Interaction between &lt;i&gt;Helicobacter pylori and &lt;/i&gt;Human Gastric Epithelial Cells in Culture: Effect of Antiulcer Drugs

Wagner, Siegfried; Beil, Winfried; U, Mai; Bokemeyer, C.; Meyer, H J; Manns, M.P.

doi:10.1159/000139238

Cited by 32 publications

(29 citation statements)

References 19 publications

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“…For this, cultures were established as indicated above using, as for MedC*, conditioned media from different sources: rat Reuber hepatoma cells (ATCC no. CRL-1600), HepG2 human hepatoma cells (HB-8065), human embryonic kidney cells HEK-293 (CRL-1573), SH-SY5Y human neuroblastoma cells (CRL-2266), CHO chinese hamster ovary cells (CCL-61), MMN02 human gastric epithelial adenocarinoma cells (Wagner et al 1994), HeLa human epidermoid carcinoma cells (CCL-2), HEC1B human endometrial carcinoma cells , and the macrophage like cell line U937 (CRL-1593.2). As shown in Fig.…”

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confidence: 99%

Long-term in vitro cultivation of Echinococcus multilocularis metacestodes under axenic conditions

Spiliotis

Tappe

Sesterhenn

et al. 2004

Parasitology Research

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We report here on the development of an in vitro system for the long-term cultivation of Echinococcus multilocularis larvae under axenic conditions. In the absence of feeder cells from the host, long-term survival of the parasite depended strictly on low oxygen conditions and the presence of reducing agents in the medium. Host serum supported survival of the parasite but the growth of metacestode vesicles and differentiation towards the protoscolex stage only occurred in the presence of culture medium that was preconditioned by hepatoma cells or several other immortal cell lines. On the basis of this in vitro system, future analyses on the identification of host-derived growth factors for E. multilocularis will be greatly facilitated.

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confidence: 99%

Long-term in vitro cultivation of Echinococcus multilocularis metacestodes under axenic conditions

Spiliotis

Tappe

Sesterhenn

et al. 2004

Parasitology Research

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“…Human gastric epithelial cells were isolated from gastric tissue specimens obtained during gastric surgery as described recently [46,47]. Informed consent was obtained from each subject, according to the revised version of the declaration of Helsinki.…”

Section: Isolation Of Human Gastric Epithelial Cellsmentioning

confidence: 99%

“…The viability of cultured cells was assessed by trypan blue exclusion and by photometric determination of released lactate dehydrogenase (LDH) enzyme activity using a commercial test (Boehringer Mannheim). Our previous work has shown that short-term incubation (12 h) of isolated human gastric epithelial cells results in the selective attachment of gastric mucous cells, leaving the chief cells and parietal cells to be recovered from the culture medium [46]. Thus these incubation conditions enabled us to cultivate a partially enriched cell fraction of gastric mucous cells without prior use of specific cell-separation procedures.…”

Section: Cell Culturementioning

confidence: 99%

“…Characterization of isolated and cultured human gastric epithelial cells Isolated and cultured gastric epithelial cells were morphologically characterized by a panel of immunocytochemical staining methods and by transmission electron microscopy as described previously [33,46,47]. The following staining methods were used: Janus green and indirect immunofluorescence (parietal cells), alcian blue pH 2.5/periodic acid-Schiff [AB2.5-PAS: AB2.5 detects acidic mucins (mucous neck cells); PAS detects neutral mucins (surface mucous cells)] and fluorescein-isothiocyanate-labelled (FITC-labelled) lectins (mucous cells), Nile blue (chief cells), cytokeratin (epithelial cells) and vimentin (non-epithelial cells).…”

Section: Cell Culturementioning

confidence: 99%

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Morphological and molecular characterization of human gastric mucous cells in long-term primary culture

Wagner¹,

Enss

Cornberg³

et al. 1998

Pfl�gers Archiv European Journal of Physiology

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A primary cell culture of human gastric mucous cells was developed using enzymatic treatment of surgically obtained gastric mucosal specimens. Preferential attachment of gastric mucous cells during a preincubation step resulted in the enrichment of mucous cells [over 90% stained with periodic acid-Schiff (PAS) and mucin-type lectins] in the primary cell culture. Gastric mucous cells could be maintained in culture for 10 days. DNA synthesis peaked during the first 2 days in culture (8+/-1% bromodeoxyuridine-positive cells). During the entire culture period gastric mucous cells released high-molecular-weight glycoproteins into the medium, as determined by gel chromatography on a Sepharose CL-4B column and by metabolic labelling with [14C]-N-acetylglucosamine. Gastric mucin was verified by gas chromatographic analysis of the carbohydrate composition and fractionation of the void-volume fraction by density gradient centrifugation. Determination of the terminal glycosylation of the secreted glycoproteins by a lectin-ELISA revealed that there was a high quantity of alpha-l-fucose. Prostaglandin E2 significantly stimulated glycoprotein secretion during the entire cultivation period by 29-60%. Analysis of mucin-encoding MUC mRNA expression by reverse transcriptase polymerase chain reaction revealed that gastric mucous cells predominantly express MUC1 and MUC5AC, and to a lesser extent MUC6, which reflects the expression pattern obtained following analysis of biopsied samples of gastric mucosa. This primary cell culture model enables the regulation of mucin secretion and mucin gene expression in man to be investigated.

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“…The determination of PKC translocation in gastric HM02 cells, which are derived from a human well-differentiated mucusproducing gastric carcinoma (Wagner et al, 1994), were grown for 3 days in RPMI-1640 medium supplemented with 10% fetal calf serum (FCS) at 370C in a 5% carbon dioxide atmosphere. The cell monolayers were then kept for 1 day in FCS-free medium and trypsinated.…”

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confidence: 99%

The Helicobacter pylori fatty acid cis-9,10-methyleneoctadecanoic acid stimulates protein kinase C and increases DNA synthesis of gastric HM02 cells

Beil

Obst²,

Wagner³

et al. 1998

Br J Cancer

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Summary Protein kinase C (PKC) has been implicated in the control of epithelial proliferative activity and in the process of malignant transformation. Helicobacter pylori (H.p.) infection is associated with increased gastric epithelial cell proliferation and has been linked with gastric carcinoma. In the present study, we report that the H.p. fatty acid cis-9,1 0-methyleneoctadecanoic acid (MOA) directly activates PKC (Ka 3.3 gM). Keywords: Helicobacterpylori; fatty acid; protein kinase C; cell proliferation Protein kinase C (PKC) is a family of enzymes that plays a pivotal role in transmembrane signalling, cell growth and cell division (Nishizuka, 1986;Clemens et al, 1992). Diacylglycerol (DAG), a breakdown product of polyphosphoinositides, is a physiological effector of this(ese) enzyme(s). Tumour-promoting phorbol esters, such as 12-O-tetradecanoyl-phorbol-13-acetate (TPA), which interact with the DAG site, are activators of the enzyme and several lines of evidence indicate that tumour promotion is related to PKC activation (Martelly and Castagna, 1989). Other tumour promoters structurally unrelated to TPA have also been described as activators of PKC, such as teleocidin B (Fujiki et al, 1984), chloroform (Roghani et al, 1987) and bile acids (Huang et al, 1992).Helicobacter pylori (H.p.) is a Gram-negative bacterium that causes chronic gastritis and peptic ulcer diseases (NIH Consensus Conference, 1994). Moreover, an association between the microorganism and gastric cancer has been demonstrated by epidemiological studies (Forman, 1993), but a causal link between gastric cancer and H.p. infection has not been proven. H.p. produces unusual fatty acids such as cis-9,10-methyleneoctadecanoic acid (Goodwin et al, 1985). Cis-unsaturated fatty acids are known as tumour promoters (Bull et al, 1981) and PKC activators (McPhail et al, 1984).There is some evidence that PKC plays a crucial role in gastric epithelial proliferation. Epidermal growth factor (EGF) and transforming growth factor a (TGF-a) have been reported to be mitogenic for primary cultured gastric parietel, chief and mucous cells (Chen et al, 1991;Rutten et al, 1993). In addition to the ability of these growth factors to stimulate tyrosine kinase activity, to increase inositol trisphosphate and to interact with G proteins, EGF has been shown to activate PKC (Reynolds et al, 1993;Wang et al, 1996). Overexpression of EGF, TGF-a and their receptor genes are thought to participate in the rapid cell proliferation seen in gastric cancer (Pfeiffer et al, 1990).Based on these observations, we have examined the characteristics of cis-9,10-methyleneoctadecanoic acid (MOA)-mediated PKC activation and the effect of MOA on cellular DNA synthesis. MATERIALS AND METHODS MaterialsRat brain PKC (a mixture of alpha-, beta-and gamma-isoforms) purified to greater than 97% by the method of Allen and Katz (1991) was purchased from Biomol (Hamburg, Germany). MOA (purity 99%) was synthesized by J Holzkampf

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Interaction between <i>Helicobacter pylori and </i>Human Gastric Epithelial Cells in Culture: Effect of Antiulcer Drugs

Cited by 32 publications

References 19 publications

Long-term in vitro cultivation of Echinococcus multilocularis metacestodes under axenic conditions

Long-term in vitro cultivation of Echinococcus multilocularis metacestodes under axenic conditions

Morphological and molecular characterization of human gastric mucous cells in long-term primary culture

The Helicobacter pylori fatty acid cis-9,10-methyleneoctadecanoic acid stimulates protein kinase C and increases DNA synthesis of gastric HM02 cells

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