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Medical application of recombinant protein Semaphorin3G in prevention and treatment of retinal diseases

A recombinant protein, retinopathy technology, applied in medical preparations containing active ingredients, sensory diseases, pharmaceutical formulations, etc., can solve problems such as the molecular mechanism of vascular development in cells has not yet been elucidated, and achieve improved vascular leakage, reduced area, Fading effect

Active Publication Date: 2021-01-29
NANJING MEDICAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, so far, there are still many gaps in the study of the biological function of Sema3G, especially the role of Sema3G in the development of blood vessels and its intracellular molecular mechanism in the pathological process of ischemic retinopathy have not yet been elucidated

Method used

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  • Medical application of recombinant protein Semaphorin3G in prevention and treatment of retinal diseases
  • Medical application of recombinant protein Semaphorin3G in prevention and treatment of retinal diseases
  • Medical application of recombinant protein Semaphorin3G in prevention and treatment of retinal diseases

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] Example 1 Detection of Sema3G protein content in vitreous cavity fluid of patients with proliferative diabetic retinopathy

[0037] The patients with diabetic retinopathy involved in this experiment were clinically diagnosed as proliferative diabetic retinopathy (PDR) from stage IV to stage VI by the ophthalmologists of Jiangsu Provincial People's Hospital according to the classification criteria of diabetic retinopathy formulated by the Fundus Disease Group of the Chinese Medical Association. Pathological neovascularization in the retina. Control patients had non-vascular pathological retinopathy diseases such as idiopathic macular hole (MH) or idiopathic epiretinal membrane (ERM). Vitreous cavity fluid samples from the above PDR patients and control patients were collected during vitrectomy. All subjects in this study signed a written informed consent, and the research project has been approved by the Medical Research Ethics Committee of Jiangsu Provincial People's H...

Embodiment 2

[0054] Example 2 Construction of an adeno-associated virus vector expressing Sema3G protein

[0055] (1) Gene sequence acquisition: Sema3G (NCBI, NM_020163.3) gene sequence and plasmid vector were obtained from the database website, and the Sema3G coding gene was constructed into the BR1 serotype adeno-associated virus backbone vector by using molecular cloning technology.

[0056] (2) PCR amplification and digestion: use the gene sequence-specific primers designed in Table 1.1, and use the Sema3G-encoding gene vector as a template to perform PCR amplification on the target gene sequence to obtain Sema3G gene fragments. Amplification (Gflex PCR enzyme, takara company) program: 95°C, 10min; 95°C, 10s; 60°C, 30s; 68°C, 2min; a total of 35 cycles; 72°C, 4min. The backbone vector was digested with restriction endonucleases EcoR I and Hind III (Takara Company), and reacted in a water bath at 37° C. for 2 hours for digestion.

[0057] Table 1.1

[0058] Primer name Pri...

Embodiment 3

[0064] Example 3 Packaging of adeno-associated virus expressing Sema3G protein

[0065] During the packaging process of AAV, the packaging plasmid is responsible for encoding the target gene and two inverted terminal repeats (ITR), and the helper plasmid pxx2 contains the cap (encodes the viral capsid protein) and rep (involves in the replication of the virus) genes required for AAV packaging. p179 is a helper plasmid packaged by adeno-associated virus. The gene encoding the capsid protein of the helper plasmid we used has NRGTEWD ​​mutation, which can specifically infect the vascular endothelial cells of the central nervous system. After the three plasmids were co-transfected into 293T cells, the AAV virus began to replicate and package. The obtained virus particles were purified by ultracentrifugation, and the virus gene copy was determined by qPCR. Follow-up related experiments were carried out according to the titer of the virus. Virus packaging was completed by Heyuan Bi...

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Abstract

The invention relates to a medical application of recombinant protein Semaphorin3G in prevention and treatment of retinal diseases. According to the invention, the level of Sema3G protein in the retina is improved through adeno-associated virus mediated Sema3G protein overexpression in retinal vascular endothelial cells or injection of Sema3G recombinant protein into vitreous cavities, the pathological process of the OIR mouse retinopathy can be effectively improved, and the Sema3G protein can reduce the area of a vascular occlusion area in the retinopathy, and can also reduce the area of pathological neovascularization clusters, and relieve pathological phenotypes.

Description

technical field [0001] The invention belongs to the field of ophthalmic retinopathy treatment, and specifically relates to the medical application of recombinant protein Semaphorin3G in preventing and treating retinal diseases. Background technique [0002] Ischemic retinopathy (ischemic retinopathy) is a group of pathological angiogenesis diseases caused by retinal ischemia and hypoxia, including diabetic retinopathy (diabetic retinopathy, DR), retinopathy of prematurity (retinopathy of prematurity, ROP) and so on. DR and ROP are the leading causes of blindness in adults and infants, respectively. In diabetic patients, the reduction of oxygen transport leads to retinal hypoxia; in preterm infants, the retinal vascular network is not fully formed, and the retinal ends are still in an avascular state. Due to the lack of blood vessels in the retina to provide oxygen, the peripheral retina is hypoxic oxygen state. These pathological processes that damage the retinal vasculatu...

Claims

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Application Information

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IPC IPC(8): A61K38/17A61K9/00A61P9/10A61P27/02
CPCA61K38/1709A61K9/0048A61K9/0019A61P9/10A61P27/02
Inventor 陈丹阳孙宁赫韩峰卢应梅陈祥陆楠楠谭超胡仔仲袁松涛刘庆淮
Owner NANJING MEDICAL UNIV
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