Optimization of immune cell isolation method from mouse adipose tissue and the role of subgroups in obese mice

doi:10.3969/j.issn.1006-7795.2021.04.009

Abstract

Abstract: Objective To provide an efficient and low-damage method to isolate immune cells from adipose tissue to provide technical support for the study of immunological mechanisms in obesity. Methods We separated the adipose tissue around the epididymis of normal mice, and separated the immune cells by mechanical grinding or enzymatic digestion (including type Ⅱ/Ⅳ collagenase, Liberase enzyme), and screened the most efficient immune cell separation method by flow cytometry technology. Then, an obese mouse model was established through high fat diet (HFD), and the proportion and subgroup changes of macrophages and T lymphocytes in mouse adipose tissue were further analyzed. Results We compared different enzymatic digestion and mechanical grinding methods and found that CD45⁺ immune cells obtained through high-purity Liberase enzyme digestion had the lowest mortality rate and the highest number of live cells. The death rate of immune cells obtained by mechanical grinding was the highest, and the number of living cells was the least.Compared with mice fed a normal diet, the proportion of innate immune cells such as neutrophils and macrophages in the adipose tissue of obese mice increased, especially the proportion of M1 macrophages, while the M2 macrophages decreased. The secretion of tumor necrosis factor-α(TNF-α) by macrophages increased, and the expression of triggering receptor expressed on myeloid cells 1(TREM1) enhanced. Among the adaptive immune cells, the proportion of T lymphocytes increased, especially the ratio of Th1/Treg cells increased the most. Conclusion We have established a method to efficiently obtain viable immune cells from mouse adipose tissue, and confirmed that the proportion of macrophages and T lymphocytes in the adipose tissue of obese mice increased, and their pro-inflammatory effect were enhanced. They played an important role in inducing inflammation in adipose tissue.

Key words: adipose tissue, enzyme digestion, mechanical method, macrophages, T lymphocytes

CLC Number:

R965

Wu Yongle, Shang Hongwei, Sun Guangyong, Zhang Dong, Ding Huiguo. Optimization of immune cell isolation method from mouse adipose tissue and the role of subgroups in obese mice[J]. Journal of Capital Medical University, 2021, 42(4): 559-567.

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URL: https://journal03.magtech.org.cn/Jweb_sdykdxxb/EN/10.3969/j.issn.1006-7795.2021.04.009

https://journal03.magtech.org.cn/Jweb_sdykdxxb/EN/Y2021/V42/I4/559

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