Objective To observe the influence of down-regulation of HtrA1 expression by small interfering RNA on light-injured human retinal pigment epithelium (RPE) cells.Methods Cultured human RPE cells(8th-12th generations)were exposed to the blue light at the intensity of (2000 ± 500) Lux for 6 hours to establish the light injured model.Light injured cells were divided into HtrA1 siRNA group,negative control group and blank control group.HtrA1 siRNA group and negative control group were transfected with HtrA1 siRNA and control siRNA respectively.The proliferation of cells was assayed by CCK-8 method.Transwell test was used to detect the invasion ability of these three groups.Flow cytometry was used to detect the cell cycle and apoptosis.The expression of HtrA1 and vascular endothelial growth factor (VEGF)-A was detected by real time-polymerase chain reaction and Western blot respectively.Results The mRNA and protein level of HtrA1 in the light injured cells increased significantly compared to that in normal RPE cells (t=17.62,15.09;P＜0.05).Compared with negative control group and blank control group,the knockdown of HtrA1 in HtrA1 siRNA group was associated with reduced cellular proliferation (t=6.37,4.52),migration (t =9.56,12.13),apoptosis (t =23.37,29.08) and decreased mRNA (t=17.36,11.32,7.29,4.05) and protein levels (t=12.02,15.28,4.98,6.24) of HtrA1 and VEGF-A.Cells of HtrA1 siRNA group mainly remained in G0/G1 phase,the difference was statistically significant (t=6.24,4.93;P ＜0.05).Conclusion Knockdown of HtrA1 gene may reduce the proliferation,migration capability and apoptosis of light-injured RPE cells,and decrease the expression of VEGF-A.

[Objective]To investigate the clinical application of treating soft tissue defects of the feet、ankle and lower one third of the legs with reverse saphenous neurocutaneous concomitant vessel pedicled island flap.[Method]From March 1999 to June 2003,six cases of soft tissue defects of the feet,ankle and lower one third of the legs were treated with reverse saphenous neurocutaneous concomitant vessel pedicled island flap.The size of flaps were from 12 cm?7 cm to 5 cm?3 cm,aver aging 8 cm?5 cm.[Result]Flaps of 6 cases survived all showing good quality.After 10 months to 3 years follow-up,all of the six cases were thinked very good.[Conclusion]The reversed saphenous neurocutaneous concomitant vessel pedicled island flap treating soft tissue defects of the feet,the ankle and the lower one third of legs is easy to be performed.The flap has liable blood supply and does not sacrifice the major arteries.

Objective To investigate the influence of sulfated propylene glycol alginate(PSS) and its fractions on the immunoregulation.Methods The immunoregulation activity of PSS and its fractions were investigated by using immunocyte cultivation technique in vitro.The structure activity relationship was analysed on the basis of the structure studies of PSS' fractions.Results The experimental results showed that PSS could improve spleen cell proliferation,enhance macrophage phagocytic function and inhibit T-cell and B-cell proliferation.Conclusion PSS possessed significant immunoregulation effect,whilst the immunocompetence comparison of PSS' fractions proved that the different immunocytes had different requirements for saccharides length.

Background Myopic maculopathy leads to visual function damage.Conventional methods for the identification of choroidal neovascularization (CNV) of myopic maculopathy are fundus fluorescine angiography (FFA) and spectral-domain OCT (SD-OCT),but FFA is an invasive process and SD-OCT is incapable to image CNV well.OCT angiography (OCTA),a novel and non-invasive vascular technique,appears to be dominant in enface imaging of CNV,however,its clinical value remains to be evaluated.Objective This study was to evaluate the application of OCTA in imaging CNV of myopic maculopathy.Methods A prospective serial cases-observational study was performed.Forty-two eyes of 40 patients of myopic maculopathy were included in Ophthalmic Center,Renmin Hospital of Wuhan University from January to October 2015,with the diopter (-10.5 ± 3.74) D.Comperehensive optical and imaginal examinations were carried out in all the eyes,including fundus photography,FFA,SD-OCT and OCTA.Ranibizumab (0.5 mg/0.05 ml) was intravitreally injected in 35 eyes of 35 patients under the informed consent and SD-OCT and OCTA were examined 1 day,1 week and subsequent each month after injection.FFA was examined 1 month after injection.The patients were followed-up for 1-6 months to evaluate the clinical values of OCTA in monitoring the CNV of myopic maculopathy.This study followed the Helsinki Declaration and was approved by the Ethics Committee of Renmin Hospital of Wuhan University.Written informed consent was obtained from each patient prior to any medical examination.Results All the affected eyes showed dye leakage in CNV lesions by FFA and high reflection signal of abnormal choroidal vascular network throughout retinal pigment epithelium by SD-OCT.Clusters of high signal CNV image was clearly visible in the lesions by OCTA,and these signals were able to be displayed on both choriocapillary layer and outer layer of retina more clearly than FFA in 31 eyes of 31 patients.In 35 eyes of 35 patients received intravitreal injection of ranibizumab,CNV images on both choriocapillary layer and outer layer of retina from OCTA shrinked 1 week after injection and the lesions were stable 1 month after injection.Conclusions OCTA can display CNV of myopic maculopathy on the retinal outer layer and choriocapillary layer more clearly than FFA.Significant changes in the CNV net can be observed by OCTA 1 week after intravitreal injection of ranibizumab.OCTA plays an important role during the following-up of CNV therapy.

Objective To observe the characteristics of optical coherence tomography (OCT) angiography (OCTA) in retinal vein occlusion (RVO).Methods Prospective and observational study.Clinical examination of 81 consecutive patients (86 eyes) diagnosed with RVO were included in the study,in which the branch retinal vein occlusion in 47 eyes,central retinal vein occlusion in 39 eyes.Forty-five patients were male and 36 patients were female.Aged from 28 to 76 years old,the mean age was (55.36±10.01) years old.Comprehensive optical and imaging examination were performed,including fundus photography,fundus fluorescein angiography (FFA),spectral domain OCT,en face OCT and OCTA.The retinal blood flow imaging scan mode and the optic disc blood flow imaging scan mode were performed,the scanning region in the macular area were 3 mm × 3 mm,6 mm × 6 mm,8 mm × 8 mm respectively,around the optic disc were 3 mm × 3 mm and 4.5 mm × 4.5 mm.Each region scans 2 times.The characteristics of foveal avascular zone change,macular edema,non-perfusion and optical disc edema in OCTA and their corresponding FFA and en face OCT were observed.Results By OCTA,67 eyes (77.9％) for foveal avascular zone change,23 eyes (26.7％) for macular edema,40 eyes (46.5％) for non-perfusion,and 33 eyes (38.4％) for optical disc edema can be detected.The foveal avascular zone change can be indentified as the tranformation,destruction and even vanish of the arch in superfacial layer of retinal macular area,acompanied with the dilatation and thickening of capillary vessels,the occlusion and expanding of capillary vessels arounded the foveal avascular zone in the deep layer of macular area.Those performances were more clear than FFA.The main expression of macular edema was low signal and was not as clear as en face OCT.The tortuosity and expansion of retinal vessels,density decreasing and even occlusion or abnormal traffic branch of capillary vessels can be observed in non-perfusion.These observations were similar to FFA.However,pieces of highly signal identical with non-perfusion area can b.e detected in chroid capillary.The representation of optical disc edema was the brush-like expanding of capillary vessels aroud optical disc.Conclusions OCTA can help for observing the abnormal changing of capillary vessels in foveal avascular zone and macular edema,non-perfusion and optical disc edema.Foveal avascular zone change showed occlusion and expanding of capillary vessels around the foveal avascular zone in the deep layer of macular area.Macular edema showed the weak signal.Non-perfusion showed tortuosity and expansion of retinal vessels,density decreasing and even occlusion or abnormal traffic branch of capillary vessels.Optical disc edema showed brush-like expanding of capillary vessels around optical disc.

Objective To optimize the manufacture process for Zhibai Anshen oral liquid. Methods The orthogonal designed experiments were conducted to monitor the effects of three factors on the content of mangiferin. The three factors included the amount of water, extraction time and alcohol precipitation concentration. Six month accelerated stability study and twelve month long term stability study were performed. Results Optimum percolation process was boiling the mixture with 10 times of water for 1 hour, followed by deposition with 60% alcohol. Conclusion This optimized process can be used for mass production.

Objective To investigate the expression profile of miRNAs up-regualted in human extrahepatic and intrahepatic cholangiocarcinoma tissues and probe the effect on cell growth of four of these miRNAs in QBC939 cell line.Methods Up-regulated miRNAs in extrahepatic or intrahepatic cholangiocarcinoma tissues were analyzed by using miRNA-microarray,which was confirmed by using miRNA Real-Time PCR analysis.Based on these findings,four of these up-regulated miNRAs were chosen to perform function investigation.The specific miRNA inhibitors were transfected into QBC939 cells,respectively,and cell proliferation assay was performed by using MTT.Results 12 miRNAs were up-regulated both in two types of cholangiocarcinoma tissues,28 miRNAs and 21 miRNAs were up-regulated in extrahepatic cholangiocarcinoma and intrahepatic cholangiocarcinoma,respectively.MiR-125b and miR-19a expression levels were increased about 3.7 and 3.6 fold,compared with the matched normal bile duct tissues (P＜0.05).MiR-92a and miR-205 expression was upregulated about 4.S- and 3.5-fold,compared with the matched normal bile duct tissues (P＜0.05).MiR-125b,miR19a,miR-21,and miR 378* were inhibited in QBC939 cells,which indicated a significant inhibitory effect on cell growth.The ratio of inhibition was 71％,72％,69％,and 76％(P＜0.05)at 36 h,61％,63％,60％,and 59％(P＜0.01) at 48 h,and 61％、56％、60％ and 59％(P＜0.05) at 60 h.Conclusion The miRNAs expression patterns in human extrahepatic and intrahepatic cholangiocarcinoma tissues are different and uo-regulated miRNAs act as oncomirs on cholangiocarcinoma cell growth.

Objective To observe the angiographic features of patients with retinal vein occlusion (RVO) by ultra-wide-field fluorescein angiography (UWFA) and compare with the conventional 7 standard field (7SF) imaging. Methods This is a retrospective clinical description study. Fifty-eight eyes of 56 RVO patients were included. There were 25 males (26 eyes) and 31 females (32 eyes). The age ranged from 25 to 69 years, with a mean age of (48.12±18.56) years. The course of disease was from 2 days to 25 months, with a mean course of (12.78±11.35) months. Thirty eyes were diagnosed with central RVO (51.72%), 26 eyes were diagnosed with branch RVO (44.83%) and 2 eyes were diagnosed with hemicentral RVO (3.45%). Retinal laser photocoagulation was performed in 11 eyes (18.97%). All patients received examinations of UWFA (British Optomap 200Tx imaging system) and optical coherence tomography (OCT). Using the protocol for obtaining 7SF images as described in the Early Treatment Diabetic Retinopathy Study, 7 circular regions with a range of 30 degrees were combined as the 7SF template to determine the observation area. This template was then overlaid on the UWFA image to identify the potential viewable area of 7SF. The visualized retinal area, retinal non-perfusion area, retinal neovascularization area, and laser spot area of UWFA and 7SF were quantified by a retinal specialist. In addition, the OCT images of the affected eye were observed and analyzed to confirm the existence of macular edema. Correlation analysis was done between retinal non-perfusion, retinal neovascularization and macular edema detected by UWFA. Results The results of UWFA and 7SF examination were the same. Compared with 7SF, UWFA showed 3.53 times more retinal visual area, 3.31 times more non-perfusion area, 1.94 times more neovascularization area, and 3.59 times more laser spots (t=72.13, 4.69, 1.76, 5.78;P=0.000, 0.005, 0.102, 0.000). Lesions of 11 eyes (18.97%) were found outside the range of 7SF images. By UWFA, non-perfusion area correlated with neovascularization and macular edema (χ2=12.13, 4.82;P=0.000, 0.028;C=0.42, 0.28). Non-perfusion area anterior to the equator have significantly correlations with macular edema (χ2=6.32, P=0.012, C=0.31), but non-perfusion posterior to the globe equator have no relevance with macular edema (χ2=2.88, P=0.090, C=0.22). Conclusions UWFA can detect more peripheral retinal lesions than 7SF images. By UWFA, non-perfusion area has correlation with neovascularization and macular edema.

Objective To observe the ocular fundus features and consistency of classification of diabetic retinopathy (DR) by ultra-wide-field fluorescein angiography (UWFA) and the simulated early treatment diabetic retinopathy study (ETDRS) 7 standard field (7SF) imaging. Methods This is a retrospective clinical description study. Ninety-six eyes of 55 DR patients were included. The ages ranged from 25 to 73 years, with a mean age of (41.34±15.07) years. UWFA examination (British Optos 200Tx imaging system) using the protocol for obtaining 7SF images as described in the ETDRS, 7 circular regions with a range of 30 degrees are spliced as 7SF templates to determine the observation range. This template was then overlaid on the UWFA image to identify the potential viewable area of 7SF. And the visualized area of the retina, retinal non-perfusion (NP) area, retinal neovascularization (NV) area, and pan-retinal photocoagulation (PRP) area of UWFA and 7SF were quantified by a retinal specialist. Results UWFA imaging and 7SF imaging have a high degree of consistency in judging DR classification (kappa=0.851, P=0.000). The retinal visual area, NP area, NV area and PRP area of the UWFA imaging were 3.16, 3.38, 2.22 and 3.15 times more comparing with the simulated 7SF imaging (t=213.430, 45.013, 22.644, 142.665;P=0.000, 0.000, 0.003, 0.000). The lesions of 8 eyes were found outside the range of simulated 7SF imaging, including peripheral NP in 5 eyes, NV areas in 3 eyes, respectively. Conclusion UWFA imaging and simulated 7SF imaging are consistent to judge DR classification, but UWFA can find more peripheral retinal lesions.

Objective To establish a non-invasive method based on fluorescent tracer technique using inducible co-stimulatory molecules(ICOS) expressed on activated T cells for diagnosing acute heart graft rejection in mice. Methods The cervical heterotopic heart transplantation was used as model to establish isograft, allograft, allograft plus tacrolimus treatment, and allograft with tacrolimus ceased groups. On the 1, 3rd, 5th and 7th day after transplantation, Cy7. SE-ICOS-Ab was injected into the heart transplant mice via tail veins. The real-time fluorescent imaging changes of the graft were observed by fluorescent equipment. Flow cytometry was used to examine the expression of ICOS on spleen T cells of mice in each group. H-E staining was used to observe the pathological changes of cardiac graft. Results There was no noticeable fluorescent imaging in the grafts at the 1, 3rd, 5th and 7th day after transplantation in the isograft and allograft with tacrolimus treatment group. On the first day after transplantation, the fluorescent imaging of graft in the allograft group had no noticeable changes, but the fluorescent imaging gradually increased on the 3rd, 5th, and 7th day. The graft fluorescent imaging became stronger on the 3rd day after ceasing tacrolimus in the treated allograft group, and it became stronger at 5 and 7 days after ceasing tacrolimus. H-E staining found no noticeable rejection in isograft group and allograft plus tacrolimus treatment group at all time points. The allograft and allograft puls tacrolimus ceased group developed rejection on the 3rd day after transplantation, and the rejection became more serious on the 5th and 7th day. Flow cytometry showed that there were no significant differences in ICOS expression on spleen T cells on the 1 day after transplantation among the four groups (P>0. 05). The isograft and allograft plus tacrolimus treatment group had no ICOS expression on the T cells, and ICOS expression in the allograft and allograft with tacrolimus ceased group gradually increased on the 3rd, 5th and 7th day. Conclusion ICOS expression intensity is associated with the degree of graft rejection. Fluorescently labeled anti-ICOS can help to assess the development and severity of acute rejection after transplantation in a non-invasive way.