Establishment of mouse intestinal myofibroblast cell lines

Kawasaki, Hideyoshi

doi:10.3748/wjg.v19.i17.2629

Cited by 15 publications

(14 citation statements)

References 30 publications

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“…Mouse intestinal myofibroblast cell lines (LmcMFs) derived from mouse colonic mucosa were established by Takashi Ohama and Koichi Sato, Laboratory of Veterinary Pharmacology, Joint Faculty of Veterinary Medicine, Yamaguchi University [27]. Cells were cultured in DMEM containing 10% FBS at 37°C in 5% CO 2 atmosphere.…”

Section: Cell Lines and Cell Culturementioning

confidence: 99%

Established fibrous peritoneal metastasis in an immunocompetent mouse model similar to clinical immune microenvironment of gastric cancer

et al. 2020

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Background Peritoneal metastasis (PM) in gastric cancer (GC) is characterized by diffusely infiltrating and proliferating cancer cells accompanied by extensive stromal fibrosis in the peritoneal space. The prognosis of GC with PM is still poor regardless of the various current treatments. In order to elucidate the cause of difficulties in PM treatment, we compared the tumor immune microenvironment (TME) in primary and PM lesions in GC. In addition, a PM model with fibrous stroma was constructed using immunocompetent mice to determine whether its TME was similar to that in patients. Methods Immuno-histochemical analyses of infiltrating immune cells were performed in paired primary and PM lesions from 28 patients with GC. A C57BL/6 J mouse model with PM was established using the mouse GC cell line YTN16 either with or without co-inoculation of mouse myofibroblast cell line LmcMF with α-SMA expression. The resected PM from each mouse model was analyzed the immunocompetent cells using immunohistochemistry. Results The number of CD8+ cells was significantly lower in PM lesions than in primary lesions (P < 0.01). Conversely, the number of CD163+ cells (M2 macrophages) was significantly higher in PM lesions than in primary lesions (P = 0.016). Azan staining revealed that YTN16 and LmcMF co-inoculated tumors were more fibrous than tumor with YTN16 alone (P < 0.05). Co-inoculated fibrous tumor also showed an invasive growth pattern and higher progression than tumor with YTN16 alone (P = 0.045). Additionally, YTN16 and LmcMF co-inoculated tumors showed lower infiltration of CD8+ cells and higher infiltration of M2 macrophages than tumors with YTN16 alone (P < 0.05, P < 0.05). These results indicate that LmcMF plays as cancer-associated fibroblasts (CAFs) by crosstalk with YTN16 and CAFs contribute tumor progression, invasion, fibrosis, and immune suppression. Conclusions This model is the first immunocompetent mouse model similar to TME of human clinical PM with fibrosis. By using this model, new treatment strategies for PM, such as anti-CAFs therapies, may be developed.

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Section: Cell Lines and Cell Culturementioning

confidence: 99%

Established fibrous peritoneal metastasis in an immunocompetent mouse model similar to clinical immune microenvironment of gastric cancer

et al. 2020

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“…Mouse intestinal myo broblast cell lines (LmcMFs) derived from mouse colonic mucosa were established by Takashi Ohama and Koichi Sato, Laboratory of Veterinary Pharmacology, Joint Faculty of Veterinary Medicine, Yamaguchi University [24]. Cells were cultured in DMEM containing 10% FBS at 37°C in 5% CO 2 atmosphere.…”

Section: Cell Lines and Cell Culturementioning

confidence: 99%

Established fibrous peritoneal metastasis in an immunocompetent mouse model similar to clinical immune microenvironment of gastric cancer

Fujimori

Kinoshita

Yamaguchi

et al. 2020

Preprint

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Background Peritoneal metastasis (PM) in gastric cancer (GC) is characterized by diffusely infiltrating and proliferating cancer cells accompanied by extensive stromal fibrosis in the peritoneal space. The prognosis of GC with PM is still poor regardless of the various current treatments. In order to elucidate the cause of difficulties in PM treatment, we compared the tumor immune microenvironment (TME) in primary and PM lesions in GC. In addition, a PM model with fibrous stroma was constructed using immunocompetent mice to determine whether its TME was similar to that in patients. Methods Immuno-histochemical analyses of infiltrating immune cells were performed in paired primary and PM lesions from 28 patients with GC. A C57BL/6J mouse model with PM was established using the mouse GC cell line YTN16 either with or without co-inoculation of mouse myofibroblast cell line LmcMF with a-SMA expression. The resected PM from each mouse model was analyzed the immunocompetent cells using immunohistochemistry.Results The number of CD8+ cells was significantly lower in PM lesions than in primary lesions (P<0.01). Conversely, the number of CD163+ cells (M2 macrophages) was significantly higher in PM lesions than in primary lesions (P=0.016). Azan staining revealed that YTN16 and LmcMF co-inoculated tumors were more fibrous than tumor with YTN16 alone (P<0.05). Co-inoculated fibrous tumor also showed an invasive growth pattern and higher progression than tumor with YTN16 alone (P=0.045). Additionally, YTN16 and LmcMF co-inoculated tumors showed lower infiltration of CD8+ cells and higher infiltration of M2 macrophages than tumors with YTN16 alone (P<0.05, P<0.05). These results indicate that LmcMF plays as cancer-associated fibroblasts (CAFs) by crosstalk with YTN16 and CAFs contribute tumor progression, invasion, fibrosis, and immune suppression.Conclusions This model is the first immunocompetent mouse model to accurately reflect the similar to TME of human clinical PM with fibrosis. By using this model, new treatment strategies for PM, such as anti-CAFs therapies, may be developed.

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“…IMFs were isolated as previously described (23). LmcMF, a mouse colon myofibroblast cell line, was established in our previous study (23).…”

Section: Cell Culture and Conditioned Medium (Cm) Collection Mousementioning

confidence: 99%

“…IMFs were isolated as previously described (23). LmcMF, a mouse colon myofibroblast cell line, was established in our previous study (23). An adult mouse colon epithelial cell line, aMoC1, was provided by Dr Mamoru Totsuka (Tokyo university) (24).…”

Section: Cell Culture and Conditioned Medium (Cm) Collection Mousementioning

confidence: 99%

“…To investigate how KRasV12 overexpression in epithelial cells affects the functions of IMFs, we treated IMFs with the CM from Mock-aMoC1 cells (Mock-CM) or KRasV12-aMoC1 cells (KRasV12-CM) and then examined IMF differentiation, proliferation, and migration. IMF differentiation was evaluated based on the expression level of myofibroblast markers (α-SMA and vimentin) as previously described (23). In primary IMFs, KRasV12-CM exerted little effect on differentiation and proliferation ( Fig.…”

Section: Krasv12 Overexpression Converts Amoc1 Cells Into Tumor Cellsmentioning

confidence: 99%

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Regulation of intestinal myofibroblasts by KRas-mutated colorectal cancer cells through heparin-binding epidermal growth factor-like growth factor

et al. 2017

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In colorectal cancer, gain-of-function mutations in KRas play a critical role in malignant transformation. Tumor growth in colorectal cancer is known to be promoted by the intestinal myofibroblasts (IMFs) that localize adjacent to the cancer cells, but the mechanisms of interaction between KRas-mutated cancer cells and the myofibroblasts remain unclear. Here, we investigated the effects of KRas-mutated cells on the behavior of myofibroblasts by using mouse primary IMFs and cells of an IMF cell line (LmcMF) and a mouse colon epithelial cell line (aMoC1). Conditioned medium (CM) was collected from aMoC1 cells overexpressing a control vector or KRasV12 vector (KRasV12-CM), and the effects of KRasV12-CM on IMFs were analyzed by performing proliferation assays, wound-healing assays, Boyden chamber assays, and western blotting. Whereas KRasV12-CM exerted little effect on the differentiation and proliferation of primary IMFs, the CM promoted migration of both primary IMFs and LmcMF cells. In KRasV12-overexpressing aMoC1 cells, mRNA expression of heparin-binding epidermal growth factor-like growth factor (HB-EGF) was higher than in mock-transfected aMoC1 cells, and HB-EGF promoted the migration of primary IMFs and LmcMF cells. Moreover, KRasV12-CM-induced IMF migration was suppressed by dacomitinib, an inhibitor of HB-EGF receptors. Notably, in LmcMF cells, both KRasV12-CM and HB-EGF activated extracellular signal-regulated kinase (ERK) and c-jun N-terminal kinase (JNK), whereas KRasV12-CM-induced migration of IMFs was suppressed following treatment with either an ERK inhibitor (FR180204) or a JNK inhibitor (SP600125). These results suggest that HB-EGF secreted from KRas-mutated colorectal cancer cells promotes IMF migration through ERK and JNK activation, which, in turn, could support cancer progression.

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Establishment of mouse intestinal myofibroblast cell lines

Abstract: We established 2 novel IMF cell lines from mouse colonic mucosa, namely, LmcMF and SmcMF, both of which were able to respond to LPS.

Cited by 15 publications

References 30 publications

Established fibrous peritoneal metastasis in an immunocompetent mouse model similar to clinical immune microenvironment of gastric cancer

Established fibrous peritoneal metastasis in an immunocompetent mouse model similar to clinical immune microenvironment of gastric cancer

Established fibrous peritoneal metastasis in an immunocompetent mouse model similar to clinical immune microenvironment of gastric cancer

Regulation of intestinal myofibroblasts by KRas-mutated colorectal cancer cells through heparin-binding epidermal growth factor-like growth factor

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