Objective To investigate the ingestion, metabolism and subcellular localization of indocyanine green (ICG) in human retinal epithelial (R PE) cells.Methods RPE cells were incubated with 0.25 mg/ml ICG under the condition of 37oC in the camera. The ICG granule and ultrastructure of RPE cells were observed under the electron microscopy after 1, 4, and 24hour incubation, and the ICG autofluorescence was detected by fluorescence microscopy after the incubation for 1, 2, 4, 8, 12, 24, and 48 hours, respectively. The ab sorbency (A value) of ICG solution was measured at 805 nm with ultraviol et/v isible specrtrometer. The standard curve of concentration of ICG was drawn and the related equation of concentration of ICG and the A value was calculated. After being incubated for 1, 2, 4, 8, 12, 24, 48, and 72 hours, respectively, the A value of supernatant fluid was calculated according to the equation. Aft er incubated with ICG for 24 hours, one sample was observed under electron microscope and fluorescence microscope per week to evaluate the metabolizable period of ICG .Results ICG granules were distributed evenly after entering the RPE cells. After incubated with 0.25 mg/ml ICG for 24 hours, no significant change of the ultrastructure of the RPE cells was found. ICG granules accu mulated in the cells as the time goes by and reached the peak after 24 hours, and then they decreased because of the slowdown of the metabolism. Few ICG was still remained in the cells 1 week later Conclusions RPE cells may take in ICG actively. ICG metabolizable period in RPE cells is long, which may be one of the mechanisms of the toxicity of ICG to the retina in the vitreous operation.(Chin J Ocul Fundus Dis，2004，20：179-181)
Objective To establish a method for primary culture of iris pigment epithelial cells(IPE). MethodsEnzyme-Assisted microdissection was used to isolate and cultivate the IPE cells.An identification was made with microscopic and immunohistochemical observations.Results IPE were successfully sultured and showed on differences with RPE in primary culture and subculture.ConclusionEnzyme-Assisted microdissection is a reliable and quick method for the isolation of IPE.
Insufficient supply of organ for allotransplantation made the study on finding new organ resources from animal progress. Pig is regarded as one of the optimal donor animals for human. The major obstacle in this field is hyperacute reaction (HAR), which is triggered after the xenogenic natural antibodies preexisting in recipient blood combine to the antigens on the surface of the endothelium and activate the complement system. alpha-Galactose residues (alpha-Gal) on the endothelial cell have been identified as the major xenoantigens. NJZ Pig has been closely breed since 1938, whose family history is clear. Tissue samples from heart, liver, kidney, pancreas, lung, small intestine, skin, spleen, thymus and lymph node were obtained and embedded in paraffin. The sections were performed the immunohistochemical staining with the sera from health volunteers (including all the blood types) as the primary antibodies as well as the biotin labeled bandeirae simplicifolia I isolectin B4 (BS I-B4), which has specific affinity to alpha-galactose. All the staining sections were compared with the tissues digested with alpha-galactosidase. There was no difference between the antigens recognized by sera of different blood types. alpha-Gal was still the major xenoantigen on the endothelial cells. There might exist non-alpha-Gal antigens on the distal convoluted tubules and collecting tubules of the kidney. There was no alpha-Gal distributing on the secreting part of pancreas, either the islet cells or the matrix cells, but surely on pancreatic duct and vessels. All the antigenity was destroyed after the enzyme digestion except that the small intestine gland still positive with the BS I-B4. alpha-Gal is the major xenogenic antigen in NJZ Pigs. There exist some unknown antigens on the distal convoluted tubules and collecting ducts of the kidney. The blood type of recipient is not the first affair to be considered in pig-to-human xenotransplantation. The specificity of BS I-B4 for the alpha-galactose needs more detail research.
Objective To observe whether apoptosis was involved in cells of aspiration fluid from vitrectomy for proliferative vitreoretinopathy(PVR),and whether there was an association with expression of Fas antigen(Fas )and Fas ligand (FasL). Methods Cytocentrifuge slides of 11 fresh vitreous specimenｓ of PVR were prepared to be stained by TUNEL met hod for detection of apoptosis and by immunohistochemical technique for detection of Fas,FasL，and cytokeratin (CK),a cell-type specific antigen. Results Fas and FasL were expressed in normal human retina.Fas,FasL,CK，and apoptosis were found in all preparations.TUNEL-positive cells were 20.53% in total cells.70.35%,51.58%，and 82.97% of cells highly expressed Fas,FasL，and CK,respectively.The linear correlation coefficient of Fas and apoptosis was 0.99(Plt;0．001). Conclusion Vitrectomy specimens of PVR showed expression of Fas，FasL，and apoptosis.Prominent Fas and FasL expressions may be associated with apoptosis of proliferating retinal pigment epithelial cells in the vitreous of PVR. (Chin J Ocul Fundus Dis,1999,15:78-80)
Objective To investigate the inhibitive effect of E2F decoy oligodeoxynucleotides (E2F decoy ODNs) on cultured human retinal pigment epithelial (HRPE) cells.Methods E2F decoy ODNs or scramble decoy ODNs at varied concentrations were put into the HRPE cells mediated by lipofectamineTM2000. The proliferative activity of HRPE was detected by methythiazolyl-terazollium assay, and the competitive combinative activity of E2F decoy ODNs and transcription factor E2F was detected by electrophoresis mobility-shift assay. Results The proliferation of HRPE was inhibited markedly by E2F decoy ODNs at the concentration of 0.2 μmol/L (P=0.002) in a dose-dependent manner but not by scrambled decoy. The results of electrophoresis mobility-shift assay showed that the combinative activity of transcription factor E2F was abolished completely by E2F decoy ODNs. Conclusions E2F decoy ODNs may sequence-specifically inhibit the combinative activity of transcripti on factor E2F,and inhibit the proliferation of HRPE cells.(Chin J Ocul Fundus Dis，2004，20：182-185)
Objective To observe the effect of continuous elastic outside distraction on the change of collagen content in female mini pig’s ni pples and their supporting tissues, and to investigate the mechanism of continuous elastic outside distraction correcting inverted ni pples. Methods Three 3-month-old female mini pigs (weighing 18.5-22.0 kg), which had 12 nipples, were employed. Four nipples of each minipig were not treated as control group (n=12), and the other nipples were continuously distracted with inverted nipple correction instruments as experimental group (n=24). The nipple specimens were harvested at 2, 4, 8, and 12 weeks after distraction and HE staining was performed to observe the change oftheir tissue structure. And saturated picric acid sirius red staining was used to observe the distribution and content of collagen types I and III, image analysis software for quantitative analysis. Results The control group had normal structure of epidermis at all time points. In experimental group, the epidermis thickened; basal cells, fibroblasts, and capillary significantly prol iferated along with the times; and the content and the density of collagen types I and III increased gradually. There were significant differences in collagen type I at 4, 8, and 12 weeks, and in collagen type III at 2, 4, 8, and 12 weeks between 2 groups (P lt; 0.01). There were significant differences in the ratio of collagen type I to III at 2 and 4 weeks between 2 groups (P lt; 0.05). Conclusion Continuous elastic outside distraction can increase the quantity of collagen types I and III in the tissue, the thickness of the dermis, and the height of the nipple, which may be one of key mechanisms of correction the inverted nipple by continuous elastic outside distraction.
OBJECTIVE:To investigate the index of the rejection of lJle retinal pigment epithelium(RPE)cells transplantation. METHOD:Allogenic RPE transplantation on rahbits by transcleral technique, the changes of interleukin-2 (IL-2) activity in peripheral blood and the effect of immunoinhibitor (methylprednisonlone)were detected. RESLILTS:In the group of simple transplantation,the IL-2 activity in peripheral blood begin to rise in the first day after operation. The peak value occured in the third day,and is still much higher than that of the control group in the 14th day,whereas in the group treated with immunoinhibitor ,there was no obvious difference in the first day after operatlon,in the third day,the IL-2 activity rises slightly,and returned to normal level in the 7th day. CONCLUSION: After RPE transplantation, the level of IL-2 activity in peripheral blood might serve as an important index to determining and detecting the rejective response. （Chin J Ocul Fundus Dis,1996,12: 239-241）
Objective To explore a better method in obtaining iris pigment epithelium(IPE) specimen for autologous transplantation in rabbits． Methods IPE was obtained from 20 black rabbits with method A，i.e.surgical peripheral iridectomy at 12:00 position obtaining a triangle iris tissue with the hemline of 4-5 mm in left eyes，and method B,i.e.surgical peripheral iridectomy at 11:00 and 1:00 positions obtaining two triangle iris tissues with the hemlines of 2-2.5 mm in right eyes . The IP E cells were isolated precisely with enzyme microdissection-enzyme isolation method, cultured in vitro, observed with light and electronic microscope, and ident ified with immunocytochemical staining.ResultsThe success ra te of cells culture were 65% for method A and 95% for method B. After 3-4 generations of culturing，the amount of IPE cells was enough for transplantation, and most of the functions of primary clutured IPE cells were kept still. Viability of IPE cells was 85%-93%. Conclusion The success rate of cells culture for method B is higher than that for method A. The third generation of cultured cells is available for autologous transplantation.(Chin J Ocul Fundus Dis，2003，19：201-268)
Animal localization and trajectory tracking are of great value for the study of brain spatial cognition and navigation neural mechanisms. However, traditional optical lens video positioning techniques are limited in their scope due to factors such as camera perspective. For pigeons with excellent spatial cognition and navigation abilities, based on the beacon positioning technology, a three-dimensional (3D) trajectory positioning and tracking method suitable for large indoor spaces was proposed, and the corresponding positioning principle and hardware structure were provided. The results of in vitro and in vivo experiments showed that the system could achieve centimeter-level positioning and trajectory tracking of pigeons in a space of 360 cm × 200 cm × 245 cm. Compared with traditional optical lens video positioning techniques, this system has the advantages of large space, high precision, and high response speed. It not only helps to study the neural mechanisms of pigeon 3D spatial cognition and navigation, but also has high reference value for trajectory tracking of other animals.
Objective To investigate the features of ocular fundus of retinal pigment epithelial detachment (PED) in Chinese patients more than 50. Methods The clinical data of 31 continuous patients (34 eyes) with PED diagnosed by ocular fundus photochromy, fundus fluorescein angiography (FFA) and indocyanine green angiography ( ICGA ) from Oct, 2001 to Aug, 2004 were analyzed retrospectively. Results In 34 eyes with PED, the results of FFA showed serous PED in 18 (52.9%), hemorrhagic PED in 8 (23.5%), and serosanguineous PED in 8 (23.5%); the results of ICGA revealed PED associated with choroidal neovascularization (CNV) in 12 (35.3%), PED associated with ploypoidal choroidal vasculopathy (PCV) in 17 (50.0%), PED associated with both CNV and PCV in 1 (2.9%), and avascular PED in 4 (11.8%). Conclusions PED in Chinese patients more than 50 can be associated with CNV, PCV or other avascular diseases, and PCV is the most common intercurrent choroidal vascular disease. (Chin J Ocul Fundus Dis, 2006, 22: 224-227)