葫芦巴碱调控SPHK1/S1P/S1PR3通路对子宫内膜癌细胞增殖和致瘤性的影响

doi:10.3969/j.issn.1006-5725.2026.14.007

摘要/Abstract

摘要：

目的探讨葫芦巴碱（Trig）调控鞘氨醇激酶1（SPHK1）/1-磷酸鞘氨醇（S1P）/1-磷酸鞘氨醇受体3（S1PR3）通路对子宫内膜癌（EC）细胞增殖和致瘤性的影响。方法 Ishikawa细胞分为：空白组、不同浓度Trig（Trig-低、Trig-高）组、SPHK1组、Trig-高 + 空载组、Trig-高 + SPHK1组，通过克隆形成、CCK-8检测细胞增殖；qRT-PCR检测SPHK1 mRNA表达水平；免疫印迹检测抗原ki67、细胞癌基因Myc（c-Myc）以及SPHK1/S1P/S1PR3通路相关蛋白表达；建立裸鼠移植瘤实验，分离瘤体并称重，免疫印迹检测移植瘤组织中ki67、c-Myc以及SPHK1/S1P/S1PR3通路相关蛋白表达；HE、免疫组化分别检测移植瘤组织病理变化及ki67、c-Myc表达。结果与空白组相比，Trig-低组、Trig-高组克隆数、OD、ki67、c-Myc、SPHK1 mRNA及蛋白、S1P、S1PR3表达降低，SPHK1组克隆数、OD、ki67、c-Myc、SPHK1 mRNA及蛋白、S1P、S1PR3表达增加（P < 0.05）；与Trig-高 + 空载组相比，Trig-高 + SPHK1组克隆数、OD、ki67、c-Myc、SPHK1 mRNA及蛋白、S1P、S1PR3表达增加（P < 0.05）；与SPHK1组相比，Trig-高 + SPHK1组克隆数、OD、ki67、c-Myc、SPHK1 mRNA及蛋白、S1P、S1PR3表达降低（P < 0.05）。体内实验显示，不同剂量Trig可降低移植瘤瘤重增加，抑制增殖相关蛋白以及SPHK1/S1P/S1PR3通路相关蛋白表达（P < 0.05）。结论 Trig抑制EC细胞增殖，抑制肿瘤生长，其机制与抑制SPHK1/S1P/S1PR3通路有关。

关键词: 葫芦巴碱, 子宫内膜癌, SPHK1/S1P/S1PR3通路, 增殖, 致瘤性

Abstract:

Objective To investigate the effects of trigonelline （Trig） on the regulation of the sphingosine kinase 1 （SPHK1）/sphingosine-1-phosphate （S1P）/sphingosine-1-phosphate receptor 3 （S1PR3） pathway and its impact on the proliferation and tumorigenicity of endometrial cancer （EC） cells. Methods Ishikawa cells were divided into the following groups： The blank group， groups treated with different concentrations of Trig （low-Trig and high-Trig groups）， the SPHK1 group， the high-Trig + empty vector group， and the high-Trig + SPHK1 group. Cell proliferation was detected by colony formation assay and CCK-8 assay. The expression level of SPHK1 mRNA was measured by qRT-PCR. Western blot analysis was performed to detect the expression of antigen ki67， Myc proto-oncogene （c-Myc）proteins and proteins related to the SPHK1/S1P/S1PR3 pathway. A nude mouse xenograft tumor model was established. The tumors were excised and weighed. Western blotting was used to measure the expressions of ki67， c-Myc， and proteins related to the SPHK1/S1P/S1PR3 pathway in the xenograft tumor tissues. Hematoxylin and eosin （HE） staining and immunohistochemistry were used to detect the pathological changes of the xenograft tumor tissues and the expressions of ki67 and c-Myc， respectively. Results Compared with the blank group， the clone number， optical density （OD）， expression levels of ki67， c-Myc， SPHK1 mRNA and protein， as well as the levels of S1P and S1PR3 were lower in both the Trig-low group and the Trig-high group， while these parameters were higher in the SPHK1 group （P < 0.05）. Compared with the Trig-high + empty group， the clone number， OD， expression levels of ki67， c-Myc， SPHK1 mRNA and protein， and the levels of S1P and S1PR3 were higher in the Trig-high + SPHK1 group （P < 0.05）. Compared with the SPHK1 group， the clone number， OD， expression levels of ki67， c-Myc， SPHK1 mRNA and protein， and the levels of S1P and S1PR3 were lower in the Trig-high + SPHK1 group （P < 0.05）. In vivo experiments demonstrated that different doses of Trig could reduce the increase in the weight of transplanted tumors and inhibit the expression of proliferation-related proteins and proteins related to the SPHK1/S1P/S1PR3 pathway （P < 0.05）. Conclusions Trig effectively inhibits the proliferation of EC cells and suppresses tumor growth. The underlying mechanism is closely associated with the inhibition of the SPHK1/S1P/S1PR3 pathway.

Key words: trigonelline, endometrial cancer, SPHK1/S1P/S1PR3 pathway, proliferation, tumorigenicity

中图分类号:

R285.5

陈菁,杨修香,王晓彬,张克. 葫芦巴碱调控SPHK1/S1P/S1PR3通路对子宫内膜癌细胞增殖和致瘤性的影响[J]. 实用医学杂志, 2026, 42(14): 2542-2549.

Jing CHEN,Xiuxiang YANG,Xiaobin WANG,Ke ZHANG. The effects of trigonelline on the proliferation and tumorigenicity of endometrial cancer cells by regulating the SPHK1/S1P/S1PR3 pathway[J]. The Journal of Practical Medicine, 2026, 42(14): 2542-2549.

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组别	克隆数/个	OD	ki67/β-actin	c-Myc/β-actin
空白组	124.08 ± 12.55	0.97 ± 0.10	1.18 ± 0.12	0.89 ± 0.09
Trig-低组	95.34 ± 9.97^*	0.71 ± 0.08^*	0.83 ± 0.09^*	0.62 ± 0.07^*
Trig-高组	65.37 ± 8.86^*&	0.49 ± 0.06^*&	0.64 ± 0.07^*&	0.45 ± 0.06^*&
SPHK1组	186.34 ± 18.74^*	1.37 ± 0.14^*	1.55 ± 0.17^*	1.29 ± 0.13^*
Trig-高 + 空载组	66.05 ± 8.94	0.51 ± 0.06	0.61 ± 0.07	0.47 ± 0.06
Trig-高 + SPHK1组	95.44 ± 9.99^#△	0.75 ± 0.08^#△	0.86 ± 0.09^#△	0.68 ± 0.08^#△
F值	96.469	79.055	67.216	82.714
P值	< 0.001	< 0.001	< 0.001	< 0.001

组别	S1PR3/β-actin	S1P/β-actin	SPHK1/β-actin	SPHK1 mRNA
空白组	0.75 ± 0.08	0.83 ± 0.10	0.68 ± 0.07	1.02 ± 0.11
Trig-低组	0.49 ± 0.05^*	0.55 ± 0.07^*	0.44 ± 0.05^*	0.72 ± 0.08^*
Trig-高组	0.22 ± 0.03^*&	0.37 ± 0.04^*&	0.25 ± 0.03^*&	0.56 ± 0.06^*&
SPHK1组	1.03 ± 0.11^*	1.25 ± 0.13^*	0.94 ± 0.10^*	1.52 ± 0.16^*
Trig-高 + 空载组	0.28 ± 0.03	0.35 ± 0.04	0.29 ± 0.03	0.52 ± 0.06
Trig-高 + SPHK1组	0.50 ± 0.05^#△	0.59 ± 0.07^#△	0.49 ± 0.05^#△	0.74 ± 0.08^#△
F值	130.383	103.603	111.135	87.315
P值	< 0.001	< 0.001	< 0.001	< 0.001

组别	瘤重/g	ki67阳性表达/%	c-Myc阳性表达/%	ki67/β-actin	c-Myc/β-actin
EC组	0.85 ± 0.10	15.34 ± 1.57	18.05 ± 1.91	0.77 ± 0.08	0.69 ± 0.08
低剂量Trig组	0.64 ± 0.07^*	9.05 ± 1.02^*	11.74 ± 1.21^*	0.52 ± 0.06^*	0.41 ± 0.05^*
高剂量Trig组	0.41 ± 0.05^*&	5.37 ± 0.71^*&	6.88 ± 0.81^*&	0.31 ± 0.04^*&	0.22 ± 0.03^*&
SPHK1组	1.25 ± 0.13^*	24.13 ± 2.55^*	27.34 ± 2.88^*	1.05 ± 0.11^*	0.99 ± 0.10^*
高剂量Trig+空载组	0.42 ± 0.05	5.44 ± 0.72	6.92 ± 0.83	0.33 ± 0.04	0.27 ± 0.03
高剂量Trig+SPHK1组	0.68 ± 0.07^#△	9.11 ± 1.04^#△	11.89 ± 1.24^#△	0.59 ± 0.06^#△	0.46 ± 0.05^#△
F值	82.606	257.821	224.325	163.910	215.655
P值	< 0.001	< 0.001	< 0.001	< 0.001	< 0.001

组别	S1PR3/β-actin	S1P/β-actin	SPHK1/β-actin
EC组	0.88 ± 0.10	0.71 ± 0.08	0.91 ± 0.10
低剂量Trig组	0.59 ± 0.07^*	0.51 ± 0.06^*	0.68 ± 0.08^*
高剂量Trig组	0.37 ± 0.05^*&	0.31 ± 0.04^*&	0.42 ± 0.05^*&
SPHK1组	1.18 ± 0.13^*	0.97 ± 0.10^*	1.37 ± 0.15^*
高剂量Trig+空载组	0.31 ± 0.04	0.34 ± 0.04	0.39 ± 0.05
高剂量Trig+SPHK1组	0.53 ± 0.06^#△	0.57 ± 0.06^#△	0.67 ± 0.08^#△
F值	165.732	135.978	157.264
P值	< 0.001	< 0.001	< 0.001