Objective To investigate the effect of exogenous erythropoietin (EPO) on the denervated muscle atrophy. Methods Twenty-four SD male rats, weighting 200-220 g were made the models of denervated gastrocnemius muscle after sciatic nerves were transected under the piriform muscle at the right lower leg, and were randomly divided into two groups (n=12). rhEPO (2 500 U/kg) was injected daily into the denervated gastrocnemius muscle in EPO group, and normal sal ine was injected into the denervated gastrocnemius muscle in control group. To observe the general state of health of the experimental animal, the muscle wet weight, the muscle cell diameter, the cross section area, the protein amount, thepercentage of the apoptotic muscle cells, and the Na+-K+-ATPase and Ca2+-ATPase activities were measured 2 and 4 weeks after operation. Results All experimental animals were survived during experiment without cut infection, and all animals could walk with pull ing the right knee. At 4 weeks after operation, 7 cases showed ulcer in the right heel, inculding 5 in the control group and 2 in the EPO group. At 2 and 4 weeks after operation, the muscle wet weight in EPO group was (885.59 ± 112.35) and (697.62 ± 94.74) g, respectively; in control group, it was (760.63 ± 109.05) and (458.71 ± 58.76) g, respectively; indicating significant differences between two groups (P lt; 0.01). The protein amount in EPO group was (77.37 ± 5.24) and (66.37 ± 4.87) mg/mL, respectivly；in control group, it was (65.39 ± 4.97) and (54.62 ± 6.32) mg/mL；indicating significant differences between two groups (P lt; 0.01). At 2 and 4 weeks after operation, the myofibrillar shapes were nearly normal in EPO group while there were muscle fiber atrophy, some collapse and obviously hyperblastosis between muscle bundle. There were significant differences in the muscle cell diameter and the cross section between two groups (P lt; 0.01). However, the percentage of the apoptotic muscle cells was 11.80% ± 1.74% and 28.47% ± 1.81% in control group, respectively, which was significantly smaller than that in EPO group (21.48% ± 2.21% and 55.89% ± 2.88%, P lt; 0.01). At 2 and 4 weeks after operation, Na+-K+-ATPaseand Ca2+-ATPase activities in EPO group were higher than those in control group (P lt; 0.01). Conclusion EPO can delay the denervated muscle atrophy.
ObjectiveTo observe the peripapillary atrophy (PPA) and peripapillary choroidal vascularity index (CVI) in patients with different degrees of myopia and to analyze their correlations. MethodsA cross-sectional clinical study. From September 2021 to December 2021, 281 mypoic patients of 281 eyes treated in Eye Hospital of Wenzhou Medical University at Hangzhou were included in this study, and the right eye was used as the treated eye. There were 135 eyes in 135 males and 146 eyes in 146 females. The age was 28.18±5.78 years. The spherical equivalent refraction (SE) was －5.13±2.33 D. The patients were divided into three groups: low myopia group (group A, －3.00 D ＜SE≤－0.50 D), moderate myopia group (group B, －6.00 D≤SE≤－3.00 D);high myopia group (group C, SE＜－6.00 D). The spherical equivalent refraction was statistically different among the three groups (H=241.353, P＜0.05). All of the affected eyes were examined by swept-source optical coherence tomography. Combined with B-scan image,assessment and area measurement of β area, γ area (β-PPA and γ-PPA) were carried out on the en-face image. After binarization of the collected images, the nasal, superior, temporal and inferior CVI of the optic disc were calculated. For comparison between groups, one-way ANOVA was used for continuous variables with normal distribution, Kruskal-Wallis test was used for continuous variables with abnormal distribution, and categorical variables were used χ2 inspection. Linear regression analysis was used for the relationship between β-PPA and γ-PPA area and peripapillary CVI of different regions. Linear regression analysis was used to evaluate the relationships between the area of peripapillary atrophy and peripapillary choroidal vascularity index in different regions. ResultsThere was no statistical difference in the incidence of β-PPA among the three groups (χ2=4.672, P=0.097). The incidence of γ-PPA in group A was lower than that in group B anc C, and the difference was statistically different (χ2=33.053, P＜0.001), in which both group A was lower than group B and C. Among the three groups, the area of β-PPA and γ-PPA was statistically significant (H=36.535, 39.503; P＜0.001, 0.001); the β-PPA area of group A and B was lower than that of group C; the γ-PPA area was group A＜group B＜group C. Peripapillary CVI of different regions in group A, group B and group C was statistically significant (F=11.450, 5.037, 6.018, 4.489; P＜0.05). The temporal CVI in group C was lower than that in group A and B; The inferior CVI of group C was lower than that of group A, and the superior and nasal CVI of group B and C were lower than that of group A. In multivariate analysis, SE (β=0.374, P＜0.001), temporal CVI (β=－0.299, P＜0.001) were correlated with the area of β-PPA (adjusted R2=296, P＜0.001); AL (β=0.452, P＜0.001), temporal CVI (β=－0.220, P＜0.001) were correlated with the area of γ-PPA (adjusted R2=0.309, P＜0.001). ConclusionsThe incidence and area of γ-PPA are increased in the higher degree of myopia group. The area of γ-PPA is positively correlated with the axial length, and both the area of β-PPA and γ-PPA are negatively correlated with temporal CVI.
Leber hereditary optic neuropathy is an optic neuropathy associated with mitochondrial DNA. The disease affects young men mainly, which is considered to be due to denaturation of the retinal nerve ganglion cell and axonal loss of optic nerve, leading to optic atrophy. Nowadays, there are some development in studying Leber hereditary optic neuropathy by optical coherence tomography (OCT) and optical coherence tomography angiography (OCTA). It is great help to know the disease, forecast the progression of disease, and take action on intervention. In addition, there is a lack of in-depth study on OCT and OCTA characteristics among different mutation sites of LHON, different genders of the same site, different families of the same site or even different branches of the same family. It is expected to be improved in the future work.
Objective To find the new mutations of Leber's hereditary optic neuropathy (LHON). Methods Two LHON families were enrolled in this study. The probands and all maternal members in this two families were underwent ophthalmologic examinations. The ages of probands were seven and 14 years old respectively. A total of 358 healthy adults were enrolled in this study as control group. The genomic DNA from whole blood of participants were extracted. The entire mitochondrial genome of probands were PCR amplified and sequenced in 24 overlapping fragments using primers as designed. At the same time, the mtDNA of maternal relatives and 358 controls were also detected. Fourteen primate species were selected from GenBank to analyzed the phylogenetics of mitochondrial sequence. Results There was no ND4 G11778A, ND1 G3460A, ND6 T14484C mutational site in all maternal members. Molecular analysis of mtDNA in this two families identified the homoplasmic tRNAGluA14683G mutation and distinct set of variants belonging to the Asian haplogroup F1a1 and G2. The site was at theTpsi;C stem oftRNAGlu and extremely conserved among 14 primate species. It was anticipated that the A14683G increased the highly conserved C-G basepairing. Furthermore, the A14683G was absence in control group. Conclusion The tRNAGluA14683G mutation is likely a new mutation associated with LHON.
Objective To study the effect of the competitive inhibitor of nitric oxide synthase NG-nitro-L-arginine methyl ester (LNAME) on thedenervated muscle atrophy. Methods A model of the denervated gastrocnemius atthe right lower limb was established in 36 SD adult rats. The rats were randomly divided into two groups: the L-NAMEgroup (Group A) and the control group(Group B). L-NAME 10 mg/ kg daily was injected into the denervated gastrocnemius inGroup A, and normal saline was injected into the denervated gastrocnemius in Group B. At 2, 4 and 8 weeks after operation, the rate of the muscle wet weight preservation, the cross section area of the myocyte, the protein amount, and the percentage of the apoptotic muscle cells were measured respectively and the ultramicrostructure of the myocyte was observed. Results At 2 and 4 weeks after operation, the rate of the muscle wet weight preservation, the cross section area of themyocyte, and the protein amount were significantly greater in Group A than in Group B; however, the percentage of the apoptotic muscle cells was significantly smaller in Group A than in Group B. The observation of the ultramicrostructure of themyocyte showed that an injection of L-NAME could protect the ultramicrostructure of themyocyte. At 8 weeks after operation, there was no significant difference between the two groups in the abovementioned parameters. Conclusion The nitric oxide synthase inhibition can delay the denervated muscle atrophy.
ObjectiveTo investigate the feasibility of immoribund skin fibroblast cell line derived from Leber′s hereditary optic neuropathy (LHON) patients as a cell model. MethodsA basic research. Two LHON patients and 2 healthy volunteers were recruited from Department of Ophthalmology of Genetic Clinic of Henan Provincial Eye Hospital. The skin tissue of participants was obtained, and the 4 immortalized skin fibroblasts were constructed by SV40 virus infection, including 2 LHON patient cells (LHON-1 and LHON-2 cells) and 2 healthy volunteers cells (NC-1 and NC-2 cells). Mitochondrial morphology in cells was observed by electron microscope. The levels of reactive oxygen species (ROS), nicotinamide adenine dinucleotide-oxidation state (NAD+), nicotinamide adenine dinucleotide-reduction state (NADH) and adenosine triphosphate (ATP) in fibroblasts were detected. Cellular oxygen consumption was measured by seahorse mitochondrial pressure assay. Cell viability was detected using cell counting kit-8 (CCK8). One-way ANOVA was performed to compare the levels of ROS, NAD+, NADH and ATP in LHON and NC cells, as well as basal oxygen consumption, maximal oxygen consumption, ATP-coupled oxygen consumption, and cell viability. ResultsCompared with NC-1 and NC-2, the number of mitochondrial crest in LHON fibroblasts was significantly reduced, indicating abnormal mitochondrial morphology. Biochemical analysis showed that ROS levels in LHON cells increased, but NAD+/NADH and ATP levels decreased, and the oxygen consumption was significantly inhibited, indicating the presence of mitochondrial damage and respiratory dysfunction. The results of CCK-8 detection showed that the survival ability of LHON-1 and LHON-2 cells was worse under stress conditions. ConclusionImmortalized skin fibroblast cell lines from LHON patients presented mitochondrial dysfunction.
Leber's hereditary optic neuropathy (LHON) is a rare hereditary optic nerve disease. At present, the understanding of its etiology and pathogenesis is relatively clear. With the emergence of new drugs such as idebenone and the possibility of gene therapy for LHON, it has brought hope for patients to recover. However, because genetic testing technology has not been widely developed in China, clinical misdiagnosis of LHON as optic neuritis still occurs from time to time. How to make timely identification and correct diagnosis of LHON still poses certain challenges for Chinese ophthalmologists. In addition, in terms of treatment, the choice of treatment methods and treatment costs in the pre-onset (gene mutation carriers) and different periods after the onset of LHON are also huge challenges for patients and their families.
Objective To investigate the characteristics and risk factors of optic nerve atrophy in eyes with complicated retinal detachment after silicone oil tamponade during the procedure of vitreoretinal operation. Methods The clinical data of 97 patients with complicated retinal detachment who had optic nerve atrophy after silicone oil tamponade during the procedure of vitreoretinal operation were an alyzed retrospectively. Logistic regression analysis by SPSS statistical software was used to analyze the factors like age, disease history, primary diseases, preoperative ocular condition, complications in and after the operation, the time taking out the silicone oil, and emulsification of the silicone oil, and Ple;0.05 was considered to be the symbol of significant difference. Results All of the affected eyes had optic discs with clear border, including paler optic disc in 65 eyes, pale one in 21 eyes, and paler optic disc with enlargement of the cup/disc (ge; 0.6) in 11 eyes. The result of logistic regression analysis showed that the intraocular pressure (P=0.022) and the visual acuity (P=0.001) during the silicone oil removal were in the equation. Conclusion The risk factor of optic nerve atrophy is the chronic increase of intraocular pressure after silicone oil tamponade. (Chin J Ocul Fundus Dis, 2006, 22: 305-307)
Objective To investigate the effect of the neuromuscular pedicle transplantation in prevention against atrophy in the denervated muscle. Methods Fortyeight SD rats were used to establish the right side tibialis anterior muscle denervation model. The long peroneal muscle neuromuscular pedicle was made as a treatment in 12 rats (Group A); the nerve shaft embedding was used in 12 rats (Group B); no treatment was used in 12 rats(Group C); the remaining 12 rats were used as normal controls (Group D). The gait analysis, electromyogram，muscle wet weight, and muscle fiber crosssectional area were used to determine and compare the effect of the operation at 6 and 12 weeks postoperatively. ResultsAt 6 weeks postoperatively, the parameters tested in Group A about the gait analysis (peroneal function index, PFI, －47.20±12.30), electromyogram, muscle wet weight (0.384 0±0.024 6 g)and muscle fiber cross-sectional area (1 040.98±120.54 μm2) were significantly better than those in Group C (PFI, -114.40±14.84; muscle wet weight, 0.173 0±0.019 1 g; muscle fiber cross-sectional area, 585.08±182.93 μm2,Plt;0.05), and the final two parameters were significantly better than those in Group B (0.294 0±0.056 4 g,763.92±82.68 μm2,Plt;0.05). At 12 weeks postoperatively, the musclefiber crosssectional area in Group A(1 360.10±261.45 μm2) had no significant difference from that in Group D (1 544.57±266.92 μm2，Pgt;0.05)，and most of the parameters tested in Group A were better than those in Groups B and C. Conclusion Neuromuscular pedicle transplantation has an excellent effect in prevention against atrophy in the denervated muscle, and the effect of neuromuscular pedicle transplantation is better than that of the nerve shaft embedding.
Objective To study the quantitative changes of ubiquitin l igase MAFbx mRNA and protein expression, muscle atrophy and muscle function following free muscle transplantation and to explore relationshi ps among them. Methods Thirty-six female SD rats, SPF grade, weighing (250 ± 25) g, were used. One hind l imb of the rat was randomly selected as experimental side to receive in situ free gracil is muscle transplantation, and the counterlateral hind l imb underwent no operation serving as control side. General condition of the rats was observed after operation. Muscle contractivecapacity and muscle wet weight maintenance rate of the experimental and the control side were detected 1, 2, 4, 10, 15, and 30 weeks after operation, and 6 rats were killed at each time point. Meanwhile, HE staining was performed to observe muscle fibre cross-sectional area, real-time quantitative PCR was appl ied to detect relative expression of MAFbx/Atrogin-1 mRNA, and Western blot test was used to observe MAFbx protein expression. Results All rats survived till the end of the experiment, all incisions healed well, and no dysfunction occurred in the experimental sides. The value of muscle contractive capacity, muscle wet weight maintenance rate, muscle’s maximal force of single contraction, and muscle’s maximal force of tetanic contraction in the experimental sides dramatically decreased in the first 4 weeks after operation and increased gradually over 4 to 30 weeks. The MAFbx mRNA expression of the experimental sides peaked and was seven times greater than the control sides 2 weeks after operation, then the value gradually decreased over 15 to 30 weeks after operation and was 1.1 to 1.5 times greater than the control sides, and significant difference was evident between the experimental sides and the control sides at each time point (P lt; 0.05). Significant difference was evident between the experimental sides and the control sides in terms of MAFbx protein expression of the muscle 1 to 15 weeks after operation according to the Western blot result (P lt; 0.05), and no significant difference was noted at 30 weeks (P gt; 0.05). The correlation coefficient between muscle wet weight maintenance rate and muscle’s maximal force of single contraction maintenance rate was 0.95, between muscle wet weight maintenance rate and muscle’s maximal force of tetanic contraction maintenance rate was 0.75, between muscle fibre cross-sectional area recovery rate and muscle’s maximal force of single contraction maintenance rate was 0.93, and between muscle fibre cross-sectional area recovery rate and muscle’s maximal force of tetanic contraction maintenance rate was 0.68 (P lt; 0.05). The correlation coefficient between MAFbx mRNA expression and the parameter of muscle wet weight maintenance rate, muscle fibre cross-sectional area recovery rate, muscle’s maximal force of single contraction maintenance rate, and muscle’s maximal force of tetanic contraction maintenance rate was — 0.62 (P lt; 0.05), — 0.45 (P gt; 0.05), — 0.72 (P lt; 0.05) and — 0.78 (P lt; 0.05), respectively; the correlation coefficient between MAFbx protein relative expression and the parameter of muscle wet weight maintenance rate, muscle fibre cross-sectional area recovery rate, muscle’s maximal force of single contraction maintenance rate, and muscle’s maximal force of tetanic contraction maintenance rate was — 0.95 (P lt; 0.05), — 0.82 (P lt; 0.05), — 0.89 (P lt; 0.05), and — 0.54 (P gt; 0.05), respectively. Conclusion Decrease of muscle function after transplantation correlates closely with muscle atrophy. The high expression of MAFbx mRNA and protein, especially their persistent increases from 4 to 15 weeks after nerve reinnervation, is a junction between the muscle atrophy and thedecrease of muscle function.