The radiosensitizing effect of ginsenoside Rg3 on lung cancer cells by inhibiting mTOR pathway-mediated pentose phosphate pathway

doi:10.11958/20222018

Abstract

Abstract:

Objective To investigate the influence of ginsenoside Rg3 on the radiosensitivity of lung cancer cells by inhibiting the pentose phosphate pathway (PPP) mediated by mammalian target of rapamycin (mTOR) pathway. Methods A549 lung cancer cells were treated with 0, 10, 20, 40, 60, 80 mg/L ginsenoside Rg3. The proliferation of A549 cells was detected by MTT method. Cells were divided into the control group (normal culture, no irradiation), the radiation group (X-ray irradiation), the ginsenoside Rg3 group (60 mg/L ginsenoside Rg3, no irradiation), the combination group (X-ray irradiation +60 mg/L ginsenoside Rg3) and the activator group (X-ray irradiation +60 mg/L ginsenoside Rg3+100 nmol/L mTOR pathway activator MHY1485). All of groups were irradiated by 8 GyX ray after 48 h of culture with corresponding drugs. Plate cloning experiment was applied to detect the formation rate of cell clones in each group. Enzyme-linked immunosorbent assay (ELISA) was applied to determine levels of glucose-6-phosphate dehydrogenase (G6PD) and reduced nicotinamide adenine dinucleotide phosphate (NADPH) in cell supernatant of each group. DCFH-DA fluorescent probe method was applied to detect the level of intracellular reactive oxygen species (ROS). Cell apoptosis was detected by flow cytometry. γ-H2AX immunofluorescence staining was applied to analyze DNA damage repair. Western blot assay was applied to detect expression levels of mTOR, p-mTOR, proliferating cell nuclear antigen (PCNA), Bcl-2-associated X protein (Bax), caspase-3 and γ-H2AX protein in cells. Results Ginsenoside Rg3 inhibited the proliferation of A549 cells in a dose-dependent manner (P<0.05). Compared with the control group, the cell clone formation rate, G6PD, ROS, NADPH levels, p-mTOR/mTOR and PCNA protein expressions were significantly decreased in the radiation group and the ginsenoside Rg3 group, and the cell apoptosis rate, γ-H2AX foci number, Bax, caspase-3, γ-H2AX protein expressions were significantly increased (P<0.05). Compared with the radiation group and the ginsenoside Rg3 group, the cell clone formation rate, G6PD, ROS, NADPH levels, p-mTOR/mTOR, and PCNA protein expressions were significantly decreased in the combination group, the cell apoptosis rate, γ-H2AX foci number, Bax, caspase-3, γ-H2AX protein expressions were significantly increased (P<0.05). Compared with the combination group, the cell clone formation rate, G6PD, ROS, NADPH levels, p-mTOR/mTOR and PCNA protein expressions were significantly increased in the activator group, and the cell apoptosis rate, γ-H2AX foci number, Bax, caspase-3, γ-H2AX protein expressions were significantly decreased (P<0.05). Conclusion The inhibitory effect of ginsenoside Rg3 on lung cancer cells may be realized through inhibition of PPP mediated by mTOR.

Key words: Ginsenoside Rg3, lung neoplasms, radiation tolerance, radiation-sensitizing agents, pentose phosphate pathway, mammalian target of rapamycin

CLC Number:

R285，R734.2

Figures/Tables 5

References 25

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组别	G6PD/（U/L）	ROS/AU	NADPH/（ng/L）
对照组	3.56±0.42	37.94±2.86	98.64±7.51
放射组	2.87±0.34^a	29.65±2.43^a	76.52±6.64^a
人参皂苷Rg3组	2.31±0.25^a	21.52±2.14^a	63.18±5.62^a
联合组	1.12±0.13^bc	15.61±1.84^bc	33.84±5.16^bc
激活剂组	2.54±0.32^d	26.87±2.23^d	56.87±6.12^d
F	50.664^＊＊	78.867^＊＊	87.996^＊＊

组别	G6PD/（U/L）	ROS/AU	NADPH/（ng/L）
对照组	3.56±0.42	37.94±2.86	98.64±7.51
放射组	2.87±0.34^a	29.65±2.43^a	76.52±6.64^a
人参皂苷Rg3组	2.31±0.25^a	21.52±2.14^a	63.18±5.62^a
联合组	1.12±0.13^bc	15.61±1.84^bc	33.84±5.16^bc
激活剂组	2.54±0.32^d	26.87±2.23^d	56.87±6.12^d
F	50.664^＊＊	78.867^＊＊	87.996^＊＊

组别		p-mTOR/mTOR		PCNA
对照组		0.86±0.08		1.68±0.15
放射组		0.53±0.04^a		1.15±0.11^a
人参皂苷Rg3组		0.48±0.04^a		0.87±0.08^a
联合组		0.24±0.03^bc		0.26±0.04^bc
激活剂组		0.61±0.07^d		0.98±0.08^d
F		97.851^＊＊		160.573^＊＊
组别	Bax		caspase-3		γ-H2AX
对照组	0.32±0.04		0.34±0.05		0.28±0.01
放射组	0.71±0.06^a		0.75±0.08^a		0.73±0.07^a
人参皂苷Rg3组	1.08±0.10^a		1.13±0.11^a		0.86±0.09^a
联合组	1.53±0.12^bc		1.62±0.13^bc		1.71±0.14^bc
激活剂组	0.72±0.08^d		0.81±0.09^d		0.75±0.06^d
F	172.975^＊＊		148.533^＊＊		224.736^＊＊

组别		p-mTOR/mTOR		PCNA
对照组		0.86±0.08		1.68±0.15
放射组		0.53±0.04^a		1.15±0.11^a
人参皂苷Rg3组		0.48±0.04^a		0.87±0.08^a
联合组		0.24±0.03^bc		0.26±0.04^bc
激活剂组		0.61±0.07^d		0.98±0.08^d
F		97.851^＊＊		160.573^＊＊
组别	Bax		caspase-3		γ-H2AX
对照组	0.32±0.04		0.34±0.05		0.28±0.01
放射组	0.71±0.06^a		0.75±0.08^a		0.73±0.07^a
人参皂苷Rg3组	1.08±0.10^a		1.13±0.11^a		0.86±0.09^a
联合组	1.53±0.12^bc		1.62±0.13^bc		1.71±0.14^bc
激活剂组	0.72±0.08^d		0.81±0.09^d		0.75±0.06^d
F	172.975^＊＊		148.533^＊＊		224.736^＊＊