Hypoxia increases endothelin-1 mRNA expression but not immunoreactive endothelin in the medium of T98G glioblastoma cells under cytokine treatment.
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.
Zhang
Yan
Y
Department of Applied Physiology and Molecular Biology, Tohoku University School of Medicine, Sendai, Miyagi 980-8575, Japan.
Li
Yin
Y
Totsune
Kazuhito
K
Kikuchi
Kumi
K
Murakami
Osamu
O
Shibahara
Shigeki
S
Takahashi
Kazuhiro
K
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
D017077:Culture Media, Conditioned; D016207:Cytokines; D019332:Endothelin-1; D012333:RNA, Messenger