Gastric cancer cell supernatant causes apoptosis and fibrosis in the peritoneal tissues and results in an environment favorable to peritoneal metastases, in vitro and in vivo.
ABSTRACT In this study, we examined effects of soluble factors released by gastric cancer cells on peritoneal mesothelial cells in vitro and in vivo.
HMrSV5, a human peritoneal mesothelial cell line, was incubated with supernatants from gastric cancer cells. Morphological changes of HMrSV5 cells were observed. Apoptosis of HMrSV5 cells was observed under a transmission electron microscope and quantitatively determined by MTT assay and flow cytometry. Expressions of apoptosis-related proteins (caspase-3, caspase-8, Bax, bcl-2) were immunochemically evaluated.
Conspicuous morphological changes indicating apoptosis were observed in HMrSV5 cells 24 h after treatment with the supernatants of gastric cancer cells. In vivo, peritoneal tissues treated with gastric cancer cell supernatant were substantially thickened and contained extensive fibrosis.
These findings demonstrate that supernatants of gastric cancer cells can induce apoptosis and fibrosis in HMrSV5 human peritoneal mesothelial cells through supernatants in the early peritoneal metastasis, in a time-dependent manner, and indicate that soluble factors in the peritoneal cavity affect the morphology and function of mesothelial cells so that the resulting environment can become favorable to peritoneal metastases.
Full-textDOI: · Available from: Zhi-Feng Miao, Apr 29, 2014
[Show abstract] [Hide abstract]
ABSTRACT: 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.Tumor Biology 05/2014; 35(8). DOI:10.1007/s13277-014-2093-8 · 2.84 Impact Factor
[Show abstract] [Hide abstract]
ABSTRACT: Pleural dissemination is commonly associated with metastatic advanced lung cancer. The injury of pleural mesothelial cells (PMCs) by soluble factors, such as transforming growth factor-beta1 (TGF-β1), is a major driver of lung cancer pleural dissemination (LCPD). In this study, we examine the effects of TGF-β1 on PMC injury and the ability of TGF-β1 inhibition to alleviate this effect both in vitro and in vivo. PMCs were co-cultured with the high TGF-β1-expressing lung cancer cell line A549 and with various TGF-β1 signaling inhibitors. Expression of cleaved-caspase 3, cleaved-caspase 9, p21, and p16 were evaluated by Western blot and immunofluorescent confocal imaging. Apoptosis was measured by 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltrazoliumbromide assay and AnnexinV-propidium iodide (PI) staining. PMC senescence was assessed by staining for senescence-associated β-galactosidase (SA-β-Gal). The ability of lung cancer cells (LCCs) to adhere to injured PMCs was investigated using an LCC-PMC adhesion assay. In our mouse model, PMC injury status was monitored by hematoxylin-eosin (H&E) and Masson's trichrome staining. LCCs expressing high levels of TGF-β1 induce apoptosis and senescence of PMCs in a co-culture system. Injured PMCs adhere to LCCs, which may further promote LCPD. Importantly, PMC monolayer injury could be reversed with TGF-β1 inhibitors. This was consistent with our in vivo data showing that the TGF-β1 inhibitor SB-431542 attenuated PMC barrier injury induced by A549 culture medium in our mouse model. Our study highlights the importance of TGF-β1 signaling in LCPD and establishes this signaling pathway as a potential therapeutic target in the disease.Tumor Biology 11/2014; 36(4). DOI:10.1007/s13277-014-2888-7 · 2.84 Impact Factor
[Show abstract] [Hide abstract]
ABSTRACT: 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.Tumor Biology 03/2014; 35(6). DOI:10.1007/s13277-014-1808-1 · 2.84 Impact Factor