Objective To study the effects of D-Ala2-Leu5-enkephalin(DADLE) preconditioning on rabbit donor heart preservation and protective mechanism.Methods 24 rabbits were randomly divided into 4 groups. In group B, after preconditioning with DADLE(1mg?kg -1 ), the donor hearts were then arrested and preserved with Krebs-Henseleit buffer at 4℃ for 4h,and group A without conditioning as controls. Then the donor hearts were transplanted into the abdomen of recipient rabbits. Recovery situation of contraction of the donor heart was compared. The left ventricular tissues were obtained from the donor hearts after 2h of transplantation.The contents of free radicals and ATPase activity were determined and cardiac ultrastructure were observed.Results After 4h cold storage, the heart undergoing DADLE preconditioning in group B showed better recovery of left ventricular development pressure (LVDP).The activity of sodium-potassium ATPase in group B were higher than that in group A. DADLE increased free radical production 2-fold versus group A. Group B showed slighter injury in transmission electron microscopy observation than that in group A.Conclusion Our study demonstrated opioid preconditioning has protective function to rabbit heart injury caused by long term cold storage. The protective mechanism maybe related to increase of free radical content.

A method was established for identifying the chemical components of a traditional Chinese medicinal formula Baihe Zhimu Tang and its single herbs by combining high performance liquid chromatography and electrospray ionization mass spectrometry(HPLC-ESI-MS). The molecular ions of compounds in both negative and positive modes were observed for molecule mass information, and the potential structures were identified by attentive studying on the mass spectra of compounds and comparing with Reference data and some of standards. The results show that in MS detection, saponins in Baihe Zhimu Tang and its single herbs are easily to become positive ions in the electrospray ionization procedure, and they have strong responses, but the mass spectrometric signals of flavonoids and phenolic glucosides are week. 38 compounds in Baihe Zhimu Tang including 3 flavonoids, 4 phenolic glucosides and 31 saponins were identified through analyzing and comparing the total ion chromatograms(TIC) and mass spectra of Baihe Zhimu Tang and its single herbs. This method has the advantages of simple operation, rapid measurement and it is a powerful tool for identification of chemical components in Baihe Zhimu Tang.

Objective To investigate whether recombinant human endostatin can create a time window of vascular normalization prior to vascular pruning to alleviate hypoxia in Lewis lung carcinoma in mice. Methods Kinetic changes in morphology of tumor vasculature in response to recombinant human endostatin were detected under a confocal microscope with immunofluorescent staining in Lewis lung carcinomas in mice. The hypoxic cell fraction of different time was assessed with immunohistochemical staining . Effects on tumor growth were monitored as indicated in the growth curve of tumors . Results Compared with the control group vascularity of the tumors was reduced over time by recombinant human endostatin treatment and significantly regressed for 9 days. During the treatment, pericyte coverage increased at day 3, increased markedly at day 5, and fell again at day 7. The vascular basement membrane was thin and closely associated with endothelial cells after recombinant human endostatin treatment, but appeared thickened, loosely associated with endothelial cells in control tumors. The decrease in hypoxic cell fraction at day 5 after treatment was also found. Tumor growth was not accelerated 5 days after recombinant human endostatin treatment. Conclusions Recombinant human endostatin can normalize tumor vasculature within day 3 to 7, leading to improved tumor oxygenation. The results provide important experimental basis for combining recombinant human endostatin with radiation therapy in human tumors.

Objective To explore the clinical efficacy of laparoscopic left hemihepatectomy for the treatment of intrahepatic bile duct stones.Methods The clinical data of 30 patients with left intrahepatic bile duct stones who were admitted to the Second Affiliated Hospital of Nancbang University from June 2013 to June 2014 were retrospectively analyzed.All the patients underwent laparoscopic left hemihepatectomy by the Glisson intra-and extra-pedicles vascular inflow occlusion techniques together with the removal of choledocholithiasis and right bile duct stones,and T tube placement or laparoscopic primary suture of common bile duct were selected according to the condition of bile duct.All the 30 patients were readmitted to hospital and detected by color Doppler ultrasound (CDUS),computed tomography (CT) and T tube cholangiography at postoperative month 1,and then received CDUS reexamination every 3 months.CT and MRI reexaminations were applied to patients with complication of residual stones if necessary.All the patients were followed up till July 2014.Results All the 30 patients were treated by laparoscopic hepatectomy with left hemihepatic vascular inflow occlusion,including 5 with conversion to open surgery and 25 with successful operation.The Glisson extra-and intra-pedicel vascular inflow occlusion techniques were used in 11 and 14 patients,respectively.The operation time and volume of blood loss were (158 ± 85) minutes and (405 ± 215) mL.Two patients received intraoperative blood transfusion.There were no residual stones in the 8 patients with choledocholithiasis by intraoperative choledochoscope,and primary suture of bile duct and T tube placement were done in 5 and 3 patients,respectively.No patients died.After operation,there were 2 patients with bile leakage and 1 with pleural effusion,and they were cured though drainage.One patient with subphrenic effusion was cured by B ultrasound-guided puncture and drainage.One patient had bleeding with the volume of blood loss of 500 mL,and was cured by conservative treatment.The duration of hospital stay in all the patients was (8.5 ± 2.3)days.No bile leakage and abdomen infection were detected by outpatient examination.The time of followup was 1-12 months,without recurrence of stones.Conclusion Laparoscopic left hemihepatectomy for the treatment of left intrabepatic bile duct stones is safe and feasible with satisfactory outcome.

Objective At present, there is no study on effect of levetiracetam(LEV) on the gray matter structure remodeling in benign epilepsy children with central temporal spikes(BECTS).The purpose of this study was to study the influence of LEV on the gray matter structure in BECTS and to evaluate the mechanism of LEV on the brain structure of BECTS through using voxel-based MRI morphological(VBM) methods.Methods From January 2014 to September 2016, twenty-four BECTS treated with LEV(LEV group), twenty-four drug-na?ve BECTS(untreated group) and twenty-four normal children(normal group) consulted in department of Neurology, Nanjing Children′s Hospital and the Nanjing Military Region, Nanjing General Hospital were continuously included to receive three-dimensional T1-weighted imaging with 3T MRI and the gray matter volume was calculated by VBM.We compared the difference of grey matter volumes of the three groups and analyzed their correlation with epilepsy duration, age of onset and medication time and other clinical index.Results Compared with the normal group, the grey matter volume of bilateral thalamus were decreased, and the volume of bilateral Rolandic areas, anterior insula/frontal operculum/frontal triangle, left supplementary motor area, paracentral lobule, precentral gyrus, superior frontal gyrus and right middle frontal gyrus were increased in the untreated group, but the grey matter volume of the bilateral Rolandic areas, frontal operculum and left supplementary motor area were decreased in the LEV group.Compared with the untreated group, the grey matter volume of bilateral supplementary motor, left paracentral lobule, precentral gyrus, bilateral anterior insula/frontal operculum/frontal triangle, left superior frontal gyrus and right middle frontal gyrus in the LEV group were decreased.The grey matter volume of left anterior insula/frontal operculum areas was negatively correlated with the medication time in LEV group(r=-0.527, P<0.01).Conclusion T The mainly representations of BECTS are thalamic gray matter damage and epileptic-related cortical area irritation structural abnormalities, but the LEV could reshape the epilepsy-related cortical area and the gray matter in the brain area associated with clinical symptoms.

Objective To investigate the diagnostic value of calcification and cystic lesion of CT findings in differentiating pancreatic head ductal carcinoma (PHDA)from mass-forming chronic pancreatitis (MFCP)of the pancreatic head.Methods The clinic data and CT findings of 30 cases with PHDA and 24 cases with MFCP of the pancreatic head,which were confirmed by surgery and pathology were analyzed retrospectively.The images were reviewed independently by two expert radiologists with a double-blind method.An independent sample t test and chi-square test were used to compare the data of imaging findings between two groups.Results ① Calcification was found in 14 cases (58.33%)with MFCP and in 3 cases (10%)with PHDA (P＜0.001).The percentage of patchy,punctate and mixed calcification were 28.57% (n=4),14.29% (n=2)and 57.14% (n=8)in MFCP,0% (n=0),66.67% (n=2)and 33.33% (n=1) in PHDA,respectively.② Necrotic cyst was founded in 7 cases (29.17%)with MFCP and 18 cases (60%)with PHDA(P＜0.05). Pseudocysts were demonstrated in 14 cases (58.33%)with MFCP and in 3 cases (10%)with PHDA (P＜0.001).Honeycombed change with tension within or around the lesion were demonstrated only in patients with MFCP.In addition,normal tissue of the pancreas was found within the lesion in 11 cases (45.83%)of MFCP and none in PHDA,which showed significant difference between two groups.Conclusion Mixed calcification and honeycomb with tension of CT findings are of significant value in differentiating PHDA from MFCP of the pancreatic head.

Objective To investigate the expression of human leucocyte antigen G (HLA-G) in urothelial carcinoma after renal transplantation,and to analyse the relationship between HLA-G expression and the various clinical and pathological parameters.Methods 29 patients with urothelium carcinoma after renal transplantation for the first time from January 2005 to June 2016 were selected as the experimental group,the age range was 32-70 years,with an average of (55.5 ± 8.1) years.29 non-transplanted patients with urothelial carcinoma as the control group 1,the age range was 36-74 years,with an average of (57.9 ± 8.2) years.15 cases of normal urinary tract epithelial were from cystoscopy biopsy as the control group 2.Immunohistochemical method was used to detect the difference of HLA-G expression between the three groups.The clinical and pathological data of patients with urothelial carcinoma after renal transplantation were analyzed.Results The expression rate of HLA-G was 79.3％ (23/32) in patients with urothelial carcinoma after renal transplantation,37.9％ (11/32) in non-transplanted group and 0 (0/15) in normal urinary tract epithelium group.The expression rate of HLA-G in non-transplanted group was significantly higher than that in normal urinary tract epithelium group (P ＜ 0.05).The expression rate of HLA-G in patients with urothelial carcinoma after renal transplantation was significantly higher than that in nontransplanted group and normal urinary tract epithelium group (P ＜ 0.05).Conclusions HLA-G is associated with the occurrence of urothelial carcinoma after renal transplantation.It may provide a new idea for the prevention and treatment of urinary tract epithelium after renal transplantation.

Objective:To explore the changes of brain activity in drug-resistant or drug-controlled medial temporal lobe epilepsy patients by the method of functional connectivity density (FCD), and to analyze their correlation with the course of the disease.Methods:According to the definition of drug-resistant epilepsy by the International League Against Epilepsy in 2010, 146 patients with medial temporal lobe epilepsy who were clearly diagnosed as unilateral hippocampal sclerosis in Jinling Hospital, Nanjing University School of Medicine from July 2009 to February 2019 were divided into drug control group ( n=73) and drug-resistant group ( n=73). The 3.0 T resting state functional magnetic resonance scan was performed on all subjects to compare the difference in FCD between the two groups, and calculate the correlation between the FCD value of the brain area and the course of the disease between the two groups of patients. Results:There was significant difference between the two groups in FCD. Compared with the drug control group, the drug-resistant group had significantly lower FCD values in the insula, lenticular nucleus, thalamus, hippocampus and precentral gyrus on the side of the epileptogenic focus. The FCD value of the precuneus on the side of the epileptogenic focus in the drug-resistant group was negatively correlated with the duration ( r=-0.30, P=0.01). Conclusions:The FCD of patients with drug-resistant medial temporal lobe epilepsy was lower than that of the drug control group. In addition, there may be progressive damage to the brain. The difference is helpful for exploring the pathophysiological mechanisms related to drug resistance in patients with medial temporal lobe epilepsy, and finding reliable neuroimaging markers related to drug resistance.

To explore the effect of Mlk3 (mixed lineage kinase 3) deficiency on blood pressure, Mlk3 gene knockout (Mlk3KO) mice were generated. Activities of sgRNAs targeted Mlk3 gene were evaluated by T7 endonuclease I (T7E1) assay. CRISPR/Cas9 mRNA and sgRNA were obtained by in vitro transcription, microinjected into zygote, followed by transferring into a foster mother. Genotyping and DNA sequencing confirmed the deletion of Mlk3 gene. Real- time PCR (RT-PCR), Western blotting or immunofluorescence analysis showed that Mlk3KO mice had an undetectable expression of Mlk3 mRNA or Mlk3 protein. Mlk3KO mice exhibited an elevated systolic blood pressure compared with wild-type mice as measured by tail-cuff system. Immunohistochemistry and Western blotting analysis showed that the phosphorylation of MLC (myosin light chain) was significantly increased in aorta isolated from Mlk3KO mice. Together, Mlk3KO mice was successfully generated by CRISPR/Cas9 system. MLK3 functions in maintaining blood pressure homeostasis by regulating MLC phosphorylation. This study provides an animal model for exploring the mechanism by which Mlk3 protects against the development of hypertension and hypertensive cardiovascular remodeling.
Animals ; Mice ; Mice, Knockout ; CRISPR-Cas Systems ; Blood Pressure ; Gene Knockout Techniques ; Zygote

Bladder cancer is the most common malignancy of the urinary system. Compound Kushen Injection (CKI) is a Chinese medicinal preparation that has been widely used in the treatment of various types of cancers in the past two decades. However, the pharmacological effect of CKI on bladder cancer is not still completely understood. In the current study, network pharmacology combined with bioinformatics was used to elucidate the therapeutic mechanism and potential targets of CKI in bladder cancer. The mechanism by which CKI was effective against bladder cancer was further verified in vitro using human bladder cancer cell line T24. Network pharmacology analysis identified 35 active compounds and 268 target genes of CKI. Bioinformatics data indicated 5500 differentially expressed genes associated with bladder cancer. Common genes of CKI and bladder cancer suggested that CKI exerted anti-bladder cancer effects by regulating genes such as MMP-9, JUN, EGFR, and ERK1. Functional enrichment analysis indicated that CKI exerted therapeutic effects on bladder cancer by regulating certain biological processes, including cell proliferation, cell migration, and cell apoptosis. In addition, Kyoto Encyclopedia of Genes and Genomes enrichment analysis implicated pathways related to cancer, bladder cancer, and the PI3K-Akt signaling pathway. Consistently, cell experiments indicated that CKI inhibited the proliferation and migration of T24 cells, and induced their apoptosis. Moreover, RT-qPCR and Western blot results demonstrated that CKI was likely to treat bladder cancer by down-regulating the gene and protein expression of MMP-9, JUN, EGFR, and ERK1. CKI inhibited the proliferation and migration, and induced the apoptosis of T24 bladder cancer cells through multiple biological pathways and targets. CKI also exhibited significant effects on the regulation of key genes and proteins associated with bladder cancer. Overall, our findings provide solid evidence and deepen current understanding of the therapeutic effects of CKI for bladder cancer, and further support its clinical use.
Computational Biology ; Drugs, Chinese Herbal ; Humans ; Network Pharmacology ; Phosphatidylinositol 3-Kinases ; Urinary Bladder Neoplasms/genetics*