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Tuberin-deficiency downregulates N-cadherin and upregulates vimentin in kidney tumor of TSC patients.

Liang S, Salas T, Gencaslan E, Li B, Habib SL - Oncotarget (2014)

Bottom Line: In addition, cells treated with rapamycin showed a significant increase in p-Akt and a decrease in p-p70S6K that was associated with a decrease expression of vimentin and a slight increase expression in N-cadherin.On the other hand, cells treated with Akt inhibitor revealed a significant decrease in p-Akt and p-p70S6K that was associated with a significant decrease in vimentin and an increase in N-cadherin expression.Kidney tumors from TSC patients showed significant decrease in N-cadherin and significant increased in vimentin protein expression compared to control kidney tissues.

View Article: PubMed Central - PubMed

Affiliation: South Texas Veterans Health Care System, San Antonio, TX.

ABSTRACT
Angiomyolipomas (AMLs) are associated with cell fibrosis in kidney of Tuberous Sclerosis Complex patients. The mechanism by which the fibrotic proteins accumulated in AMLs has not been explored. In the present study, we investigated the role of Akt/tuberin/mTOR pathway in the regulation cell fibrosis proteins. AML cells that expressed low levels of tuberin showed less expression of N-cadherin and higher of vimentin proteins compared to HEK293 cells. AML cells infected with Ad-tuberin showed a significant decrease in vimentin and an increase in N-cadherin protein expression. In addition, cells treated with rapamycin showed a significant increase in p-Akt and a decrease in p-p70S6K that was associated with a decrease expression of vimentin and a slight increase expression in N-cadherin. On the other hand, cells treated with Akt inhibitor revealed a significant decrease in p-Akt and p-p70S6K that was associated with a significant decrease in vimentin and an increase in N-cadherin expression. In addition, cells transfected with DN-Akt or DN-S6K show significant increase expression in N-cadherin and a decrease in vimentin. Moreover, cells transfected with siRNA against rictor or siRNA against raptor resulted in a decrease in vimentin and an increase N-cadherin expression. Kidney tumors from TSC patients showed significant decrease in N-cadherin and significant increased in vimentin protein expression compared to control kidney tissues. These data comprise the first report to provide the role of Akt/tuberin/mTORC1/2 in the regulation of N-cadherin and vimentin that are involved in the progression of fibrosis in kidney tumor of TSC patients.

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Inhibition of Akt resulted in significant decrease in vimentin and increase in N-cadherin expression in AML cellsAML cells were treated with different concentrations of Akt inhibitor IV (0-10μM) for 24h. Western blot analysis was performed in cell lysates using p-Akt, Akt, p-p70S6K, p70S6k, vimentin, N-cadherin and GAPDH antibodies. Cells treated with Akt inhibitor showed significant decrease in (A) P-Akt at Ser473 and (B) p-p70S6K at Thr389 compared to non-treated cells. Decrease in Akt phosphorylation is associated with significant decrease in (C) vimentin expression in cells treated with lower dose 2.5μM of Akt inhibitor. (D) Cells treated with lower dose of Akt inhibitor (2.5-5 μM) did not show any alteration in N-cadherin while cells treated with higher dose (10 μM) showed significant increase in N-cadherin expression compared to no-treated cells. GAPDH was used as a loading control.
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Figure 3: Inhibition of Akt resulted in significant decrease in vimentin and increase in N-cadherin expression in AML cellsAML cells were treated with different concentrations of Akt inhibitor IV (0-10μM) for 24h. Western blot analysis was performed in cell lysates using p-Akt, Akt, p-p70S6K, p70S6k, vimentin, N-cadherin and GAPDH antibodies. Cells treated with Akt inhibitor showed significant decrease in (A) P-Akt at Ser473 and (B) p-p70S6K at Thr389 compared to non-treated cells. Decrease in Akt phosphorylation is associated with significant decrease in (C) vimentin expression in cells treated with lower dose 2.5μM of Akt inhibitor. (D) Cells treated with lower dose of Akt inhibitor (2.5-5 μM) did not show any alteration in N-cadherin while cells treated with higher dose (10 μM) showed significant increase in N-cadherin expression compared to no-treated cells. GAPDH was used as a loading control.

Mentions: In order to confirm the role of Akt in regulating the expression of N-cadherin and vimentin, AML cells were treated with different concentrations of Akt inhibitor IV (0-10μM) for 24 hrs. Protein was extracted and Western blot analysis was performed and blotted against p-Akt (Ser473), Akt, p-p70S6K (Thr389), p70S6k, vimentin, N-cadherin antibodies. AML cells treated with Akt inhibitor showed significant decrease in P-Akt at Ser473 (Fig. 3A) and p-p70S6K at Thr389 (Fig. 3B) compared to non-treated cells. The expression of vimentin was also significantly decreased at very low doses of Akt inhibitor (2.5μM) and abolished at higher dose (5-10μm) (Figure 3C). AML cells treated with lower doses of (2.5-5μM) did not show any changes in N-cadherin expression while cells treated with higher dose (10μM) showed significant increase in N-cadherin expression compared to non-treated cells (Figure 3D). These data indicated that the change in both protein expressions is dose-dependent for Akt inhibitor. Together, these data indicates that Akt appositely regulates the expression of N-cadherin and vimentin in AML cells.


Tuberin-deficiency downregulates N-cadherin and upregulates vimentin in kidney tumor of TSC patients.

Liang S, Salas T, Gencaslan E, Li B, Habib SL - Oncotarget (2014)

Inhibition of Akt resulted in significant decrease in vimentin and increase in N-cadherin expression in AML cellsAML cells were treated with different concentrations of Akt inhibitor IV (0-10μM) for 24h. Western blot analysis was performed in cell lysates using p-Akt, Akt, p-p70S6K, p70S6k, vimentin, N-cadherin and GAPDH antibodies. Cells treated with Akt inhibitor showed significant decrease in (A) P-Akt at Ser473 and (B) p-p70S6K at Thr389 compared to non-treated cells. Decrease in Akt phosphorylation is associated with significant decrease in (C) vimentin expression in cells treated with lower dose 2.5μM of Akt inhibitor. (D) Cells treated with lower dose of Akt inhibitor (2.5-5 μM) did not show any alteration in N-cadherin while cells treated with higher dose (10 μM) showed significant increase in N-cadherin expression compared to no-treated cells. GAPDH was used as a loading control.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4196174&req=5

Figure 3: Inhibition of Akt resulted in significant decrease in vimentin and increase in N-cadherin expression in AML cellsAML cells were treated with different concentrations of Akt inhibitor IV (0-10μM) for 24h. Western blot analysis was performed in cell lysates using p-Akt, Akt, p-p70S6K, p70S6k, vimentin, N-cadherin and GAPDH antibodies. Cells treated with Akt inhibitor showed significant decrease in (A) P-Akt at Ser473 and (B) p-p70S6K at Thr389 compared to non-treated cells. Decrease in Akt phosphorylation is associated with significant decrease in (C) vimentin expression in cells treated with lower dose 2.5μM of Akt inhibitor. (D) Cells treated with lower dose of Akt inhibitor (2.5-5 μM) did not show any alteration in N-cadherin while cells treated with higher dose (10 μM) showed significant increase in N-cadherin expression compared to no-treated cells. GAPDH was used as a loading control.
Mentions: In order to confirm the role of Akt in regulating the expression of N-cadherin and vimentin, AML cells were treated with different concentrations of Akt inhibitor IV (0-10μM) for 24 hrs. Protein was extracted and Western blot analysis was performed and blotted against p-Akt (Ser473), Akt, p-p70S6K (Thr389), p70S6k, vimentin, N-cadherin antibodies. AML cells treated with Akt inhibitor showed significant decrease in P-Akt at Ser473 (Fig. 3A) and p-p70S6K at Thr389 (Fig. 3B) compared to non-treated cells. The expression of vimentin was also significantly decreased at very low doses of Akt inhibitor (2.5μM) and abolished at higher dose (5-10μm) (Figure 3C). AML cells treated with lower doses of (2.5-5μM) did not show any changes in N-cadherin expression while cells treated with higher dose (10μM) showed significant increase in N-cadherin expression compared to non-treated cells (Figure 3D). These data indicated that the change in both protein expressions is dose-dependent for Akt inhibitor. Together, these data indicates that Akt appositely regulates the expression of N-cadherin and vimentin in AML cells.

Bottom Line: In addition, cells treated with rapamycin showed a significant increase in p-Akt and a decrease in p-p70S6K that was associated with a decrease expression of vimentin and a slight increase expression in N-cadherin.On the other hand, cells treated with Akt inhibitor revealed a significant decrease in p-Akt and p-p70S6K that was associated with a significant decrease in vimentin and an increase in N-cadherin expression.Kidney tumors from TSC patients showed significant decrease in N-cadherin and significant increased in vimentin protein expression compared to control kidney tissues.

View Article: PubMed Central - PubMed

Affiliation: South Texas Veterans Health Care System, San Antonio, TX.

ABSTRACT
Angiomyolipomas (AMLs) are associated with cell fibrosis in kidney of Tuberous Sclerosis Complex patients. The mechanism by which the fibrotic proteins accumulated in AMLs has not been explored. In the present study, we investigated the role of Akt/tuberin/mTOR pathway in the regulation cell fibrosis proteins. AML cells that expressed low levels of tuberin showed less expression of N-cadherin and higher of vimentin proteins compared to HEK293 cells. AML cells infected with Ad-tuberin showed a significant decrease in vimentin and an increase in N-cadherin protein expression. In addition, cells treated with rapamycin showed a significant increase in p-Akt and a decrease in p-p70S6K that was associated with a decrease expression of vimentin and a slight increase expression in N-cadherin. On the other hand, cells treated with Akt inhibitor revealed a significant decrease in p-Akt and p-p70S6K that was associated with a significant decrease in vimentin and an increase in N-cadherin expression. In addition, cells transfected with DN-Akt or DN-S6K show significant increase expression in N-cadherin and a decrease in vimentin. Moreover, cells transfected with siRNA against rictor or siRNA against raptor resulted in a decrease in vimentin and an increase N-cadherin expression. Kidney tumors from TSC patients showed significant decrease in N-cadherin and significant increased in vimentin protein expression compared to control kidney tissues. These data comprise the first report to provide the role of Akt/tuberin/mTORC1/2 in the regulation of N-cadherin and vimentin that are involved in the progression of fibrosis in kidney tumor of TSC patients.

Show MeSH
Related in: MedlinePlus