ABSTRACT
Three plasmids (pGenesil-P1, pGenesil-P2, pGenesil-P3) with different p27Kip1-shRNA sequences were designed and synthesized. Their effects on the proliferation of bovine corneal endothelial cells (bCEC) were investigated. Plasmid expressing irrelevant shRNA with a random combination was used as negative control (pGenesil-HK). The recombination of four plamids was confirmed by restrictive enzyme digestion and sequence analysis. The expression of mRNA and protein of p27Kip1 was detected by RT-PCR and Western blotting after stable transfection. The expressions of p27Kip1 mRNA and p27Kip1 protein of pGenesil-P1 group, pGenesil-P2 group and pGenesil-P3 group were all lower than those in the pGenesil-HK group and the blank group (non-transfected group). pGenesil-P3 had the strongest inhibitory effect and was selected for the next steps. The proliferation rates of the pGenesil-P3 group, the pGenesil-HK group and the blank group were assessed by MTT. The influence of shRNA-p27Kip1 on bCEC cell cycle was detected by flow cytometry (FCM). Compared with the control groups, the proliferation rate of the pGenesil-P3 group was increased significantly, and the ratio of S-phase also increased. It is concluded that shRNA-p27Kip1 could down-regulate the expression of p27Kip1 effectively and increase the proliferation of bCEC. RNA interference (RNAi) may be an effective means to promote the proliferation of CEC.
Subject(s)
Cell Proliferation , Cornea/cytology , Cyclin-Dependent Kinase Inhibitor p27/metabolism , Endothelial Cells/cytology , Endothelial Cells/metabolism , Gene Expression Regulation , Models, Biological , Plasmids/metabolism , RNA Interference , RNA, Messenger/metabolism , RNA, Small Interfering/metabolism , Reverse Transcriptase Polymerase Chain Reaction , Tetrazolium Salts/pharmacology , Thiazoles/pharmacology , TransfectionABSTRACT
The diagnosis and treatment of the lacrimal passage obstruction with lacrimal endoscope was investigated and its subsidiary surgical procedures were evaluated. Ninety-three patients (109 eyes) with lacrimal passage obstruction, including presaccal canalicular obstruction (PSCO) and na-solacrimal duct obstruction (NLDO), were examined under a lacrimal endoscope, and the obstruc-tions were treated with laser or micro-drill. All patients were followed up after the operation for 3-6 months. The difference between the laser and the micro-drill treatment was observed. During the pe-riod of follow-up, the curative rate was 82.57%. The healing rate in PSCO group and NLDO was 80.36% and 84.91% respectively (P>0.05). After treatment with the laser, the healing rate was 93.33% in the PSCO group and 66.67% in the NLDO group respectively (P<0.05). After treatment with the micro-drill, the healing rate in PSCO and NLDO groups was 65.39% and 94.28% respec-tively (P<0.01). The lacrimal passage obstruction can be observed and treated directly through the lacrimal endoscope. Choosing different surgical procedures in operation according to the locations of the obstruction is helpful to improve the effectiveness.
ABSTRACT
The diagnosis and treatment of the lacrimal passage obstruction with lacrimal endoscope was investigated and its subsidiary surgical procedures were evaluated. Ninety-three patients (109 eyes) with lacrimal passage obstruction, including presaccal canalicular obstruction (PSCO) and nasolacrimal duct obstruction (NLDO), were examined under a lacrimal endoscope, and the obstructions were treated with laser or micro-drill. All patients were followed up after the operation for 3-6 months. The difference between the laser and the micro-drill treatment was observed. During the period of follow-up, the curative rate was 82.57%. The healing rate in PSCO group and NLDO was 80.36% and 84.91% respectively (P>0.05). After treatment with the laser, the healing rate was 93.33% in the PSCO group and 66.67% in the NLDO group respectively (P<0.05). After treatment with the micro-drill, the healing rate in PSCO and NLDO groups was 65.39% and 94.28% respectively (P<0.01). The lacrimal passage obstruction can be observed and treated directly through the lacrimal endoscope. Choosing different surgical procedures in operation according to the locations of the obstruction is helpful to improve the effectiveness.
Subject(s)
Young Adult , Endoscopy/methods , Follow-Up Studies , Lacrimal Duct Obstruction/diagnosis , Lacrimal Duct Obstruction/surgery , Laser Therapy/methodsABSTRACT
In order to investigate whether cultured normal human lens epithelial cells (LEC) express transforming growth factor beta (TGF-beta), reverse transcriptase polymerase chain reaction (RT-PCR) and immunohistochemical methods were used for detection of TGF-beta mRNA and protein in cultured normal human LEC. The results showed that a single RT-PCR amplified product about 310bp was obtained, and the sequence was homologous to the known sequence. TGF-beta immunostain was positive in the plasma of LEC. It was suggested that normal human LEC could produce TGF-beta, and LEC could be affected by TGF-beta through autocrine action.
Subject(s)
Humans , Cells, Cultured , Epithelial Cells , Cell Biology , Metabolism , Lens, Crystalline , Cell Biology , Metabolism , RNA, Messenger , Genetics , Transforming Growth Factor beta , GeneticsABSTRACT
In order to investigate whether cultured normal human lens epithelial cells (LEC) express transforming growth factor beta (TGF-beta), reverse transcriptase polymerase chain reaction (RT-PCR) and immunohistochemical methods were used for detection of TGF-beta mRNA and protein in cultured normal human LEC. The results showed that a single RT-PCR amplified product about 310bp was obtained, and the sequence was homologous to the known sequence. TGF-beta immunostain was positive in the plasma of LEC. It was suggested that normal human LEC could produce TGF-beta, and LEC could be affected by TGF-beta through autocrine action.