Insulin Potentiates AVP-induced AQP2 Expression in Cultured Renal Collecting Duct Principal Cells
| Title: | Insulin Potentiates AVP-induced AQP2 Expression in Cultured Renal Collecting Duct Principal Cells |
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| Authors: | Bustamante, Mauro; Hasler, Udo; Kotova, Olga; Chibalin, Alexander V.; Mordasini, David; Rousselot, Martine; Vandewalle, Alain; Martin, Pierre-Yves; Feraille, MD, PhD Privat Docent, Eric |
| Publisher: | American Journal of Physiology: Renal Physiology |
| Date Published: | October 19, 2004 |
| Reference Number: | 675 |
In the renal collecting duct (CD), water reabsorption depends on the presence of aquaporin-2 (AQP2) in the apical membrane of principal cells. AQP2 expression and subcellular repartition are under the control of AVP. Some pieces of experimental evidence indicate that additional hormonal factors, including insulin, may also control AQP2 expression and thereby CD water permeability. We have previously shown that AVP induces endogenous AQP2 expression in cultured mouse mpkCCD(cl4) CD principal cells (23). In the present study, we investigated the effect of insulin on AQP2 expression in mpkCCD(cl4) cells. Addition of insulin to the basal medium of cells grown on filters slightly increased AQP2 mRNA and protein expression, whereas insulin potentiated the effect of AVP. The potentiation of AVP-induced AQP2 expression by insulin was abolished by actinomycin D, a transcriptional inhibitor. Analysis of AQP2 protein expression under conditions of AVP washout and/or in the presence of chloroquine, a lysosomal degradation inhibitor, revealed that insulin did not significantly alter AQP2 protein degradation. Inhibition of ERK, p38 kinase, and phosphatidylinositol 3'-kinase (PI 3-kinase) activities prevented the insulin-induced stimulation of AQP2 expression, whereas inhibition of PKC has no effect. Taken together, our results indicate that insulin increased AQP2 protein expression mostly through increased AQP2 mRNA levels in cultured mpkCCD(cl4) cells. This effect most likely relies on increased AQP2 gene transcription in response to MAPK and PI 3-kinase activation.
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This translation by the NDI Foundation is to assist the lay reader. To provide a clear, accessible interpretation of the original article, we eliminated or simplified some technical detail and complicated scientific language. We concentrated our translation on those aspects of the article dealing directly with NDI. The NDI Foundation thanks the researchers for their work toward understanding and more effectively treating this disorder.
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By performing a thorough series of experiments using laboratory cell cultures, the research team arrived at results that indicate the insulin significantly increases AVP-dependent AQP2 content by increasing the AQP2 mRNA levels in the cells.
More specifically:
- Insulin significantly increased AVP-induced AQP2 abundance in the laboratory cell cultures 8 hours after being introduced, indicating that insulin strengthens AVP’s ability to increase AQP2 content.
- Insulin's ability to increase AVP-induced AQP2 expression is dependent on the amount of insulin used in the experiment, and the amount needed corresponds to the maximal amount of insulin circulating normally in the body.
- Insulin by itself is not a strong AQP2 inducer, but rather increases AVP’s ability to instigate an increase in AQP2 in the cell.
- Insulin enhances the effect of AVP on AQP2 expression by a means other than making the laboratory cell cultures more sensitive to AVP.
- The insulin receptors (proteins on the cell membrane to which the insulin hormone attaches itself in order to affect the cell) are located solely on the basolateral membrane (bottom and side part of the cell) of the laboratory cell cultures.
- The primary manner in which insulin increases AVP-induced AQP2 expression in the laboratory cell cultures may be by increasing AQP2 mRNA levels through enhanced transcription of the AQP2 gene. (Transcription is an essential step in the protein synthesis process.)
- The ability of insulin to enhance AVP-induced AQP2 expression has nothing to do with a decrease in the breakdown of AQP2 protein.
- Stimulation of the enzyme phosphatidylinositol 3' kinase (PI 3-kinase) is required for increased AQP2 expression in response to insulin.
- The effect of insulin on AQP2 expression is also dependent on the activity of extracellular regulated kinase (ERK) and p38 kinase.
- Protein kinase C (PKC) isozyme activation is not involved in the insulin-induced increase in AVP dependent AQP2 expression in the laboratory cell culture.