Tumor-associated mesothelial cells are negative prognostic factors in gastric cancer and promote peritoneal dissemination of adherent gastric cancer cells by chemotaxis

Peritoneal dissemination is highly frequent in gastric cancer. Damage to human peritoneal mesothelial cell (HPMC) barriers provokes gastric cancer peritoneal dissemination (GCPD), the key events during GCPD, is characterized by fibroblastic development. In this study, we have studied the association between fibroblast activation protein (FAP) expression in peritoneum and the pathological features of the primary tumor. The clinical prognosis of gastric cancer patients was evaluated according to FAP expression. In a gastric cancer cell-HPMC co-culture system, expression of E-cadherin, α-smooth muscle actin, and FAP were evaluated by Western blotting. Gastric cancer cell migration and adhesion to HPMC were also assayed. Our results showed positive peritoneal staining of FAP in 36/86 cases (41.9 %), which was associated with a higher TNM stage in primary gastric cancer and higher incidence of GCPD (both p?<?0.05). Survival analysis showed FAP expression was an independent prognostic factor of poor survival (p?=?0.02). Peritoneum of FAP-positive expression exhibited a distinct fibrotic development and expressed higher level of the mesenchymal marker α-SMA, which was confirmed by the in vitro Western blot assay. In HPMC and gastric cancer cell adherence assay, SGC-7901 cells preferentially adhered to TA-HPMC at different cell densities (both p?<?0.05). Additionally, SGC-7901 cells were more prone to chemotaxis by FAP-expressed tumor-associated–human peritoneal mesothelial cells (TA-HPMC) compared with HPMC co-cultured with normal gastric glandular epithelial cells in a time-dependent manner (both p?<?0.05). Our study indicated a positive correlation between peritoneum FAP expression and GCPD. FAP-expressed TA-HPMC might be an important cellular component and instigator of GCPD.     

Gli-1信号通路对TGF-β1诱导的人胃癌细胞上皮间质转化和侵袭的影响

目的:探讨核转录因子神经胶质瘤关联癌基因同源物1（glioma associated oncogene homolog 1,Gli-1）对转化生长因子-β1(transforming growth factor-β1,TGF-β1)诱导的人胃癌SGC-7901细胞发生上皮间质转化（epithelial-mesenchymal transition,EMT）的作用机制。方法:体外采用10 ng/ml TGF-β1对胃癌SGC-7901细胞进行处理,使用倒置显微镜观察细胞形态变化,RT-PCR和Western blot检测EMT上皮表型蛋白E-cadherin和间质表型蛋白Vimentin的表达水平;Transwell细胞侵袭实验检测细胞侵袭能力变化,检测TGF-β1 对SGC-7901细胞发生EMT 的影响;同时采用RT-PCR和Western blot检测Gli-1的mRNA和蛋白表达水平。随后进一步采用Gli-1基因特异性阻断剂GANT 61阻断Gli-1表达,并使用TGF-β1（10 ng/ml）对SGC-7901细胞进行处理,RT-PCR 和Western blot检测Gli-1、E-cadherin和Vimentin mRNA 及其蛋白表达水平的改变,并使用Transwell细胞侵袭实验检测阻断Gli-1对SGC-7901细胞侵袭能力的影响。结果:TGF-β1可以诱导人胃癌SGC-7901细胞发生上皮间质转化并促进细胞侵袭。TGF-β1可以在mRNA和蛋白水平下调上皮表型蛋白E-cadherin的表达、提高Gli-1和间质表型蛋白Vimentin的表达。TGF-β1可以明显提高SGC-7901细胞侵袭,而阻断Gli-1后可以抑制TGF-β1诱导的人胃癌SGC-7901细胞上皮间质转化和细胞侵袭。结论:胃癌SGC-7901细胞中Gli-1 可能参与TGF-β1 介导的EMT 的发生,Gli-1 可能作为胃癌基因治疗中的有效靶点发挥作用。     

The Angiotensin II type 1 receptor blocker candesartan suppresses proliferation and fibrosis in gastric cancer

Gastric cancer with peritoneal dissemination has poor clinical prognosis because of the presence of rich stromal fibrosis and acquired drug resistance. Recently, Angiotensin II type I receptor blockers such as candesartan have attracted attention for their potential anti-fibrotic activity. We examined whether candesartan could attenuate tumor proliferation and fibrosis through the interaction between gastric cancer cell line (MKN45) cells and human peritoneal mesothelial cells. Candesartan significantly reduced TGF-β1 expression and epithelial-to-mesenchymal transition-like change, while tumor proliferation and stromal fibrosis were impaired. Targeting the Angiotensin II signaling pathway may therefore be an efficient strategy for treatment of tumor proliferation and fibrosis.     

Induction of gastric cancer cell adhesion through transforming growth factor-beta1-mediated peritoneal fibrosis

Background

Peritoneal dissemination is one of the main causes of death in gastric cancer patients. Transforming growth factor-beta1 (TGF-β1), one of the most potent fibrotic stimuli for mesothelial cells, may play a key role in this processing. The purpose of this study is to elucidate the effects of TGF-β1 on regulation of gastric cancer adhesion to mesothelial cells.

Methods

Peritoneal tissues and peritoneal wash fluid were obtained for hematoxylin and eosin staining or ELISA to measure fibrosis and TGF-β1 levels, respectively. The peritoneal mesothelial cell line, HMrSV5, was used to determine the role of TGF-β1 in regulation of gastric cancer cell adhesion to mesothelial cells and expression of collagen, fibronectin, and Smad 2/3 by using adhesion assay, western blot, and RT-PCR.

Results

The data showed that TGF-β1 treatment was able to induce collagen III and fibronectin expression in the mesothelial cells, which was associated with an increased adhesion ability of gastric cancer cells, but knockdown of minimal sites of cell binding domain of extracellular matrix can partially inhibit these effects.

Conclusion

Peritoneal fibrosis induced by TGF-β1 may provide a favorable environment for the dissemination of gastric cancer.     

二甲双胍通过AMPK/Smad3信号通路抑制卵巢癌腹腔间皮细胞的间皮-间质转化

目的:探讨二甲双胍对人腹腔间皮细胞（human peritoneal mesothelial cells,HPMC）间皮-间质转化的影响及其可能机制,以及进一步对卵巢癌细胞系SKOV3系膜清除能力的影响。方法:搜集2018年02月至2021年02月襄阳市第一人民医院妇产科正常网膜组织和高级别浆液性卵巢癌网膜转移灶标本进行免疫组织化学检测Calretinin表达情况;分离、培养原代人腹腔间皮细胞并进行Calretinin流式鉴定;Western blot和Transwell实验检测二甲双胍或肿瘤上清处理正常患者来源HPMC后其间皮-间质转化相关蛋白（E-cadherin、Vimentin、α-SMA）及迁移能力改变情况;Western blot检测二甲双胍、Smad3抑制剂及AMPK干扰处理前后正常来源原代人腹腔间皮细胞中MMP2、E-cadherin、Vimentin、p-AMPK和p-Smad3的表达情况;SKOV3球体的系膜清除实验验证二甲双胍预处理人腹膜间皮细胞系HMrSV5对SKOV3系膜清除能力的影响。结果:正常网膜中Calretinin表达于腹腔侧表面的腹腔内皮细胞,卵巢癌网膜转移灶表面Calretinin表达缺失,肿瘤间质中高表达Calretinin;分离培养的肿瘤来源原代人腹腔间皮细胞流式检测表现为CD326-CD45-CD31-Calretinin+;二甲双胍能够抑制肿瘤上清诱导的正常来源原代HPMC的间皮-间质转化及迁移;二甲双胍能够抑制肿瘤上清诱导的正常来源原代HPMC中Smad3/MMP2通路活化和E-cadherin表达,并依赖于AMPK信号活化;二甲双胍预处理HMrSV5通过AMPK信号的介导抑制了HMrSV5的间皮-间质转化进而间接抑制了SKOV3的系膜清除能力。结论:卵巢癌转移灶中的间质成分部分来源于腹腔间皮细胞,二甲双胍通过AMPK调控的Smad3信号通路抑制其间皮-间质转化,进而抑制卵巢癌肿瘤细胞的系膜清除。     

The role of human peritoneal mesothelial cells in the fibrosis and progression of gastric cancer

Peritoneal dissemination is the most frequent metastatic pattern of scirrhous gastric cancer. However, despite extensive research effort, disease outcomes have not improved sufficiently. Tumor progression and metastasis result from interactions between cancer and various cells in the stroma, including endothelial cells, immune cells and fibroblasts. Fibroblasts have been particularly well studied; they are known to change into carcinoma-associated fibroblasts (CAFs) and produce transforming growth factor β (TGF-β), which mediates cancer-stroma interactions. Here, we investigated whether TGF-β derived from cancer cells in the peritoneal microenvironment activates human peritoneal mesothelial cells (HPMCs), leading to the progression and fibrosis of gastric cancer. We found that activated HPMCs (a-HPMCs) took on a spindle shape formation, decreased the expression of E-cadherin and increased that of α-SMA. Furthermore, a-HPMCs became more invasive and upregulated proliferation of human gastric cancer-derived MKN45 cells following direct cell-cell contact. Notably, MKN45 cells co-cultured with a-HPMCs also acquired anchorage-independent cell growth and decreased expression of E-cadherin in vitro. To measure the effects of the co-culture in vivo, we developed a mouse xenograft model into which different culture products were subcutaneously injected. The largest tumors were observed in mice that had been given MKN45 cells co-cultured with a-HPMCs. Furthermore, these tumors contained HPMC-derived fibrous tissue. Thus, the epithelial-mesenchymal transition (EMT) of HPMCs appears to drive peritoneal dissemination and tumor fibrosis.     

乳斑肿瘤相关巨噬细胞诱导人腹膜间皮HMR-sv5细胞损伤的机制研究

  目的   研究胃癌细胞对巨噬细胞的诱导作用, 分析活化巨噬细胞对间皮细胞的损伤作用及机制。   目法   人正常胃腺上皮细胞系GES-1及人低分化胃癌细胞系SGC-7901与人单核巨噬细胞THP-1共培养, 诱导后者分化, 研究后者对人腹膜间皮细胞HMR-sv5的损伤作用及分子机制。   结果   胃癌细胞诱导THP-1形成肿瘤相关巨噬细胞(TAM), M1型巨噬细胞表面抗原的表达显著下调, 而M2表型的表面抗原明显上调, 细胞形态也发生明显改变。TAM显著抑制正常间皮细胞生长, 促进间皮细胞凋亡和上皮间质转化。   结论   胃癌细胞诱导巨噬细胞发生表型和功能转化, 进而导致间皮细胞发生EMT和凋亡, 促进形成腹膜转移癌。      

The cytotoxic effect of TGF-β1 on mesothelial cells via apoptosis in early peritoneal carcinomatosis

Peritoneal dissemination is one of the main causes of death in gastric cancer patients. We have previously reported that gastric cancer cells can induce peritoneal apoptosis, lead to damage of peritoneum integrity, and therefore promote peritoneal metastasis. However, the soluble factors secreted by cancer cells to trigger the damaging cascade remain unclear. TGF-β1, a cytokine known for its capacity to induce proliferative and transformative changes of cells is found in significantly higher quantities correlated with peritoneal metastasis and TNM stages of gastric cancer. High levels of TGF-β1 in the subperitoneal milieu may affect the morphology and function of mesothelial cells, so that the resulting environment becomes favorable for peritoneal metastases. We observed apoptosis induced by TGF-β1 in mesothelial cells in peritoneal carcinomatosis. Knockdown of the smad2 gene by siRNA silencing can partially inhibit these effects. TGF-β1 could upregulate the expressions of Bax and suppress Bcl-2 in mesothelial cells. We conclude that TGF-β1 could induce apoptosis of mesothelial cells, which involves the smad2 signaling pathway in peritoneal carcinomatosis. Bcl-2 and Bax may contribute to this phenomenon.     

Hepatocyte growth factor (HGF) produced by peritoneal fibroblasts may affect mesothelial cell morphology and promote peritoneal dissemination

Mesothelial cell monolayers have been reported to prevent infiltration of cancer cells into the peritoneum. We have previously reported that peritoneal fibrosis induced by gastric cancer cells prior to metastatization may provide a congenial environment for peritoneal metastases. In this study, we investigated the effects of peritoneal fibroblasts on peritoneal mesothelial cell morphology. Human gastric cancer (OCUM-2MD3), peritoneal fibroblast (NF-2P) and mesothelial (MS-1) cell lines were established in our laboratory. Histology of the peritoneum was investigated following intraperitoneal inoculation of serum-free conditioned media (SF-CM) from OCUM-2MD3 cells into nude mice. SF-CM from peritoneal fibroblasts was added to monolayer-cultured mesothelial cells, and their morphology was examined by phase-contrast microscopy. This experiment was conducted in the presence and absence of neutralizing antibodies against various factors. Mesothelial cells exposed to fibroblasts proliferation became hemispherical and separated from each other, while unexposed mesothelium remained as a flat monolayer. Cultured-mesothelial cells rounded up or exhibited a fibroblast-like shape following the addition of peritoneal fibroblast SF-CM. Anti-hepatocyte growth factor (HGF) neutralizing antibody partly inhibited this effect. We suggest that soluble factors, such as HGF, produced by peritoneal fibroblasts affect the morphology of mesothelial cells in monolayers so that the resulting environment may become prone to the peritoneal dissemination of cancer cells. © 1996 Wiley-Liss, Inc.     

胃癌细胞与腹膜间皮相互作进用促腹膜纤维化致腹膜转移的研究

  目的   研究间皮细胞对胃癌细胞的抵御作用, 模拟游离癌细胞在接触腹膜前通过其分泌物导致的腹膜增厚、纤维化的过程, 同时观察了间皮细胞对胃癌细胞迁移及侵袭能力的反作用。   方法  用荧光显微镜观察共培养时间皮细胞对胃癌细胞的抵御作用; 体内实验观察腹膜变化; 电镜、光镜下观察体外实验中间皮层在胃癌-腹膜相互作用中的损伤和细胞骨架变化; 采用Millicell小室共培养观察间皮细胞对胃癌细胞迁移及侵袭能力的反作用。   结果  正常间皮细胞可防止肿瘤细胞对于腹膜的粘附, 受损脱落后胃癌细胞可轻易粘附。体内外实验均显示接触胃癌细胞上清后腹膜间皮细胞受损、凋亡, 并且受损残余的间皮可以反作用于癌细胞, 使其迁移转移力提高。   结论  正常间皮细胞可以抵御胃癌细胞的侵袭, 受损伤刺激后的间皮细胞可以反作用于胃癌细胞促进其迁移及侵袭。      

Importance of human peritoneal mesothelial cells in the progression,fibrosis, and control of gastric cancer: inhibition of growth and fibrosis by tranilast

Background

Scirrhous gastric cancer is an intractable disease with a high incidence of peritoneal dissemination and obstructive symptoms (e.g., ileus, jaundice, and hydronephrosis) arising from accompanying marked fibrosis. Microenvironmental interactions between cancer cells and cancer-associated fibroblasts are the suggested cause of the disease. We elucidated the mechanisms of tumor growth and fibrosis using human peritoneal mesothelial cells (HPMCs) and investigated the effects of tranilast treatment on cells and a xenograft mouse model of fibrosis.

Methods

HPMCs were isolated from surgically excised omentum and their interaction with MKN-45 gastric cancer cells was investigated using co-culture. Furthermore, a fibrosis tumor model was developed based on subcutaneous transplantation of co-cultured cells into the dorsal side of nude mice to form large fibrotic tumors. Mice were subsequently treated with or without tranilast.

Results

The morphology of HPMCs treated with transforming growth factor (TGF)-β1 changed from cobblestone to spindle-type. Moreover, E-cadherin was weakly expressed whereas high levels of α-smooth muscle actin expression were observed. TGF-β-mediated epithelial–mesenchymal transition-like changes in HPMCs were inhibited in a dose-dependent manner following tranilast treatment through inhibition of Smad2 phosphorylation. In the mouse model, tumor size decreased significantly and fibrosis was inhibited in the tranilast treatment group compared with that in the control group.

Conclusions

Tranilast acts on the TGF-β/Smad pathway to inhibit interactions between cancer cells and cancer-associated fibroblasts, thereby inhibiting tumor growth and fibrosis. This study supports the hypothesis that tranilast represents a novel strategy to prevent fibrous tumor establishment represented by peritoneal dissemination.

     

HMGA2在胃癌细胞SGC-7901上皮间质转化中的作用

目的 探讨TGF-β1调控人胃癌细胞SGC-7901由上皮向间质细胞转化过程中,HMGA2的表达及其作用。方法 用TGF-β1处理人胃癌细胞SGC-7901(0,3,6,24,48,72h),在显微镜下观察SGC-7901细胞形态的变化,蛋白质印记法检测HMGA2、E-cadherin和Vimentin蛋白的表达,Transwell小室实验检测细胞侵袭能力。结果 (1)随着TGF-β1作用时间的延长,SGC-7901细胞逐渐变长,变大,细胞融合;(2)在TGF-β1作用下,SGC-7901均不同程度地表达HMGA2、E-cadherin和Vimentin。HMGA2蛋白从0h开始表达,到6h表达最强,24h以后表达稍有下降。E-cad蛋白从0h开始表达,到3h表达最高,随后开始降低。Vimentin蛋白48h表达最高,72h有降低的趋势;(3)Transwell小室实验显示SGC-7901在TGF-β1处理后,3h时细胞穿膜最多,从6h起穿过的细胞开始降低,到72h最低。结论 在TGF-β1诱导人胃癌细胞SGC-7901细胞上皮间质转化的过程中,HMGA2蛋白起到重要作用,HMGA2可作为Snail的启动子,结合Smads,促进上皮间质转化HMGA2还可能通过调控Wnt/β-Catenin信号通路对肿瘤细胞的生长和凋亡发挥作用。     

Adhesion molecules and TGF-β1 are involved in the peritoneal dissemination of NUGC-4 human gastric cancer cells

Peritoneal dissemination frequently occurs after surgery in patients with gastric cancer. The presence of peritoneal metastasis after surgery affects prognosis. Very little is known about the biochemical processes involved in the initial attachment of gastric cancer cells to peritoneal mesothelial cells. We conducted in vitro and in vivo studies to assess the role of adhesion molecules and TGF-β1 in this process, using 4 cell lines derived from human gastric cancers. NUGC-4 cells, which disseminate early after inoculation into the abdominal cavity of nude mice, predominantly express CD44H and β₁ integrin. We found that NUGC-4 cells adhered to monolayers of mesothelial cells more firmly than to other cell lines. Adhesion of NUGC-4 cells to mesothelial cells was partially inhibited by antibodies against CD44H or the β₁ subunit of integrin and was completely blocked by a combination of these 2 antibodies. Treatment with ligands for CD44H and β₁ integrin also inhibited adhesion. In the NUGC-4 cell culture medium, larger amounts of TGF-β₁ were detected in relation to the increase in cancer cells than in the other cell lines. TGF-β1 increased the expression of CD44H in NUGC-4 cells and in mesothelial cells and augmented adhesion and implantation of NUGC-4 cells to mesothelial cells accompanied by accumulation of extracellular matrix (ECM) components. Treatment with antibodies against both CD44H and β₁ integrin inhibited the dissemination of NUGC-4 cells in the peritoneal cavity of nude mice and prolonged their survival time. Our findings suggest that CD44H and integrins mediate the initial attachment of gastric cancer cells to mesothelial cells and that TGF-β1 participates in the promotion of the disease. Increased expression of CD44H and of the amount of ligands for CD44H and integrins induced by TGF-β1 promotes early development of peritoneal dissemination. Int. J. Cancer 70:612–618. © 1997 Wiley-Liss Inc     

胃癌细胞促进网膜脂肪干细胞分化的机制研究

  目的  本实验主要研究在胃癌条件培养基（conditioned medium,CM）诱导下网膜脂肪干细胞（omental-adipose stromal cells,O-ASCs）是否能分化为癌相关成纤维细胞（carcinoma-associated fibroblasts,CAFs）,及ERK信号通路在其中的作用。  方法  通过诱导分化成骨、成脂及流式细胞鉴定O-ASCs,将O-ASCs与MGC803和SGC7901 CM共培养,通过RT-PCR和Western-blot检测O-ASCs细胞CAFs标志物α-SMA、FSP-1、vimentin,旁分泌因子VEGFA、TGFβ-1、FAP、SDF-1的表达水平。将O-ASCs分为对照组,SGC7901-CM实验组,SGC7901-CM+U0126处理组,12 h后收集细胞。Western blot检测O-ASCs细胞CAFs标志物α-SMA、FSP-1及ERK1/2、p-ERK1/2的表达水平。  结果  经鉴定原代培养出的细胞为O-ASCs,在SGC7901 CM和MGC803 CM作用下,CAFs标志物α-SMA、FSP-1、vimentin及旁分泌因子SDF-1、VEGFA、TGFβ-1、FAP表达均有明显增加（P < 0.05）。与对照组比较SGC7901-CM组α-SMA、FSP-1、p-ERK1/2表达明显增加（P < 0.05）,ERK表达未见明显变化（P > 0.05）。SGC7901-CM+U0126组与SGC7901-CM组比较,α-SMA、FSP-1及p-ERK1/2的蛋白表达水平明显降低（P < 0.05）,ERK表达变化无统计学意义（P > 0.05）。  结论  O-ASCs通过分化为CAFs及旁分泌作用参与胃癌腹膜转移,ERK信号通路在该过程中发挥了重要作用。      

IRX1 influences peritoneal spreading and metastasis via inhibiting BDKRB2-dependent neovascularization on gastric cancer

The overexpression of IRX1 gene correlates with the growth arrest in gastric cancer. Furthermore, overexpression of IRX1 gene suppresses peritoneal spreading and long distance metastasis. To explore the precise mechanisms, we investigated whether restoring IRX1 expression affects the angiogenesis or vasculogenic mimicry (VM). Human umbilical vein endothelial cells (HUVECs) and chick embryo and SGC-7901 gastric cancer cells were used for angiogenesis and VM analysis. Small interfering RNA was used for analyzing the function of BDKRB2, a downstream target gene of IRX1. As results, the remarkable suppression on peritoneal spreading and pulmonary metastasis of SGC-7901 cells by IRX1 transfectant correlates to reduced angiogenesis as well as VM formation. Using the supernatant from SGC-7901/IRX1 cells, we found a strong inhibiting effect on angiogenesis both in vitro and in chick embryo. SGC-7901/IRX1 cells revealed strong inhibiting effect on VM formation too. By gene-specific RNA interference for BDKRB2, or its effector PAK1, we got an effective inhibition on tube formation, cell proliferation, cell migration and invasion in vitro. In conclusion, enforcing IRX1 expression effectively suppresses peritoneal spreading and pulmonary metastasis via anti-angiogenesis and anti-VM mechanisms, in addition to previously found cell growth and invasion. BDKRB2 and its downstream effector might be potential targets for anti-cancer strategy.     

Peritoneal dissemination is inhibited by treatment with antibodies against CD44H, beta(1) integrin, and carcinostatic agents in NUGC-4 human gastric cancer cells

Peritoneal dissemination frequently occurs after surgery in patients with gastric cancer. The presence of peritoneal metastasis after surgery affects the prognosis, therefore, a way must be found to effectively prevent the development of peritoneal dissemination. Very little is known about the biochemical processes involved in the initial attachment of gastric cancer cells to peritoneal mesothelial cells. We conducted in vitro and in vivo studies to assess the role of adhesion molecules and TGF-beta 1 in this process, using 4 gastric cancer cell lines. NUGC-4 cells, which disseminate early after inoculation into the abdominal cavity of nude mice, predominantly expressed CD44H and beta(1) integrin. We found that NUGC-4 cells adhered to monolayers of mesothelial cells more firmly than other cell lines. Adhesion of NUGC-4 cells to mesothelial cells was partially inhibited by antibodies against CD44H or the beta(1) subunit of integrin, and was completely blocked by a combination of these 2 antibodies. Treatment with ligands for CD44H and beta(1) integrin also inhibited this adhesion. In the NUGC-4 cell culture medium, larger amounts of transforming growth factor beta 1(TGF-beta 1) was detected in proportion to the increase in cancer cells than in the other cell lines. TGF-beta 1 increased the expression of CD44H in NUGC-4 cells and in mesothelial cells, and augmented the adhesion and implantation of NUGC-4 cells to mesothelial cells accompanied by accumulation of extracellular matrix (ECM) components. Carcinostatic agents decreased the expression of CD44H but increased the expression of E-cadherin in NUGC-4 cells. Treatment with carcinostatic agents and antibodies against CD44H and beta(1) integrin inhibited the dissemination of NUGC-4 cells in the peritoneal cavity of nude mice, and prolonged their survival time. These findings suggest that CD44H and integrins mediate in the initial attachment of gastric cancer cells to mesothelial cells, and TGF-beta 1 participates in the promotion of the disease. It is possible that a treatment strategy that interferes with CD44H or integrins function and increases the functions of E-cadherin immediately after surgery may result in the decreased intra-abdominal spread of gastric cancer.     

Correlation of peritoneal dissemination and integrin alpha 3 expression in gastric cancer

To clarify the interrelationship between alpha 3 integrin, subunit/E-cadherin expression and peritoneal dissemination in gastric cancer, alpha 3 integrin subunit and E-cadherin expression gas were immunohistochemically examined and were correlated with clinicopathologic parameters. Among 150 primary gastric cancers, alpha 3 integrin subunit and E-cadherin were strongly expressed in 96 (65%) and 88 (59%) tumors, respectively. Integrin alpha 3 expression was closely associated with peritoneal dissemiantion, but there was no relationship between integrin alpha 3 expression and histologic type, nodal status, macroscopic type or wall invasion. Furthermore, 22 (84%) of 26 tumors which recurred in peritoneum overexpressed integrin alpha 3 expression in primary tumors. All seven peritoneal foci obtained from peritoneal dissemination expressed integrin alpha 3 expression despite no integrin alpha 3 expression in two of these 7 primary tumors. Reduced E-cadherin expression in primary tumors was intimately associated with large tumor size (>6 cm), nodal involvement, peritoneal dissemination and positive serosal invasion. Peritoneal dissemination was most frequently found in the tumors with positive integrin alpha 3 expression and reduced E-cadherin expression. Patients with these type of tumor [E-cadherin expression (-), and integrin alpha 3 subunit (+)] showed the poorest prognosis as compared with the other groups of patients. These results indicate that upregulation of integrin alpha 3 expression and down regulation of E-cadherin might have an important role in the formation of peritoneal dissemination. These tumors have characteristics of easy detachment from the serosal surface via downregulation of E-cadherin and strong adhesion capacity to the peritoneum via up-regulation of integrin alpha 3 expression. The immunohisto-chemical combination analysis of E-cadherin and integrin alpha 3 expression on the primary gastric cancer may be a good screening method to predict peritoneal recurrence.     

川陈皮素抑制胃癌SGC-7901细胞侵袭能力的机制探讨

目的:观察川陈皮素（nobiletin,Nob）对胃癌SGC-7901细胞侵袭能力的影响,并探讨其可能的作用机制。方法:不同浓度的川陈皮素（0、30、60、120 mg/L）体外处理SGC-7901细胞。CCK8法检测细胞增殖能力;Transwell小室法检测细胞侵袭能力的改变;免疫印迹法检测细胞上皮间质转化（epithelial-mesenchymal transition,EMT)标志物（E-cadherin、Vimentin、Fibronectin）蛋白、MMP-9、STAT3、p-STAT3蛋白表达的改变。结果:CCK8和Transwell小室法检测结果显示,与对照组（0 mg/L）比较,川陈皮素可显著抑制SGC-7901细胞的增殖和侵袭能力;免疫印迹结果显示,随着川陈皮素浓度的增加,SGC-7901细胞E-cadherin蛋白表达逐渐上调,同时Vimentin、Fibronectin蛋白的表达量降低,MMP-9蛋白表达下调,STAT3总量未发生变化,但p-STAT3表达逐渐降低。结论:川陈皮素可降低胃癌SGC-7901细胞侵袭能力,其机制可能为通过抑制STAT3通路,继而抑制EMT。     

胃癌细胞中COX-2通过NF-κB/Snail调控E-cadherin表达的机制

目的 探讨COX-2调控E-cadherin表达的相关信号通路及分子机制,揭示COX-2与胃癌侵袭转移的关系及作用机制.方法 采用不同浓度的选择性COX-2抑制剂塞来昔布干预人胃癌细胞株SGC-7901不同时间后,应用实时荧光定量RT-PCR法和Western blot法检测SGC-7901细胞中COX-2、NF-κB、Snail及E-cadherin mRNA和蛋白的表达情况.结果 随着塞来昔布作用剂量的增大和干预时间的延长,COX-2、NF-κB和Snail mRNA及蛋白的表达量均显著下降(P＜0.05),呈剂量和时间依赖性降低;E-cadherin mRNA及蛋白的表达量上升(P＜0.05),呈剂量和时间依赖性升高.采用Spearman相关分析,显示COX-2与NF-κB及COX-2与Snail蛋白表达呈正相关性(r =0.881,P＜0.01;r =0.839,P＜0.01);COX-2与E-cadherin蛋白表达呈负相关性(r=-0.814,P＜0.01).结论 COX-2可能通过NF-κB/Snail信号通路调控E-cadherin的表达,进而参与胃癌的浸润转移过程.     

Induction of apoptosis in human peritoneal mesothelial cells by gastric cancer cell supernatant promotes peritoneal carcinomatosis

This study was carried out to evaluate the effects of gastric cancer cell supernatant on human peritoneal mesothelial cell viability and apoptosis and to investigate the mechanism of action of gastric cancer in a mesothelial cell line (HMrSV5). Cell viability was measured using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazoliumbromide (MTT) assay. Mesothelial cells treated with gastric cancer cell supernatant were stained with acridine orange/ethidium bromide and subjected to fluorescence microscopy. C57BL/6 mice were used to establish a cancer invasion model. Morphological changes and exfoliation occurred, and naked areas appeared in both cultured mesothelial cells and the parietal peritoneum after treatment with gastric cancer cell supernatant. Cell supernatant from gastric cancer cells induced apoptosis of mesothelial cells in a time-dependent manner. Obvious morphological changes of cell apoptosis were detected, such as condensation of chromatin, nuclear fragmentations, and apoptotic ladders. These findings demonstrate that gastric cancer cells induce apoptosis of human peritoneal mesothelial cells through supernatants in early peritoneal metastasis.