Journal: Cell Death Discovery

Article Title: Inhibition of TGF-beta signaling protects from alpha-synuclein induced toxicity

doi: 10.1038/s41420-025-02901-2

Figure Lengend Snippet: Quantification of LDH release into the cell culture medium after treatment of LUHMES cells with TGFb1 ( A ), TGFb2 ( B ), or TGFb3 ( C ). The treatment of TGFb ligands led to an increase in aSyn-induced toxicity, as compared to untreated aSyn-overexpressing cells. The knockdown of ALK5 , SMAD2 , and DYNLRB1 by siPOOL siRNA significantly protected TGFb ligand-treated aSyn-overexpressing cells. Data are normalized to the LDH release of untreated aSyn-overexpressing cells. D Representative images of the ICC staining against DAPI (blue), GFP (green, if any), aSyn (red), and beta-III tubulin (cyan) in aSyn-overexpressing cells. Corresponding network analysis for each representative figure is shown below the staining, in which black lines outline the neurites. E Quantification of the number of quadruple points per cell as a measure for changes in the density of neuronal network showed that the treatment of TGFb1 led to a reduction in the density of neuronal network in aSyn-overexpressing cells. The knockdown of ALK5 , SMAD2 , or DYNLRB1 by siPOOL siRNA significantly increased the density of the neuronal network in TGFb1-treated aSyn-overexpressing cells. TGFb1 ligand treatment did not result in significant changes in the density of neuronal network in untransduced cells (UTCs) or GFP-expressing cells. In addition, no significant changes in the density of the neuronal network resulted from the knockdown of ALK5 , SMAD2 , and DYNLRB1 in TGFb1-treated UTCs or GFP-expressing cells. F Quantification of the relative mRNA expression of SNCA after the treatment of TGFb1, TGFb2, or TGFb3 in UTCs, aSyn-overexpressing cells, and GFP-expressing cells. TGFb2 and TGFb3 treatment led to a significant increase in the expression of SNCA only in aSyn-overexpressing cells. UTCs: untransduced cells; aSyn/+aSyn: cells overexpressing alpha-Synuclein; +GFP: cells expressing GFP; ALK5 siPOOL: siPOOL siRNA against ALK5 ; neg. ctrl. siPOOL: negative control siPOOL siRNA; SMAD2 siPOOL: siPOOL siRNA against SMAD2 ; DYNLRB1 siPOOL: siPOOL siRNA against DYNLRB1 ; untreated: cells that were not treated with TGFb ligands; untransfected: cells that were not transfected with siPOOL siRNAs. # p < 0.05, ### p < 0.001 against untreated UTCs; * p < 0.05, ** p < 0.01, *** p < 0.001 against untreated aSyn; § p < 0.05, §§§ p < 0.001 against aSyn+TGFb ligand; ^ p < 0.05 against untreated GFP; ns: not significant.

Article Snippet: The following ligands and inhibitors were used: Recombinant human TGFb1 protein (R&D Systems), recombinant human TGFb2 protein (R&D Systems), recombinant human TGFb3 protein (R&D Systems), tideglusib (Sigma-Aldrich), GSK3B-inhibitor VIII (Sigma-Aldrich), or SM16 (Tocris, Minneapolis, MN, USA)

Techniques: Cell Culture, Knockdown, Staining, Expressing, Negative Control, Transfection

Journal: Cell Death Discovery

Article Title: Inhibition of TGF-beta signaling protects from alpha-synuclein induced toxicity

doi: 10.1038/s41420-025-02901-2

Figure Lengend Snippet: Quantification of the relative gene expression of TGFB1 ( A ), TGFB2 ( B ), TGFB3 ( C ), and TGFBR2 ( D ) was analyzed in untransduced cells (UTCs), aSyn-overexpressing cells, and GFP-expressing cells. TGFB1 and TGFB2 were found to be up-regulated in aSyn-overexpressing cells only. # p < 0.05, ## p < 0.01, ### p < 0.001 against UTCs; +++ p < 0.001: p < 0.001 against GFP-expressing cells; ns: not significant.

Article Snippet: The following ligands and inhibitors were used: Recombinant human TGFb1 protein (R&D Systems), recombinant human TGFb2 protein (R&D Systems), recombinant human TGFb3 protein (R&D Systems), tideglusib (Sigma-Aldrich), GSK3B-inhibitor VIII (Sigma-Aldrich), or SM16 (Tocris, Minneapolis, MN, USA)

Techniques: Gene Expression, Expressing