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)
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 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)
In order to study the influence of biliary tract obstruction on enteromicroflora,we ligate the canine biliary tract to observe the acrobic and anerobic bacteria in the duodenum and ileum at intervals of post-ligation(the 10th,20th,30th days),and to study the pathogenesis and ultramicroscopic of the ileal mucosa at the same intervals.The results showed that:the population and species of enteroflora in small intestine gradually increased after biliary obstruction.Bacteria(especialy E.coli) ascended to the upper part of small intestine,from their normal habitant of lower part of small intestine.Therefore the radio of general aerobia and E.coli risen obviously in duodenum.The longer the obstruction,the more pathologic changes were observed in ileal mucosa.such as edema,leukocytes infiltration and destruction of epithelial villi.All of those changed may be the causative factor of biliary tract infection.So that,in the programs of preventing enterogenic infection at the state of biliary tract obstruction,the protection and adjusting of normal enteroflora should be adventently considered.
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.
OBJECTIVE: To study the characteristics of, morphology histology and ultrastructure of anterior cruciate ligament(ACL) autograft and two-step cryopreserved ACL allograft after transplantation. METHODS: Sixty New Zealand rabbits and sixty Japanese rabbits were randomly divided into two groups: ACL autograft group and two-step cryopreserved ACL allograft group. Immunosuppressant were not used after transplantation. The histology and ultrastructure of the ACL of transplantation and normal knee were observed after 4 weeks and 12 weeks, respectively. RESULTS: The rate of remodeling process was faster in ACL autograft than in two-step cryopreserved ACL allograft, but there was similar remodeling process between two groups 12 weeks after transplantation. The proportions of large-diameter fibers(gt; or = 80 nm) of ACL autograft and cryopreserved ACL allograft were 6% and 24% in the 4th week, and were 0 and 2% in the 12th week, respectively. The proportions of small-diameter of fibers(lt; 80 nm) of ACL autogrft and cryopreserved ACL allograft were 94% and 76% in the 4th week, and 100% and 98% in the 12th week, respectively. Histologic incorporation in ACL autograft was similar to that in cryopreserved ACL allograft. CONCLUSION: Two-step cryopreserved bone-ACL-bone allograft were similar to bone-ACL-bone autograft cryopreserved in remodeling process and histology. The rate of remodeling process was faster in ACL autograft than in cryopreserved ACL allograft.
Objective To investigate whether the agonist of delta opoid receptor D-Ala(2),D-Leu(5) enkephalin (DADLE) has the effect of decreasing myocardial injury during ischemia-reperfusion of adult rabbits’ myocardium，so that a new mehanism and way to myocardial protection could be found. Methods Langendorff model was used during the experiment. Thirty rabbits were divided into three groups randomly (each group 10 rabbits). Control group: St.Thomas Ⅱ cardioplegic solution was used; group 1: St.Thomas Ⅱ cardioplegic solution and DADLE (1mg/kg) were used; group 2: St.Thomas Ⅱ cardioplegic solution and naloxone(3mg/kg) were used to induce the hearts to arrest respectively. After arrest the hearts were reperfused respectively. Data of left ventricle development pressure(LVDP) was recorded before and after ischemia. Biochemical indicators of myocardium, lactate dehydrogenase(LDH) were detected before and after ischemia. Some myocardial tissues were used to explore the changes of the tissue of ultrastructure with electron microscope，when the experiment was over. Still some myocardial tissues were to be detected by flow cytometer to evaluate the apoptosis of the myocardium. Results The LDH and LVDP showed significant difference among three groups after ischemia(Plt;0.05); LVDP in group 1 was higher than those in group 2 and control group(69.8±5.8 mmHg vs. 23.4±3.9 mmHg; 69.8±5.8 mmHg vs. 37.9±4.7 mmHg; Plt;0.05), the LDH in group 1 was lower than those in group 2 and control group(1 272.6±59.1 U/L vs. 2 764.4±27.7 U/L, 1 272.6±59.1 U/L vs. 1 884.4±37.5 U/L; Plt;0.05). The apoptosis rate in group 1 was lower than those in group 2 and control group. As could be shown from the ultrastructure: mitochondria structure was nearly normal in group 1; mitochondria structure was injuried severely in group 2; there was a minor injury in control group. Conclusion Agonist of δ opoid receptor DADLE in cardioplegic solution could induce hibernation, which has myocardial protection effect during ischemia-reperfusion injury.
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 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 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)