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Preparation method of kidney sertoli cells and special mediums thereof

A culture medium and podocyte technology, which is applied to the preparation of kidney podocytes and the field of special culture medium, can solve the problems of low transformation efficiency of iPSCs, unclear reprogramming mechanism of oncogenes, and limited application of iPSCs cells, etc. Easy to operate effect

Active Publication Date: 2015-11-25
THE INST OF BASIC MEDICAL SCI OF CHINESE ACAD OF MEDICAL SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, due to the low transformation efficiency of iPSCs, the unclear reprogramming mechanism and the activation of oncogenes during the reprogramming process, the further application of iPSCs in clinical treatment is also limited.

Method used

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  • Preparation method of kidney sertoli cells and special mediums thereof
  • Preparation method of kidney sertoli cells and special mediums thereof
  • Preparation method of kidney sertoli cells and special mediums thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0035] Embodiment 1, the preparation of culture medium

[0036] DMEM / F12 medium (also known as DF12 medium) was purchased from GIBCO Company. Fetal bovine serum (FBS) was purchased from GIBCO. Human activin A (activinA) was purchased from PeproTech, UK, with a catalog number of 120-14. Retinoic acid was purchased from Sigma Company, the product catalog number is R2625. Bone morphogenic protein 7 (bonemorphogeneticprotein7, BMP7) was purchased from PeproTech Company, the product catalog number is 120-03.

[0037] Medium 1 is made up of DMEM / F12 medium and solute; Said solute and its concentration in medium 1 are as follows: volumetric concentration is 2% fetal calf serum, 10ng / mLactivinA and 10 -5 mol / L retinoic acid.

[0038] Medium 2 is made up of DMEM / F12 medium and solute; Said solute and its concentration in medium 2 are as follows: volumetric concentration is the fetal bovine serum of 2%, 10ng / mLactivinA and 10 -7 mol / L retinoic acid, 20ng / ml BMP7.

Embodiment 2

[0039] Example 2, Induction and Culture of Kidney Podocytes

[0040] 1. Preparation of the third generation of mesenchymal stem cells

[0041] 1. Aseptically collect adipose tissue from adults after liposuction (the source is human), wash twice with cell washing solution containing penicillin and streptomycin in a biological safety cabinet, remove blood cells and anesthetics, and centrifuge at 800rpm for 3min.

[0042] 2. Transfer the adipose tissue to a new 50ml centrifuge tube, add 0.05% collagenase P to digest the tissue, and incubate at 37°C with a shaker for 30 minutes.

[0043] 3. Filter the digested adipose tissue through a 100-mesh sieve and collect the filtrate, centrifuge at room temperature at 1200rpm for 10 minutes to collect cells.

[0044] 4. Culture the cells in step 3 in an incubator at 37°C, 5% CO2, and 95% relative air humidity. When the cells reach 70%-80% confluence, wash them twice with D-Hank's solution and use 1g / L trypsin (Gibco Company) was routinel...

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Abstract

The invention relates to a preparation method of kidney sertoli cells and special mediums thereof, in particular provides a method of inducing mesenchymal stem cells into the kidney sertoli cells, and the method comprises the following steps: (1) inoculating the mesenchymal stem cells into a medium I; and (2) transferring the mesenchymal stem cells done in step (1) into a medium II so as to obtain the kidney sertoli cells, wherein the medium I is composed of a zooblast basic medium and solutes, and the solutes comprise 1.9 percent to 2.1 percent of fetal calf serum, 9.75ng / mL to 10.25ng / mL of activin A and 0.95*10 to 5mol / L to 1.05*10 to 5mol / L of tretinoin; and the medium II is composed of a zooblast basic medium and solutes, and the solutes comprise 1.9 percent to 2.1 percent of fetal calf serum, 9.75ng / mL to 10.25ng / mL of activin A, 0.95*10 to 7mol / L to 1.05*10 to 7mol / L of tretinoin and 19ng / ml to 21ng / ml of BMP7. An induction culture method of the invention is suitable for the mesenchymal stem cells of various sources, and is free of virus introduction, easy in operation and high in efficiency.

Description

technical field [0001] The invention relates to a method for inducing differentiated cells from mesenchymal stem cells, in particular to a preparation method for renal podocytes and a special culture medium thereof. Background technique [0002] The kidney is one of the most important organs of the human body. Its main physiological function is to remove metabolites, excess substances and poisons entering the body through the production of urine, and at the same time retain water and other useful substances through the reabsorption function to realize the recovery of body fluids, The regulation of electrolyte and acid-base balance maintains the stability of the internal environment of the body. [0003] Kidney podocytes, namely visceral epithelial cells of the renal capsule, are important components to maintain the normal structure and function of the glomerular filtration barrier, and play an important role in maintaining the permeability of the glomerulus and resisting the...

Claims

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Application Information

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IPC IPC(8): C12N5/071A61K35/22A61P13/12
Inventor 赵春华张丽娜李康华
Owner THE INST OF BASIC MEDICAL SCI OF CHINESE ACAD OF MEDICAL SCI
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