Neuropilins与肿瘤淋巴管生成和淋巴道转移的研究进展

淋巴道转移是肿瘤转移的重要途径,也是判断预后的重要依据。如何预防和抑制肿瘤转移一直是肿瘤研究的重点和热点。大量研究表明,恶性肿瘤和瘤周组织中存在新生的淋巴管,能够促进肿瘤的转移扩散,而且其密度与淋巴结转移密切相关。近年来研究发现,Neuropilins表达可能与肿瘤淋巴管新生和淋巴结转移密切相关,以Neuropilins为新型治疗靶点对恶性肿瘤淋巴管生成和转移进行靶向治疗也取得初步的研究成果。以上为揭示恶性肿瘤淋巴道转移的机制和开发淋巴管靶向治疗提供了重要的研究途径。

Neuropilins and liver

Neuropilins(NRPs) are highly conserved transmembrane glycoproteins that possess pleiotropic functions. Neuropilin-1(NRP1) and its homologue neuropilin-2 interact as coreceptors with both class 3 semaphorins and vascular endothelial growth factor and are involved in neuronal guidance and angiogenesis, respectively. The contribution of NRPs to tumor angiogenesis has been highlighted in previous studies, leading to the development of NRP antagonists as novel anti-angiogenesis therapies. However, more recent studies have demonstrated that NRPs have a much broader spectrum of activity in the integration of different pathways in physiological and pathological conditions. A few studies investigated the role of NRPs in both malignant and non-neoplastic liver diseases. In normal liver, NRP1 is expressed in hepatic stellate cells and liver sinusoidal endothelial cells. NRP1 expression in hepatocytes has been associated with malignant transformation and may play an important role in tumor behavior. A contribution of NRPs in sinusoidal remodeling during liver regeneration has been also noted. Studies in chronic liver diseases have indicated that, besides its influence on angiogenesis, NRP1 might contribute to the progression of liver fibrosis owing to its effects on other growth factors, including transforming growth factor β1. As a result, NRP1 has been identified as a promising therapeutic target for future antifibrotic therapies based on the simultaneous blockade of multiple growth factor signaling pathways. In this review, the structure of NRPs and their interactions with various ligands and associated cell surface receptors are described briefly. The current understanding of the roles of the NRPs in liver diseases including tumors, regeneration and fibrogenesis, are also summarized.

Neuropilin与肿瘤血管新生

Neuropilin作为轴突导向分子collapsin/semaphorin的受体在神经发育过程中引导轴突选择正确途径以成功到达靶区 ,与此同时又可作为血管内皮生长因子受体在血管新生中发挥作用。近年neuropilin在神经系统中的作用已基本得以阐明 ,而有关其在血管新生方面的研究亦有所进展 ,并且指出neuropilin参与调节肿瘤血管新生 ,在肿瘤生长转移中发挥作用。

Neuropilin1单克隆抗体对胶质瘤大鼠Bcl-2/Bax表达的影响

目的:研究Neuropilin1单克隆抗体对胶质瘤大鼠Bcl-2/Bax表达的影响。方法:将36只SD大鼠随机分为假手术组、模型组和治疗组,每组12只;假手术组只行颅骨开窗手术,其余各组均建立胶质细胞瘤模型。模型组腹腔注射0.9%氯化钠,治疗组腹腔注射NRP-1 MAb,均每日1次。干预14天后取材。对肿瘤组织称重,免疫组化检测Bax和Bcl-2表达,Western blot检测NRP-1蛋白、Bax蛋白和Bcl-2蛋白表达量,qPCR检测Bax mRNA和Bcl-2 mRNA表达情况,TUNEL检测细胞凋亡情况。结果:假手术组无肿瘤,治疗组肿瘤组织重量显著小于模型组,差异具有统计学意义(P<0.05)。免疫组化显示:与假手术组比较,模型组和治疗组Bax表达显著下降,Bcl-2表达显著上升,差异具有统计学意义(P<0.05);与模型组比较,治疗组Bax表达上升,Bcl-2表达下降,差异具有统计学意义(P<0.05)。Western blot检测显示:与假手术组比较,模型组和治疗组NRP-1蛋白和Bcl-2蛋白表达量显著上升,Bax蛋白表达量显著下降,差异具有统计学意义(P<0.05);与模型组比较,治疗组NRP-1蛋白和Bcl-2蛋白表达量显著下降,Bax蛋白表达量显著上升,差异具有统计学意义(P<0.05)。qPCR检测显示:与假手术组比较,模型组和治疗组Bax mRNA表达显著下降,Bcl-2mRNA表达显著上升,差异具有统计学意义(P<0.05);与模型组比较,治疗组Bax mRNA表达上升,Bcl-2mRNA表达下降,差异具有统计学意义(P<0.05)。TUNEL检测显示:与假手术组比较,模型组和治疗组的细胞凋亡率显著上升,差异具有统计学意义(P<0.05);与模型组比较,治疗组的细胞凋亡率显著下降,差异具有统计学意义(P<0.05)。结论:NRP-1MAb可通过抑制NRP-1表达而上调Bax蛋白表达,下调Bcl-2蛋白表达促进胶质细胞瘤凋亡。

Silencing Neuropilin 1 gene reverses TGF-β1-induced epithelial mesenchymal transition in HGC-27 gastric cancer cell line

Objective The aim of this study was to determine Neuropilin 1(NRP1)contribution to transforming growth factorβ1(TGF-β1)-induced epithelial mesenchymal transition(EMT)of HGC-27 gastric cancer cells and study its mechanism.Methods In this study,TGF-β1 was used to induce EMT in HGC-27 cells.Further,these cells were stably transfected with siRNA targeting NRP1.Wound healing and transwell assays were used to measure cell migration and invasion,respectively.NRP1 and EMT markers were measured using quantitative real time reverse transcription polymerase chain reaction and western blotting.Results Exposure of TGF-β1 conferred a fibroblastic-like shape to cancer cells and significantly increased the expression of NRP1 in HGC-27 cells.TGF-β1 subsequently promoted migration and invasion of HGC-27 cells.Furthermore,silencing NRP1 inhibited the invasion and migration of TGF-β1-induced cells undergoing EMT.Conclusion Silencing NRP1 can inhibit cell migration,invasion,and metastasis and reverse the TGF-β1-induced EMT process of gastric cancer.

Guinea pigs as an animal model for sciatic nerve injury

The overwhelming use of rat models in nerve regeneration studies is likely to induce skewness in treatment outcomes.To address the problem,this study was conducted in 8 adult guinea pigs of either sex to investigate the suitability of guinea pig as an alternative model for nerve regeneration studies.A crush injury was inflicted to the sciatic nerve of the left limb,which led to significant decrease in the pain perception and neurorecovery up to the 4th weak.Lengthening of foot print and shortening of toe spread were observed in the paw after nerve injury.A 3.49 ± 0.35 fold increase in expression of neuropilin 1（NRP1） gene and 2.09 ± 0.51 fold increase in neuropilin 2（NRP2） gene were recorded 1 week after nerve injury as compared to the normal nerve.Ratios of gastrocnemius muscle weight and volume of the experimental limb to control limb showed more than 50% decrease on the 30 th day.Histopathologically,vacuolated appearance of the nerve was observed with presence of degenerated myelin debris in digestion chambers.Gastrocnemius muscle also showed degenerative changes.Scanning electron microscopy revealed loose and rough arrangement of connective tissue fibrils and presence of large spherical globules in crushed sciatic nerve.The findings suggest that guinea pigs could be used as an alternative animal model for nerve regeneration studies and might be preferred over rats due to their cooperative nature while recording different parameters.

Correlation of Sema3A/Nrp1 pathway in gingival crevicular fluid of chronic periodontitis with inflammatory response and bone destruction

Objective: To study the correlation of Sema3A/Nrp1 pathway in gingival crevicular fluid of chronic periodontitis with inflammatory response and bone destruction. Methods: A total of 49 patients who were diagnosed with chronic periodontitis in the First People's Hospital of Jiangxia District, Wuhan City between July 2014 and January 2017 were selected as CP group, and 55 volunteers who received physical examination during the same period and had complete dentition were selected as control group. Gingival crevicular fluid was collected to detect the protein levels of Sema3A/Nrp1, inflammatory response indexes, bone metabolites and bone metabolism molecules. Results: Sema3A, Nrp1, PICP, Wnt1, β-catenin, and OPG protein levels in gingival crevicular fluid of CP group were significantly lower than those of control group while YKL40, CXCL16, ICAM1, HMBG1, SDF1α, ICTP, NTX, CTX, RANKL, 5-LOX and LTB4 protein levels were significantly higher than those of control group;Sema3A and Nrp1 protein levels in gingival crevicular fluid of CP patients were negatively correlated with YKL40, CXCL16, ICAM1, HMBG1, SDF1α, ICTP, NTX, CTX, RANKL, 5-LOX and LTB4 protein levels, and positively correlated with PICP, Wnt1, β-catenin and OPG protein levels. Conclusion: The inhibition of Sema3A/Nrp1 pathway in gingival crevicular fluid of chronic periodontitis can aggravate inflammatory response and promote bone destruction.

NETO2 promotes melanoma progression via activation of the Ca^(2+)/CaMKⅡ signaling pathway

Melanoma is the most aggressive cutaneous tumor.Neuropilin and tolloid-like 2(NETO2)is closely related to tumorigenesis.However,the functional significance of NETO2 in melanoma progression remains unclear.Herein,we found that NETO2 expression was augmented in melanoma clinical tissues and associated with poor prognosis in melanoma patients.Disrupting NETO2 expression markedly inhibited melanoma proliferation,malignant growth,migration,and invasion by downregulating the levels of calcium ions(Ca^(2+))and the expression of key genes involved in the calcium signaling pathway.By contrast,NETO2 overexpression had the opposite effects.Importantly,pharmacological inhibition of CaMKII/CREB activity with the CaMKII inhibitor KN93 suppressed NETO2-induced proliferation and melanoma metastasis.Overall,this study uncovered the crucial role of NETO2-mediated regulation in melanoma progression,indicating that targeting NETO2 may effectively improve melanoma treatment.

神经纤毛蛋白1对非小细胞肺癌放射敏感性的影响

目的 探讨神经纤毛蛋白1（neuropilin 1,NRP1）在不同种类非小细胞肺癌（NSCLC）细胞系的表达情况及其与肿瘤放射敏感性的关系,方法 通过Western blot技术检测不同来源的5种人NSCLC细胞系（H358、H460、H1299、A549、SK-MES-1） NRP1的基础表达水平,MTT方法检测经不同剂量X射线照射后肺癌细胞的存活情况,初步筛选5种细胞中辐射敏感和辐射抵抗的2种细胞;采用克隆形成实验及流式细胞术分别检测细胞存活分数及凋亡率的变化,分析2种肺癌细胞的放射敏感性,采用Western blot检测2种细胞受X射线作用后NRP1的表达变化,通过克隆形成实验检测靶向抑制NRP1对A549细胞放射敏感性的影响,结果 5种肿瘤细胞NRP1表达量由高到低依次为：A549＞ SK-MES-1＞ H358＞ H460＞ H1299;X射线对肿瘤细胞的抑制能力呈剂量依赖性增强,其抑制率由高到低依次为：H460＞ H1299＞H358＞ SK-MES-1＞A549,因此选取A549细胞和H460细胞做后续研究.克隆形成实验显示,A549在2 Gy照射下的存活分数（SF2）是0.887,而H460细胞为0.313。A549细胞受照后的凋亡率为对照组的122.54％,明显低于H460细胞的238.88％,辐射可以诱导A549细胞NRP1的表达上调达40％,同时靶向抑制NRP1则可以增强A549细胞的放射敏感性,结论 在NSCLC细胞中A549细胞具有较强的辐射抗性,且辐射抗性的形成与其NRP1表达上调有关。