Altered autophagy response in huntington ‘s disease (hd) human neural stem cells is context-dependent

JUAN CRUZ CASABONA; ISABEL FARIAS; FERNANDO JUAN PITOSSI; KOTLER

Riveirao Preto

Congreso; ISSCR International Congress.; 2023

ISSCR

Abstract TitleALTERED AUTOPHAGY RESPONSE IN HUNTINGTON ‘S DISEASE (HD) HIPSC DERIVED NEURAL STEM CELLS IS CONTEXT-DEPENDENT1.Abstract ObjectiveTo evaluate the effect of sphingosine 1-phosphate receptor (S1Pr) activation in human induced pluripotent stem cells (hiPSc)-derived neural stem cells (NSC) from HD patients on apoptosis, mitochondrial network and autophagy2.Abstract BackgroundAlthough S1P signaling has been involved in several pathways of neurodegenerative diseases such as HD, data regarding a protective or detrimental role S1Pr stimulation still remains controversial. In this sense, the NCS from HD patients retain several characteristics of HD and  have been proposed as a promising strategy to test the functional role of S1Pr.3.Abstract MethodhiPS from controls or HD patients were generated from fibroblasts and differentiated to NSC (Figure 1). >95% of the NSCs expressed Sox1, Sox2, Pax6 and Nestin markers, as expected for bona fide NSCs (Figure 2A and 2B). One NSC line from each group were incubated with or without growth factors for 24 h and treated with different dilutions of a S1Pr modulator (BAF312) or media. Under these conditions, cell viability was studied by MTS assay, apoptosis was evaluated by measuring Cleaved Caspase 3 and DAPI-stained apoptotic nuclei counting and autophagy activity was analyzed by LC3B  immunofluorescence. Changes in morphology of the mitochondrial network were studied by staining with MitoTracker Red CMXRos.4.Abstract ResultsThe incubation of cells with BAF312  showed no changes in cell viability (Figure  3), apoptosis (Figure 4) or on mitochondrial network in any cell line used (Figure 5A and 5B), with or without growth factors deprivation. Interestingly, growth factors deprivation elicited autophagy downregulation in control NSC, but not in HD NSC. In the presence of growth factors, S1Pr stimulation at 1/10000 dilution increased autophagy in control NSC, but not in HD NSC. At different dilutions (1/1000 and 1/100.000) this treatment increased autophagy in both cell types (Figure 6A and 6B).5.Abstract ConclusionIn HD NSCs autophagy depends on the context, dose of S1Pr modulator and growth factors presence. These variables add to the ones previously reported (compartment and cell-type) to affect S1Pr effects, keeping the controversy open.Topic:Disease ModellingKeyword 1Huntington´s disease Keyword 2BAF312 S1P Receptor ModulatorKeyword 3Autophagy