scholarly journals A Novel Methodology For Isolation and Primary Culture of Swine Gastric Epithelial Cells.

2020 ◽  
Author(s):  
Henry Bautista-Amorocho ◽  
Jorge Alexander Silva-Sayago ◽  
Diego A. Goyeneche-Patino ◽  
Tania Liseth Pérez-Cala ◽  
Fabio Macías-Gómez ◽  
...  
Keyword(s):  
Growth Factors ◽  
Epithelial Cells ◽  
Protease Inhibitors ◽  
Swine Model ◽  
Type I ◽  
Fundic Gland ◽  
Gastric Epithelial Cells ◽  
Gland Area ◽  
Fundic Gland Area ◽  
Functional Phenotype

Abstract Background: Culture of primary epithelial cells has a great advantage over tumor-derived or immortal cells lines since functional phenotype and genetic makeup are preserved. Swine model has proved to be helpful and reliable as a surrogate model in human diseases. Several porcine cell lines have been established from a variety of tissues and shown to extensively contribute to the current understanding of several pathologies, including cancer. However, few protocols for the isolation and culture swine gastric epithelial cells with phenotype preservation have been described. Therefore, the objective of this research was to develop a new methodology for establishing a primary cell culture from the fundic gland area of the porcine stomach.Results: Enzymatic disaggregation of gastric tissue by using a combination of collagenase type I and dispase II, protease inhibitors (soybean trypsin inhibitor and bovine serum albumin), and antioxidants (Dithiothreitol) allowed the isolation of gastric epithelial cells from the fundic gland area with viability > 90% during the incubation period. Gastric epithelial cells cultured in RPMI 1640, DMEM HG, and DMEM/F12 media did not lead to cell adhesion, cluster formation and cell proliferation. By contrast, Williams’ medium supplemented with growth factors supports the confluence and proliferation of a pure epithelial cell monolayer after 10 days of incubation at 37oC in a 5% CO2 incubator. Mucin-producing cell phenotype of primary isolates was confirmed by PAS staining as well as the expression of MUC1 and MUC20 genes by RT-PCR and DNAc sequencing. Swine Gastric epithelial cells also showed origin-specific markers such as epithelial membrane antigen (EMA), cytokeratin cocktail (AE1/AE3) and cytokeratin 18 (CK-18) detected by immunohistochemistry and immunofluorescence, respectively. Conclusions: A new methodology was successfully established for the isolation of primary gastric epithelial cells from the fundic gland area in a swine model, based on a combination of tissue specific proteases, protease inhibitors, and antioxidants. The formulation of Williams’ medium with growth factors for epithelial cells supports the adherence and maintains the functional phenotype of primary cells, which was confirmed by mucin production and expression of typical epithelial markers in the long term.

scholarly journals A novel method for isolation and culture of primary swine gastric epithelial cells

2020 ◽  
Author(s):  
Henry Bautista-Amorocho ◽  
Jorge Alexander Silva-Sayago ◽  
Diego A. Goyeneche-Patino ◽  
Tania Liseth Pérez-Cala ◽  
Fabio Macías-Gómez ◽  
...  
Keyword(s):  
Cell Adhesion ◽  
Growth Factors ◽  
Epithelial Cell ◽  
Epithelial Cells ◽  
Protease Inhibitors ◽  
Cell Lines ◽  
Cell Phenotype ◽  
Swine Model ◽  
Fundic Gland ◽  
Gastric Epithelial Cells

Abstract Background: Culturing primary epithelial cells has a major advantage over tumor-derived or immortalized cell lines as long as their functional phenotype and genetic makeup are mainly maintained. The swine model has shown to be helpful and reliable when used as a surrogate model for human diseases. Several porcine cell lines have been established based on a variety of tissues, which have shown to extensively contribute to the current understanding of several pathologies, especially cancer. However, protocols for the isolation and culture of swine gastric epithelial cells that preserve cell phenotype are rather limited. We aimed to develop a new method for establishing a primary epithelial cell culture from the fundic gland region of the pig stomach.Results: Mechanical and enzymatic dissociation of gastric tissue was possible by combining collagenase type I and dispase II, protease inhibitors and antioxidants, which allowed the isolation of epithelial cells from the porcine fundic glands showing cell viability > 90% during the incubation period. Gastric epithelial cells cultured in RPMI 1640, DMEM-HG and DMEM/F12 media did not contribute enough to cell adhesion, cluster formation and cell proliferation. By contrast, William’s E medium supplemented with growth factors supports confluency and proliferation of a pure epithelial cell monolayer after 10 days of incubation at 37oC, 5% CO2. Mucin-producing cell phenotype of primary isolates was confirmed by PAS staining, MUC1 by immunohistochemistry, as well as the expression of MUC1 and MUC20 genes by RT-PCR and cDNA sequencing. Swine gastric epithelial cells also showed origin-specific markers such as cytokeratin cocktail (AE1/AE3) and cytokeratin 18 (CK-18) using immunohistochemical and immunofluorescence methods, respectively.Conclusions: A new method was successfully established for the isolation of primary gastric epithelial cells from the fundic gland zone through a swine model based on a combination of tissue-specific proteases, protease inhibitors and antioxidants after mechanical cell dissociation. The formulation of William’s E medium with growth factors for epithelial cells contributes to cell adhesion and preserves functional primary cells phenotype, which is confirmed by mucin production and expression of typical epithelial markers over time.

scholarly journals A novel method for isolation and culture of primary swine gastric epithelial cells

2021 ◽  
Author(s):  
Henry Bautista-Amorocho ◽  
Jorge Alexander Silva-Sayago ◽  
Diego A. Goyeneche-Patino ◽  
Tania Liseth Pérez-Cala ◽  
Fabio Macías-Gómez ◽  
...  
Keyword(s):  
Cell Adhesion ◽  
Growth Factors ◽  
Epithelial Cell ◽  
Epithelial Cells ◽  
Protease Inhibitors ◽  
Cell Lines ◽  
Cell Phenotype ◽  
Swine Model ◽  
Fundic Gland ◽  
Gastric Epithelial Cells

Abstract Background: Culturing primary epithelial cells has a major advantage over tumor-derived or immortalized cell lines as long as their functional phenotype and genetic makeup are mainly maintained. The swine model has shown to be helpful and reliable when used as a surrogate model for human diseases. Several porcine cell lines have been established based on a variety of tissues, which have shown to extensively contribute to the current understanding of several pathologies, especially cancer. However, protocols for the isolation and culture of swine gastric epithelial cells that preserve cell phenotype are rather limited. We aimed to develop a new method for establishing a primary epithelial cell culture from the fundic gland region of the pig stomach.Results: Mechanical and enzymatic dissociation of gastric tissue was possible by combining collagenase type I and dispase II, protease inhibitors and antioxidants, which allowed the isolation of epithelial cells from the porcine fundic glands showing cell viability > 90% during the incubation period. Gastric epithelial cells cultured in RPMI 1640, DMEM-HG and DMEM/F12 media did not contribute enough to cell adhesion, cluster formation and cell proliferation. By contrast, William’s E medium supplemented with growth factors supports confluency and proliferation of a pure epithelial cell monolayer after 10 days of incubation at 37oC, 5% CO2. Mucin-producing cell phenotype of primary isolates was confirmed by PAS staining, MUC1 by immunohistochemistry, as well as the expression of MUC1 and MUC20 genes by RT-PCR and cDNA sequencing. Swine gastric epithelial cells also showed origin-specific markers such as cytokeratin cocktail (AE1/AE3) and cytokeratin 18 (CK-18) using immunohistochemical and immunofluorescence methods, respectively.Conclusions: A new method was successfully established for the isolation of primary gastric epithelial cells from the fundic gland zone through a swine model based on a combination of tissue-specific proteases, protease inhibitors and antioxidants after mechanical cell dissociation. The formulation of William’s E medium with growth factors for epithelial cells contributes to cell adhesion and preserves functional primary cells phenotype, which is confirmed by mucin production and expression of typical epithelial markers over time.

scholarly journals A novel method for isolation and culture of primary swine gastric epithelial cells

2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Henry Bautista-Amorocho ◽  
Jorge Alexander Silva-Sayago ◽  
Diego A. Goyeneche-Patino ◽  
Tania Liseth Pérez-Cala ◽  
Fabio Macías-Gómez ◽  
...  
Keyword(s):  
Cell Adhesion ◽  
Growth Factors ◽  
Epithelial Cell ◽  
Epithelial Cells ◽  
Protease Inhibitors ◽  
Cell Lines ◽  
Cell Phenotype ◽  
Swine Model ◽  
Fundic Gland ◽  
Gastric Epithelial Cells

Abstract Background Culturing primary epithelial cells has a major advantage over tumor-derived or immortalized cell lines as long as their functional phenotype and genetic makeup are mainly maintained. The swine model has shown to be helpful and reliable when used as a surrogate model for human diseases. Several porcine cell lines have been established based on a variety of tissues, which have shown to extensively contribute to the current understanding of several pathologies, especially cancer. However, protocols for the isolation and culture of swine gastric epithelial cells that preserve cell phenotype are rather limited. We aimed to develop a new method for establishing a primary epithelial cell culture from the fundic gland region of the pig stomach. Results Mechanical and enzymatic dissociation of gastric tissue was possible by combining collagenase type I and dispase II, protease inhibitors and antioxidants, which allowed the isolation of epithelial cells from the porcine fundic glands showing cell viability > 90% during the incubation period. Gastric epithelial cells cultured in RPMI 1640, DMEM-HG and DMEM/F12 media did not contribute enough to cell adhesion, cluster formation and cell proliferation. By contrast, William’s E medium supplemented with growth factors supports confluency and proliferation of a pure epithelial cell monolayer after 10 days of incubation at 37 °C, 5% CO2. Mucin-producing cell phenotype of primary isolates was confirmed by PAS staining, MUC1 by immunohistochemistry, as well as the expression of MUC1 and MUC20 genes by RT-PCR and cDNA sequencing. Swine gastric epithelial cells also showed origin-specific markers such as cytokeratin cocktail (AE1/AE3) and cytokeratin 18 (CK-18) using immunohistochemical and immunofluorescence methods, respectively. Conclusions A new method was successfully established for the isolation of primary gastric epithelial cells from the fundic gland zone through a swine model based on a combination of tissue-specific proteases, protease inhibitors and antioxidants after mechanical cell dissociation. The formulation of William’s E medium with growth factors for epithelial cells contributes to cell adhesion and preserves functional primary cells phenotype, which is confirmed by mucin production and expression of typical epithelial markers over time.

Gastric ulcer and carcinoma in fundic gland area

1975 ◽  
Vol 10 (4) ◽  
pp. 290-299
Author(s):  
Hiroshi Yamagiwa ◽  
Akinori Ishihara ◽  
Minoru Hamazaki
Keyword(s):  
Gastric Ulcer ◽  
Fundic Gland ◽  
Gland Area ◽  
Fundic Gland Area

Intestinal metaplasia of the stomach confined to the fundic gland area. Report of two cases

1987 ◽  
Vol 17 (4) ◽  
pp. 276-280
Author(s):  
Masashi Hirono ◽  
Koichi Mandai ◽  
Tetsuya Toge ◽  
Minoru Niimoto ◽  
Takao Hattori ◽  
...  
Keyword(s):  
Intestinal Metaplasia ◽  
Fundic Gland ◽  
Gland Area ◽  
Fundic Gland Area

Clinical and histochemical studies on early and early simulating advanced carcinoma located in the fundic gland area of the stomach.

1989 ◽  
Vol 22 (10) ◽  
pp. 2338-2343
Author(s):  
Michio SOWA ◽  
Yasuyuki KATO ◽  
Masanori NISHIMURA ◽  
Toshiaki KUBO ◽  
Hitoshi MAEKAWA ◽  
...  
Keyword(s):  
Fundic Gland ◽  
Advanced Carcinoma ◽  
Gland Area ◽  
Fundic Gland Area

The Further Purification of Motilin, a Gastric Motor Activity Stimulating Polypeptide from the Mucosa of the Small Intestine of Hogs

1971 ◽  
Vol 49 (5) ◽  
pp. 399-405 ◽  
Author(s):  
John C. Brown ◽  
Victor Mutt ◽  
Jill R. Dryburgh
Keyword(s):  
Small Intestine ◽  
Motor Activity ◽  
Glutamic Acid ◽  
Duodenal Mucosa ◽  
The Other ◽  
Fundic Gland ◽  
Terminal Residue ◽  
Gastric Motor Activity ◽  
Gland Area ◽  
Fundic Gland Area

A polypeptide has been isolated from the duodenal mucosa of hogs and has been named motilin. Motilin stimulates motor activity in both antral and fundic gland area pouches of the stomach of dogs. It will stimulate pepsin output, with no change in H+ secretion, from fundic gland area pouches. Motilin differs from the other characterized gastrointestinal polypeptides both chemically and in its physiological actions. In particular, the presence of phenylalanine as N-terminal residue, the absence of histidine, and presence of large amounts of glutamic acid and glutamine distinguish it from cholecystokinin–pancreozymin and secretin.

scholarly journals Regulation of lung cell proliferation by polypeptide growth factors

1989 ◽  
Vol 257 (2) ◽  
pp. L23-L38 ◽  
Author(s):  
R. J. King ◽  
M. B. Jones ◽  
P. Minoo
Keyword(s):  
Growth Factors ◽  
Respiratory Failure ◽  
Epithelial Cells ◽  
Acute Respiratory Failure ◽  
Traumatic Injury ◽  
Alveolar Epithelial Cells ◽  
Metabolic Effects ◽  
Cell Populations ◽  
Type I ◽  
Polypeptide Growth Factors

In the last 10 years there has been an increased appreciation of the changes in lung cell populations that occur in association with the acute respiratory failure often induced by traumatic injury. Early events result in an accumulation in the lung of platelets, neutrophils, monocytes, and macrophages, a release of substances having potent cardiopulmonary and metabolic effects, and an ensuing edema and transudation of materials from the interstitium and capillaries into the alveoli. Further progression of the injury results in significant decreases in the number of endothelial and type I epithelial cells and a subsequent hyperplasia of fibroblasts and type II-like epithelial cells. Major sequelae of the latter stage of the disease are interstitial and interalveolar fibrosis, probably resulting from the increased number of fibroblasts present. The activity and composition of pulmonary surfactant are often perturbed. This review will discuss mechanisms that may be involved in these processes, with major emphasis on cell-cell interactions mediated through polypeptide growth factors. We describe the properties of certain growth factors commonly associated with inflammatory and wound healing reactions, discuss their cellular origins, and speculate on their possible roles in mediating the structural and physiological responses seen in the lung during acute respiratory failure. The majority of work done with lung cells has concentrated on interactions between macrophages and fibroblasts, and it is evident that macrophages are capable of producing mitogens affecting the proliferation of fibroblasts. However, from the results of studies that are less developed, it is possible that epithelial cells and immunologically stimulated cells could also be involved in these actions. We conclude that the homeostasis of lung cell populations may be influenced by both growth-stimulating and growth-inhibiting substances and potentially could involve interactions through growth factors of fibroblasts, macrophages, lymphocytes, alveolar epithelial cells, endothelial cells, and platelets. At this time the information on these purported interactions is quite limited and there are far more questions than answers.

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