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Suppression of Klotho expression by protein-bound uremic toxins is associated with increased DNA methyltransferase expression and DNA hypermethylation.

Sun CY, Chang SC, Wu MS - Kidney Int. (2012)

Bottom Line: The expression of DNA methyltransferases 1, 3a, and 3b isoforms in HK2 cells treated with indoxyl sulfate or p-cresyl sulfate was significantly increased.Specific inhibition of DNA methyltransferase isoform 1 by 5-aza-2'-deoxycytidine caused demethylation of the Klotho gene and increased Klotho expression in vitro.Thus, inhibition of Klotho gene expression by uremic toxins correlates with gene hypermethylation, suggesting that epigenetic modification of specific genes by uremic toxins may be an important pathological mechanism of disease.

View Article: PubMed Central - PubMed

Affiliation: School of Medicine, Chang Gung University, Taoyuan, Taiwan.

ABSTRACT
The expression of the renoprotective antiaging gene Klotho is decreased in uremia. Recent studies suggest that Klotho may be a tumor suppressor, and its expression may be repressed by DNA hypermethylation in cancer cells. Here we investigated the effects and possible mechanisms by which Klotho expression is regulated during uremia in uninephrectomized B-6 mice given the uremic toxins indoxyl sulfate or p-cresyl sulfate. Cultured human renal tubular HK2 cells treated with these toxins were used as an in vitro model. Injections of indoxyl sulfate or p-cresyl sulfate increased their serum concentrations, kidney fibrosis, CpG hypermethylation of the Klotho gene, and decreased Klotho expression in renal tubules of these mice. The expression of DNA methyltransferases 1, 3a, and 3b isoforms in HK2 cells treated with indoxyl sulfate or p-cresyl sulfate was significantly increased. Specific inhibition of DNA methyltransferase isoform 1 by 5-aza-2'-deoxycytidine caused demethylation of the Klotho gene and increased Klotho expression in vitro. Thus, inhibition of Klotho gene expression by uremic toxins correlates with gene hypermethylation, suggesting that epigenetic modification of specific genes by uremic toxins may be an important pathological mechanism of disease.

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DNA methyltransferase 1 (DNMT 1) inhibitor demethylated the Klotho gene and increased Klotho expression in vivo. (a) Methylation-specific PCR (MSP) with mouse Klotho set 1 primers showed that simultaneous treatment with 5-aza-2′-deoxycytidine (5Aza-2dc) in indoxyl sulfate (IS)- and p-cresyl sulfate (PCS)-injected mice significantly demethylated the Klotho gene. (b) Western blotting showed that 5Aza-2dc significantly increased the Klotho expression in IS- and PCS-injected mice.
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fig4: DNA methyltransferase 1 (DNMT 1) inhibitor demethylated the Klotho gene and increased Klotho expression in vivo. (a) Methylation-specific PCR (MSP) with mouse Klotho set 1 primers showed that simultaneous treatment with 5-aza-2′-deoxycytidine (5Aza-2dc) in indoxyl sulfate (IS)- and p-cresyl sulfate (PCS)-injected mice significantly demethylated the Klotho gene. (b) Western blotting showed that 5Aza-2dc significantly increased the Klotho expression in IS- and PCS-injected mice.

Mentions: MSP analysis showed that inhibiting DNMT 1 by using 5Aza-2dc significantly decreased the DNA methylation of Klotho gene in IS- and PCS-injected mice when compared with the mice treated with IS or PCS alone (Figure 4a). In addition, the results of western blotting revealed that treatment with 5Aza-2dc significantly increased Klotho expression in IS- and PCS-injected mice when compared with the mice treated with IS or PCS alone (Figure 4b).


Suppression of Klotho expression by protein-bound uremic toxins is associated with increased DNA methyltransferase expression and DNA hypermethylation.

Sun CY, Chang SC, Wu MS - Kidney Int. (2012)

DNA methyltransferase 1 (DNMT 1) inhibitor demethylated the Klotho gene and increased Klotho expression in vivo. (a) Methylation-specific PCR (MSP) with mouse Klotho set 1 primers showed that simultaneous treatment with 5-aza-2′-deoxycytidine (5Aza-2dc) in indoxyl sulfate (IS)- and p-cresyl sulfate (PCS)-injected mice significantly demethylated the Klotho gene. (b) Western blotting showed that 5Aza-2dc significantly increased the Klotho expression in IS- and PCS-injected mice.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3306006&req=5

fig4: DNA methyltransferase 1 (DNMT 1) inhibitor demethylated the Klotho gene and increased Klotho expression in vivo. (a) Methylation-specific PCR (MSP) with mouse Klotho set 1 primers showed that simultaneous treatment with 5-aza-2′-deoxycytidine (5Aza-2dc) in indoxyl sulfate (IS)- and p-cresyl sulfate (PCS)-injected mice significantly demethylated the Klotho gene. (b) Western blotting showed that 5Aza-2dc significantly increased the Klotho expression in IS- and PCS-injected mice.
Mentions: MSP analysis showed that inhibiting DNMT 1 by using 5Aza-2dc significantly decreased the DNA methylation of Klotho gene in IS- and PCS-injected mice when compared with the mice treated with IS or PCS alone (Figure 4a). In addition, the results of western blotting revealed that treatment with 5Aza-2dc significantly increased Klotho expression in IS- and PCS-injected mice when compared with the mice treated with IS or PCS alone (Figure 4b).

Bottom Line: The expression of DNA methyltransferases 1, 3a, and 3b isoforms in HK2 cells treated with indoxyl sulfate or p-cresyl sulfate was significantly increased.Specific inhibition of DNA methyltransferase isoform 1 by 5-aza-2'-deoxycytidine caused demethylation of the Klotho gene and increased Klotho expression in vitro.Thus, inhibition of Klotho gene expression by uremic toxins correlates with gene hypermethylation, suggesting that epigenetic modification of specific genes by uremic toxins may be an important pathological mechanism of disease.

View Article: PubMed Central - PubMed

Affiliation: School of Medicine, Chang Gung University, Taoyuan, Taiwan.

ABSTRACT
The expression of the renoprotective antiaging gene Klotho is decreased in uremia. Recent studies suggest that Klotho may be a tumor suppressor, and its expression may be repressed by DNA hypermethylation in cancer cells. Here we investigated the effects and possible mechanisms by which Klotho expression is regulated during uremia in uninephrectomized B-6 mice given the uremic toxins indoxyl sulfate or p-cresyl sulfate. Cultured human renal tubular HK2 cells treated with these toxins were used as an in vitro model. Injections of indoxyl sulfate or p-cresyl sulfate increased their serum concentrations, kidney fibrosis, CpG hypermethylation of the Klotho gene, and decreased Klotho expression in renal tubules of these mice. The expression of DNA methyltransferases 1, 3a, and 3b isoforms in HK2 cells treated with indoxyl sulfate or p-cresyl sulfate was significantly increased. Specific inhibition of DNA methyltransferase isoform 1 by 5-aza-2'-deoxycytidine caused demethylation of the Klotho gene and increased Klotho expression in vitro. Thus, inhibition of Klotho gene expression by uremic toxins correlates with gene hypermethylation, suggesting that epigenetic modification of specific genes by uremic toxins may be an important pathological mechanism of disease.

Show MeSH
Related in: MedlinePlus