Effects of hypoxia exposure on proliferation and apoptosis of mouse spleen lymphocytes

doi:10.11958/20220649

Abstract

Abstract:

Objective To investigate the relationship between lymphocyte number reduction, cell proliferation and apoptosis induced by hypoxia exposure. Methods Lymphocytes were isolated from spleen of C57BL/6 mice and cultured in hypoxia (1%O₂) and normoxia (21%O₂) for 12, 24 and 48 h, respectively. T and B lymphocytes were assessed by flow cytometry. Proliferation and apoptosis of lymphocytes were detected by CFSE and AnnexinⅤ-FITC/PI method, respectively. The morphology of lymphocytes was observed by scanning electron microscope. The mRNA and protein expression levels of apoptosis-related factor bcl-2 homologous antagonist killer protein (Bak), murine B-cell lymphoma/leukemia-2 (bcl-2) and cysteine aspartate protease-3 (caspase-3) were detected by real-time quantitative reverse transcription PCR (qPCR) and Western blot assay. Results Hypoxic exposure for 12, 24 and 48 h can reduce the number of T and B lymphocytes, inhibit the proliferation of lymphocytes and promote cell apoptosis. Scanning electron microscopy showed that at the same time, the characteristic morphological changes of apoptosis appeared earlier in the hypoxic group. After 12, 24 and 48 h hypoxia exposure, the mRNA and protein expression levels of Bak and caspase-3 in lymphocytes were up-regulated, while the mRNA and protein expression levels of bcl-2 were down-regulated. Conclusion Hypoxic exposure mediates the decline of lymphocyte numbers by inhibiting lymphocyte proliferation and promoting apoptosis.

Key words: hypoxia, lymphopenia, cell proliferation, apoptosis, bcl-2 homologous antagonist-killer protein, caspase 3

CLC Number:

R392.12

Figures/Tables 10

References 27

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基因名称	引物序列（5′→3′）	产物大小（bp）
β-actin	上游：CATCCGTAAAGACCTCTATGCCAAC	171
β-actin	下游：ATGGAGCCACCGATCCACA	171
Bak	上游：GGTCTTTCGAAGCTACGTTTTT	243
Bak	下游：ATCTTGGTGAAGAGTTCGTAGG	243
bcl-2	上游：GATGACTTCTCTCGTCGCTAC	156
bcl-2	下游：GAACTCAAAGAAGGCCACAATC	156
caspase-3	上游：GGCCTGAAATACCAAGTCAGGAA	128
caspase-3	下游：CCATGGCTTAGAATCACACACACA	128

基因名称	引物序列（5′→3′）	产物大小（bp）
β-actin	上游：CATCCGTAAAGACCTCTATGCCAAC	171
β-actin	下游：ATGGAGCCACCGATCCACA	171
Bak	上游：GGTCTTTCGAAGCTACGTTTTT	243
Bak	下游：ATCTTGGTGAAGAGTTCGTAGG	243
bcl-2	上游：GATGACTTCTCTCGTCGCTAC	156
bcl-2	下游：GAACTCAAAGAAGGCCACAATC	156
caspase-3	上游：GGCCTGAAATACCAAGTCAGGAA	128
caspase-3	下游：CCATGGCTTAGAATCACACACACA	128

组别	12 h		24 h		48 h
组别	T淋巴细胞	B淋巴细胞	T淋巴细胞	B淋巴细胞	T淋巴细胞	B淋巴细胞
常氧组	40.17±1.90	33.47±2.38	32.65±4.60	27.96±1.50	26.30±1.11	24.62±2.88
低氧组	32.26±3.30	27.42±2.73	23.77±2.61	23.39±2.31	23.16±1.33	18.71±1.31
t	3.603^＊	2.895^＊	2.904^＊	2.873^＊	3.140^＊	3.229^＊

组别	12 h		24 h		48 h
组别	T淋巴细胞	B淋巴细胞	T淋巴细胞	B淋巴细胞	T淋巴细胞	B淋巴细胞
常氧组	40.17±1.90	33.47±2.38	32.65±4.60	27.96±1.50	26.30±1.11	24.62±2.88
低氧组	32.26±3.30	27.42±2.73	23.77±2.61	23.39±2.31	23.16±1.33	18.71±1.31
t	3.603^＊	2.895^＊	2.904^＊	2.873^＊	3.140^＊	3.229^＊

组别	12 h	24 h	48 h
常氧组	31.56±1.63	38.50±0.31	43.32±1.35
低氧组	27.31±2.05	34.74±0.82	40.66±0.49
t	4.749^＊＊	11.324^＊＊	5.566^＊＊