Objective To explore the effects of Neurogenesin 1 (Ng1) gene on functional recovery after spinal cord injury (SCI) and its mechanism. Methods Thirty-six rats (aging 4 months, weighing 230 g and being male or female), were randomly divided into two groups: experimental group (n=18) and control group (n=18). After spinal cord contusive injury at T10 level was made in all these rats using modified Allen’s method, Ng1 recombinant plasmid and blank plasmid were transfectedinto the damaged areas of exprimental group and control group respectively by Alzet pumps. At 1 day, 1 week, 2 weeks, 3 weeks, and 4 weeks after SCI, Basso-Beattle-Bresnahan (BBB) Rating Scale was used to observe the recovery of motor function. At 1 week after injury, the expressions of Ng1 mRNA and protein in injured spinal cord were detected by RT-PCR and Western blot techniques. And at 2 and 4 weeks, double immunofluorescence and histopathologic examinations were performed to study the prol iferation of the adult endogenous neural stem cells and pathological change after SCI. Results At 1-4 weeks after SCI, the BBB scores in the exprimental group was significantly higher than that in control group (P lt; 0.05), and at 4 weeks the BBB score of the experimental group (16.80 ± 1.79) was significantly higher than that of the control group (9.60 ± 1.67), (P lt; 0.01). RTPCR and Western blot showed that the mRNA and protein expressions of Ng1 were observed in the exprimental group and no expression was seen in the control group. Histologic observation showed that the morphology of spinal cord and neurons in the exprimental group was better than that in the control group and was close to the normal tissue. The mean number of Nestin+/ BrdU+ newborn endogenous neural stem cells in the exprimental group was significantly more than that in control group (P lt; 0.05). Conclusion Ng1 gene could promote the prol iferation of endogenous neural stem cells and protect the injured neurons, which enhances the repair of the motor function after SCI.
Epigenetic modifications such as DNA methylation, histone post-translational modifications, non-coding RNA are reversible, heritable alterations which are induced by environmental stimuli. Major risk factors of diabetes and diabetic complications including hyperglycemia, oxidative stress and advanced glycation end products, can lead to abnormal epigenetic modifications in retinal vascular endothelial cells and retinal pigment epithelium cells. Epigenetic mechanisms are involved in the pathogenesis of macular edema and neovascularization of diabetic retinopathy (DR), as well as diabetic metabolic memory. The heritable nature of epigenetic marks also playsakey role in familial diabetes mellitus. Further elucidation of epigenetic mechanisms in DR can open the way for the discovery of novel therapeutic targets to prevent DR progression.
ObjectiveTo identify and observe disease-causing gene variants and clinical phenotypes in a Han Chinese family with Leber congenital amaurosis (LCA). MethodsA retrospective study. A patient with LCA10 and his parents who had presented at Department of Ophthalmology of Henan Provincial People's Hospital on May 2022 were selected as the study subject. Detailed medical and family histories were recorded, fundus photography and flash electroretinogram (F-ERG) were performed. Peripheral venous blood samples (3 ml) of the proband and his parents were collected to extract whole genomic DNA, then whole exome sequencing (WES) and mitochondrial DNA (mtDNA) sequencing were carried out for the proband to determine the disease-causing gene and variants. All variants were annotated by bioinformatics analysis. According to the American College of Medical Genetics and Genomics (ACMG) guidelines, the pathogenicity of all detected variants were evaluated. Candidate variants were verified by Sanger sequencing, and in vitro minigene assay were performed to evaluate the impact of the missense variant with insufficient evidence on mRNA splicing. ResultsThe proband, male, 7-month-old, presented with an inability to follow light or objects, eye poking, photophobia, nystagmus, partial loss of retinal pigment epithelium around the fovea of the macula. At the age of 2 years old, F-ERG revealed severe reduction, elongation, or even no waveform of a-wave and b-wave in both eyes. No obvious abnormality was found in the clinical phenotype of his parents. The result of WES revealed that the proband carried two variants in exon 40 and exon 2 of CEP290, a frameshift variant c.5515_5518del (p.Glu1839Lysfs*11) (V1) and a novel missense variant c.74C>T (p.Ala25Val) (V2), respectively. The result of mitochondrial DNA sequencing was negative. Sanger sequencing confirmed that the heterozygous frameshift variant was inherited from his father and the heterozygous novel missense variant was inherited from his mother, which constituted compound heterozygous variants. In vitro minigene splicing assay confirmed that V2 created a new splicing donor at exon 2, leading to the in-frame deletion of 30bp fragment during transcription and loss of 10 amino acid residues in the protein. The two variants were pathogenic (V1) and likely pathogenic (V2) based on ACMG guidelines, respectively. ConclusionsThe c.5515_5518del and novel c.74C>T compound heterozygous variants of the CEP290 gene probably are the cause of LCA10 in this family, which lead to the production of a truncated protein and aberrant splicing of pre-mRNA, respectively. LCA is characterized by early onset, severe impairment of visual function, and a wide range of disease-causing variations.
Objective lt;brgt;To evaluated the effect of transpupillary thermotherapy (TTT) on age-related macular degeneration (AMD). lt;brgt; lt;brgt;Methods lt;brgt;Sixty-two cases (62 eyes) of exudative AMD were managed with TTT. Before treatment, 58 cases underwent fundus fluorescein angiography(FFA),42 cases underwent simultaneous indocyanine green angiography (ICGA), and 56 cases underwent optic coherence tomography (OCT).TTT was delivered using a 810 nm diode laser with variable spot sizes 0.5-3.0 mm and power range 60-40 mW,60 seconds duration. Sixty-two cases were followed up for 1-10 months with 4.8 months average. lt;brgt; lt;brgt;Results lt;brgt;The visual acuities of last visit were compared with those before the treatment. The visual acuity was unchanged in 43 cases (69.3%), improved in 15 cases (24.2%), and declined in 4 cases (6.5%). OCT was re-done in 51 cases and compared with OCT images before TTT treatment. The height of macular edema was unchanged in 29 cases (56.9%), decreased in 18 cases (35.3%), and increased in 4 cases (7.8%). The amelioration of visual acuity was compatible with that of macular configuration in the majority of cases (74.5%). Only in 13 cases (25.5%) the amelioration of visual acuity lagged behind that of macular configuration. The re-treatment was performed in 18 cases (29.1%), probably due to insufficiency of laser power. No side-effect was found. lt;brgt; lt;brgt;Conclusion lt;brgt;TTT makes most of the cases of exudative AMD retaining or improving their visual acuity. The employment is secured. Further exploration is needed in order to obtain the parameters of the laser treatment. (Chin J Ocul Fundus Dis, 2002, 18: 180-183)
PURPOSE:To investigate the spatial and temporal relation of fibronectin(Fn),basic fibroblast growth factor(b-FGF)and astrocytes with the retinal vascular developmemt of human fatuses. METHODS:The retinas of 86 human fetuses from 13th week to 40th week were studied by immunohistochemical methods and light microscopy. RESULTS:Fn immunoreativity was localized in spindle cells ,vascular endothelial cells and extracellular matrix ahead of the spindle cells,vascular endothelial cells,ganglion cells and cone cells were b-FGF immunopositive. The b-FGF immunoreactivity in ganglion cells and cone cells appeared earlier than the vascularization nearby.Astrocytes migrated to ora serrata in close association with the spindle cells.and sent numerous processes to ensheath the blood vessels formed in two processes of retinal vascuiarlzation. CONCLUSION:These results suggest that Fn ,b-FGF and astrocytes were involved in modulating both of two processes of retinal vascularizalion. （Chin J Ocul Fundus Dis,1996,12:180-182）
Objective To analyse the indocyanine green angiographic findings in contralateral eyes of patients with unilateral exudative age-related macular degeneration(AMD). Methods Fundus photograph,fundus fluorescein angiography(FFA) and indocyanine green angiography(ICGA) were performed in a series of 70 patients with unilateral AMD and drusens and pigmentary changes in the macular region in contralateral eyes.The findings of fluoroangiograms were observed and analysed. Results ICGA revealed the characteristics of the contralateral eyes as follows:(1)Drusen could be hypofluorescent,hyperfluorescent or normal fluorescent;(2)14 eyes revealed plaque-like late hyperfluorescent;(3)13 eyes revealed choroidal filling defect;(4)18 eyes revealed pindot-like clusters of late hyperfluorescence. Conclusion ICGA is useful in evaluating the lesions and circulation disturbance of the contralateral eye,and may help to find the risk factors of developing future exudative changes. (Chin J Ocul Fundus Dis, 1999, 15: 216-218)
Objective To investigate the relationship between diabetic retinopathy (DR) and insertion/deletion (a/b) polymorphism of a 27 base pair variable number tandem repeat (VNTR) in intron 4 of the endothelial nitric oxide synthase (eNOS) gene. Methods 321 patients of type 2 diabetes mellitus with over 10 years duration (case group) and 146 normal subjects (control group) were enrolled in this study. All the clients are Han Chinese. The case group was divided into DR subgroup (154 patients) and non-DR (NDR) subgroup (167 patients) according to the results of indirect ophthalmoscope and fundus fluorescent angiography. The VNTR polymorphism in eNOS gene was determined by polymerase chain reaction (PCR) combined with 8% agarose gel electrophoresis. Then the b, a allele frequency and b/b, a/a, b/a allele frequency of two groups were compared, and its correlation with diseases were analyzed. Results The b allele frequency of the VNTR in intron 4 of eNOS gene in the DR group was significantly higher than that in the NDR group（chi;2=4.745，P=0.029；OR=1.685，95%CI=1.050-3.905）and control group（chi;2=6.958，P=0.008；OR=1.891，95%CI=1.172-4.437）; b/b allele frequency in the DR group was also significantly higher than that in the NDR group（chi;2=4.811，P=0.028；OR=1.790，95%CI=1.060-4.645）and control group（chi;2= 5.203，P=0.023；OR=1.859，95%CI=1.087-4.952）. Conclusions The b allele and b/b genotype in intron 4 of eNOS gene in the Han Chinese are closely related to DR.
Objective To study the expression and clinic significance of nm23 gene (product of uncleoside ciphosphate kinase) in the pancreatic adenocarcinoma tissues. MethodsSP immunohistochemical method was used to examine the expression of nm23/NDPK in 40 pancreatic adenocarcinoma and 14 normal pancreas tissues.ResultsTwentysix of 40(65%) pancreatic adenocarcinoma showed b immunoreactivity for NDP kinase, whereas 4 of 14 (28.5%) normal pancreatic tissues showed weak immunoreactivity. Significant difference was found between the two groups (P＜0.05). The nm23/NDPK expression levels in pancreatic adenocarcinoma of lower differentiation was higher than those in pancreatic adenocarcinoma of higher differentiation (10/11，90.9%; 2/8, 25%; P＜0.05). Positive staining was associated with higher incidence of lymph node metastasis (10/14, 71.4%) than negative staining (6/19,31.5%, P＜0.05). These results suggested that nm23/NDPK expression was positively associated with lymph node metastasis and aggressiveness. They also suggested that nm23/NDPK expression had negative correlation with the extent of histologic differentiation. Conclusion nm23/NDPK can serve as a marker for malignant potentiality and indicate the prognosis of pancreatic adenocarcinoma.
Objective To explore effects of several immunosuppressants on cytokine expressions after repair for a sciatic nerve injury in a rat model. Methods The sciatic nerves of 42 rats were cut and suturedend to end. After operation, the rats were divided into 6 groups. Group A(n=9) was served as a control with no medicines given. Group B (n=9) was given methylprednisolone 20 mg/(kg·d) for 2 days. Groups C(n=9) and D(n=3) were given FK506 1 mg/(kg·d) for 2 weeks and 4 weeks respectively, and were given the same doses of methylprednisolone as Group B. Groups E and F were given CsA 2 mg/(kg·d) for 2 weeks and 4 weeks respectively, and were given the same doses of methylprednisolone as Group B. The sciaticnerves were sampled at 1, 2 and 4 weeks postoperatively. And immuneohistochemistry stainings of interleukin 1β(IL-1β)， tumor necrosis factor α(TNF-α), interferon γ(IFN-γ) and macrophage migration inhibitory factor(MIF) were performed. The staining results were compared and analyzed. Results The expression peaks of IL-1β and IFN-γ were found at the 1st week postoperatively in Group A. Then, the expression decreased rapidly at the 2nd week and disappeared at the 4th week. As for TNF-α and MIF, they were only found to have a low expression until the 1st week in Group A. In groups C-F, the expression peaks of IL-1β, TNF-α and IFN-γ were found at the 2nd week, while the expression peak of MIF was still at the 1st week, and the expression of all the cytokines extended to the 4th week. The expressions of these cytokines in Group B were just between the expression levels of Group A and Groups C-F. Conclusion Immunosuppressants can delay the expression peaks and significantly extend the expression time of IL-1β, TNF-α, IFN-γ and MIF after repair for a sciatic nerve injury in a rat model.
OBJECTIVE: To study the effect of simvastatin on the expression of bone morphogenetic protein-2 (BMP-2) and alkaline phosphates (ALP) activity in the primary cultured bone marrow stromal cells, and to elucidate the mechanism of the anabolic osteogenetic effect of simvastatin. METHODS: Bone marrow stromal cells in femur and tibia of adult mouse were cultured in vitro. after treated with different concentrations of simvastatin (0, 0.1, 0.2, 0.5 and 1.0 mumol/L) or recombinant human BMP-2 for 72 hours, ALP activity of bone marrow stromal cells was determined. BMP-2 expression of bone marrow stromal cells was analyzed by using immunocytochemistry and Western blotting. RESULTS: After treated with simvastatin for 72 hours, BMP-2 expression increased, while little BMP-2 expression could be observed in the control group. ALP activity also increased in a dose-dependent manner; t-test showed that ALP activity in the group which concentrations of simvastatin were 0.5 mumol/L (t = 2.35, P = 0.041), 1.0 mumol/L (t = 2.348, P = 0.041) had significant difference when compared with control group. CONCLUSION: Simvastatin lead to high expression of BMP-2 in bone marrow stromal cells, via the increased auto- or para-crine of BMP-2, and ALP activity increased. These may be parts of the mechanism on the anabolic osteogenetic effect of simvastatin.