experimental group, and (18.42 ± 2.21) ng/ ml in normal group (P > 0.05 ). Topical propranolol hydrochloride gel (3%) has no irritation to nude mice's skin.

CONCLUSIONSTopical application of 3% propranolol hydrochloride gel is effective and safe for the treatment of infantile hemangioma.

Animals ; Gels ; administration & dosage ; Hemangioma ; drug therapy ; pathology ; Humans ; Immunohistochemistry ; Mice ; Mice, Nude ; Propranolol ; administration & dosage ; Skin Neoplasms ; drug therapy ; pathology ; Tumor Burden ; drug effects

Objective To explore the differential liver plasma membrane (PM) proteins that may be related to the occurrence,development and reversal process of hepatitis and to understand the pathogenesis of hepatitis and the new drug targets by performing a comparative proteomics research of liver PM between hepatitis B surface antigen (HBsAg) transgenic mice and wild-type C57 mice.Methods A 6-month-old HBsAg transgenic mouse model was established.The pathological examination was performed to observe the pathological changes of transgenic mice and wild-type C57 mice.The PM from liver tissue of 6-month-old transgenic mouse and the control mouse were purified through twice sucrose density grade centrifugation combined with second antibody magnetic bead enrichment.The purity of extracted PM was verified by Western blot.Differential proteome expression analysis was performed by using two-dimensional electrophoresis (2-DE) and ImageMaster software analysis.The differentially expressed proteins were lysed by trypsin and identified by liquid chromatography combined with tandem mass spectrometry (LC-MS/MS) analysis.Results The pathological examination results showed that hepatitis was observed in the transgenic mouse group,while no abnormity was found in the controls.The PM was successfully enriched and the mitochondria contamination was reduced by sucrose density grade centrifugation combined with second antibody magnetic bead purification treatment.Thirty differential mice liver PM protein spots were visualized,in which 11 non-redundant proteins were successfully identified by LC-MS/MS in transgenic mouse group,including 9 up regulated protein spots and 2 down-regulated protein spots.These differentially expressed proteins included keratin,cardiac Ca2+ release channel,cytochrome B5,ATP synthase subunit alpha,etc.Conclusions A batch of HBsAg gene expression related differential proteins are identified in mouse liver plasma.These proteins might be new drug targets for anti-HBV treatment.This study will guide further investigation on the mechanism of HBV infection induced hepatitis.

Objective To study the effect of brucine on the growth of a hepatocellular carcinoma cell line in vitro. Methods Brucine was added into a liver cancer cell line of SMMC-7721 in vitro, at drug concentration of brucine from 2. 5 μg/ml to 400 μg/ml. The inhibition rate of cell growth was measured by MTT technique after the cells were cultured for 72 hours. The protein and mRNA expression of PCNA,cyclin D1 and FAS were respectively assayed with Western blotting and fluorescent quantitation RT-PCR techniques at 24, 48, 72 h. Results The inhibition rate of liver cancer cell was near 100% when the brucine concentration was at 320 μg/ml. The protein and mRNA expression of FAS were of no significant difference at 24 h vs 48 h ( seperately F = 2. 547,1. 582, all P ＞ 0. 05 ), and significant difference existed at 24 h vs 72 h( seperately F = 1. 036, 1. 137, all P ＜ 0. 05 ). The protein and mRNA expression of PCNA,Cyclin D1 were of no significant difference between various time period( seperately PCNA F = 3.612,2. 174,3.029;Cyclin D1 F=2.361,2.915,1.976,all P＞0.05). Conclusions Brucine inhibits the growth of liver cancer cells, by inducing increased apoptosis of the cells probably through FAS overexpression.

Objective: To investigate the effects of Chinese herbal drug-containing serum, prepared by administration of Chinese herbal medicine for activating blood (Xiongshao Capsule, XS) or for activating blood and detoxifying (Xiongshao Capsule plus Huanglian Capsule, XSHL) in rats, on cell viability, oxidative damage and apoptosis of human umbilical vein endothelial cells (HUVECs) induced by oxidized low-density lipoprotein (ox-LDL). Methods: Thirty-two rats were randomly divided into 4 groups: control group, positive control group (simvastatin 1.8 mg/kg), activating blood (XS, 0.135 g/kg) group, and activating blood and detoxifying (XS Capsule 0.135 g/kg and Huanglian Capsule 0.135 g/kg, XSHL) group. Corresponding drugs were continuously administered to the rats for 7 days and then drug-containing serum was harvested 1 hour after the last administration. HUVECs isolated from newborn children by collagenase digestion were stimulated by ox-LDL (100 μg/L) and incubated with corresponding drug-containing serum for 24 hours. Untreated HUVECs were also used as a normal control. The morphology and structure of HUVECs were observed by an inverted microscope. Cell viability was measured by methyl thiazolyl tetrazolium method, and cell membrane damage was determined by lactate dehydrogenase (LDH) leakage. Activity of superoxide dismutase (SOD) was examined by spectrophotometry, and content of malondialdehyde (MDA) in the cell lysate was examined by thiobarbituric acid assay. HUVECs were stained with Annexin V-fluorescein isothiocyanate and propidium iodide and analyzed on a flow cytometry to determine apoptosis. Results: Compared with the normal HUVECs, the cell viability and the activity of SOD were significantly decreased while the content of MDA and apoptosis rate were significantly increased after 24-hour ox-LDL stimulation (P<0.01, P<0.05). Simvastatin-, XS-, and XSHL-containing serum significantly promoted the ox-LDL-stimulated HUVEC viability and inhibited early apoptosis (P<0.01, P<0.05), while had no significant effect on LDH leakage. Simvastatin-containing serum and XS-containing serum also showed significant decrease in MDA content and increase in SOD activity, while XSHL-containing serum showed no significant effects. There was no significant difference between the XS-containing serum group and the XSHL-containing serum group. Conclusion: Both sera containing XSHL and XS show protective action against the oxidative damage and apoptosis of HUVECs induced by ox-LDL.

Objective To observe the clinical significance and effect of dysfunction of dendritic cell( DC) in colorectal cancer with liver metastasis.Methods Peripheral blood were respectively collected from healthy adult donors (30 cases), preoperative and postoperative coloreatal cancer patients without liver metastasis (30 cases) , and 30 postoperative coloreatal cancer patients with liver metastasis from Jan 2008 to Jun 2010.Peripheral blood mononuclear cells were separated, GM-CSF( 1000 U/ml) , IL-4( 1000 U/ml) and TNF-α ( 1000 U /ml) were added into cell culture fluid to induce the mononuclear cells for mature dendritic cells.There were two subgroups, and in antigen processing subgroup the lysate of HT29 colonic carcinoma cells (100 μg/ml) were added into the cell culture fluid.The T lymphocytes from healthy adults were added into two subgroups by ratio of 1∶10 ( DCs∶ T cells) , cocultured for 7 days.The level of INF-γ and IL-10 in cell culture fluid was assayed with ELISA method.The optical density (OD) of CCK8 ans LDH was assayed with ELIASA to indirectly measure the reproductive activity and the killing efficacy of T lymphocytes.Results The IL-10 level in cocultured fluid of peripheral blood DCs in postoperative colorectal carcinoma patients with liver metastasis and T lymphocytes of healthy adults was significantly higher than that of preoperative patients of colorectal carcinoma and health adults without tumor antigenic stimulation(11.9±1.3) pg/ml vs.(29.6±9.7) pg/ml, (23.4±8.0) pg/ml, F =4.475, P ＜0.05).The IFN-γ level in cocultured fluid of peripheral blood DCs in postoperative colorectal carcinoma patients with liver metastasis and T lymphocytes of healthy adults was significantly higher than that of postoperative patients of colorectal carcinoma and healthy adults with or without tumor antigenic stimulation ( 34 ± 9) pg/ml vs.(26 ± 12 ) pg/ml, (24 ± 6) pg/ml, F = 5.206, P ＜ 0.05).The killing activity of healthy adults T lymphocytes induced by HT29 colonic carcinoma cells in postoperative colorectal carcinoma patients with liver metastasis was significantly higher than that of preoperative patients of colorectal carcinoma and healthy adults (30.6 ±8.6) pg/ml vs.(12.1 ±2.4) pg/ml, (14.9 ± 1.7) pg/ml, F =4.147, P ＜ 0.05).Conclusions T lymphocytes produce IL-10 when indued by DCs from patients with colorectal carcinoma under stimulation of tumor antigen leading to tumor immune escape and liver metastasis.The killing activity of T lymphocytes can be enhanced when stimulated by exogenous tmuor antigen.

BACKGROUND: Magnalium which is potential to be the medical biodegraded metal implant is more and more interesting,but it must be well biocompatibility to human body.OBJECTIVE: To evaluate the sensitization of magnalium (AZ31B).METHODS: A total of 35 guinea pigs were randomly divided into saline group (negative control group,n=10),5% volume of formaldehyde (positive control group,n=10),and AZ31B group (n=15).Sensitization test at the maximal dosage was performed according to "Biological evaluation of medical devices-Part 10: Tests for irritation and delayed-type hypersensitivity",including intracutaneous induction,local induction,and provocation.Patch was removed after 6,24,48,and 72 hours,and the skin response was classified accordingto Magnusson and Kligman criteria.Patch was removed after 72 hours,and skin was performed with biopsy,stained with FIE staining,and observed under optic microscope.RESULTS AND CONCLUSION: Sensitization response was not tested in both negative control group and AZ31B group at 24,48,and 72 hours after patch removal; however,moderate erythema was observed in the positive control group.Optic microscope demonstrated that criteria of allergy such as spongiosis,edema,and diffuse as well as perivascular mononuclear infiltration was not observed in the AZ31B group,but a few basophilic calls ware observed.This suggested that AZ31B was biologically safe for sensitization.

Objective To observe the therapeutic effect of brucine nanoparticles on hepatocellular carcinoma.Methods Brucine nanoparticles with block copolymer of carboxylation polyethylene glycol and polylactic acid were manufactured by ultraphonic emulsification.The effect brucine nanoparticles on growth of SMMC-7721 cell line was observed in vitro.The protein and mRNA levels of Fas were measured with Western blotting and FQ-PCR respectively after the brucine nanoparticles were added into cell culture fluid for 72 hours.Results The mean diameter,the carried drug rate and the entrapment rate of brucine nanopaticles was 146±96 nm,4.2％ and 67％,respectively.The growth inhibition of liver cancer cells was enhanced significantly with the increasing drug dose.The IC50 of growth inhibition of 5-FU,brucine and brucine nanoparticles was 16.7 μg/ml,90.3 μg/ml and 164.9 μtg/ml,respectively.There was significant difference among them (P＜0.05).The protein and mRNA expression of Fas in brucine nanoparticles treated SMMC-7721 cells increased significantly compared with that in blank control group(P＜0.05).Conclusion Brucine nanoparticles may potentially be a novel therapeutic drug for hepatocellular carcinoma.