J Korean Soc Endocrinol.  2004 Oct;19(5):501-510.

The Abundance and Nuclear Distribution of TonEBP in Response to Changes of Tonicity in Hyperglycemic Cells

• Park WK

Abstract

BACKGROUND: TonEBP (Tonicity-responsive enhancer binding protein) regulates the transcription of tonicity responsive genes, such as sodium-myo-inositol and the sodium- chloride-betaine co- transporters (SMIT & BGT1), heat shock protein 70 (HSP70) and aldose reductase (AR). To characterize the signals that activate TonEBP in hyperglycemic human retinal pigment epithelial (hRPE) cells, the abundance and nuclear distribution of TonEBP were studied in response to changes of tonicity in culture media.
METHODS
After the cultures reached confluence, the hRPE cells were exposed for 3 days to 25 mM glucose and 100 mM NaCl, both with and without 20 M tolrestat. The expressions of AR, SMIT and HSP 70 were determined by northern blot, and the abundances of TonEBP by western blot. The nuclear distributions of TonEBP were observed by fluorescence microscopy, after immuno staining.
RESULTS
The AR and SMIT mRNA levels in hyperglycemic and hypertonic media were decreased compared to those in hypertonic media alone. These decreased AR and SMIT mRNA expressions were also observed to be significantly prevented in those cells incubated with tolrestat. Stimulation of TonEBP in hypertonic medium occurs due to a combination of an increased abundance of TonEBP and an increased distribution into the nucleus from the cytoplasm. However, the expressions and nuclear distributions of TonEBP in hyperglycemic and hypertonic media were not different from those in hypertonic media alone.
CONCLUSION
The expressions of AR and SMIT genes that may influence the development of diabetic complication were down-regulated by the intracellular accumulation of sorbitol in sustained hyperglycemia. TonEBP does not play a key role in the hypertonicity-induced transcriptional regulation of AR and SMIT in hyperglycemic cells, due to the intracellular accumulation of sorbitol and depletion of myo-inositol

MeSH Terms

Aldehyde Reductase
Blotting, Northern
Blotting, Western
Culture Media
Cytoplasm
Diabetes Complications
Glucose
HSP70 Heat-Shock Proteins
Humans
Hyperglycemia
Microscopy, Fluorescence
Retinaldehyde
RNA, Messenger
Sorbitol
Aldehyde Reductase
Culture Media
Glucose
HSP70 Heat-Shock Proteins
RNA, Messenger
Retinaldehyde
Sorbitol