To investigate the effects of pyrrolidine dithiocarbamate (PDTC) on the proliferation and PCNA (proliferating cell nuclear antigen) expression of cultured human retinal pigment epithelium cells, human retinal pigment epithelium cells (RPE) were cultured from normal adults who, died accidentally. The effects of PDTC on the proliferation of RPE cells were examined by using methyl thiazlyl tetrazolium (MTT) assay. The effects of PDTC on the PCNA expression of RPE cells were immunohistochemically examined by employing biological image analysis system (BIAS). After treatment with PDTC of various of concentration ranging from 0.062 to 1 g/L for 24 h, or concentrations ranging from 0.031 to 1 g/L, the proliferation of RPE cells decreased in a dose-dependent manner. After treatment with PDTC of concentration varying from 0.062 to 1 g/L for 24 h, the PCNA expression was also suppressed in a dose-dependent manner. It is concluded that PDTC can inhibit the proliferation of RPE cells in vitro in a dose-and time-dependent manner, at least in part, by down-regulating the expression of PCNA. PDTC may be used to prevent and treat the proliferative vitreoretinopathy (PVR).

AIM: To investigate the efficacy and safety of frequency of intense pulsed light(IPL)in the treatment of meibomian gland dysfunction.METHODS: In this retrospective study, a total of 108 patients(216 eyes)with meibomian gland dysfunction admitted to our hospital from January 2021 to June 2022 were included. They were divided into two groups, with 54 cases(108 eyes)IPL group(energy density 13.0J/cm2, pulse width 6ms, delay time 50ms), and 54 cases(108 eyes)in advanced optimal pulsed technology(AOPT)group(energy density 10.0-16.0J/cm2, pulse width 7-4-4 ms in unequal-division mode). The clinical effects of the two groups were observed and compared, including ocular surface symptoms, corneal fluorescein staining score(FL), tear film lipid layer thickness(LLT), ocular surface disease index(OSDI)score, mean non-invasive tear film break-up time(NIBUTav)and first non-invasive tear film break-up time(NIBUTf), tear meniscus height, score of meibomian gland secretion and its secretion traits, and the incidence of adverse effects was also calculated.RESULTS: The effective rate of the AOPT group(106 eyes, 98.1%)was higher than that of the IPL group(90 eyes, 83.3%, P<0.05), as well as OSDI score, FL score, score of meibomian gland secretion and its secretion traits, LLT NIBUTav, NIBUTf and tear meniscus height(all P<0.001). However, the incidence of adverse effects of the AOPT group(18 eyes, 16.7%)was higher than that of the IPL group(4 eyes, 3.7%; P<0.05).CONCLUSION: With significant improvement in the ocular surface symptoms and function, AOPT has a better therapeutic effect on the treatment of meibomian gland dysfunction, but it has more adverse reactions. Therefore, optimal treatment plan should be fully selected in combination with the actual clinical situation.

In order to explore the difference of intraocular pressure (IOP) at different points of cornea before and after laser in situ keratomileusis (LASIK), IOP was measured by Tono-Pen Tonometer at central cornea, pericentral cornea and limbus respectively and analyzed statistically. After LASIK, IOP was dropped significantly at central cornea and pericentral cornea (P<0.05), while no statistically significant change occurred at limbus (P>0.05). There was no statistically significant difference in IOP at different points before LASIK (F=0.110, P=0.896), but statistically significant difference was found after LASIK (F=7.375, P=0.001). It was suggested that reliable IOP after LASIK could be obtained from the limbus by Tono-Pen tonometer.

The efficiency and safe range of Lipofectamine2000 (LF2000)/bcl-xl applied in human keratocytes, the optimal ratio of LF2000/bcl-xl and the bcl-xl gene expression in human keratocytes were investiaged. By using trypan-blue staining, the effects of LF2000 and bcl-xl on the survival rate of the cultured human keratocytes were measured respectively. By using semi-quantitative RT-PCR, the efficiency and the expression of LF2000-mediated bcl-xl transfection into keratocytes were examined. The results showed that the survival rate of human keratocytes had no signficant change in the presence of LF2000 (20 microg/ml) or bcl-xl (10 microg/ml) for 24 h. LF2000 could effectively mediate the transfection of exogenous gene bcl-xl into human keratocytes. The best transfection efficiency could be obtained when the ratio of bcl-xl/LF2000 was 1:8. One day after transfection, the positive cells for bcl-xl could be detectable, and the positive rate reached the peak-on the posttransfection day 3 (48.3%), then gradually decreased. Fifteen days after transfection, there were few positive cells. It was suggested that LF2000 could effectively transfer the exogenous gene bcl-xl into human keratocytes without obvious toxicity during a concentration range. LF2000/bcl-xl may be likely to play an important role in gene therapy of human keratocytes.
Cations/administration & dosage ; Cornea/cytology ; Gene Therapy ; Keratinocytes/cytology ; Keratinocytes/*metabolism ; Liposomes ; Transfection ; bcl-X Protein/biosynthesis ; bcl-X Protein/*genetics

Sonic hedgehog (Shh) signaling has recently been shown to be involved in the pathological angiogenesis in response to tissue hypoxia and ischemic injury. Hypoxia/ischemia is considered to play an important role in the development of choroidal neovascularization (CNV). This study was aimed to examine the effect of blockade of the Shh signaling pathway on CNV and the underlying mechanism. A total of 64 male Brown-Norway (BN) rats were used in this study. One eye of each rat underwent laser photocoagulation. The other eye served as normal control. After the laser treatment, the 64 rats were divided into four groups (n=16 in each group): Blank control group, in which no intravitreal administration was given; cyclopamine group, recombinant Shh N-terminals protein (rShh) group and phosphate-buffered saline (PBS) group, in which cyclopamine (a Shh inhibitor), rShh (a Shh activator) and PBS were intravitreally injected into the laser-treated eyes respectively every other day for a total of four intravitreal injections immediately after the laser treatment. Fourteen days after the intravitreal administration, the changes of CNV-related variables, including positive CNV lesion percentage, CNV membrane area and CNV membrane thickness, were evaluated by fluorescein angiography, indocyanine green angiography and pathological examinations. The mRNA and protein expression of PTCH1, Gli1, HIF-1(α), VEGF and DLL4 in each group on 14 days of CNV model was detected by real-time quantitative PCR and western blot analysis, and the relationship between the Shh cascade and the HIF-1(α)-VEGF-DLL4 cascade in CNV was analyzed. The results showed that the CNV membrane area and the CNV membrane thickness were decreased by 62.5% and 41.9% in the cyclopamine group and increased by 85.7% and 64.3% in the rShh group in comparison to those in the blank control group (P<0.01 for each). There was no significant difference in the CNV membrane area and thickness between the blank control group and PBS group (P=0.102 and P=0.063, respectively). Real-time quantitative PCR revealed a 5.23-, 4.14-, 2.97-, 2.78- and 2.39-fold up-regulation of the mRNA expression of PTCH1, Gli1, HIF-1(α), VEGF and DLL4 genes in the laser-treated eyes compared with the normal control eyes in the control group. In the cyclopamine group, the mRNA and protein expression of Gli1, HIF-1(α), VEGF and DLL4 was significantly down-regulated (P<0.05 for each) while the expression of PTCH1 showed no significant changes at the mRNA (P=0.293) and protein level (P=0.304). The mRNA expression and protein expression (P=0.001 and P=0.021, respectively) of PTCH1, Gli1, HIF-1(α), VEGF and DLL4 was significantly increased in the rShh group when compared with the control group. The expression level of these genes was related to the severity of the CNV. It was concluded that intravitreal administration of cyclopamine can effectively inhibit the formation of laser-induced experimental CNV by down-regulating the expression of the HIF-1(α)-VEGF-DLL4 cascade in CNV. The Shh signaling pathway as an upstream signaling pathway of HIF-1(α)-VEGF-DLL4 cascade is implicated in the development of experimental CNV.

With relatively low rejection rate and better visual prognosis, Descemet membrane endothelial keratoplasty (DMEK) has become the mainstream surgery for the treatment of endothelial dysfunction in some developed countries, but it has not been applied widely in China due to technical difficulties, the long learning curve, shallow anterior chamber of Chinese people, and the fact that domestic corneal endothelial lesions are often accompanied with other complex eye diseases.In this review, the indications, donor graft preparation including donor selection, graft preparation techniques and visualization of graft, key surgical techniques including the implantation, unwrapping and positioning of graft, postoperative complications including graft detachment, high intraocular pressure, rejection, endothelial cell loss, graft survival rate, and visual prognosis of DMEK were reviewed.

Objective: To develop an automatic warning software system for MEWS, and apply the MEWS system and SBAR communication mode to the early warning of surgical patients to evaluate its implementation effect. Methods: From November 2017 to November 2018, 400 patients in the People′s Hospital of Guigang City, Guangxi, with vital signs and critical illness after surgery were divided into 200 patients in the control group and 200 in the study group according to the random number table. The control group: routinely calculated MEWS scores and reported abnormal values to the doctor to treat the patient′s condition. The research group: Combining MEWS assignment with computer technology, developing MEWS automatic disease warning software system and combining it with SBAR communication mode for early warning of surgical patients′ condition, comparing the two groups′ disease evaluation time, treatment response speed, communication completion rate and Whether patients and doctors have different job satisfaction with nurses. Results: The disease assessment time and treatment response rate of the study group were (45.89±1.528) seconds and (1.22±0.57) minutes, respectively. The control group was (58.01±5.123) seconds and (3.19±1.56) minutes respectively. The difference between the two groups was significant (t=32.078, -7.899, P<0.01). The communication completion rate of the study group, the patient's job satisfaction to the nurses, and the doctor's job satisfaction to the nurses was 96.0% (192/200), 91.5% (183/200) and 97.0% (194/200) respectively. The rate in the control group was 61.5% (123/200), 79.0% (158/200), and 78.0% (156/200) respectively, the difference between the two groups was significant (χ²=71.126, 12.426, 33.006, P < 0.01). Conclusion The MEWS automatic warning software system and SBAR communication mode can be used to quickly assess the patient′s condition, improve the accuracy of medical communication, alert patients to potential risks and severity of illness, and improve nurses' work efficiency and patient and doctor's satisfaction with nurses' work degree.

Objective To investigate the killing effect of herpes simplex virus thymidine kinase/gancyclovir (HSV-tk/GCV) suicide gene therapy system on retinoblastoma (Rb) cells and the mechanism of bystander effect. Methods By using liposome, pCMV/hytk-IREShrGFP plasmid was transferred into HXO-Rb44 cells. A fluorescence microscope was used to detect the transduction effeciency. The positive cell clones were selected by hygromycin and were named HXO-Rb44/tk. RT-PCR was resorted to demonstrate the sucessful transduction and transcription of hytk gene in the HXO-Rb44/tk cells. The morphologic features and growth patterns of HXO-Rb44/tk were compared with those of HXO-Rb44. Then MTT assay was used to determined the killing effect of GCV on HXO-Rb44/tk and the mixture of HXO-Rb44/tk and HXO-Rb44 in different ratios ("bystander effect"). The mechanism of bystander effect was studied by the experiment of supernatant shifting. Results The transduction effeciency was 20%. 530bp hytk gene strand was seen through HXO-Rb44/tk RT-PCR. There were no differences in the morphologic features or the growth patterns between HXO-Rb44/tk and HXO-Rb44. HXO-Rb44/tk was more sensitive to GCV than was HXO-Rb44. The cytotoxicity of HXO-Rb44/tk was dose and time dependent. An obvious "bystander effect" was seen even with low proportions of HXO-Rb44/tk, but this effect disappeared when transferring GCV containing supernatant of HSV-tk-positive cells to the negative cells. Conclusion The transfer of the HSV-tk gene into Rb cells followed by the administration of GCV could serve as a model for gene therapy for retinoblastoma, the "bystander effect" in HSV-tk/GCV-mediated gene therapy occurs by transfer of GCV metabolite from cell to cell through gap junction.

This study examined the corneal permeability of topical eye drop solutions added with various corneal penetrating accelerators and gadolinium-diethylene triamine pentaacetic acid (Gd-DTPA) by nuclear magnetic resonance imaging (MRI). Twenty-four New Zealand rabbits were randomly divided into 3 groups according to the random digits table: Gd-DTPA group, in which the rabbits received 23.45% Gd-DTPA; hyaluronic acid group, in which 23.45% Gd-DTPA plus 0.2% hyaluronic acid was administered; azone group, in which 23.45% Gd-DTPA with 0.2% azone was given. Fifty microliters of the eye drops was instilled into the conjunctive sac every 5 min, for a total of 6 applications in each group. Contrast medium signals in the cornea, anterior chamber, posterior chamber, and vitreous body were scanned successively by MRI. The morphology and cell density of the corneal endothelium were examined before and 24 h after the treatment. The results showed that the residence time of Gd-DTPA in the conjunctival sac in the hyaluronic acid and azone groups was longer than that in the Gd-DTPA group. The signals in the anterior chamber of the Gd-DTPA and hyaluronic acid groups were increased slightly, and those in the azone group strengthened sharply. The signal intensity continuously rose over 80 min before reaching plateau. The strengthening rate of signals in the anterior chamber was 19.63% in the Gd-DTPA group, 53.42% in the sodium hyaluronate group, and 226.94% in the azone group. No signal was detected in the posterior chamber or vitreous body in all the 3 groups. Corneal morphology and cell density did not show any significant changes after the treatment in all the 3 groups. It was concluded that azone can significantly improve the corneal permeability of drugs that are similar to Gd-DTPA in molecular weight and molecular size, and MRI is a noninvasive technique that can dynamically detect eye drop metabolism in real time.

Background Endothelin-1 (ET-1) is an active regulator of intraocular pressure.The ET-1 level in aqueous humor is elevated in primary open-angle glaucoma,normal intraocular tension glaucoma and the animal model of glaucoma.There is now accumulating evidence for a role of ET-1 in the pathogenesis of glaucoma.However,the effect of ET-1 on the phagocytic function in trabecular meshwork cells (TMCs) is unclear.ObjectiveThis study is to observe the effect of ET-1 on the phagocytic function in cultured human TMCs.Methods Human trabecular meshwork tissue was obtained from healthy donator and cultured and subcultured in vitro by the explant culture method.The third passage of human TMCs were incubated with fluoresent red-labeled latex beads for 0,4,8,12,24,48 and 72 hours.The phagocytic kinetics of human TMCs were continuously evaluated by counting the number of latex beads in TMCs using a fluorescence microscope.Depending on the concentrations of ET-1 in culture medium,the TMCs were divided into control group (without ET-1),low-dose ET-1 (10~(-9)mol/L) treatment group,middle-dose ET-1 (10~(-8)mol/L) treatment group and high-dose ET-1 (10~(-7) mol/L) treatment group.In addition,based on the addition of endothelin receptor (ETAR) antagonist,the TMCs were divided into control group (without ETAR antagonist),ET-1 (10~(-8)mol/L) treatment group,ETAR antagonist (1×10~(-7)mol/L BQ123+10~(-8)mol/L ET-1) treatment group and ETBR antagonist(1×10~(-7) mol/L BQ788+10~(-8) mol/L ET-1)treatment group.TMCs of each group were incubated with latex beads,and the numbers of latex beads in TMCs were counted under a fluorescent microscope.Results Cultured HTM cells showed positive reactions for FN,LN,NSE and negative response for FⅧRag.The phagocytic kinetics test revealed that the latex beads were detected 4 hours after incubation.The density of latex beads was gradually increased with the delay of incubation duration and peaked at 24 hours.The number of the latex beads saturated after 48 hours of incubation.However,the number of latex beads in TMCs was significantly reduced after the addition of ET-1 in a dose-dependent manner (F=28.91,P＜0.05).The number of latex beads in the ET-1 group was less than that in the control group and the ETAR receptor antagonist group (q=13.7228,q=9.4312,P＜0.05).No significant difference was found in latex beads number between the ET-1 group and the ETBR antagonist group (q=1.1600,P＞0.05).Conclusion ET-1 inhibits the phagocytic function of human TMCs and ETAR plays a partial role in the phagocytic function of human TMCs.