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Method for promoting brown adipose differentiation through SFRP4 and application

A brown adipose tissue and cell differentiation technology, applied in the field of basic medical experiments, can solve problems that have not yet attracted people's attention, and achieve the effect of promoting differentiation and increasing expression levels

Active Publication Date: 2020-08-18
XIAN MEDICAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

At present, the research on SFRP4 mainly focuses on bone metabolism, cancer and cell apoptosis, etc., and its regulation in adipose tissue and adipocyte differentiation has not yet attracted people's attention

Method used

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  • Method for promoting brown adipose differentiation through SFRP4 and application
  • Method for promoting brown adipose differentiation through SFRP4 and application
  • Method for promoting brown adipose differentiation through SFRP4 and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0054] Embodiment 1, the temporal expression of SFRP4 in the differentiation process of BAT adipocytes

[0055] Primary brown adipocytes were isolated from the scapular triangle of 3-week-old C57BL / 6J mice and grown in culture flasks. Mouse brown adipocyte induction medium was added to the complete medium to induce differentiation into mature adipocytes, brown adipocytes The cells were induced to differentiate to the 8th day, the cells reached terminal differentiation, and the cells formed clusters of lipid droplets. The results of Oil Red O staining were as follows: image 3 shown by image 3 It can be seen that with the increase of culture time, the number of lipid droplets increased significantly, indicating that preadipocytes differentiated into mature adipocytes. Total cellular RNA was collected and extracted on days 2, 0, 2, 4, 6, and 8 of brown adipocyte differentiation. After the RNA purity and integrity were tested by Nanodrop instrument, brown adipocyte marker genes...

Embodiment 2

[0056] Example 2, overexpression of SFRP4 gene promotes differentiation of brown preadipocytes

[0057] In order to study the regulatory function of SFRP4 gene in the differentiation process of brown preadipocytes, the present invention adopts SFRP4 recombinant active protein. Brown preadipocytes were treated with different concentrations (0, 1, 10 and 100 ng / mL) of SFRP4 recombinant active protein and incubated for 24 hours, then continued to culture to induce differentiation to terminal differentiation. from Image 6 It can be seen that after SFRP4 active protein treatment, oil red O staining was used to analyze lipid accumulation in adipocytes, and high concentration of SFRP4 (10 and 100 ng / mL) treatment promoted lipid production in brown preadipocytes. According to the oil red O staining results obtained above, brown adipocytes were collected on the 8th day of induced differentiation, and real-time quantitative PCR was used to analyze the expression of lipid production-re...

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Abstract

The invention discloses a method for promoting brown adipose tissue differentiation through SFRP4 and an application. The method comprises treating precursor adipose cells for differentiation by usingSFRP4 active proteins (with different concentrations of 0, 1, 10, 100ng / mL), when the cells grow to 80%, culturing with a cell differentiation induction culture medium I until the cells are fused, adding an induction culture medium II into the cells, inducing for 2-8 days, and replacing the culture medium in the culture bottle with an induction culture medium III, inducing for 2-8 days, observingunder a microscope, after dyeing with oil red O, observing under the microscope again, collecting cells with different differentiation days, extracting total RNA of the cells, detecting expression levels of marker genes UCP-1 and SFRP4 of brown adipose cells by using a real-time quantitative PCR analysis technology after the purity and integrity of the RNA are detected to be qualified, and finally, carrying out statistical analysis. The experimental process is uniform in sampling, and the result reliability is high.

Description

technical field [0001] The invention belongs to the field of basic medical experiments, and specifically relates to a method for promoting brown fat differentiation through SFRP4, and also relates to the application of the above-mentioned SFRP4 in promoting brown fat tissue differentiation. Background technique [0002] Brown adipose tissue can perform oxidative respiration uncoupling with mitochondria, so that ATP can be converted into thermal energy, thereby realizing the conversion of thermal energy and consuming stored energy. The best way to treat obese patients is to increase the amount of brown adipose tissue, and this can also be of great help in the treatment of patients with metabolic syndrome. Secreted frizzled-related protein 4 (SFRP4), as a member of the SFRPs family, is one of the newly discovered adipocyte cytokines. At present, the research of SFRP4 mainly focuses on bone metabolism, cancer and cell apoptosis, etc., but its regulation in adipose tissue and a...

Claims

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Application Information

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IPC IPC(8): C12N5/077
CPCC12N5/0653C12N2501/01C12N2501/33C12N2501/395C12N2501/30C12N2501/998
Inventor 关华黄政豪余琦张云婷
Owner XIAN MEDICAL UNIV
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