激光周边虹膜成形术对角膜内皮细胞影响的临床观察

目的观察激光周边虹膜成形术对角膜内皮细胞的影响并分析相关影响因素。方法对16例(30眼)高褶虹膜综合征及闭角型青光眼患者行激光周边虹膜成形术。于术前及术后1周应用非接触角膜内皮显微镜进行观察及对比分析,测量指标包括:角膜中央区域内测量的最大细胞面积(MAX)、最小细胞面积(MIN)、平均细胞面积(AVE)、细胞面积标准差(SD)、变异系数(CV)及细胞密度(CD)。结果手术前后比较,患者角膜内皮细胞在术后1周时,角膜内皮细胞的丢失率为2.13%±9.57%。角膜内皮细胞密度和平均细胞面积均有变化,差异具有统计学意义(t=2.42,-2.24,P<0.05)。经统计学分析,单次激光能量、激光击射次数、激光总能量与角膜内皮细胞丢失率呈正相关(r=0.59,0.69,0.66;P<0.05)。结论激光周边虹膜成形术可能对角膜内皮细胞造成一定损伤,并损伤与单次激光能量的大小、激光击射次数和激光总能量呈正相关。

不同方法获取Descemet膜-角膜内皮细胞复合体的效果比较

目的评价不同方法获取Descemet膜-角膜内皮细胞复合体(DM+CECs)的效果。方法将20只兔(40只眼)根据手术方法的不同随机分成4组:镊子剥离组、水分离组、大气泡法组和机械分离组,每组10只眼。通过活体染色和组织切片分析评价DM+CECs。结果成功获取DM+CECs镊子剥离组2眼,水分离组8眼,大气泡法组5眼,机械分离法组3眼,4个组之间的成功率差异有统计学意义(P<0.05)。水分离法组(98.03±2.18)%和大气泡法组(97.14±3.04)%内皮细胞存活率(ESR)明显高于镊子剥离法组(80.16±5.21)%和机械分离法组(71.56±9.57)%(F=436.04,P<0.05)。苏木精-伊红染色显示水分离法和大气泡法对内皮细胞损伤小。结论水分离法或大气泡法是安全、有效地获取DM+CECs的技术。

抗磷脂抗体损害子宫蜕膜血管内皮细胞血管形成的机制研究

目的:研究抗磷脂抗体(aPL)对蜕膜组织血管内皮细胞(DVEC)增殖、迁移与血管形成能力的影响。方法:采用细胞免疫荧光鉴定DVEC细胞;CCK-8法检测aPL对DVEC细胞增殖的影响;细胞划痕实验与血管形成实验检测aPL对DVEC细胞迁移与血管生成的影响;实时荧光定量PCR(RT-PCR)检测aPL对DVEC细胞基质金属蛋白酶2(MMP-2)和血管内皮生长因子(VEGF)mRNA表达的影响;Western blot检测aPL对DVEC细胞中p-ERK、t-ERK、VEGF与MMP-2蛋白表达的影响。结果:aPL能抑制DVEC细胞增殖、迁移与血管形成能力;aPL能抑制DVEC细胞中p-ERK、VEGF与MMP-2表达水平,加ERK激动剂后DVEC细胞中p-ERK、VEGF与MMP-2表达水平明显升高。结论:aPL可通过调控ERK通路减少DVEC细胞中VEGF与MMP-2表达,进而抑制DVEC的增殖、迁移与血管形成能力。

银杏叶提取物对体外培养心肌膜微血管内皮细胞分泌NO和ET-1的影响

研究银杏叶提取物(EGb761)对微血管内皮细胞分泌NO和ET-1的影响。采用体外培养大鼠心肌膜微血管内皮细胞(RMMMVECs)模型,测定其一氧化氮(硝酸还原酶法)和内皮素-1(ELISA法)分泌量的变化。EGb761能显著性提高RMMMVECsNO和ET-1的分泌量,且NO/ET-1值没有显著性变化。

Effect of acetyl L-carnitine on human retinal pigment epithelium-19 cells in hypoxic conditions

AIM:To investigate the effect of acetyl-L-carnitine(ALCAR)on cell viability,morphological integrity,and vascular endothelial growth factor(VEGF)expression in human retinal pigment epithelium(ARPE-19)cells using a hypoxic model.METHODS:In the first set of experiments,the optimal CoCl_(2) dose was determined by exposing ARPE-19 cell cultures to different concentrations.To evaluate the effect of ALCAR on cell viability,five groups of ARPE-19 cell culture were established that included a control group,a sham group(200μM CoCl_(2)),and groups that received 1,10 and 100 mM doses of ALCAR combined with 200μM CoCl_(2),respectively.The cell viability was measured by MTT assay.The morphological characteristics of cells were observed by an inverted phase contrast microscope.The levels of VEGF and HIF-1α secretion by ARPE-19 cells were detected by enzyme linked immunosorbent assay(ELISA)assay.RESULTS:ARPE-19 cells were exposed to different doses of CoCl_(2) in order to create a hypoxia model.Nevertheless,when exposed to a concentration of 200μM CoCl_(2),a notable decrease in viability to 83% was noted.ALCAR was found to increase the cell viability at 1 mM and 10 mM concentrations,while the highest concentration(100 mM)did not have an added effect.The cell viability was found to be significantly higher in the groups treated with a concentration of 1 mM and 10 mM ALCAR compared to the Sham group(P=0.041,P=0.019,respectively).The cell viability and morphology remained unaffected by the greatest dose of ALCAR(100 mM).The administration of 10 mM ALCAR demonstrated a statistically significant reduction in the levels of VEGF and HIF-1α compared with the Sham group(P=0.013,P=0.033,respectively).CONCLUSION:The findings from the current study indicate that ALCAR could represent a viable therapeutic option with the potential to open up novel treatment pathways for retinal diseases,particular relevance for age-related macular degeneration(AMD).However,to fully elucidate ALCAR’s application potential in retinal diseases,additional investigation is necessary to clearly define the exact mechanisms involved.

Isolation, Cuture and Biological Characteristic Analysis of Stromal and Glandular Epithelial Cells of Buffalo (Bubalus bubalis) Endometrium

[Objective] The experiment aimed to set up a method for isolating and culturing endometrial stromal cells (BESC) and endometrial glandular epithelial cells(BEGEC) of buffalo as well as laid foundation for studying biological mechanism of embryo implantation and uterine diseases. [Method] The enzymatic digestion method, scraping method, serial filtration and differential velocity adherent technique were used to isolate BESC and BEGEC, then immunocytochemical method and TRYPAN-Blue assay were used to determine the purity and survival rate of isolated cells. [Result] The BESC and BEGEC were successfully isolated and cultured while immunocytochemical method and cell count method demonstrated that the purity was over 90%. The result of TRYPAN-Blue assay shown that survival rate of BESC and BEGEC was 91% and 78% respectively. [Conclusion] The enzymatic digestion method, scraping method, serial filtration and differential velocity adherent technique could isolate BESC and BEGEC with high purity.

Origin and fate of the nucleolar channel system of normal human endometrium^1

Human normal endometrium was examined in ultrathin sections. Nucleolar channel system (NCS) appeared in the endometrial epithelial cells during the early and mid secretory phase of menstrual cycle. The NCS was a hollow ball like structure of different sizes and was composed of 2 to 5 rows of tubules embedded in an amorphous matrix. On its surface there were numerous electron dense particles resembling ribosonies. It was usually located within or associated with the nucleolus. Sometimes, it was close to the nuclear envelope or protruding out from the nucleus. On occasion, NCS with simplified structure was found in the perinuclear cytoplasm. Concepts concerning the genesis, involution and function(s) of the NCS were discussed.

The mitochondrial Na^＋/Ca^2＋ exchanger may reduce high glucose-induced oxidative stress and nucleotide-binding oligomerization domain receptor 3 inflammasome activation in endothelial cells

Background The mitochondrial Na^＋/Ca^2＋ exchanger, NCLX, plays an important role in the balance between Ca2. influx and efflux across the mitochondrial inner membrane in endothelial ceils. Mitochondrial metabolism is likely to be affected by the activity of NCLX because Ca^2＋ activates several enzymes of the Krebs cycle. It is currently believed that mitochondria are not only centers of energy produc- tion but are also important sites of reactive oxygen species （ROS） generation and nucleotide-binding oligomerization domain receptor 3 （NLRP3） inflammasome activation. Methods ＆ Results This study focused on NCLX function, in rat aortic endothelial cells （RAECs）, induced by glucose. First, we detected an increase in NCLX expression in the endothelia of rats with diabetes mellitus, which was induced by an injection of streptozotocin. Next, colocalization of NCLX expression and mitochondria was detected using confocal analysis. Suppression of NCLX expression, using an siRNA construct （siNCLX）, enhanced mitochondrial Ca^2＋ influx and blocked efflux induced by glucose. Unexpectedly, silencing of NCLX expression induced increased ROS generation and NLRP3 inflammasome activation. Conclusions These findings suggest that NCLX affects glucose-dependent mitochondrial Ca^2＋ signaling, thereby regulating ROS generation and NLRP3 in- flammasome activation in high glucose conditions. In the early stages of high glucose stimulation, NCLX expression increases to compensate in order to self-protect mitochondrial maintenance, stability, and function in endothelial cells.

Cytokine orchestration in post-operative peritoneal adhesion formation

Peritoneal adhesions are a near inevitable occurrence after laparotomy and a major cause of both patient and physician misery. To date, clinical attempts at their amelioration have concentrated on manipulating the physical factors that affect their development despite a wealth of experimental data elucidating the molecular mechanisms that underlie their initiation, development and maturation. However, the advent of targeted, specific anti-cytokine agents as directed therapy for inflammatory and neoplastic conditions raises the prospect of a new era for anti-adhesion strategies. To harness this potential will require considerable cross-disciplinary collaboration and that surgeon-scientists propel themselves to the forefront of this emerging fi eld.

Duodenal intraepithelial T lymphocytes in patients with functional dyspepsia

AIM： To quantify the intraepithelial lymphocytes （IELs） and to document the membrane expression of CD4, CD8, TCRγδ and adhesion and/or activation-associated molecules （CD103, CD28, CD44, CD69, HLA-DR, CD95/ Fas） in the duodenal mucosa of patients with functional dyspepsia （FD） in order to provide arguments for an immunological process in FD. METHODS： Twenty-six FD patients according to Rome Ⅱ criteria （20 were H pylori negative） were studied and compared to 12 healthy adults. IELs were isolated from five duodenal biopsy samples, then quantified by microscopy and flow cytometry while the membrane phenotypes were determined by cytofluorometry. RESULTS： Duodenal histological examination was normal. In H pylori negative patients, the number of IELs was not different from that in healthy controls. Median percentage expression of CD4, CD8, or TCRy8 and CD103, CD44, CD28, CD69 on CD3＋ IELs, among the adhesion/activation associated molecules tested, was not different from that in healthy controls. In contrast, the median percentage expression of CD95/ Fas [22 （9-65） vs 45 （19-88）, P = 0.03] and HLADR expressing CD3＋ IELs [4 （0-30） vs 13 （4-42）, P = 0.04] was significantly lower in the H pylori negative FD group than in healthy controls, respectively. The number of IELs was significantly greater in H pylori positive FD patients than in healthy controls [median ratio for 100 enterocytes 27.5 （6.7-62.5） vs 10.8 （3-33.3）, P = 0.02] due to a higher number of CD8＋ CD3＋ IELs. CONCLUSION： In H pylori negative FD patients, the phenotypic characterization of IELs suggests that we cannot exclude a role of IELs in FD.

组织工程的一个初步研究混合多尿烷血管移植(英文)

本文的目的是研究半多孔内皮细胞的包膜(SPEUs)的效应 以PTMEG1000为基础的SPEU的多孔性为51.58%(±5.86%) 与天然脉管相比较,例如与一头6个月小牛胸部的上部下来的主动脉大血管相比较,SPEU的拉伸强度是足够强的 脉管的典型脉动压力服从静脉管压力(1.5～2%Δd/d),此压力小于动脉管压力(9～15%Δd/d) 在种植EC之前,观察了在SPEUs上培育的人皮的成纤维细胞(HDF) SPEU的DNA数量与HDF在一天之内种植;有71.9%(±11.01%)细胞被种植在SPEUs上 LDH试验表明:在有控制的和实验群数据之间没有显著的差别,说明没有细胞毒素 因此,EC在SPRUs上具有良好的粘附性,这是所期望的。

DIFFERENT RESPONSES OF CHORIOCAPILLARY ENDOTHELIAL CELLS AND RETINAL CAPILLARY ENDOTHELIAL CELLS TO MITOGENIC AND VASOACTIVE FACTORS

The responses of choriocapillary endothelial cells(CCE) and retinal capillary endothelial cells(RCE) in culture,in terms of phosphoinositide(PI) breakdown and cellular mitogenesis, to retinal pigment epithelial cell(RPE)-conditioned medium and vasoactive agents have been compared. RPE-conditioned medium did not induce PI breakdown in either type of cell. However, it stimulated DNA synthesis in CCE but not in RCE. Bradykinin(BDK) acted as both a fast signaling and a slow mitogenic factor on CCE, but BDK did not affect PI turnover or DNA synthesis in RCE. In contrast, thrombin stimulated PI turnover in RCE but not in CCE, though it did not induce 3H-thymidine incorporation into either type of cell. These differences in cellular functions between CCE and RCE following stimulation suggest that induction of DNA synthesis and receptor-mediated PI turnover by external factors is determined, at least in part, by the origin of the capillary endothelial cell. Therefore, extrapolation to CCE pathophysiology from experiments using endothelial cells from other capillary origins may not be valid.

A study of chitosan blend film as a carrier of corneal endothelial cells

In order to evaluate the cytocompatibility and biocompatibility of a new kind of chitosan blend film as a carrier of corneal endothelial cell, rabbit corneal endothelial cells cultured in vitro were breeded onto the film. After a cell monolayer formed, the scanning electron micrography was performed. After inplanted into anterior chamber, slit lamp observation, thickness metering, specular microscopy and HE staining were performed at random time after operation to evaluate the biocompatibility. Inflmmnation in anterior, thickness of cornea, cell density, hexagonality and cell size of the surgical cornea were taken as the indexes of biocompatibility. The cultured cells exhibited a confluent monolayer 10 days after incubation, which proved the satisfactory cytocompatibility of this film. Biocompatibility assay results suggested the implantation feasibility of the film as a carder of corneal endothelial cells.

In Vitro Model of Human Choroidal Neovascular

Choroidal capillary endothelia cell （CEC） plays a critical role in the development of choroidal neovascularization which is one of the major causes of blindness. An effective method for CEC cultivation was proposed. The isolation of human choroidal CECs using micro dissection followed by the use of superparamagnetic beads （Dynabeads） coated with the CD 31, which selectively binds to the endothelial cell surface. Cells bound to beads were isolated using a magnetic particle concentrator. The CECs were planted into type Ⅳ collagen coated 24 well plates. The results show that the primary cultured CEC is induced to tube formation in collagen Ⅳ coated environment, which can be presented as an in vitro model of choroidal neovascularization.

脑膜瘤的CT表现、组织学类型及临床间的相关性(附40例报告)

脑膜瘤源发于蛛网膜内皮细胞，大多数屑良性肿瘤，少数可恶变，目前随着诊疗水平的提高，脑膜瘤的谚断、手术垒切除率也不断提高，为进一步提高疗效，本文对手术后的40例脑膜瘤的CT表现、组织学类型及临床间关系的相关性研究报告如下。

Inhibition of angiogenesis by DADAG in vivo and in vitro

1,2,5,6-Dianhydro-3,4-diacetyl-galactitol （DADAG）, an alkylating sugar alcohol derivative, has been shown effective against tumor growth. In this research, we explored the effect of DADAG on angiogenesis in chick chorioallantoic membrane （CAM） model and on the proliferation and migration of human umbilical vein endothelial cells （HUVECs）. We also studied the possible mechanism of the anti-angiogenesis effect of DADAG. The results showed that DADAG （100, 500 and 1000μnol/L） inhibited angiogenesis in CAM model dose-dependently. Sulforhodamine B （SRB） assay indicated that DADAG （45, 90, 135, 180 and 225 μmol/L） suppressed HUVECs proliferation in a dose-dependent and time-dependent manner. High Content Screening （HCS, Cellomics） assay, in which the influence of cell proliferation on migration could be excluded, indicated that DADAG （45, 135 and 225 ~xmol/L） directly inhibited the motility ofHUVECs. Immunofiuorescence assay suggested that DADAG inhibited angiogenesis possibly by decreasing vascular endothelial growth factor （VEGF） expression in HUVECs. Our findings reveal that DADAG show anti-angiogenic activity in vivo and in vitro, which is related to the downregulation of VEGF expression in endothelial cells.

Expression of carbonic anhydrase Ⅳ in rabbit corneal endothelial cells

Objective To demonstrate the molecular expression of carbonic anhydrase Ⅳ (CA Ⅳ) in rabbit corneal endothelium.Methods Reverse transcriptase polymerase chain reaction (RT-PCR) was performed using cultured and fresh rabbit corneal endothelial total RNA and specific primers for CA Ⅳ. The RT-PCR product was subcloned and sequenced. Immunoblotting and indirect immunofluorescence staining were performed to detect protein expression and distribution of CA Ⅳ using fresh and cultured rabbit corneal endothelium and rat anti-CA Ⅳ polyclonal antibody. Results RT-PCR screening gave positive bands at the predicted size for CA Ⅳ from fresh and cultured rabbit corneal endothelium. Sequencing further confirmed the identity of CA Ⅳ in corneal endothelium. Immunoblotting analysis showed a single band at 52 kDa for freshly isolated and cultured endothelial cells. Indirect immunofluorescence staining revealed an apparent positive staining in cultured endothelial cells.Conclusion Carbonic anhydrase Ⅳ is expressed in rabbit corneal endothelium, which could contribute to the transendothelial HCO 3 - flux that is necessary to maintain corneal hydration and transparency.

Punicalagin protects bovine endometrial epithelial cells against lipopolysaccharide-induced inflammatory injury

Objective：Bovine endometritis is one of the most common reproductive disorders in cattle.The aim of this study was to investigate the anti-inflammation potential of punicalagin in lipopolysaccharide（LPS）-induced bovine endometrial epithelial cells（bE ECs）and to uncover the underlying mechanisms.Methods：bE ECs were stimulated with different concentrations（1,10,30,50,and 100μg/ml）of LPS for 3,6,9,12,and 18 h.MTT assay was used to assess cell viability and to identify the conditions for inflammatory injury and effective concentrations of punicalagin.Quantitative real-time polymerase chain reaction（q RT-PCR）was used to assess gene expression of pro-inflammatory cytokines.Western blotting was used to assess levels of inflammation-related proteins.Results：Treatment of b EECs with 30μg/ml LPS for 12 h induced cell injury and reduced cell viability.Punicalagin（5,10,or 20μg/ml）pretreatment significantly decreased LPS-induced productions of interleukin（IL）-1β,IL-6,IL-8,and tumor necrosis factor-α（TNF-α）in bE ECs.Molecular research showed that punicalagin inhibited the activation of the upstream mediator nuclear factor-κB（NF-κB）by suppressing the production of inhibitorκBα（IκBα）and phosphorylation of p65.Results also indicated that punicalagin can suppress the phosphorylation of mitogen-activated protein kinases（MAPKs）including p38,c-Jun N-terminal kinase（JNK）,and extracellular signal-regulated kinase（ERK）.Conclusions：Punicalagin may attenuate LPS-induced inflammatory injury and provide a potential option for the treatment of dairy cows with Escherichia coli endometritis.

Therapeutic efficiency of tissue-engineered human corneal endothelium transplants on rabbit primary corneal endotheliopathy

To evaluate the therapeutic efficiency of tissue-engineered human corneal endothelia (TE-HCEs) on rabbit primary corneal endotheliopathy (PCEP),TE-HCEs reconstructed with monoclonal human corneal endothelial cells (mcHCECs) and modified denuded amniotic membranes (mdAMs) were transplanted into PCEP models of New Zealand white rabbits using penetrating keratoplasty.The TE-HCEs were examined using diverse techniques including slit-lamp biomicroscopy observation and pachymeter and tonometer measurements in vivo,and fluorescent microscopy,alizarin red staining,paraffin sectioning,scanning and transmission electron microscopy observations in vitro.The corneas of transplanted eyes maintained transparency for as long as 200 d without obvious edema or immune rejection.The corneal thickness of transplanted eyes decreased gradually after transplanting,reaching almost the thickness of normal eyes after 156 d,while the TE-HCE non-transplanted eyes were turbid and showed obvious corneal edema.The polygonal corneal endothelial cells in the transplanted area originated from the TE-HCE transplant.An intact monolayer corneal endothelium had been reconstructed with the morphology,cell density and structure similar to those of normal rabbit corneal endothelium.In conclusion,the transplanted TE-HCE can reconstruct the integrality of corneal endothelium and restore corneal transparency and thickness in PCEP rabbits.The TE-HCE functions normally as an endothelial barrier and pump and promises to be an equivalent of HCE for clinical therapy of human PCEP.