Fig. 1: Identification of the upregulated sphingosine biosynthesis pathway in TSC2-deficient cells. | Cell Death & Disease

Fig. 1: Identification of the upregulated sphingosine biosynthesis pathway in TSC2-deficient cells.

From: Targeting SPHK1/S1PR3-regulated S-1-P metabolic disorder triggers autophagic cell death in pulmonary lymphangiomyomatosis (LAM)

Fig. 1

A Immunohistochemistry staining of ASAH1, SPHK1 and phospho-S6(Ser235/236) was performed on rat-derived ELT3-V3 (TSC2-deficient, V3) and ELT3-T3 (TSC2-expression, T3) cell xenograft tumors of SCID mice. Representative images of five mice in each group. B Immunoblot analysis of ASAH1, SPHK1, S1PR3, p-ERK, pS6(235/236) protein levels in 621-101 cells treated with vehicle control or 10 nM estrogen for 24 h. β-actin was used as the loading control. C–F were treated with vehicle control or 20 nM rapamycin for 24 h. The mRNA levels of TSC2, ASAH1, and SPHK1 were detected by RT-qPCR in (C) 621-101 and 621-103 cells, (E) Mouse embryo fibrosis cells (TSC2− MEFs and TSC2 + MEFs). Data showed the mean of three sets of independent samples. Protein expressions of tuberin, SPHK1, pS6(ser235/236) and ASAH1 were determined by immunoblot analysis in (D) 621-101 and 621-103 cells, (F) Mouse embryo fibrosis cells (TSC2- MEFs and TSC2+ MEFs). β-actin was used as loading control. G, H TSC2-addback cells (621-103) were transfected with human TSC2 siRNAs or control siRNA for 24 h. Gene expressions of TSC2, SPHK1, and ASAH1 were determined by RT-qPCR, and protein levels of Tuberin, SPHK1, pS6(ser235/236) and ASAH1 were detected by immunoblotting. β-actin was used as loading control. Student’s t test, *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001.

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