Objective To investigate changes in the expression of prepro-orexin and orexin receptor-1 ( OX1R) following permanent middle cerebral artery occlusion ( MCAO ) with or without preconditioning through electrical stimulation of the cerebellar fastigial nucleus (FNS). Methods Wistar rats were subjected to permanent MCAO and randomly divided into 5 groups: a sham-operated control group (PO), an FNS preconditioning + shamoperated control group (FNS-PO) , an ischemia group, an FNS preconditioning + ischemia group (FNS-PI) and a cerebellar fastigial nucleus injury + FNS preconditioning + ischemia group (FNL-FNS-PI). Each group was divided into 5 subgroups according to the time at which the animals were sacrificed after the MCAO ( 1, 3, 6, 12 and 24 h).RT-PCR was used to detect expression of OX1R mRNA, and ELISA to measure the levels of orexin-A in the hypothalamus and plasma. Results The immunoreactivity of prepro-orexin decreased significantly in the PI groups, with further decreases over time. At the 12th h after MCAO, the immunoreactivity of prepro-orexin reached a minimum.There were significant differences between the rats in the PO and FNS-PO groups. On the contrary, the immunoreactivity of OX1R increased significantly in the PI groups, with further increases continuing over time, peaking at 12 h after the MCAO. There were significant differences between the PO and FNS-PO groups. In the rats with FNS preconditioning (PI-FNS) , the decrease in prepro-orexin and the increase in OX1R were significantly inhibited compared to the PI subgroups at the 6th and 12th hour. There was no significant difference between the FNL-PIFNS group and the PI group. The expression of OX1R mRNA increased significantly in the PI group, with further increases continuing over time, peaking at 24 hours. The plasma levels of orexin-A were not significantly different among the groups, but the levels of orexin-A in the hypothalamus decreased significantly in the PI and FNL-PI-FNS groups, with further decreases continuing over time. At the 12th h after the MCAO the levels were significantly different compared with the PO and PO-FNS groups. While in the rats with FNS preconditioning (PI-FNS) , the decrease in orexin-A level was reversed and there was no significant difference compared with PO and PO-FNS groups. Conclusions The orexinergic system is altered following cerebral ischaemia. FNS preconditioning may be able to regulate these changes.

Objective To investigate the characteristics of coronary vessel re-endothelialization after placement of drug-eluting stents (DES), and to provide clinical evidence for the double anti-platelet treatment. Methods Optical coherence tomography (OCT) was performed in 43 patients in 1 year after DES implantation. Characteristics of re-endothelialization and percentage of neointimal coverage of stent struts were evaluated by OCT. Results The rate of stent struts intimal coverage was 90.70%, and the remain was lack of endothelial coverage; The ratio of neointimal thickness (NIT) between 0-99, 100-199 and above 200 microns was 19.92%, 37.55% and 42.53%, respectively. The rate of neointimal coverage was higher and the degree of neointimal hy-perplasia was more extensive in patients with DM and in patients with ACS than those of patients without DM and of patients with stable angina pectoris. Conclusion One year after stent placement, most of the stent struts were covered with neointima and few struts obtained poor coverage of endothelial. DM and ACS may be impact factors for the progress of re-endothelialization after DES placement.

Objective: To compare the percutaneous transluminal septal myocardial ablation (PTSMA) and percutaneous transluminal septal tunnel myocardial ablation (PTSTMA) on cardiac function in experimental canines.
Methods: According to CAG determined coronary septal branches, a total of 25 hybrized canines were divided into 2 groups:PTSMA group, n=13 canines with the bigger septal branches and PTSTMA group, n=12 canines with the smaller or uneven septal branches. Alcohol ablation model was established. Electrocardiograph (ECG) at before and after the operation, biomarkers for myocardial injury, echocardiography and hemodynamic changes were recorded. The animals were scariifes at 1 week after operation, the pathological changes in ventricular septal were observed by HE and Masson staining.
Results: Myocardial infarction (MI) could be induced by either PTSMA or PTSTMA and the thickness of septal was decreased. LVEDd, LVEF and hemodynamic indexes were similar between 2 groups. The alcohol volume used in operation, EKG and echocardiography ifndings were similar between 2 groups, P>0.05. Pathological staining indicated that there was a well-demarcate between the ablation focal and normal myocardium, merging area had neutrophiles invasion, infarcted cells were partially having the ghost cell sample and they were gradually replaced by ifbrous tissue. There was nest-like necrosis in ablated lumen and the normal vessel wall disappeared. PTSMA group had vessel lumen conifguration in septal branch and the necrosis limited inside the lumen;while in PTSTMA group, the vessel wall of was discontinued and some necrosis materials move out to from lumen.
Conclusion: Both PTSMA and PTSTMA were effective for alcohol septal ablation in different coronary septal branches, the impacts on cardiac function and hemodynamic changes were similar in experimental canines.

Background: To evaluate the safety and effectiveness of empagliflozin in routine clinical settings, we collected and assessed the clinical profiles of Korean patients with type 2 diabetes mellitus. Methods: This was a post-marketing surveillance study of empagliflozin 10 and 25 mg. Information on adverse events and adverse drug reactions (ADRs) was collected as safety data sets. Available effectiveness outcomes, including glycosylated hemoglobin (HbA1c) level, fasting plasma glucose, body weight, and blood pressure, were assessed. Results: The incidence rate of ADRs was 5.14% in the safety dataset (n=3,231). Pollakiuria, pruritis genital, and weight loss were the most common ADRs. ADRs of special interest accounted for only 1.18%, and there were no serious events that led to mortality or hospitalization. In the effectiveness data set (n=2,567), empagliflozin significantly reduced the mean HbA1c level and body weight during the study period by –0.68%±1.39% and –1.91±3.37 kg (both P<0.0001), respectively. In addition, shorter disease duration, absence of dyslipidemia, and higher baseline HbA1c levels were identified as the clinical features characteristic of a “responder” to empagliflozin therapy. Conclusion Empagliflozin is a safe and potent glucose-lowering drug in routine use among Korean patients with type 2 diabetes mellitus. It is expected to have better glycemic efficacy in Korean patients with poorly controlled type 2 diabetes mellitus.

Objective: To isolate a bacteriophage against pan-drug resistant Klebsiella pneumoniae in a burn patient, and to study its biological characteristics, genomic information, and effects on bacterial biofilm. Methods: (1) In 2018, pan-drug resistant Klebsiella pneumoniae UA168 (hereinafter referred to as the host bacteria) solution isolated from the blood of a burn patient in Ruijin Hospital Affiliated to Shanghai Jiao Tong University School of Medicine (hereinafter referred to as Ruijin Hospital) was used to isolate and purify the bacteriophage against pan-drug resistant Klebsiella pneumoniae from the sewage of Ruijin Hospital with sewage co-culture method, drip plate method, and double-agar plate method. The bacteriophage was named as phage KP168 and the plaque morphology was observed. (2) The phage KP168 solution was taken for cesium chloride density gradient centrifugation and dialysis, and then the morphology of phage KP168 was observed through transmission electron microscope after phosphotungstic acid negative staining. (3) The phage KP168 solution was taken to determine the lytic ability of the phage KP168 against 20 strains of pan-drug resistant Klebsiella pneumoniae isolated from the burned patients′ blood in Ruijin Hospital by the drip plate method, and then the lysis rate was calculated. (4) The phage KP168 solution at a initial titer of 9.3×10¹¹ plaque-forming unit (PFU)/mL (400 μL per tube) and the host bacteria solution at a concentration of 1×10⁹ colony-forming unit (CFU)/mL (4 mL per tube) were conventionally shaking cultured together for 4 hours at multiplicity of infection (MOI) of 10.000, 1.000, 0.100, 0.010, or 0.001, respectively (1 tube per MOI). The titer of phage KP168 was measured by the double-agar plate method (the measurement method was the same below) to select the optimal MOI. The experiment was repeated three times. (5) The host bacteria solution at a concentration of 1×10⁹ CFU/mL (4 mL per tube) and the phage KP168 solution at an adjusted titer of 5×10⁷ PFU/mL (400 μL per tube) were mixed at the MOI of 0.005. The plaques were counted 0 (immediately), 1, 2, 3, 4, 5, 15, and 30 minutes (1 tube at each time point) after mixing by the double-agar plate method (the counting method was the same below), and the percentage of adsorbed phages was calculated to screen for the optimal adsorption time. The experiment was repeated three times. (6) The host bacteria solution at a concentration of 1×10⁹ CFU/mL (300 μL per tube) and the phage KP168 solution at a titer of 5×10⁸ PFU/mL (60 μL per tube) were mixed at MOI of 0.005 and conventionally shaking cultured after standing for the optimal adsorption time. The phage KP168 titer was measured 0 (immediately), 10, 20, 30, 40, 50, 60, 70, 80, 90, and 100 minutes after culture, and a one-step growth curve was drawn. The experiment was repeated three times. (7) The phage KP168 solution at a titer of 2.5×10¹⁰ PFU/mL was left to stand for 1 hour at 37, 40, 50, 60, or 70 ℃ (3 tubes at each time point, 1 mL per tube) for counting the plaques, and then the thermal stability curve was drawn. SM buffer at a pH values of 5.0, 6.0, 7.0, 7.4, 8.0, 9.0, or 10.0 were added to the phage KP168 solution at a titer of 3.0×10¹⁰ PFU/mL, respectively. The mixed solution was left to stand for 1 hour at 37 ℃ (3 tubes of each pH, each tube containing 100 μL phage KP168 solution and 900 μL SM buffer), and then the plaques were counted, and an acid-base stability curve was drawn. (8) The phage KP168 solution was taken for DNA extraction and sequencing after dialysis as in experiment (2). The whole genome was annotated with Prokka to obtain the coding sequence of phage KP168. Nucleotide′s BLAST function was used to proceed nucleic acid sequence alignment for finding a known phage with the highest similarity to the phage KP168 nucleic acid sequence, and Blastx function was used to translate the coding sequence into protein for its function prediction. The comparison with Antibiotic Resistance Genes Database and Virulence Factors Database was proceeded. (9) In a 96-well plate, at a MOI of 1.000, 0.100, 0.010 or 0.001 (3 wells per MOI), 20 μL phage KP168 solution at a initial titer of 5.8×10¹⁰ PFU/mL was added to 200 μL host bacteria solution at a concentration of 1.5×10⁸ CFU/mL (the same concentration below) for co-cultivation for 48 hours. After 200 μL host bacteria solution was left to stand for 48 hours, 20 μL phage KP168 solution at a titer of 1×10^6, 1×10^7, 1×10^8, 1×10^9, or 1×10¹⁰ PFU/mL (3 wells per titer) was added respectively for action for 4 hours. In both experiments, 200 μL host bacteria solution added with 20 μL SM buffer (3 wells) acted as a negative control, and 220 μL LB culture medium (3 wells) acted as a blank control. Absorbance values were measured by a microplate reader, and inhibition/destruction rates of biofilm were calculated. The experiments were both repeated three times. Results: (1) The plaques of phage KP168 successfully isolated and purified were transparent and round, and its diameter was approximately 1.5 mm. (2) The phage KP168 has a regular polyhedron structure with a diameter of about 50 nm and without a tail. (3) The phage KP168 could lyse 13 of 20 strains of Klebsiella pneumoniae from burned patients, with a lysis rate of 65.0%. (4) When MOI was 1.000, the titer was the highest after co-culturing the phage KP168 with the host bacteria for 4 hours, which was the optimal MOI. (5) After the mixing of the phage KP168 with the host bacteria for 4 minutes, the percentage of the adsorbed phage reached the highest, which was the optimal adsorption time. (6) The one-step growth curve showed that during the lysis of the host bacteria by phage KP168, the incubation period was about 10 minutes, and the lysis period was about 40 minutes. (7) With the condition of 40 ℃ or pH 7.4, the number of plaques and the activity of phage KP168 reached the highest. (8) The genome of phage KP168 was a linear double-stranded DNA with a length of 40 114 bp. There were 48 possible coding sequences. It had the highest similarity to Klebsiella phage_vB_Kp1. The most similar known proteins corresponding to the translated proteins of coding sequences contained 23 hypothetical proteins and 25 proteins with known functions. No resistance genes or virulence factor genes were found. The GeneBank accession number was KT367885. (9) After 48 hours of co-cultivation of the phage KP168 and the host bacteria at each MOI, the inhibition rates of biofilm were similar, with an average of about 45%. After the phage KP168 with a titer of 1×10⁹ PFU/mL acted on the biofilm formed by the host bacteria for 4 h, the destruction rate of biofilm was the highest, reaching an average of 42%. Conclusions In this study, a bacteriophage against pan-drug resistant Klebsiella pneumoniae from a burn patient, phage KP168, is isolated from sewage, which belongs to the tailless phage. It has a wide host spectrum, short adsorption time, and short incubation period, with certain thermal and acid-base stability. Its genomic information is clear, and it does not contain resistance genes or virulence factor genes. It also has an inhibitory effect on the formation of bacterial biofilm and a destructive effect on the formed bacterial biofilm.

Objective To investigate the clinical, biochemical, pathological, disease course, and prognostic features of drug-induced liver injury (DILI) patients with different types of bile duct injury. Methods Four patients who were diagnosed with bile duct injury-type DILI by liver biopsy in Shijiazhuang Fifth Hospital, from March 2015 to October 2010 were selected, and related data were collected, including clinical data, laboratory examinations, radiological examination, and prognosis.The semi-quantitative score was determined for liver pathological morphology, and each indicator was compared between the four patients. Results Bile duct injury-type DILI was more common in female patients, and most patients tended to have a good prognosis.Clinical symptoms, liver biochemical parameters, and prognosis varied with the site, grade, scope, regeneration, and repair of bile duct injury. Conclusion Liver biopsy is still the gold standard for making a definite diagnosis of bile duct injury-type DILI, understanding the condition of lesions, and judging the prognosis of this disease.

Cabozantinib, mainly targeting cMet and vascular endothelial growth factor receptor 2, is the second-line treatment for patients with advanced hepatocellular carcinoma (HCC). However, the lower response rate and resistance limit its enduring clinical benefit. In this study, we found that cMet-low HCC cells showed primary resistance to cMet inhibitors, and the combination of cabozantinib and mammalian target of rapamycin (mTOR) inhibitor, rapamycin, exhibited a synergistic inhibitory effect on the in vitro cell proliferation and in vivo tumor growth of these cells. Mechanically, the combination of rapamycin with cabozantinib resulted in the remarkable inhibition of AKT, extracellular signal-regulated protein kinases, mTOR, and common downstream signal molecules of receptor tyrosine kinases; decreased cyclin D1 expression; and induced cell cycle arrest. Meanwhile, rapamycin enhanced the inhibitory effects of cabozantinib on the migration and tubule formation of human umbilical vascular endothelial cells and human growth factor-induced invasion of cMet inhibitor-resistant HCC cells under hypoxia condition. These effects were further validated in xenograft models. In conclusion, our findings uncover a potential combination therapy of cabozantinib and rapamycin to combat cabozantinib-resistant HCC.
Anilides/pharmacology* ; Animals ; Carcinoma, Hepatocellular/drug therapy* ; Cell Line, Tumor ; Cell Proliferation ; Endothelial Cells/metabolism* ; Humans ; Liver Neoplasms/drug therapy* ; Pyridines/pharmacology* ; Sirolimus/pharmacology* ; Xenograft Model Antitumor Assays