基于肿瘤神经新生学说、p75NTR信号通路探讨电针瘤周围刺抑瘤分子机制

Neoplastic neurogenesis plays an important role in the development and metastasis of tumor. Tumor cells promote neurogenesis by secreting nerve growth corresponding factors and expressing related receptor. Meanwhile, newborn nerves are involved in the regulation of tumor angiogenesis, and can promote tumor growth through by releasing neurotransmitters and neuropeptides. p75NTR is a neurotrophic factor receptor, which can activate NF-κB and JNK-p53-Bax signaling pathway, promoting cell apoptosis. Surround needling is regarded as a main external treatment of inhibiting tumor in Traditional Chinese Medicine (TCM). Our previous studies had confirmed that needling surround tumor can inhibit angiogenesis and tumor growth. Meanwhile, it was found that acupuncture promoted tumor neurogenesis and expression of p75NTR, however, the mechanisms were unclear. With the building of 4T1 mouse breast cancer xenograft model in this study, the expression of nerve growth associated protein can clarify the effect mechanism of acupuncture on tumor neurogenesis. The expression of Ach, NE, substance P, dopamine can illuminate the secretion fuction of tumor nerve. The expression of proNGF, NGF, TrKA, p75NTR, Sortilin protein and detecting key genes of downstream pathway can expound the antitumor mechanisms of acupuncture by nerve, so as to provide a scientific basis of antitumor by surround needling.

肿瘤神经新生在肿瘤发展、转移中扮演重要的角色。肿瘤细胞分泌神经生长相关因子，表达相应受体，促进神经新生；新生神经参与调控肿瘤血管生成，并能通过释放神经递质及神经肽促进肿瘤的生长；p75NTR是神经营养因子的受体，激活NF-κB、JNK-p53-Bax等信号通路，促进细胞凋亡。针灸围刺是中医外治治疗肿瘤主要手段，我们既往研究已经证实，瘤周围刺能够抑制肿瘤血管生成，抑制肿瘤生长，同时发现针刺促进肿瘤神经新生及p75NTR的表达，但是其中的机理尚不清楚。本研究拟用4T1小鼠乳腺癌移植瘤模型，通过检测神经生长相关因子的表达情况，阐明针刺对肿瘤神经新生的作用机制；通过Ach、NE、P物质、多巴胺等的表达情况，阐明肿瘤新生神经的分泌功能；通过检测proNGF、NGF、TrKA、p75NTR、Sortilin等蛋白表达及激活通路的关键基因，阐明针刺通过神经调控肿瘤生长的作用机制，为围刺抑瘤提供科学依据。

肿瘤神经新生在肿瘤发展、转移中扮演重要的角色。p75NTR是神经营养因子的受体，激活下游信号通路，促进细胞凋亡。针灸围刺是中医外治治疗肿瘤主要手段，我们既往研究已经证实，瘤周围刺能够抑制肿瘤血管生成，抑制肿瘤生长，同时发现针刺促进肿瘤神经新生及p75NTR的表达。本研究拟用4T1小鼠乳腺癌移植瘤模型，通过检测神经支配、神经递质、神经生长相关因子，阐明针刺对肿瘤神经新生的作用机制；通过p75NTR抑制剂、非参转录测序，阐明针刺通过神经调控肿瘤生长的作用机制。研究发现乳腺癌肿瘤间质及包膜均可见神经支配，电针围刺具有促进神经新生、抑制PNI和交感神经形成的作用。电针围刺对血清神经递质、NTs影响相对较小，而对肿瘤组织神经递质（ACH、AD、NE）、NTs（NGF、proNGF、NT-3）的影响较显著，体现了电针围刺治疗肿瘤的组织靶向性特点。电针围刺上调p75NTR表达水平，其高亲和力结合pro-NGF，上调p75NTR/proNGF信号轴，诱导癌细胞凋亡而发挥抑瘤作用。此外电针围刺经由p75NTR介导以抑制PNI和交感神经支配，下调肿瘤组织NE、ACH水平，达到调控肿瘤神经微环境而发挥间接抑瘤作用。肿瘤组织的非参转录组测序显示，相较于TG组，ETG组共获得116个差异基因，其中上调基因66个，下调基因50个，GO功能注释分析显示电针围刺相关的差异基因主要与免疫反应、细胞死亡及稳态等生物学过程相关，KEGG通路富集分析发现电针围刺相关的差异表达基因主要与神经、免疫反应、物质与能量代谢相关，其中在神经的生物学过程中，包含四个神经递质兴奋性突触通路和一个神经轴突导向通路，涉及11个差异基因，其中5-HT2、VGCC、COX、TRPC1、Ablim下调，Gi/o、PLA2、GLS、CREB、Boc、Netin-G2上调，涉及胞吐作用、钙离子内流、神经兴奋性、神经递质释放的反馈抑制调节、轴突导向等生物学功能。电针围刺可能通削弱胞吐作用、抑制钙离子内流、调控神经兴奋性等作用抑制神经递质释放，以及通过促进神经保护和轴突导向促进神经轴突生成，以调控肿瘤微环境。