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Novel biological preparation for resisting renal fibrosis, of human umbilical cord MSC exosome and preparation method

A technology of anti-fibrosis and biological agents, applied in biochemical equipment and methods, medical preparations containing active ingredients, drug combinations, etc., can solve the problem of uncontrollable dose safety, no substantial improvement of etiology, and existence of ethics, etc. problems, to achieve the effect of inhibiting renal interstitial fibrosis and protecting renal function

Pending Publication Date: 2019-07-23
JIANGSU UNIV
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  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] To sum up, the problems existing in the existing technology are: to clarify the mode of action, dose efficacy, and mechanism of action of mesenchymal stem cell-derived exosomes (hucMSC-exosomes) in inhibiting renal interstitial fibrosis and delaying diabetic nephropathy
There are many toxic and side effects in the existing technology, and it is only used to control the disease without substantial improvement in the etiology; or there are ethical, uncontrollable dosage and safety issues

Method used

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  • Novel biological preparation for resisting renal fibrosis, of human umbilical cord MSC exosome and preparation method
  • Novel biological preparation for resisting renal fibrosis, of human umbilical cord MSC exosome and preparation method
  • Novel biological preparation for resisting renal fibrosis, of human umbilical cord MSC exosome and preparation method

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preparation example Construction

[0028] Such as figure 1 As shown, the preparation method of the novel anti-fibrosis biological preparation of human umbilical cord MSC exosomes provided by the embodiments of the present invention comprises the following steps:

[0029] S101: Centrifuge at 300g for 10min, 2000g for 10min, 10000g for 30min by ultracentrifugation, collect the supernatant and centrifuge at 100000g for 70min;

[0030] S102: Remove the supernatant by chemical reagent precipitation method, collect the precipitate and dissolve it in PBS;

[0031] S103: Extracting exosomes in the stem cell culture supernatant by ultrafiltration and centrifugation.

[0032] Further, the component of the exosome is a nanoscale biofilm vesicle, called exosome. It plays an important role in the transmission of substances and information between cells. It can intervene in the occurrence and development of various diseases by delivering specific proteins, messenger RNA (mRNA) and various biologically active molecules to t...

Embodiment 1

[0035] Example 1 Extraction and identification of MSC exosomes

example 1

[0036] The main materials and sources used in example 1 are as follows respectively:

[0037] MSC culture reagents: α-MEM, fetal bovine serum (Gibco company product), trypsin (Sigma company product), carbon dioxide incubator (Forma company), serum-free medium (Shanghai Yikesai company; inverted microscope, fluorescence microscope, Biological microscopes, electron microscopes, ultra-clean benches, desktop centrifuges, and ultracentrifuges (Beckman Corporation, USA).

[0038]Exosome extraction reagents: heavy water (D2O, Shanghai Chuangsai Company), analytically pure sucrose (Guangzhou Chemical Reagent Factory), rabbit anti-human CD9 antibody (US Bioworld Technology Company), rabbit anti-human CD63 antibody (US Epitomics Company), rabbit anti-human CD81 antibody (Epitomics, USA), horseradish peroxidase (HRP)-labeled goat anti-rabbit IgG secondary antibody (Beijing Kangwei Century Company), HRP chemiluminescent substrate, 100-kDa MWCO ultrafiltration centrifuge tube, 0.22 μm Ste...

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Abstract

The invention belongs to the technical field of a biological preparation, and discloses a novel biological preparation for resisting renal fibrosis, of human umbilical cord MSC exosome and a preparation method. The novel biological preparation for resisting renal fibrosis, of human umbilical cord MSC exosome has main component of cup-shaped membrane-shaped vesicles which are prepared through combination of intracellular multivesicular bodies with cell membranes, are secreted outside cells and are about 100nm in size. The preparation method of the novel biological preparation for resisting renal fibrosis, of human umbilical cord MSC exosome comprises an ultracentrifugation method, a chemical reagent precipitation method, an ultrafiltration and centrifugation method and a magnetic bead immunization method. When the MSC cell exosome is extracted and used for treatment of an animal model suffering from diabetic nephropathy, the inventor finds that MSC-Ex has the treatment effects of restraining YAP activity, allevaiting renal interstitial fibrosis and delaying diabetic nephropathy.

Description

technical field [0001] The invention belongs to the technical field of biological preparations, and in particular relates to a novel anti-fibrosis biological preparation and a preparation method of human umbilical cord MSC exosomes. Background technique [0002] At present, the commonly used prior art in the industry is as follows: diabetes mellitus (DM) is a kind of metabolic disease characterized by hyperglycemia caused by insulin secretion defect or insulin action disorder. The latest data from the International Diabetes Federation (IDF) shows that in 2017, there were 425 million diabetic patients aged 20-79 in the world; as my country's aging population intensifies, the national prevalence rate of diabetes in China has risen to 10.96%, and the number of patients is 114 million, which is the highest in the world. Countries with the fastest growing and highest numbers of DM patients. Diabetic kidney disease (DKD) is one of the most important microvascular complications of ...

Claims

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Application Information

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IPC IPC(8): A61K35/28A61P13/12A61P3/10C12N9/12
CPCA61K35/28A61P13/12A61P3/10C12N9/1205
Inventor 纪成钱晖许文荣张家慧尹思琪
Owner JIANGSU UNIV
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