Objective
To observe the ultrastructural characteristics of human retinal progenitor cells cultured in vitro.
Methods
Six 5-month-old human fetuses（12 eyes）without eye diseases were selected.
Retinal
progenitor cells from the retina of one eye of each fetus were cultured in vitro，and observed by transmission electronic microscopy（TEM）; while those from the other eye were directly observed by TEM. Results Abundant heterochromatin were found in the karyon of 5-month embryonic retinal neuroepithelial cells，and the figure of the karyons was irregular.A few scattered initial cells were seen in retinal neuroepithelial layer with large karyon，smooth surface，abundant euchromatin，and distinct nucleolus.The human retinal progenitor cells cultured in vitro had the same ultrastructural characteristics as the initial cells:with huge karyon which almost occupied the whole cell，little cytoplasm，distint nucleolus，abundant euchromatin，and little heterochromatin.The cells clung to each other in the neural globoid cell mass.The size of the outer cells was large，and karyokinesis could be found.
Conclusion
The cultured human retinal progenitor cells are provided with the same ultrastructure characteristics as the initial cells.
(Chin J Ocul Fundus Dis, 2006, 22: 185-187)
Objective：To observe the histochemical changes of retinal photochemical damage in rats. Methods：The changes of retinal ultrastructure were observed.The concentration of malondaldehyde(MDA) was tested and the activity the histochemical change of cytochrome oxidase (CCO) and (Mg ++ -ATPasw) were evaluated on the retnal photochemical damage in SD rats. Results：At the 6th hour after light exposure,the swelling appwared at the nuclei of photoreceptor,the mitochondria of inner segment.The apical microvilli of RPE disappeared and lysosomes increased in RPE.On the 6th day after light exposure,the changes became more obvious.While on the 14th day after light expose the nuclei of photoreceptors and the inner segments renewed but the arrangement of the disk was lose;and the microvilli appeared of the disk was lose;and the microvilli appeared at the tip of RPE.The Activity of CCO and Mg ++ -ATPase decreased and MDA increased in retina at the 6th hour and on the 6th day and they recovered on the 14th day after light exposure. Conclusion：Lipd peroxidation that broke the cell membrane system of photoreceptor which induced changes of the cell ultrastru cture abd the activity of enzyme might relate to pathogenesis in retinal photochemical damage. (Chin J Ocul Fundus Dis,1998,14:38-40)
Objective To investingate the ultrastructural changes of retinal pigment epithelium(RPE) and its permeability in spontaneously hypertensive rats(SHR)and explore the relation between these changes and hypertensive retinopathy.MethodsThe ultrastructure of RPE cells in the SHR aged five,six,seven months wasobserved with transmission electronmicroscope and compared to its normotensive control strain(WKY) with the same age.Then,lanthanum tracer procedures were carried out to investigate pathological changes of the blood-retinal barrier.Results (1)In SHR the main pathological changes involved swelling of mitochondria,enlargement of endoplasmic reticula,decrease of RPE cell infolding,and sparseness of microvilli.These degenerations were more serious in older rats with higher blood pressure.(2)The breakdown of outer blood-retinal barrier with permeation of lanthanum tracers were evident in SHR aged six or seven month,however,in WKY and five-month SHR the traces were prevented from passing by tight junctions.ConclusionThe degeneration of RPE owing to ischemia and anoxia arises in early periosd of hypertensive retinopathy.The pathological changes of ultrastructure and permeability might interact with the damage of visual cells and play a main role in the hypertensive retinopathy.
Objective
To observe the morphological changes of dendrite and soma in retinal ganglion cells (RGCs) which subsisted in early diabetic rats.
Methods
The RGCs of 3-months-course diabetic rats and coeval normal rats were marked by gene gun techniques. To collect RGCs photographs by Leica microscope with Z axis and CCD camera；to observe the changes of diameter, variance of structural features in dendritic field and somata after classification which according to the size and morphology. Thy-1 antibody marks on the retinal RGCs, taking a photograph under fluorescent microscope, counting the changes of retinal RGCs density in early diabetic rat.
Results
In three-month diabetic rats,the density of retinal RGCs was decreased obviously. Morphological changes of RGCs in the dendritic fields were observed with gene gun technique. There was no severe variation in all kinds of the bole of cell dendrite, in which some only showed crispation partially and sparseness also twisting in the dendritic ramus. The mean diameter of dendritic field and soma in class A of diabetic rats was (401plusmn;86) mu;m, the mean diameter of dendritic field in control group was (315plusmn;72) mu;m,compared with each other, there is statistically significant differences (t=21.249，Plt;0.001); the mean diameter of soma in class A of diabetic rats was (24plusmn;6) mu;m, the mean diameter of soma in control group was (22plusmn;5) mu;m, compared with each other, there is no statistically significant differences (t=0.927，Pgt;0.05); the mean diameter of dendritic field and soma in class B of diabetic rats were (170plusmn;36)、(14plusmn;2) mu;m respectively, in control group were (165plusmn;36)、(16plusmn;2) mu;m, the mean diameter of dendritic field and soma in class C of diabetic group were(265plusmn;78)、(17plusmn;5) mu;m respectively, in control group were (251plusmn;57)、(17plusmn;4) mu;m , compared with each other, there are on statistically significant differences(t=1.357，0.798，0.835，1.104，Pgt;0.05).
Conclusions
In short-term diabetes, the survived RGCs show good plasticity in adult diabetic rats, especially in class A. The changes of dendrites were more sensitive than the soma, which could be the leading index of the morphologic changes of RGCs in the early stage. The good plasticity showed by the RGCs and the time window from changing in dendrite to cell death provide us many evidences not only for the research but also for the nerve protection in clinic.
(Chin J Ocul Fundus Dis,2008,24:249-254)
ObjectiveTo investigate the medium and long-term influence of silicon oil versus heavy silicone oil on rabbit retinas.
Methods28 health standard rabbits were randomly divided into A, B and C groups, with 12, 12 and 4 rabbits respectively. All rabbits received routine vitrectomy and tamponade with silicone oil (group A), or heavy silicone oil (group B) or balanced salt solution (group C). After 4, 8, 12 and 24 weeks, the retinal b-wave amplitude was measured by ERG, posterior retinal thickness was measured by optical coherence tomography (OCT). Retinal ultrastructure and tissue morphology were observed by transmission electron microscopy and optical microscopy.
ResultsCompare to group C, the b-wave amplitude decreased at 4 weeks after surgery, and decreased at 8 weeks after surgery for group B, and decreased at 8 weeks after surgery, and decreased at 24 weeks after surgery for group A. The decreases were greater in group B than group A at 8, 12, 24 weeks after surgery, the difference was statistically significant (P < 0.05). The posterior retinal thickness of group A and B was thinner than group C at 24 weeks after surgery (P < 0.05). The decreases were greater in group B than group A, the difference was statistically significant (P < 0.05). Transmission electron microscopy and optical microscopy revealed severe pathological changes of retinal ultrastructure and morphology in group A and B rabbit eyes, at 12 weeks and 8 weeks after surgery respectively. The changes were more severe in group B and group A, including edema and necrosis in cone/rod cells, in disk membranes, mitochondria, cytoplasm, nucleus and other organelles. The morphological changes were also more severe in group B and group A, including degenerations of ganglion cell layer, inner nuclear layer changes. Those changes became more severe when the tamponade time extended.
ConclusionThe heavy silicone influence on visual function, ultrastructures, histomorphology of rabbit retinas is much worse than the silicon oil, and the effect is more significant with its time prolong.
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.
Objective To observe the retinal ultrastructure of the human fetal at the age of 9 months, and to investigate the clinical significance of the observation on retinal neuron development during the prenatal period.Methods Four human fetal eyes of 2 fetus at the gestational age of 9 months, including 1 at 35 and the other at 36 weeks, were obtained after termination of pregnancy due to trauma. The gestational ages of the fetus were estimated according to both last menstrual period (LMP) of the pregnant women and the weight/crownheel length of fetus at the delivery. From each eyeball, 4 pieces of retina at the posterior pole were obtained and observed after specimens handling according to the procedure of routine electron microscopy. Eight pieces of retina which were randomly selected from total of 16 pieces of retina in each group were processed and observed by scanning electron microscopy (SEM) and transmission electron microscopy (TEM). Permissions from pregnant women and family members were guaranteed.Results At the gestational age of 9 months, the outer nuclear layer of fetal retina contained 5 to 6 layers of photoreceptor cells (PRC), and sphericallike membrane structures were found outside of the outer limiting membrane (OLM). Among many tightaligned inner segments of PRCs there was zonula adherens of OLM, mitochondrias at inner side of OLM, and cilium at outer side of OLM. Outer segment of PRCs were short and contained a few irregularly arranged disc membrane. Some PRC had a multishaped nucleus in which equal amount of euchromatin and heterochromatin. There were only few and thin axon branches from photoreceptor cells, and very few axons contacted with inner nuclear layer (INL) and no typical synapse was found. The INL contained 4 to 5 layers of cell bodies, in which many cellular nuclear had uneven density of euchromatin and heterochromatin; some were lobulated nucleus with clear karyotheca. In inner plexiform layer (IPL), the nerve cells had small branches, and only little connection among the synapses and few synapse structures were found. Although not many retinal ganglion cells (RGC) existed,RGC had both intact cell membrane and some rough endoplasmic reticulum (RER).The karyotheca of RGC had double-layers structures, and the nucleus was mainly consisted of euchromatin. Internal limiting membrane (ILM) had doublelayer membrane structures, and the wellarranged nerve fiber layer was located at the outer side of ILM, with some micropores on the surface. Conclusions At the gestational age of 9 months, all layers of the human retinal has been formed, but some cell structure and cell connections are not yet mature, suggesting that at this time of period, human retina is still at an important stage of developing and remodeling.
ObjectiveTo investigate relationship between ultrastructural changes and expression of basic fibroblast growth factor of diabetic retinopathy in rats.MethodsDiabetes was induced in rats with a single injection of streptozotocin (STZ) and divided into normal control group and 1- , 3- and 5- month diabetes group. The paraffin slide was observed by in-situ hybridization and immunohistochemistry, and retinal ultrastructure was examined by transmission electron microscopy.ResultsNo change of retinal ultrastructure was found in the control group. Different degrees of ultrastructure lesion were found in 1-month diabetic rats with fragmental increase of thickness of basement membrane, swelling of endothelial cells and obvions fingerlike processes in the capillary cavity, disconcentration of heterochromatin both in endothelium and pericyte, and swelling and degeneration of mitochondrion. The edema of endothelial cells of 3-month diabetic rats was more serious than that of 1month ones, and the capillary cavity was nearly occluded. In 5-month diabetic rats, the basement membrane was unevenly thickened, or obviously split. The positive rate of in-situ hybridization in 3-month diabetic rats was 77.8% while the positive rate of immunohistochemical stain was 55.6%, which increased to 88.9% in 5-month diabetic rats.ConclusionsThe occurrence of the ultrastructural changes in STZ rats with diabetic retinopathy is earlier than that of the expression of bFGF.(Chin J Ocul Fundus Dis, 2003,19:348-351)
Objective
To observe the enzymic histochemical and ultrastructral changes of cryopreserved human retina.
Methods
To compare the activity of lactate dehydrogenase (LDH)， succinate dehydrogenase (SDH) and ATPase in cryopreserved retina with those in fresh retina and to observe the histological and ultrastructural changes of cryopreserved retina.
Results
There was no statistical difference between the activity of LDH，SDH and ATPase in fresh and in cryopreserved retina. Histologically, in the cryopreserved retina, fluid in neural fiber and outer plexiform layers, as well as in cone and rod layer, was sligthly more than normal. The ultrastructure is normal except that the mitochondria was swollen in different degree.
Conclusion
Cryopreservation may be an effective method for keeping the retinal cells alive for a long period and might free the transplantation from dependance on aviability of fresh dornor tissue.
(Chin J Ocul Fundus Dis，2000，16：139-212)
ObjectiveTo investigate the histological characteristics of autogenous hamstring grafts after anterior cruciate ligament (ACL) reconstruction.MethodsThe patients who underwent arthroscopic single-bundle ACL reconstruction with autogenous hamstring tendons and were followed up at least 4 years and also underwent second-look arthroscopy between March 2017 and December 2017 and met the selection criteria were considered for enrollment. Graft quality under arthroscopy was evaluated as good remodeling group (GRG, the total scores were 4-6) and poor remodeling group (PRG, the total scores were 1-3) according to synovial and vascular coverage, the apparent tension of the grafts, the thickness and retear of the grafts. During the second-look arthroscopic procedures, ACL graft biopsies were performed. Normal ACL tissues harvested from the patients under 60 years old who underwent total knee arthroplasty were designated as normal controls. Graft vascularity, cellular morphology, cellular metabolism, and collagen fibril distribution were analyzed.ResultsThe 18 specimens (11 cases of GRG group and 7 cases of PRG group) and 9 native ACL biopsied tissue sample were enrolled into the study. Arthroscopy scores were 2-6 (mean, 4.7). The biology under light microscopy of GRG group was similar to that of native ACL in control group. There was no significant difference in the scores of graft vascularity and cellular morphology between GRG group and control group (P>0.05), while PRG group was significantly lower than the other two groups (P<0.05). Transmission electron microscope evaluation showed that GRG group and control group had better collagen fibril distribution and lower levels of cellular metabolism than PRG group (P<0.05). There was no significant difference in cellular metabolism between GRG and control groups (P>0.05), while collagen fibril distribution score of GRG group was significantly lower than that of control group (P<0.05).ConclusionWhile good remodeling grafts under arthroscopy in histological maturation period was proved to be more similar to normal ACL on ultrastructure properties under light and electron microscope, ultra structural differences regarding collagen fibril distribution still persist.
