Knockdown of GSG2 inhibits the development and progression of non-small cell lung cancer in vitro and in vivo

Figures & data

Figure 1. Elevated GSG2 was detected in NSCLC tumor tissues relative to para-carcinoma tissues by immunohistochemical staining analysis.

Table 1. Expression patterns of GSG2 in lung cancer tissues and para-carcinoma tissues revealed in immunohistochemistry analysis.

GSG2 expression	Tumor tissue		Para-carcinoma tissue		P value
	Cases	Percentage	Cases	Percentage
Low	253	47%	27	100%	<0.001
High	285	53%	0	-

Table 2. Relationship between GSG2 expression and tumor characteristics in patients with lung cancer.

Features	No. of patients	GSG2 expression		P value
		low	high
All patients	538	253	285
Age (years)				0.576
<59	259	126	133
≥59	279	129	150
Gender				0.305
Male	393	181	212
Female	145	74	71
Grade				0.022
1	34	21	13
2	232	97	135
3	109	39	70
Tumor infiltrate				<0.001
T1	52	35	17
T2	378	185	193
T3	93	30	63
T4	15	5	10
Lymphatic metastasis (N)				<0.001
N0	256	143	113
N1	220	88	132
N2	55	22	33
N3	7	2	5
Stage				<0.001
I	217	134	83
II	167	68	99
III	154	53	101

Table 3. Relationship between GSG2 expression and tumor characteristics in patients with lung cancer.

		GSG2
Stage	Spearman correlation	0.235
	Signification (double-tailed)	0.000
	N	538
Grade	Spearman correlation	0.118
	Signification (double-tailed)	0.022
	N	375
Lymphatic metastasis (N)	Spearman correlation	0.157
	Signification (double-tailed)	0.000
	N	538
T infiltrate	Spearman correlation	0.183
	Signification (double-tailed)	0.000
	N	538

Figure 2. NSCLC cell models were established through infecting shGSG2 and shCtrl lentiviruses. (a) The fluorescence levels in A549 and NCI-H1299 cells were measured after 72 h-infection. Magnificationtimes: 200 ×. (b, c) The expression of GSG2 protein and mRNA in A549 and NCI-H1299 cell lines after infection was detected by western blot (b) and qRT-PCR (c). Results were presented as mean ± SD. ** P < 0.01, ***P < 0.001.

Figure 3. GSG2 knockdown inhibited cell proliferation, migration and arrested cell cycle. (a) The cell proliferation level was evaluated in A549 and NCI-H1299 cell lines after infection by MTT assay. (b) The effects of GSG2 knockdown on A549 and NCI-H1299 cell cycle were assessed by flow cytometry. (c, d) The migration rate of cells was detected in NSCLC cell lines after infection by wound-healing assay (c) and transwell assay (d). (e) The effects of GSG2 knockdown on cell apoptosis of A549 and NCI-H1299 cells were examined by flow cytometry. The red-R-fluorescence on the X-axis represented the percentage of apoptotic cells. The Y-axis being marked Green-B fluorescence represented the fluorescence signal of the GFP label on the lentivirus used to infect A549 and NCI-H1299 cells. Results were presented as mean ± SD. ** P < 0.01, ***P < 0.001.

Figure 4. GSG2 knockdown suppressed NSCLC tumor growth in vivo. (a) A nude mice model of GSG2 knockdown was constructed. Before sacrificing the mice, the fluorescence intensity was obtained by injecting D-luciferase. (b, c) The tumor volume (b) and body weight (c) were monitored throughout the feeding period until the mice were sacrificed. (d, e) After removing tumors, the tumors were weighed (d) and photographed (e). (f) The expression levels of Ki-67 in tumor sections were detected by IHC. Magnification times: 200×. Results were presented as mean ± SD. * P < 0.05.

Figure 5. The mechanism of GSG2 regulating NSCLC was investigated. (a) The expression of apoptosis-related proteins in NCI-H1299 cells infected with shGSG2 and shCtrl was measured by ECL with Human Apoptosis Antibody Array. The results circled in red represented that the protein expression was up-regulated and P < 0.05. (b) The expression levels of apoptosis-related proteins were presented in grayscale and visualized by R studio. (c) The levels of apoptosis-associated protein were detected by western blot. (d) The expression of cancer-associated elements, Akt, P-Akt, Cyclin D1, PI3K and CDK6, was detected by western blot. Results were presented as mean ± SD. Results were presented as mean ± SD. * P < 0.05.