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)
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 investigate whether recombinant human serum albumin (rHSA) can replace traditional B27 as a basic medium for differentiation of human pluripotent stem cells (hPSCs) into cardiomyocytes. Methods hPSCs were seeded at a cell density of 1.2×104/cm2; until up to 75% confluency hPSCs were induced by differentiation medium containing various concentration of rHSA (0, 50, 100, 200 g/L). Light microscope and fluorescence microscope recorded the whole process of stem cells differentiating into myocardium. Flow cytometry was used to detect the cardiac differentiation efficiency at different concentrations of rHSA. Immunofluorescence staining was used to detect the cardiac specific protein α-actinin and troponin T (cTnT) and electron microscope to observe the ultrastructure of human pluripotent stem cell-derived cardiomyocytes (hPSC-CM) and beating rates of hPSC-CMs response to drugs. Results A large number of spontaneous beating cardiomyocytes were observed 9 days after induction and differentition. The percentage of colonies showing beating cardiomyocytes was 60.4% at the concentration of 200 g/L of rice derived-rHSA. Beating cardiomyocytes were α-actinin and cTnT positive. Ultrastructural analysis showed scattered sarcomeres and mitochondrial. hPSC-CMs were dose-dependent on isopropyl adrenaline and verapamil. Conclusion Using such simple media to differentiate hPSCs into functional cardiomyocytes is cost-effective and highly efficient, and can be used in the clinical research.
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.
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.
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.
ObjectiveTo evaluate the correlation between the Mohawk (MKX) expression level and the collagen fiber diameter of autologous hamstring tendon graft during the stable graft remodeling phase after anterior cruciate ligament (ACL) reconstruction.MethodsBetween January 2018 and August 2018, patients who underwent arth-roscopic single-bundle anatomical ACL reconstruction with autologous hamstring tendons for at least 48 months and also underwent second-look arthroscopy were enrolled in study. During the second-look arthroscopic procedures, ACL graft biopsies were performed from the surface of central part of the ligament. MKX expressions of ACL grafts were analysed by real-time fluorescent quantitative PCR (qRT-PCR). The ultrastructure of collagen fibers of grafts were evaluated by transmission electron microscopy, which included average diameter of collagen fibers (Dc), average diameter of large-diameter collagen fibers (DL), average diameter of small-diameter collagen fibers (DS), and large-small collagen fibers ratio (RL/S). The correlation between MKX expression level and graft collagen fiber diameter was calculated.ResultsTwenty-six patients met the selection criteria and their ACL graft specimens were enrolled in the study. The interval between ACL reconstruction and second-look arthroscopy was 52-128 months, with an average of 78.6 months. Arthroscopic graft remodeling score was 3-6 (mean, 4.8). There were 17 cases of excellent remodeling and 9 cases of fair remodeling. All ACL grafts showed typical bimodal distributions of both large-diameter collagen fibers and small-diameter collagen fibers, but the ultrastructural characteristics of the graft collagen fibers were different according to different remodeling status under arthroscopy. The DC, DL, DS, and RL/S of the graft specimens were (65.2±9.3) nm, (91.6±10.5) nm, (45.7±8.6) nm, and 0.73±0.12, respectively. The relative expression level of MKX was 1.42±0.11, which was positively linearly correlated with DC, DL, and RL/S, and the correlation coefficient was statistically significant (r=0.809, P=0.000; r=0.861, P=0.000; r=0.942, P=0.000), while there was no significant correlation between DS and relative expression level of MKX (r=0.147, P=0.238). Regression analysis showed that the relative expression level of MKX could predict the DC, DL, and RL/S results of the ACL graft specimens (P<0.05).ConclusionAfter autologous hamstring tendon grafts stepped into stabilized remodeling phase, MKX expression level could predict the diameter measurement results of collagen fibers and be used as an important evaluation basis for graft collagen anabolic metabolism.
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 study on the ultrastructural characteristic of segments of photoreceptors from neonatal retinas for supporting donor retina choice of retinal transplantation． Methods Photoreceptors from neonatal calf and adult calf were analysed by scanning electron microscopy and transmission electron microscopy． Results Segments of photoreceptors from neonatal calf appeared the mushroom pattern, in which, distal end of outer segment which was ball-shaped formed the head with mushrooms appearance, and the inner segments along with some of outer segments formed the body with mushrooms appearance． Within the outer segment, plasma membranes of adjacent evaginations form a disk subsequently． The a rray of most disks were vertical to the entire length of segments, but some were parallel and slope to．Owing to the incomplete formation, some rim of disk near distal end of outer segment revealed step-shaped appearance． The distal end of outer segment displays some processes consisted of membranous discs, much vesicular material and mitochondria, much rough endoplasmic reticulum (RER) and numerous polyso mes．Segments of photoreceptor connected with outer nuclear layer via the external limiting membrane． Conclusion The typical morphol ogical structures of outer segments suggest the immature and b gowth ability of photoreceptors of the retina of neonatal calf, and therefore the competence for donor material of retinal transplantation． (Chin J Ocul Fundus Dis, 2001,227-229)
PURPOSE:Investigating on histopathologic changes of the photoreceptors in retinitis pigmentosa. METHODS:Observation of the photoreceptors of retinitis pigmentosa in 11 eyes among 9 cases using light and electron microscope. RESULTS: The pathologic changes of the photoreceptors were found to be mostly marded at the equatorial area and less at the periphery,posterior pole and macular region of the retina. In relatively early cases,degeneration and shortening of outer segments,reduction or loss of connecting cilia,stubby inner segments and swollen mitochondria Were the predominant findings. In advanced cases,the inner and outer segments and connecting cilia were diminished with reduction of nuclei in number and disarangement,cellular degeneration and disorganization. The outer limiting membrane adhered to RPE or Bruch membrane. The spaces left over by the above pathologic changes were replaced by the displaced Muuml;ller cells and their hypertrophic processes. Also there were degeneration of the RPE cells,and some of them might migrate into the retina. CONCLUSION:Obvious invasions of pathologic processes in photoreceptors of the retina did present in patients with retinitis pigmentosa. （Chin J Ocul Fundus Dis,1996,12:160-162）