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Substrate-specific gene expression profiles in different kidney cell types are associated with Fabry disease.

Shin YJ, Jeon YJ, Jung N, Park JW, Park HY, Jung SC - Mol Med Rep (2015)

Bottom Line: The majority of the biological functions of the regulated genes were associated with fibrogenesis or epithelial‑mesenchymal transition (EMT).In the HK‑2 cells, the ADAMTS6, BEST1, IL4, and MYH11 genes were upregulated.The gene expression profiles in kidney cells following the addition of Gb3 or lyso‑Gb3 revealed substrate‑specific and cell‑specific patterns.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, School of Medicine, Ewha Womans University, Seoul 158‑710, Republic of Korea.

ABSTRACT
Fabry disease is an X-linked lysosomal storage disorder caused by mutations in the gene encoding the α-galactosidase A (α-Gal A) lysosomal enzyme, which results in globotriaosylceramide (Gb3) storage in vascular endothelial cells and different cell types throughout the body. Involvement of the kidney and heart is life threatening, and fibrosis of these organs is considered to be involved in the pathogenesis of Fabry disease. An increased concentration of deacylated Gb3 (lyso‑Gb3) in the plasma of symptomatic patients has also been suggested as a causative molecular event. To elucidate the molecular mechanisms involved in renal fibrosis in Fabry disease, the present analyzed the changes in global gene expression prior to and following Gb3 or lyso‑Gb3 treatment in two types of kidney cell lines, human proximal renal tubular epithelial (HK‑2) and mouse renal glomerular mesangial (SV40 MES 13) cells. Gb3 and lyso‑Gb3 treatment regulated the expression of 199 and 328 genes in each cell type, demonstrating a >2.0‑fold change. The majority of the biological functions of the regulated genes were associated with fibrogenesis or epithelial‑mesenchymal transition (EMT). The gene expression patterns of sphingolipid‑treated HK‑2 cells were distinguishable from the patterns in the SV40 MES 13 cells. Several genes associated with the EMT were selected and evaluated further in kidney cells and in Fabry mouse kidney tissues. In the SV40 MES 13 cells, the DLL1, F8, and HOXA11 genes were downregulated, and FOXP2 was upregulated by treatment with Gb3 or lyso‑Gb3. In the HK‑2 cells, the ADAMTS6, BEST1, IL4, and MYH11 genes were upregulated. Upregulation of the FOXP2, COL15A1, IL4, and MYH11 genes was also observed in the Fabry mouse kidney tissues. The gene expression profiles in kidney cells following the addition of Gb3 or lyso‑Gb3 revealed substrate‑specific and cell‑specific patterns. These findings suggested that Gb3 and lyso‑Gb3 lead to renal fibrosis in Fabry disease through different biochemical modulations.

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RT-qPCR analyses to determine changes in gene expression. (A) RT-qPCR analyses of the HOXA11, FOXP2, F8 and DLL1 genes in SV40 MES 13 cells, and of the (B) ADAMTS6, BEST1, IL4 and MYH11 genes in HK-2 cells. Expression levels of genes in SV40 MES 13 and HK-2 cells treated with Gb3 or lyso-Gb3 were analyzed using RT-qPCR and expressed as a ratio relative to the expression in untreated cells. Data are presented as the mean ± standard error of the mean of at least three experiments. RT-qPCR, reverse transcription-quantitative polymerase chain reaction; Gb3, globotriaosylceramide; lyso-Gb3, deacylated Gb3.
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f3-mmr-12-04-5049: RT-qPCR analyses to determine changes in gene expression. (A) RT-qPCR analyses of the HOXA11, FOXP2, F8 and DLL1 genes in SV40 MES 13 cells, and of the (B) ADAMTS6, BEST1, IL4 and MYH11 genes in HK-2 cells. Expression levels of genes in SV40 MES 13 and HK-2 cells treated with Gb3 or lyso-Gb3 were analyzed using RT-qPCR and expressed as a ratio relative to the expression in untreated cells. Data are presented as the mean ± standard error of the mean of at least three experiments. RT-qPCR, reverse transcription-quantitative polymerase chain reaction; Gb3, globotriaosylceramide; lyso-Gb3, deacylated Gb3.

Mentions: To confirm the microarray-predicted expression patterns, the present study performed RT-qPCR to analyze the candidate genes. The 'target' genes, which presented with relatively consistent patterns in the micro-array and RT-qPCR, including IL4, MYH11, ADAMTS6, BEST1, FOXP2, F8, HOXA11, DLL1 and WT1 were identified. In the SV40 MES 13 mesangial cells treated with Gb3 or lyso-Gb3, the expression of FOXP2 was upregulated and the expression levels of HOXA11, F8 and DLL1 were downregulated, compared with the control cells (Fig. 3A). In the HK-2 epithelial cells, the mRNA expression levels of ADAMTS6, BEST1, IL4 and MYH11 were upregulated in the Gb3- and lyso-Gb3-treated epithelial cells, compared with the control cells (Fig. 3B).


Substrate-specific gene expression profiles in different kidney cell types are associated with Fabry disease.

Shin YJ, Jeon YJ, Jung N, Park JW, Park HY, Jung SC - Mol Med Rep (2015)

RT-qPCR analyses to determine changes in gene expression. (A) RT-qPCR analyses of the HOXA11, FOXP2, F8 and DLL1 genes in SV40 MES 13 cells, and of the (B) ADAMTS6, BEST1, IL4 and MYH11 genes in HK-2 cells. Expression levels of genes in SV40 MES 13 and HK-2 cells treated with Gb3 or lyso-Gb3 were analyzed using RT-qPCR and expressed as a ratio relative to the expression in untreated cells. Data are presented as the mean ± standard error of the mean of at least three experiments. RT-qPCR, reverse transcription-quantitative polymerase chain reaction; Gb3, globotriaosylceramide; lyso-Gb3, deacylated Gb3.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4581816&req=5

f3-mmr-12-04-5049: RT-qPCR analyses to determine changes in gene expression. (A) RT-qPCR analyses of the HOXA11, FOXP2, F8 and DLL1 genes in SV40 MES 13 cells, and of the (B) ADAMTS6, BEST1, IL4 and MYH11 genes in HK-2 cells. Expression levels of genes in SV40 MES 13 and HK-2 cells treated with Gb3 or lyso-Gb3 were analyzed using RT-qPCR and expressed as a ratio relative to the expression in untreated cells. Data are presented as the mean ± standard error of the mean of at least three experiments. RT-qPCR, reverse transcription-quantitative polymerase chain reaction; Gb3, globotriaosylceramide; lyso-Gb3, deacylated Gb3.
Mentions: To confirm the microarray-predicted expression patterns, the present study performed RT-qPCR to analyze the candidate genes. The 'target' genes, which presented with relatively consistent patterns in the micro-array and RT-qPCR, including IL4, MYH11, ADAMTS6, BEST1, FOXP2, F8, HOXA11, DLL1 and WT1 were identified. In the SV40 MES 13 mesangial cells treated with Gb3 or lyso-Gb3, the expression of FOXP2 was upregulated and the expression levels of HOXA11, F8 and DLL1 were downregulated, compared with the control cells (Fig. 3A). In the HK-2 epithelial cells, the mRNA expression levels of ADAMTS6, BEST1, IL4 and MYH11 were upregulated in the Gb3- and lyso-Gb3-treated epithelial cells, compared with the control cells (Fig. 3B).

Bottom Line: The majority of the biological functions of the regulated genes were associated with fibrogenesis or epithelial‑mesenchymal transition (EMT).In the HK‑2 cells, the ADAMTS6, BEST1, IL4, and MYH11 genes were upregulated.The gene expression profiles in kidney cells following the addition of Gb3 or lyso‑Gb3 revealed substrate‑specific and cell‑specific patterns.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, School of Medicine, Ewha Womans University, Seoul 158‑710, Republic of Korea.

ABSTRACT
Fabry disease is an X-linked lysosomal storage disorder caused by mutations in the gene encoding the α-galactosidase A (α-Gal A) lysosomal enzyme, which results in globotriaosylceramide (Gb3) storage in vascular endothelial cells and different cell types throughout the body. Involvement of the kidney and heart is life threatening, and fibrosis of these organs is considered to be involved in the pathogenesis of Fabry disease. An increased concentration of deacylated Gb3 (lyso‑Gb3) in the plasma of symptomatic patients has also been suggested as a causative molecular event. To elucidate the molecular mechanisms involved in renal fibrosis in Fabry disease, the present analyzed the changes in global gene expression prior to and following Gb3 or lyso‑Gb3 treatment in two types of kidney cell lines, human proximal renal tubular epithelial (HK‑2) and mouse renal glomerular mesangial (SV40 MES 13) cells. Gb3 and lyso‑Gb3 treatment regulated the expression of 199 and 328 genes in each cell type, demonstrating a >2.0‑fold change. The majority of the biological functions of the regulated genes were associated with fibrogenesis or epithelial‑mesenchymal transition (EMT). The gene expression patterns of sphingolipid‑treated HK‑2 cells were distinguishable from the patterns in the SV40 MES 13 cells. Several genes associated with the EMT were selected and evaluated further in kidney cells and in Fabry mouse kidney tissues. In the SV40 MES 13 cells, the DLL1, F8, and HOXA11 genes were downregulated, and FOXP2 was upregulated by treatment with Gb3 or lyso‑Gb3. In the HK‑2 cells, the ADAMTS6, BEST1, IL4, and MYH11 genes were upregulated. Upregulation of the FOXP2, COL15A1, IL4, and MYH11 genes was also observed in the Fabry mouse kidney tissues. The gene expression profiles in kidney cells following the addition of Gb3 or lyso‑Gb3 revealed substrate‑specific and cell‑specific patterns. These findings suggested that Gb3 and lyso‑Gb3 lead to renal fibrosis in Fabry disease through different biochemical modulations.

Show MeSH
Related in: MedlinePlus