Objective To investigate the spatio-temporal expression of connexin 43(Cx43) in the cultured ventricular myocardial cells of neonatal rats. Methods The techniques of Immunocytochemistry(ICC) and immuno-electron microscopy were used to detect the Cx43 expression in the cultured rat ventricular myocardial cells on the(2nd),(4th),(8th),(10th),(12th),(16th),(20th,)(26th) and(30th) days. Results Cx43 expression was detected in the cultured ventricular myocardial cells on the 2(nd) day,and the Cx43 granules were located largely in the cellular cytoplasm and membrane.The punctiform granule of the cellular cytoplasm decreased and the expression of Cx43 was located mainly in cellular membrane junction on the 4(th) day.The expression of Cx43 increased in cellular membrane junction on the 10(th) day,and the morphology of Cx43 expression was chain-and strip-like.There were not obvious changes in the following days.The expression of Cx43 on the 30(th) day was derangement.Conclusion The spatio-temporal expression of Cx43 in the cultured ventricular myocardial cells of neonatal rats changed with the cultural time in terms of location and quantity.It was in accordance with the growth and development of the cultured ventricular myocardial cells.

Objective To observe the elimination of oxygen free radicals in rat myocardium mitochondria by diazoxide(DE) during different duration of hypothermic preservation.Methods The Langendorff model of isolated rat heart was used.After stored in 4℃ Celsior solution or Celsior solution containing 30 ?mol/L of DE for different duration(0,3,6,9 or 12h),the activity of the superoxide dismutase(SOD) and the content of maleic dialdehyde(MDA) in heart mitochondrial were measured.The myocardial mitochondria ultrastructure was also evaluated.Results(1) After hypothermic preservation,the activity of SOD was decreased,the content of MDA was increased.The ultrastructure of myocardium mitochondria was also destrored in preserved rat hearts in a time-dependent manner.(2) Compared with the corresponding Celsior solution hypothermic preservation groups,DE significantly improved the above indexes(SOD,MDA and myocardial mitochondria ultrastructure).Conclusion Diazoxide can protect rat myocardial mitochondria from long-term hypothermic preservation injury by direct elimination of oxygen free radials and enhancement of the activity of SOD.

ObjectiveTo investigate the establishment of stable mice orthotopic left lung transplant model MethodsForty male Balb/c mice randomly served as recipients and donors.Each couple had the similar weight. Three-cuff technique was used to establish mice orthotopic lung transplant model.The micro CT of chest and the artery blood analysis were detected at first month after transplantation.ResultsThe achievement ratio was 95％.More than 90％ of the recipients survived after surgery.Cold ischemia time was (35.6±5.9) min,warm ischemia time was (25.3±7.2) min,donor lung back up time was (21.0±5.6) min,and the whole surgery time was (85±15)min.The long-term survival time achieved more than 30 days with functional lung graft.Micro CT showed clear left lung graft field one month after surgery,and left bronchus cuff was still open.The PaO2 of the artery blood was ( 106.9±5.8 ) mmHg before clipping right lung hlium,and after clipping that was (105.0±8.7) mmHg with the difference being not significant between them (P＞0.05).Histologically,the lung graft appeared very similar to the right lung one month after surgery.The pulmonary alveoli were aerated well.ConclusionBased on our experience of the orthotopic lung transplantation model in mice,through practicing,we independently and successfully established orthotopic lung transplantation model in mice.Compared to the method established by the other country,our method is easier to perform and more stable.This will benefit the basic research related to lung transplantation.

It was recently shown that several new synthetic 2-alkylsulfanyl-6-benzyl-3, 4-dihydropyrimidin-4(3H)-one (S-DABO) derivatives demonstrated anti-HIV-1 activity. Three of the derivatives namely RZK-4, RZK-5 and RZK-6 were used in this study to explore their inhibitory effects on a variety of HIV strains. These compounds at a concentration of 200 microg mL(-1) almost completely inhibited the activity of recombinant HIV-1 reverse transcriptase. All of the three compounds reduced replication of HIV-1 laboratory-derived strains, low-passage clinical isolated strain, and the drug resistant strain. In particular RZK-6 showed potent activity against the HIV-1 drug resistant strain. In general, the antiviral activities are similar in magnitude to nevirapine (NVP), which is a non-nucleoside reverse transcriptase inhibitor approved by FDA. The therapeutic indexes of these compounds were remarkable, ranging from 3704 to 38462 indicating extremely low cytotoxicity. These results suggest that the three S-DABO derivatives in this study have good potential for further development in anti-HIV-1 therapy. It may be particularly useful to target at the non-nucleoside reverse transcriptase inhibitors resistant HIV-1 strain.

Objective To investigate the biocompatibility between the atelocollagen scaffold and 3D-cultured myocardial cells,and to find out the condition and an optimal biomaterial scaffold that can be applied to 3D-culture myocardial tissue. Methods The primary cultured myocardial cells were purified and then inoculated into the atelocollagen scaffold.The cells in the atelocollagen scaffold were observed by light microscope,scanning electron microscope(SEM) and transmission electron microscope(TEM) at different times(8d,16d,20d). Results On the first day,cardiac cells pulsating together with the atelocollagen scaffold could be detected under the microscope,which were pulsating complex.A plenty of cells in the atelocollagen mesh were observed under the light microscope,and the cells coalesced with the scaffold.The cells were compacted to the scaffold and coalesced with it at three stages under TEM.Those cells were sticked to the atelocollagen scaffold and expanded sufficiently under SEM.On the atelocollagen scaffold surface,these cells coalesced with lamellar.Conclusion The biocompatibility of the atelocollagen scaffold is better for cultured cardiac myocyte.It can be used as a natural material for 3D-cultured cells,and is suitable for 3D-cultured cardiac cells and cardiac tissues.

Objective To investigate the clinical value of Hangzhou Criteria in selection of patients with hepatocellular carcinoma(HCC)for living donor liver transplantation(LDLT)and their effects on recipient's post-transplant survival.Methods A retrospective analysis was conducted on 43 patients with HCC who underwent LDLT in our center.All of the recipients were classified into subgroups according to Milan Criteria,Up-to-Seven Criteria and Hangzhou Criteria,and post-transplant total survival rate and tumor-free survival rate were compared among the criteria.Results The number of patients within Hangzhou Criteria was 61.5 %(8/13)more than that within Milan Criteria and 23.5 %(4/17)more than that within Up-to-Seven Criteria.The 1-,and 3-years total survival rate and tumor-free survival rate of the recipients within Milan Criteria(n=13)were 100 %,80.0 % and 84.6 %,84.6 %,those of the recipients within Up-to-Seven Criteria(n=17)were 100 %,75.2 % and 87.5 %,81.2 %,and those of the recipients within Hangzhou Criteria(n=21)were 100 %,80.0 % and 89.5 %,84.2 %,respectively.There was no significant difference among the three criteria in total survival rate and tumor-free survival rate(P>0.05).Conclusion Hangzhou Criteria further enrolled more liver transplantation candidates without decline of total survival rate and tumor-free survival.It is a more effective selection of HCC recipients for LDLT.

Objective:To explore the value of aspartate aminotransferase(AST)and platelet (PLT)ratio index(APRI)in the prognosis of liver transplantation(LT)for hepatocellular carcinoma and establish a nomogram model for evaluating its clinical application potential.Methods:From January 2015 to December 2019, retrospective review was conducted for clinical data of LT for hepatocellular carcinoma(HCC)at First Affiliated Hospital of Zhejiang University School of Medicine and Shulan(Hangzhou)Hospital(601 cases). They were randomized into two groups of modeling (399 cases)and validation(202 cases)and then divided into low and high APRI groups according to the APRI value at Month 1 post-transplantation. The independent risk factors of recurrence and prognosis post-LT were screened in modeling group using univariate and multivariate Cox regression analyses and were further used for constructing a nomogram prediction model. The receiver operating characteristic curve(ROC)and survival curve were utilized for verifying the accuracy of nomogram prediction model.Results:Univariate and multivariate Cox regression analyses revealed that independent risk factors for the prognosis of HCC-LT included cold ischemic time(CIT) >8 h, beyond Hangzhou criteria, surgical bleeding volume >1 000 ml and APRI >1.5. The AUC of HCC-LT recurrence prediction model was 0.734(95%CI: 0.681~0.787)and 0.749(95%CI: 0.671~0.817)in modeling and validation groups; the AUC of HCC-LT mortality prediction model was 0.735(95%CI: 0.679~0.790)and 0.758(95%CI: 0.682~0.834)in modeling and validation groups.Conclusions:APRI>1.5 is an independent risk factor for postoperative recurrence and mortality after HCC-LT. The nomogram prediction model based upon CIT, Hangzhou criteria, intraoperative bleeding volume and APRI can effectively predict the recurrence and overall survival of LT for HCC.

Objective:To observe the multimodal image features of inflammatory lesions and choroidal neovascularization (CNV) in multifocal choroiditis (MFC).Methods:A retrospective clinical analysis. A total of 90 eyes of 46 patients with MFC diagnosed in the Department of Ophthalmology of Yunnan University Affiliated Hospital from May 2017 to April 2021 were included in the study. Among them, there were 21 males and 25 females; the average age was 38.30±8.97 years old. Twenty-nine cases of MFC were diagnosed in the past, and they visited the doctor again due to new symptoms; 17 cases without a clear past medical history were the first visits. All eyes underwent color fundus photography, fluorescein fundus angiography (FFA), optical coherence tomography (OCT), and OCT angiography (OCTA). With reference to the literature and the results of multimodal fundus imaging examinations, MFC lesions were divided into active CNV lesions, inactive CNV lesions, active inflammatory lesions, and inactive inflammatory lesions, with 31 (34.4%, 31/90), 12 (13.3%, 12/90), 26 (28.9%, 26/90), 90 (100.0%, 90/90) eyes. Nineteen eyes were treated with anti-vascular endothelial growth factor drugs. To summarize and analyze the manifestations of inflammatory lesions and CNV lesions in different imaging examinations. The Wilcoxon rank test was used to compare the detection rate of CNV lesions between FFA and OCTA.Results:In eyes with active inflammatory lesions and active CNV lesions, yellow-white lesions, retinal hemorrhage and exudation were seen on fundus color photography; FFA examination showed fluorescein leakage in the lesions; OCT examination showed retinal pigment epithelium (RPE) layer in the lesions was uplifted, the boundary was unclear, combined with subretinal and intraretinal fluid; OCTA examination showed that there was no blood flow signal in each layer of vascular tissue in active inflammatory lesions, and blood flow signals were seen in active CNV lesions. In the eyes of inactive inflammatory lesions and inactive CNV lesions, the fundus color photography showed that the lesions had clear boundaries without bleeding or exudation; FFA examination, the lesions were fluorescently stained, and there was no fluorescein leakage; OCT examination, inactive CNV lesions manifested as raised lesions with clear boundaries, and inactive inflammation manifested as scars formed by mild RPE hyperplasia or depressions in outer structures formed by atrophy; OCTA examination, inactive inflammatory lesions showed patchy loss of blood flow signal or penetrating blood flow signal below, blood flow signal can be seen in inactive CNV lesions.Conclusion:MFC active inflammatory lesions and active CNV lesions are often accompanied by retinal hemorrhage and exudation; FFA shows fluorescein leakage; OCT shows that the boundary of raised lesions is unclear; OCTA can identify the nature of CNV or inflammatory lesions.

Objective:To observe the clinical evolution process and imaging characteristics of choroidal lesions in different subtypes of serpiginous choroiditis (SC), and to explore the clinical significance of subtype classification.Methods:A retrospective, uncontrolled and observational study. A total of 45 eyes of 25 SC patients diagnosed in Yunnan Eye Hospital from May 2009 to September 2021 were included in the study. According to the initial location of the lesion and fundus images, including fundus color photography, fundus fluorescein angiography (FFA), optical coherence tomography (OCT) and other examination results. SC was divided into peripapillary serpiginous choroiditis, macular serpiginous choroiditis and ampiginous choroiditis. According to the shape of the lesions at the first diagnosis, it can be divided into new lesions with only infiltrating edema, old lesions with only atrophy and recurrent lesions with coexistence of edema and atrophy. the imaging features, development and complications of different subtypes of ocular lesion were observed.Results:Among the 45 eyes of 25 cases, 15 cases were male and 10 cases were female, 20 cases of binocular and 5 cases of monocular, age was 42.3±5.7 years old. There were 21 eyes with active lesions, of which 5 eyes were new lesions and 16 eyes with recurrent lesions; 24 eyes were old lesions. Concurrent optic disc edema occurred in 3 eyes; mild vitreitis occurred in 5 eyes; retinal occurred vasculitis in 3 eyes; choroidal neovascularization occurred in 3 eyes. Among the 16 cases (64%, 16/25) of the peripapillary serpiginous choroiditis, 2 cases (2 eyes) were monocular, and 14 cases (28 eyes) were binocular. Active lesions were found in 16 eyes, of which patients with binocular lesions only one had active lesions. The choroidal lesions that were close to the optic disc or around the optic disc, expanded outwards centrifugally with the prolongation of the disease course, and can progress to the macula. The edge of the lesion was tortuous, with a geographic-like, amoeboid-like and finger-like, polypoid or propeller-like shape. Active lesions in FFA showed weak fluorescence in the early stage and strong fluorescence in the late stage; the old lesions showed weak fluorescence in the early stage and mottled fluorescence in the late stage, and mostly strong fluorescence on the edge. OCT showed thickening of active lesions and thinning of old lesions. Among the 4 cases (16.0%, 4/25) of macular type, 2 cases (2 monocular eyes) had active lesions; 2 cases (4 eyes) had lesion in both eyes, among them, 1 case (2 eyes) had old lesion, and the other case had alternate active lesions. The initial lesions were all located in the off-center of the macula, and most of them were disk-shaped and progressing centrifugally to the periphery. The FFA and OCT imaging findings of the lesions were similar to those of the peridisc type. Among the 5 cases (20.0%, 5/25) of ampiginous choroiditis, 1 case (1 eye) was monocular and 4 cases (8 eyes) were binocular. These lesions were multiple old lesions of varying sizes, gray-white with pigmentation, with clear borders in the posterior pole. Among them 4 eyes have new active lesions appeared near the old lesions. The old lesions showed weak fluorescence with clear borders, and the fluorescein leakage at the late edge formed a strong fluorescence ring; the active lesions showed weak fluorescent spots with blurred edges, and the fluorescence was slightly enhanced in the late stage. In old lesions, atrophy of the photoreceptor layer, RPE and choroid can be seen, and RPE hyperplasia in some areas.Conclusions:SC subtype is a classification of the location of the first lesion, but the characteristics of the repeated attack of this disease can lead to the annihilation of each subtype due to the continuous expansion of the lesion. The phenomenon that the fundus active lesions only occur in one eye that can explain the clinical manifestations of asymmetric morphology of binocular lesions. The characteristics of binocular subtype warn that the predilection site of the healthy eye should be paid attention to.

AIM: To assess the protective effect of adipose tissue-derived mesenchymal stem cells(ADSCs)exosomes on injured retinal ganglion cells(RGCs)by establishing an in vitro rat RGC pressure injury model.METHODS: ADSCs were cultured, and exosomes were extracted from the supernatant and identified. Rat RGCs were divided into a control group, pressure model groups(40, 80, 120 mmHg), and exosome-treated groups under different pressures. Cell proliferation activity was assessed using the CCK-8 assay. The mRNA expression levels of brain-derived neurotrophic factor(BDNF)and Caspase-3 in RGCs were detected by qPCR, and protein levels were measured by Western Blot.RESULTS: The CCK-8 assay showed that cell proliferation activity in the control group increased significantly at 48 h compared to 24 h(P<0.05). At 48 h, cell viability in the exosome-treated groups increased significantly compared to the 40, 80, and 120 mmHg pressure model groups(all P<0.05). qPCR results indicated that BDNF mRNA expression decreased in the 40 mmHg pressure model group without statistical significance(P>0.05), and significantly decreased in the 80 and 120 mmHg pressure model groups(all P<0.05). BDNF mRNA expression significantly increased in the 40 and 80 mmHg pressure model groups after exosome treatment(both P<0.05), and increased in the 120 mmHg pressure model group without statistical significance(P>0.05). Caspase-3 mRNA expression increased in the 40 mmHg pressure model group without statistical significance(P>0.05), and significantly increased in the 80 and 120 mmHg pressure model groups(all P<0.05). Caspase-3 mRNA expression significantly decreased in the 40 and 80 mmHg pressure model groups after exosome treatment(P<0.05), and decreased in the 120 mmHg pressure model group without statistical significance(P>0.05). Western Blot analysis showed that BDNF protein expression decreased in the 40 mmHg pressure model group without statistical significance(P>0.05), and significantly decreased in the 80 and 120 mmHg pressure model groups(all P<0.001). After exosome treatment, BDNF protein expression significantly increased compared to the pressure model groups(all P<0.05). Caspase-3 protein expression increased significantly in all pressure model groups compared to the control group(all P<0.05), and significantly decreased in all exosome-treated groups compared to the model groups(all P<0.05).CONCLUSION: ADSCs-derived exosomes enhance cell proliferation and viability in cultured rat RGCs in vitro under different pressure-induced injuries, enhance BDNF mRNA and protein expression levels, and reduce Caspase-3 mRNA and protein expression levels, suggesting that ADSCs-derived exosomes have a protective effect on pressure-injured in cultured rat RGCs in vitro.