title: Hypoxia increases endothelin-1 mRNA expression but not immunoreactive endothelin in the medium of T98G glioblastoma cells under cytokine treatment.
abstract: Endothelin-1 (ET-1) levels in the culture medium were considered to reflect the transcription of the ET-1 gene and the subsequent secretion of ET-1 from cultured cells. It has not been clarified how different ET-1 mRNA expression levels and immunoreactive (IR)-ET levels in the culture medium are in the cell culture system. We studied ET-1 mRNA expression levels and IR-ET levels in the medium of T98G glioblastoma cells treated with cytokines. T98G glioblastoma cells were cultured with cytokines (interferon-gamma 100 U/ml, tumor necrosis factor-alpha 20 ng/ml and interleukin-1beta 10 ng/ml) under normoxia or hypoxia (1% O(2)). Northern blot analysis showed that ET-1 mRNA expression levels were increased by tumor necrosis factor-alpha alone or a combination of tumor necrosis factor-alpha and interleukin-1beta, or three cytokines, and the increase was further enhanced under hypoxia. Particularly, relative expression levels of ET-1 mRNA were significantly higher under hypoxia than in normoxia in the treatment with a combination of three cytokines. IR-ET levels in the medium were increased by treatment with tumor necrosis factor-alpha, interleukin-1beta or a combination of tumor necrosis factor-alpha and interleukin-1beta, or three cytokines. In contrast to the mRNA expression levels, IR-ET levels in the medium of T98G cells treated with a combination of three cytokines were rather decreased under hypoxia compared with those in normoxia. These findings indicate that hypoxia induces ET-1 mRNA expression in the treatment of three cytokines, but IR-ET levels in the medium do not reflect this induction in T98G glioblastoma cells.
authors.affiliation: Department of Applied Physiology and Molecular Biology, Tohoku University School of Medicine, Sendai, Miyagi 980-8575, Japan.
authors.forename: Yan
authors.initials: Y
authors.lastname: Zhang
authors.forename: Yin
authors.initials: Y
authors.lastname: Li
authors.forename: Kazuhito
authors.initials: K
authors.lastname: Totsune
authors.forename: Kumi
authors.initials: K
authors.lastname: Kikuchi
authors.forename: Osamu
authors.initials: O
authors.lastname: Murakami
authors.forename: Shigeki
authors.initials: S
authors.lastname: Shibahara
authors.forename: Kazuhiro
authors.initials: K
authors.lastname: Takahashi
mesh_terms: D015687:Cell Hypoxia; D045744:Cell Line, Tumor; D017077:Culture Media, Conditioned / Q000737:chemistry; D016207:Cytokines / Q000494:pharmacology*; D019332:Endothelin-1 / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D005909:Glioblastoma / Q000188:drug therapy* / Q000378:metabolism*; D006801:Humans; D012333:RNA, Messenger / Q000378:metabolism*; D011863:Radioimmunoassay; D015854:Up-Regulation
chemical_list: D017077:Culture Media, Conditioned; D016207:Cytokines; D019332:Endothelin-1; D012333:RNA, Messenger