低强度激光通过Wnt/β-catenin信号通路对RKO结直肠癌细胞的影响

Influence of Low-level Laser Irradiation via Wnt/β-catenin Signal Pathway on RKO Colorectal Cancer Cells

Wnt/β-catenin信号通路的正常表达是调控正常细胞生长、分化的重要通路,对Wnt/β-catenin信号通路中靶点的研究也是当前肿瘤靶向治疗的热点。本文旨在探究低强度激光(LLLI)通过Wnt/β-catenin信号通路对RKO结直肠癌细胞的影响。利用632.8 nm波长的红外激光照射RKO细胞,应用半定量聚合酶链式反应(RTPCR)法筛选梯度低强度激光对RKO细胞Wnt/β-catenin信号通路中的APC、β-catenin、DVL基因表达的影响。RT-PCR结果显示:15 J/cm2能量密度的低强度激光可以降低RKO细胞中β-catenin基因的表达,可以增强APC及DVL基因的表达,且有随能量密度的递增而递增的趋势,差异有统计学意义。通过实时荧光定量聚合酶链式反应(qRT-PCR)法及蛋白质印迹(Western blot)法检测15 J/cm2能量密度的低强度激光照射后的RKO细胞相关指标的变化。qRT-PCR结果显示,Wnt5A、LRP5、TCF-4的基因表达均有不同程度的降低,差异有统计学意义。Western blot结果显示,β-catenin蛋白表达的下降相对于空白对照组的差异有统计学意义。qRT-PCR及Western blot的结果表明,15 J/cm^2能量密度的低强度激光短时间照射仍能对RKO细胞Wnt/β-catenin信号通路的过度表达起抑制作用。活细胞/死细胞试剂盒染色观察15 J/cm^2能量密度的低强度激光照射下,试验组RKO死细胞较对照组多。综上所述,15 J/cm^2能量密度的低强度激光照射能通过抑制Wnt/β-catenin信号通路过度表达以延缓RKO结直肠癌细胞的增殖,有剂量依赖性及时间累积效应,并可能额外激活细胞凋亡途径导致细胞凋亡。本研究可为结直肠癌的治疗提供一种新的思路。

Normal expression of the Wnt/β-catenin signal pathway plays an important role in regulation of normal cell growth and differentiation,and research on target sites in the Wnt/β-catenin signal pathway thus has become a hotspot on targeted anticancer therapy.Low-level laser irradiation(LLLI)was discussed for its influence on RKO colorectal cancer cells through the Wnt/β-catenin signal pathway.RKO cells were irradiated by the 632.8 nm infrared laser,and RT-PCR method was adopted for screening gene expressions of APC、β-catenin and DVL in the Wnt/β-catenin signal pathway of RKO cells irradiated by a gradient low-level laser.Results of the RT-PCR showed that the low-level laser with an energy density of 15 J/cm^2 reducedβ-catenin expression of RKO cells,and increased APC and DVL expressions,indicating both an increasing trend as the rising energy density and a statistical discrepancy.Related indicators of RKO cells were tested for their changes by qRT-PCR and Western blot methods,under the low-level laser irradiation of 15 J/cm^2.Results of qRT-PCR showed that gene expressions of Wnt5A,LRP5 and TCF-4 indicated different levels of reduction,with a statistical discrepancy.Results of Western blot showed that reduction in protein expression ofβ-catenin,relative to that of the blank control group,had a statistically significant discrepancy.Both qRT-PCR and Western blot showed that low-level laser irradiation of 15 J/cm2 for short time still had an inhibiting effect on overexpression of Wnt/β-catenin signal pathway of RKO cells.By observation on low-level laser irradiation of 15 J/cm^2 through viable and dead-cell kits staining,it can be seen that the dead RKO cells in experimental group were more than that in control group.In conclusion,low-level laser irradiation of 15 J/cm2 can delay proliferation of RKO colorectal cancer cells through inhibition on overexpression of Wnt/β-catenin signal pathway,and is dose-dependent with a time-accumulation effect;it can also activate the apoptosis pathway and therefore incur cell apoptosis.This paper,in view of the above,provided a new thinking for the treatment of colorectal cancer.