血管生成拟态与肝癌研究进展

血管生成拟态的机制主要涉及肿瘤细胞基因型的转变、肿瘤细胞与细胞外基质的相互作用、肿瘤细胞分子信号改变.近年来的一些研究表明,肝癌组织中血管生成拟态的存在可能与肝癌的侵袭转移潜能以及较差的临床预后有关.     

血管生成拟态和血管生成及其意义

血管生成拟态(vasculogenic mimicry)是近两年来提出的一种全新的肿瘤内血管生成模式.其特点为肿瘤细胞通过自身变形和基质重塑产生血管样通道,通道内无内皮细胞衬覆,通道外基底膜PAS染色为阳性.该血管样通道生成机制、管壁结构等都与传统的肿瘤内血管生成不一样,因此通过血管生成拟态构建的血管样通道有可能成为某些肿瘤新的治疗靶点.     

VEGF165在HepG2体外形成血管生成拟态中的作用

 目的 探讨VEGF₁₆₅在人肝癌细胞株HepG2体外形成血管生成拟态（Vasculogenic mimicry,VM）中的作用。方法 三维细胞培养HepG2,观察其形成血管样通道的能力;构建pcDNA3.0 VEGF165和pcDNA3.0-VEGF₁₆₅b 真核表达质粒,以Lipofectamine2000转染HepG2细胞,RT-PCR及Western blot检测转染后各组细胞VEGF165和VEGF₁₆₅b的表达;三维培养转染后HepG2细胞,观察各组细胞形成血管生成拟态的能力。结果 HepG2细胞在三维细胞培养系统中可形成血管样通道;VEGF₁₆₅及VEGF₁₆₅b真核表达质粒转染HepG2细胞后,RT-PCR及Western blot均示VEGF165和VEGF₁₆₅b表达分别上调,而转染VEGF₁₆₅b质粒组VEGF₁₆₅表达下调;三维细胞培养转染后的HepG2细胞,各组形成血管生成拟态的能力无明显区别。结论 HepG2细胞可于三维培养系统中形成管道结构,但VEGF165对HepG2细胞体外形成血管生成拟态的能力无明显影响。     

肿瘤血管生成拟态研究进展

血管生成拟态（VM）是一种不依赖于血管内皮细胞的全新肿瘤微循环模式,VM的形成通过多条信号通路的转导。存在VM的肿瘤预后不良,因此在抑制肿瘤微循环的治疗中应兼顾VM与血管生成,尤其重视VM的治疗才能获得良好疗效。     

血管生成拟态及其在胶质瘤中的研究进展

近几年来,人们先后在黑素瘤、胶质瘤等肿瘤中发现了血管生成拟态。血管生成拟态是肿瘤细胞自身形成的,是肿瘤的微循环特殊形式。血管生成拟态的调节与基质金属蛋白酶、血管上皮钙黏素等的表达相关。血管生成拟态预测低分化、高浸润和低生存率,是预后不良的标志。所以经典抗血管治疗联合抗血管生成拟态治疗在抗肿瘤治疗中成为必要。     

胶质瘤干细胞血管生成拟态现象的体外研究

背景与目的：血管生成拟态是存在于恶性肿瘤中的,由肿瘤细胞而不是内皮细胞围成的管腔样结构,管腔中有血液流通并参与肿瘤微循环。我们采用体外三维培养模型来观察对比不同来源的胶质瘤干细胞血管生成拟态现象。方法：采用悬浮克隆球形成法诱导获得胶质瘤干细胞,应用胶质瘤干细胞相关分子标记免疫荧光技术鉴定及胶质瘤干细胞裸鼠移植成瘤试验证实已诱导获取的胶质瘤干细胞;采用体外三维培养模型来观察胶质瘤干细胞体外形成血管生成拟态现象。结果：成功获取6种不同来源的胶质瘤干细胞：GSC-1、GSC-2（来源于临床胶质母细胞瘤标本）和SKMG-4G、SKMG-1G、SF-295G、SF-767G（分别来源于胶质瘤细胞系SKMG-4、SKMG-1、SF-295、SF-767）。体外三维培养模型中,拟态管腔分圆形、多边形和三角形3种。GSC-1和SKMG-4G能形成典型的拟态管腔;SKMG-1G和SF-295G只能形成过渡性的非典型的拟态管腔;而GSC-2和SF-767G不能形成拟态管腔。结论：不同来源的胶质瘤干细胞拟态血管形成的能力不同,形态也各异。     

Dickkopf-1 对结肠癌组织血管生成拟态形成的抑制作用及相关机制研究*

目的：探讨结肠癌组织中Dickkopf-1（Dkk 1）表达对血管生成拟态（vasculogenic mimicry ,VM）的抑制作用及相关机制。方法：收集2002年1 月至2004年12月间天津医科大学肿瘤医院收治的217 例结肠癌患者资料,利用CD34-PAS对结肠癌组织进行双重染色及免疫组化方法检测并分析VM与Dkk 1 表达的关系;利用体外三维培养（three dimensional culture,3Dculture）观察Dkk 1 对人结肠癌HCT 116 细胞管状结构形成能力及血管内皮钙黏蛋白（VE-cadherin）表达的影响;建立裸鼠皮下移植瘤模型进一步明确Dkk 1 对肿瘤组织内VM形成的抑制作用。结果：存在VM的结肠癌组织中Dkk 1 表达较低（P < 0.05）;Dkk 1 过表达的HCT 116 细胞形成管状结构能力明显减弱,且VE-cadherin表达降低;Dkk 1 可抑制裸鼠移植瘤组织中HCT 116 细胞形成VM。结论：Dkk 1 过表达可抑制结肠癌中VM形成。

     

血管生成拟态的形成机制及其调控因素与临床意义

血管生成拟态(VM)能促进肿瘤细胞的生长、侵袭和转移.VM的形成机制与上皮细胞激酶、脂酰肌醇-3激酶、缺氧诱导因子1等多种蛋白分子相关,局部微环境也是重要的调控因素.     

肿瘤血管生成拟态研究新进展

血管生成拟态是独立于血管生成的一种全新肿瘤血管模式。越来越多的研究发现血管生成拟态的形成机制和肿瘤细胞自身的可塑性及肿瘤干细胞关系密切,另外,肿瘤微环境如细胞外基质重塑、缺氧等在血管生成拟态形成过程中也有很重要的作用。近年来,血管生成拟态形成机制的相关研究已经取得了很大进展,为寻找抑制VM的关键作用靶点、筛选有效的治疗药物提供了新的思路和方向。本文就血管拟态的发现、形成机制及其在肿瘤治疗中的研究现状等方面做一综述。     

血管生成拟态研究进展

目的:总结肿瘤血管生成拟态(VM)的研究进展,并探讨肿瘤血管生成拟态形成机制。方法:应用PubMed和CNKI期刊全文数据库检索系统,以血管生成拟态为关键词,检索1999-01-01-2011-06-30相关文献,以三维培养模型中VM形态学、分子调节机制及抗肿瘤治疗研究为入选标准,分析文献45篇。结果:VM的生成机制比较复杂,其中VE-cad-herin/EphA2/PI3K/MMP是VM形成的关键通路,同时缺氧的微环境、cAMP和COX-2等对VM结构的形成也起一定的作用。结论:VM与肿瘤的侵袭转移潜能以及较差的临床预后有关,为抗肿瘤治疗提供了新的研究靶点。     

Long noncoding RNA n339260 promotes vasculogenic mimicry and cancer stem cell development in hepatocellular carcinoma

Vasculogenic mimicry (VM) refers to the unique capability of aggressive tumor cells to mimic the pattern of embryonic vasculogenic networks. Cancer stem cells (CSC) represent a subpopulation of tumor cells endowed with the capacity for self‐renewal and multilineage differentiation. Previous studies have indicated that CSC may participate in the formation of VM. With the advance of high‐resolution microarrays and massively parallel sequencing technology, long noncoding RNAs (lncRNAs) are suggested to play a critical role in tumorigenesis and, in particular, the development of human hepatocellular carcinoma (HCC). Currently, no definitive relationship between lncRNA and VM formation has been described. In the current study, we demonstrated that expression of the lncRNA, n339260, is associated with CSC phenotype in HCC, and n339260 level correlated with VM, metastasis, and shorter survival time in an animal model. Overexpression of n339260 in HepG2 cells was associated with a significant increase in CSC. Additionally, the appearance of VM and vascular endothelial (VE)‐cadherin, a molecular marker of VM, was also induced by n339260 overexpression. Using a short hairpin RNA approach, n339260 was silenced in tumor cells, and knockdown of n339260 was associated with reduced VM and CSC. The results of this study indicate that n339260 promotes VM, possibly by the development of CSC. The related molecular pathways may be used as novel therapeutic targets for the inhibition of HCC angiogenesis and metastasis.     

肝细胞癌中血管生成拟态的三维细胞培养及组织学研究

背景与目的:血管生成拟态(vasculogenic mimicry,VM)是新近发现的一种存在于恶性肿瘤中的血液供应方式.拟态血管由肿瘤细胞围成,无内皮细胞衬覆的管道样结构,其间有血液通过.目前对拟态血管的研究多局限在高侵袭性、双向分化的恶性肿瘤,对上皮来源的肿瘤很少涉及.本研究通过细胞三维培养,免疫组织化学方法观察肝细胞癌(hepatocellular carcinoma,HCC)中是否存在血管生成拟态,并探讨其形成过程及结构特点.方法:建立肝癌细胞株HepG2的三维培养模型,并收集15例肝癌石蜡包埋样本进行过碘酸雪夫氏(PAS)及CD31免疫组织化学双重染色以及PAS与Ferritin免疫组织化学双重染色,观察肝细胞肝癌中的血管生成拟态.结果:肝癌细胞三维培养两天时伸出细长突起,培养7天时彼此之间相互连接形成网络样、环状结构.15例肝癌免疫组织化学染色可见血管内皮CD31染色均为阳性,肝癌细胞Ferritin染色均为阳性.其中7例免疫组织化学与组织化学双重染色可见CD31阴性Ferritin阳性的肝癌细胞围成管道样结构,一层PAS阳性物质将肿瘤细胞同管腔分隔,肿瘤细胞构成的管腔中可见红细胞存在.并观察到分化良好的肝癌中血管生成拟态的形成少于分化差的肝癌,随着分化程度的降低肝癌细胞逐渐出现CD31着色阳性倾向.结论:肝癌细胞具有进行自身变形并与胶原蛋白相互作用,形成血管样通道的能力.肿瘤细胞可以通过血管生成拟态结构获得血液供应.     

Prognostic significance and mechanisms of patterned matrix vasculogenic mimicry in hepatocellular carcinoma

Vasculogenic mimicry (VM), including tubular VM and patterned matrix VM, has been generally recognized as a new pattern of tumor neovascularization. Pilot studies of tubular VM showed that it was present in hepatocellular carcinoma (HCC) and associated with poor clinical prognosis. However, whether patterned matrix VM is clinically significant in HCC is unknown. To elucidate the effects of patterned matrix VM on prognosis of HCC and the mechanisms involved in VM formation, we examined 151 cases of surgically resected human HCC by immunohistochemistry and transmission electron microscopy and conducted hypoxic experiments on human HCC cell line MHCC97-H. We observed 31 of 151 (20.5%) cases exhibited evidence of patterned matrix VM. The expression of patterned matrix VM was associated with larger tumors (P = 0.042), vascular invasion (P = 0.016), high-grade HCC (P = 0.022), and late-stage HCC (P = 0.013). Kaplan–Meier survival analysis revealed that cases of the VM group had lower overall survival (OS) rate (P < 0.001) and disease-free survival (DFS) rate (P = 0.002) than that of the non-VM group. Univariate and multivariate analysis indicated that the presence of patterned matrix VM was independent adverse prognostic factor for both OS (P = 0.004) and DFS (P = 0.011). Expression of hypoxia-inducible factor 1 alpha (HIF-1alpha), matrix metalloproteinase (MMP)-2, and MMP-9 were higher in the VM group than in the non-VM group (P = 0.001, P = 0.030, P = 0.007, respectively). After VM formation was induced by hypoxia, up-regulated expression of HIF-1α, MMP-2, and MMP-9 was also detected in cells cultured under hypoxia condition. Our results indicate that patterned matrix VM exists in HCC, and it might serve as an unfavorable prognostic factor for HCC patients. It is possible that hypoxia via induction of expression of HIF-1alpha, MMP-2, and MMP-9 may enhance VM formation in HCC.

     

Prognostic significance and mechanisms of patterned matrix vasculogenic mimicry in hepatocellular carcinoma

Vasculogenic mimicry (VM), including tubular VM and patterned matrix VM, has been generally recognized as a new pattern of tumor neovascularization. Pilot studies of tubular VM showed that it was present in hepatocellular carcinoma (HCC) and associated with poor clinical prognosis. However, whether patterned matrix VM is clinically significant in HCC is unknown. To elucidate the effects of patterned matrix VM on prognosis of HCC and the mechanisms involved in VM formation, we examined 151 cases of surgically resected human HCC by immunohistochemistry and transmission electron microscopy and conducted hypoxic experiments on human HCC cell line MHCC97-H. We observed 31 of 151 (20.5%) cases exhibited evidence of patterned matrix VM. The expression of patterned matrix VM was associated with larger tumors (P = 0.042), vascular invasion (P = 0.016), high-grade HCC (P = 0.022), and late-stage HCC (P = 0.013). Kaplan-Meier survival analysis revealed that cases of the VM group had lower overall survival (OS) rate (P < 0.001) and disease-free survival (DFS) rate (P = 0.002) than that of the non-VM group. Univariate and multivariate analysis indicated that the presence of patterned matrix VM was independent adverse prognostic factor for both OS (P = 0.004) and DFS (P = 0.011). Expression of hypoxia-inducible factor 1 alpha (HIF-1alpha), matrix metalloproteinase (MMP)-2, and MMP-9 were higher in the VM group than in the non-VM group (P = 0.001, P = 0.030, P = 0.007, respectively). After VM formation was induced by hypoxia, up-regulated expression of HIF-1α, MMP-2, and MMP-9 was also detected in cells cultured under hypoxia condition. Our results indicate that patterned matrix VM exists in HCC, and it might serve as an unfavorable prognostic factor for HCC patients. It is possible that hypoxia via induction of expression of HIF-1alpha, MMP-2, and MMP-9 may enhance VM formation in HCC.     

Metastasis-associated in colon cancer-1 promotes vasculogenic mimicry in gastric cancer by upregulating TWIST1/2

Vasculogenic mimicry (VM) is a blood supply modality that is strongly associated with the epithelial-mesenchymal transition (EMT), TWIST1 activation and tumor progression. We previously reported that metastasis-associated in colon cancer-1 (MACC1) induced the EMT and was associated with a poor prognosis of patients with gastric cancer (GC), but it remains unknown whether MACC1 promotes VM and regulates the TWIST signaling pathway in GC. In this study, we investigated MACC1 expression and VM by immunohistochemistry in 88 patients with stage IV GC, and also investigated the role of TWIST1 and TWIST2 in MACC1-induced VM by using nude mice with GC xenografts and GC cell lines. We found that the VM density was significantly increased in the tumors of patients who died of GC and was positively correlated with MACC1 immunoreactivity (p < 0.05). The 3-year survival rate was only 8.6% in patients whose tumors showed double positive staining for MACC1 and VM, whereas it was 41.7% in patients whose tumors were negative for both MACC1 and VM. Moreover, nuclear expression of MACC1, TWIST1, and TWIST2 was upregulated in GC tissues compared with matched adjacent non-tumorous tissues (p < 0.05). Overexpression of MACC1 increased TWIST1/2 expression and induced typical VM in the GC xenografts of nude mice and in GC cell lines. MACC1 enhanced TWIST1/2 promoter activity and facilitated VM, while silencing of TWIST1 or TWIST2 inhibited VM. Hepatocyte growth factor (HGF) increased the nuclear translocation of MACC1, TWIST1, and TWIST2, while a c-Met inhibitor reduced these effects. These findings indicate that MACC1 promotes VM in GC by regulating the HGF/c-Met-TWIST1/2 signaling pathway, which means that MACC1 and this pathway are potential new therapeutic targets for GC.     

Frizzled 2‐induced epithelial‐mesenchymal transition correlates with vasculogenic mimicry,stemness, and Hippo signaling in hepatocellular carcinoma

Prior observation has indicated that Frizzled 2 (FZD2)‐induced epithelial‐mesenchymal transition (EMT) could be a key step in metastasis and early recurrence of hepatocellular carcinoma (HCC). However, the mechanism underlying tumor development and progression due to aberrant FZD2 expression is poorly defined. Here, we provide evidence that FZD2 is a driver for EMT, cancer stem cell properties, and vasculogenic mimicry (VM) in HCC. We found that FZD2 was highly expressed in two cohorts of Chinese hepatitis B virus‐related HCC patients, and that high FZD2 expression was associated with poor prognosis. Concerning the mechanism, gain‐ and loss‐of‐function experiments showed the oncogenic action of FZD2 in HCC cell proliferation, apoptosis, migration, and invasion. Further investigations in vitro and in vivo suggested that FZD2 promotes the EMT process, enhances stem‐like properties, and confers VM capacity to HCC cells. Notably, integrative RNA sequencing analysis of FZD2‐knockdown cells indicated the enrichment of Hippo signaling pathway. Taken together, our data suggest for the first time that FZD2 could promote clinically relevant EMT, CD44⁺ stem‐like properties, and the VM phenotype in HCC involving a potential Hippo signaling pathway‐dependent mechanism, and should be considered as a promising therapeutic target for the treatment of HCC.     

Microbial metabolite deoxycholic acid promotes vasculogenic mimicry formation in intestinal carcinogenesis

A high‐fat diet (HFD) leads to long‐term exposure to gut microbial metabolite secondary bile acids, such as deoxycholic acid (DCA), in the intestine, which is closely linked to colorectal cancer (CRC). Evidence reveals that vasculogenic mimicry (VM) is a critical event for the malignant transformation of cancer. Therefore, this study investigated the crucial roles of DCA in the regulation of VM and the progression of intestinal carcinogenesis. The effects of an HFD on VM formation and epithelial‐mesenchymal transition (EMT) in human CRC tissues were investigated. The fecal DCA level was detected in HFD‐treated Apc ^min/+ mice. Then the effects of DCA on VM formation, EMT, and vascular endothelial growth factor receptor 2 (VEGFR2) signaling were evaluated in vitro and in vivo. Here we demonstrated that compared with a normal diet, an HFD exacerbated VM formation and EMT in CRC patients. An HFD could alter the composition of the gut microbiota and significantly increase the fecal DCA level in Apc ^min/+ mice. More importantly, DCA promoted tumor cell proliferation, induced EMT, increased VM formation, and activated VEGFR2, which led to intestinal carcinogenesis. In addition, DCA enhanced the proliferation and migration of HCT‐116 cells, and induced EMT process and vitro tube formation. Furthermore, the silence of VEGFR2 reduced DCA‐induced EMT, VM formation, and migration. Collectively, our results indicated that microbial metabolite DCA promoted VM formation and EMT through VEGFR2 activation, which further exacerbated intestinal carcinogenesis.