Objective To investigate the effect of progesterone pretreatment of focal cerebral ischemic and reperfusion injury (fCIRI) and underlying molecular mechanisms.Methods A single intraperitoneal injection of progesterone (8 mg/kg) given 1 h,48 h and 96 h before fCIRI was established in male Sprague-Dawley rats.The number of survival of neurons in hippocampal CA1 region of the ischemiaside,as well as spatial memory function,was detected on days 3-8 after fCIRI.Extracellular-signalregulated kinase 1/2 phosphorylation (p-ERK1/2) and nuclear translocation of p-ERK1/2 in hippocampal CA1 region were examined using western blot.Results The number of survival of neuronal cells was significantly increased in ischemic groups treated with progesterone at 1 h and 48 h pre-fCIRI (164.3 ± 11.0,218.5 ± 9.1 and 142.7 ± 12.1,F =29.4,P ＜ 0.01) compared with fCIRI group treated with vehicle.Likewise,the escape-latency to reach the hidden-platform recorded in day 5 of Morris water maze test was reduced markedly in fCIRI-treatment groups compared with the vehicle group(10.3 ± 11.1,19.2 ±9.6 and 32.4 ± 14.3 ;F =35.8,P ＜0.01).The level of p-ERK1/2 was elevated notably during 24 h to 48 h postprogesterone by western blot,while restored to the baseline at 96 h post-progesterone.Improved nuclear translocation of p-ERK1/2 was observed from 2 h to 48 h post-progesterone.The progesterone receptor antagonist RU486 blocked the exaltation of either intracellular level or nuclear translocation of p-ERK1/2,which was induced by progesterone.Conclusions The pretreatment with progesterone exerts a neuroprotective effect against the ischemia-induced neuronal death and ameliorates the deficits in spatial memory through enhancing the activation of ERK1/2.The neuroprotection derived from pretreatment with progesterone achieves a time window of not less than 48 h,which is progesterone receptor-mediated ERK1/2 signaling pathway-dependent.

This article was aimed to study the therapeutic effects and anti-inflammatory effects of Chinese and west-ern medicine ointment on fungal infection animals. Guinea pig models which were infected with Trichophyton rubrum or Microsporum lanosum were applied to observe the effects of antifungal infections on She-Zhi Qin-Xian (SZQX) ointment by transdermal administration. The ointment is the combination of miconazole nitrate, snake oil and Sophora flavescens which is traditional Chinese medicine (TCM). Simultaneously, the models of croton oil-induced mouse ear swelling and carrageenan-induced rat paw swelling were established to observe the curative effect of SZQX ointment. The results showed that the SZQX ointment had a protective role on both guinea pig models. The microscopic scale of localized lesions in infected guinea pigs was significantly decreased (P< 0.01). The effect of SZQX ointment was better than the single using of Chinese or western medicine. The SZQX ointment was also able to significantly reduce the croton oil-induced mouse ear swelling and carrageenan-induced rat paw swelling (P< 0.05, or P< 0.01) with distinct anti-inflammatory effect. And the effect of SZQX ointment was superior to the single using of either Chinese or western medicine. It was concluded that the SZQX ointment had significant antifungal infection and anti-inflam-matory effect.

Objective To investigate the effects of recombinant tissue plasminogen activator (rt-PA)intravenous thrombolysis on blood-brain barrier (BBB) permeability,the expressions and the activities of MMP-2 and MMP-9 after cerebral ischemia in rats.Methods A total of 40 adult male Sprague-Dawley rats were allocated into 3 groups:Sham operation group (n =10),middle cerebral artery occlusion (MCAO) group (n =18),and rt-PA thrombolysis group (n =18).A MCAO model was established by using autologous thromboembolism.The sham operation group did not inject any thromboembolus,the MCAO group only made MCAO,and the rt-PA thrombolysis group received intravenous thrombolysis with rt-PA at 3 hours after MCAO.Brain infarct volume was determined by 2,3,5-triphenyl tetrazolium chloride staining BBB permeability was measured by Evans blue dye leakage.The activities and the expressions of MMP-2 and MMP-9 in brain tissue were detected by Gelatin zymography and Western blot,respectively.Results Compared to the MCAO group,the neurological function was improved significantly in the rt-PA thrombolysis group,and the infarct volume was also reduced significantly (t =7.365,P =0.005).However,the hemorrhage score (t =-3.286,P =0.017) and BBB permeability (t =-3.947,P =0.029) were increased significantly.The activities and the expressions of MMP-2 and MMP-9 in the sham operation group were lower.The activities and the expressions of MMP-2 (t =-45.121,P =0.000; t =-11.624,P=0.000) and MMP-9 (t=-71.849,P=0.000; t=-8.992,P=0.000) in the MCAO group were increased and upregulated significantly.Compared to the MCAO group,the activities and the expressions of MMP-2 (t =-28.792,P =0.000; t =-3.809,P =0.013) and MMP-9 (t =-53.506,P =0.000; t =-2.640,P =0.046) in the rt-PA thrombolysis group were increased and upregulated significantly.Conclusions After rt-PA intravenous thrombolytic therapy,the BBB permeability was increased.The activities and the expressions of MMP-2 and MMP-9 were increased and upregulated.MMP-2 and MMP-9 might participate in the increased BBB permeability,and thus inducing hemorrhagic transformation after rt-PA intravenous thrombolytic therapy in rats with cerebral ischemia.

Objective To prepare anti-folic acid (FA) polyclonal antibody and develop a new non-balanced competing chemiluminescence analysis for clinical detection of FA.Methods Established the detection method by added FITC-FA-analogs and FAHRP-antibody in the light emitting plate,which coated with anti-FITC antibody,to form the immune response complex of FITC/antibody-FITC-FA-analogs/FA-antibody-HRP.Then methodology evaluation was performed to evaluate the method performance;and further compared the detecting results with non-FITC system detection system and Electrochemiluminescence system (Roche Elecsys 2010).Results The ELISA results showed that the prepared anti-FA antibodies can recognize serum FA specificly.The methodology evaluation indicated that the linear correlation coefficient of the standard curve was 0.990 0;the analytical sensitivity was 1.21 ng/mL;the range of linear detection was 1.21～ 38.80 ng/mL;The coefficient variability of intra-assay was ＜5 ％,which was better than the results of non-FITC detection system;and the correlation coefficient was 0.908 1 compared with the Elecsys-2010 detection system.Conclusion The established chemiluminescence immunoassay for human serum FA has a good sensitivity and specificity,and suitable for clinical serum FA quantitativedetecting.

Objective To detect the expression of BRIT1 in cervical cancer tissues and cervical noncancer tissues ,and to analyze the differences between the two tissues .Methods The expression of BRIT1 mRNA and protein in cervical cancer tissues and the paired cervical noncancer tissues was evaluated by RT‐PCR and immmunohistochemistry .Its correlation with the clinicopathological parameters including age ,tumor types ,size ,tumor pathological grade and clinical stage was analyzed .Results RT‐PCR results showed that the BRIT1 mRNA level in cervical cancer tissues was significantly lower than that in the paired cervical noncancer tis‐sues ,the difference was statistically significant (P<0 .05) .The immmunohistochemistry results showed that the BRIT 1 protein ex‐pression level in 44 cases of 63 (69 .8% ) samples wa slower than that in the paired cervical noncancer tissues ,the difference was statistically significant(P<0 .05);In high pathological grades and high clinical stages ,the decrease of BRIT1 protein expression was more significant .Conclusion The difference of the BRIT1 expression between the cervical cancer tissues and cervical noncancer tis‐sues suggests that BRIT1 may play a certain role in the occurrence and development of cervical cancer .

OBJECTIVETo prepare antibodies (pAbs) against phosphorylated Y-box binding protein 1 (pYB-1), perform qualitative detection of the ascites/pYB-1 ratio in patients with hepatocellular carcinoma with pulmonary metastasis (HCC-PM), and assess the clinical significance of the ascites/pYB-1 ratio as a diagnostic biomarker for HCC-PM.

METHODSBioinformatic prediction and chemical synthesis was used to identify and generate the YB-1 polypeptide with phosphorylation at serine position 102 (KYLRSVGDG). Rabbits were immunized with the YB-1 polypeptide coupled to a carrier protein. Protein A affinity chromatography was used to prepare highly-purified pAbs.ELISA and SDS-PAGE were used to determine concentration and purity of the pAbs. A total of 109 ascites specimens were collected from patients (36 cases of HCC,44 cases HCC-PM, and 29 cases of liver cirrhosis) and concentrated to obtain the pYB-1. Western blotting was used to qualitatively detect pYB-1 in ascites. Regression analysis and receiver operating characteristic (ROC) curve analysis were used to assess the qualitative data.

RESULTSThe prepared pAbs had a concentration of more than or equal to 1:1 * 106 and high purity. The pAbs/YB-1S102 specifically recognized endogenous pYB-1S102. The pYB-1S102 detected in ascites specimens from patients with HCC and HCC-PM, and the positive rate of detection was 30.6% and 77.3% respectively (P < 0.01).The pYB-1S102 showed sensitivity of 77.3% and a accuracy rate of 73.8% for diagnosis of HCC-PM.

CONCLUSIONDetection of pYB-1S102 in ascites could be a useful biomarker for diagnosis and metastasis monitoring in patients with HCC.

Antibodies ; Biomarkers, Tumor ; Blotting, Western ; Carcinoma, Hepatocellular ; Humans ; Liver Cirrhosis ; Liver Neoplasms ; Neoplasm Metastasis ; Phosphorylation ; ROC Curve

Objective To construct microRNA (miRNA) -mediated gene regulation of ovarian cancer by establishing regulatory networks of miRNA and long non-coding RNA (lncRNA) database based on bioinformatic analysis.Methods Ovarian cancer-related regulatory miRNAs were obtained via miRwalk 2.0and HMDD v2.0.According to the frequency of miRwalk 2.0software, we screened and identified the corresponding target gene expression.Upon a matching analysis, ovarian cancer matched lncRNAs searched from the lncRNA database were performed to determine miRNAs interacted with lncRNA, and starBase v2.0was used to comprehensively analyze the obtained miRNA and lncRNA.Finally, miRNA-mediated gene regulatory networks of ovarian cancer were mapped by the Cytoscape software.Results MiRNA-mediated gene regulation of ovarian cancer was successfully established regulatory networks of miRNA and lncRNA databases.Hsa-let-7a, hsamir-21, hsa-miR-16, hsa-mir-17, hsa-mir-19b, hsa-mir-29aand hsa-mir-92awere high frequency miRNA of ovarian cancer.According to matching analysis, lncRNA gene symbol X inactive-specific transcript (XIST) played the central role in lncRNA-miRNA interaction in regulation of ovarian cancer.Conclusion The miR-NA-mediated gene regulating network of ovarian cancer was successfully established according to bioinformatic analysis.Databases like miRwalk, HMDD, starBase could be used as effective tools to analyze relationship between miRNA and disease.