Effects of hyperosmolarity on the Na⁺-myo-inositol cotransporter SMIT2 stably transfected in the Madin-Darby canine kidney cell line

Abstract

Bissonnette P, Lahjouji K, Coady MJ, Lapointe J-Y. Effects of hyperosmolarity on the Na ϩ -myo-inositol cotransporter SMIT2 stably transfected in the Madin-Darby canine kidney cell line. Am J Physiol Cell Physiol 295: C791-C799, 2008. First published July 23, 2008 doi:10.1152/ajpcell.00390.2007 is a compatible osmolyte used by cells to compensate for changes in the osmolarity of their surrounding milieu. In kidney, the basolateral Na ϩ -MI cotransporter (SMIT1) and apical SMIT2 proteins are homologous cotransporters responsible for cellular uptake of MI. It has been shown in the Madin-Darby canine kidney (MDCK) cell line that SMIT1 expression was under the control of the tonicity-sensitive transcription factor, tonicity-responsive enhancer binding protein (TonEBP). We used an MDCK cell line stably transfected with SMIT2 to determine whether variations in external osmolarity could also affect SMIT2 function. Hyperosmotic conditions (ϩ200 mosM raffinose or NaCl but not urea) generated an increase in SMIT2-specific MI uptake by three-to ninefold in a process that required protein synthesis. Using quantitative RT-PCR, we have determined that hyperosmotic conditions augment both the endogenous SMIT1 and the transfected SMIT2 mRNAs. Transport activities for both SMIT1 and SMIT2 exhibited differences in their respective induction profiles for both their sensitivities to raffinose, as well as in their time course of induction. Application of MG-132, which inhibits nuclear translocation of TonEBP, showed that the effect of osmolarity on transfected SMIT2 was unrelated to TonEBP, unlike the effect observed with SMIT1. Inhibition studies involving the hyperosmolarity-related MAPK suggested that p38 and JNK play a role in the induction of SMIT2. Further studies have shown that hyperosmolarity also upregulates another transfected transporter (Na ϩ -glucose), as well as several endogenously expressed transport systems. This study shows that hyperosmolarity can stimulate transport in a TonEBP-independent manner by increasing the amount of mRNA derived from an exogenous DNA segment.

Deficiency in the α1 subunit of Na+/K+‐ATPase Enhances the Anti‐Proliferative Effect of High Osmolality in Nucleus Pulposus Intervertebral Disc Cells

“…This finding reinforces the previously reported idea that nucleus pulposus cells' osmoregulatory response is not mediated by ionic concentration changes, but rather depends directly on osmotic pressure (Ishihara et al, ; Mavrogonatou and Kletsas, ). In addition, our data are in agreement with studies describing an increase in sodium/myo‐inositol transport by NaCl or non‐ionic hyperosmolar solutes (Denkert et al, ; Boyd et al, ; Bissonnette et al, ), an increased Na + /K + ‐ATPase activity by both NaCl and cellobiose (Yokoyama et al, ) and an increase in the amino acid transport similarly induced by the use of sucrose or NaCl (Maallem et al, ; Bruscalupi et al, ) in disc cells, as well as in other cell models. The more pronounced effect of salt on the expression of some genes in comparison to the sorbitol solution observed at 24 h may ultimately stem from the long‐standing presence of ions, as has also been hypothesized for nucleus pulpous cells' proliferation (Mavrogonatou and Kletsas, ).…”

The diabetic rat kidney mediates inosituria and selective urinary partitioning of D-chiro-inositol

“…This increased expression is consistent with the regulation of SMIT1 by hyperosmotic conditions via a tonicity-responsive enhancer binding protein (TonEBP). 19 These findings show that reduced renal reabsorption of Figure 3 Renal expression of SMIT1 and SMIT2 in diabetes mellitus. (a) SMIT1 transcription using qPCR; (b) SMIT1 protein expression quantified by Western blotting and representative blot with ACTB as a loading control; (c) SMIT2 transcription using qPCR; (d) SMIT2 protein expression quantified by Western blotting and representative blot with ACTB as a loading control.…”

Osmoregulatory inositol transporter SMIT1 modulates electrical activity by adjusting PI(4,5)P2levels

“…Hypertonicity suffices to induce increased SMIT1 expression in several cell types (55,56). This is considered adaptive because the transporter will import more myo-inositol as an osmolyte that compensates for the raised extracellular solute.…”