Extended data Fig. 6: P-G3 enters cells to modulate mTOR and NAD signal pathways in adipocyte development.

a, Schematic experiment design (top) and Oil Red O staining (bottom) of 3T3-L1 cells after the treatments of different localizations of P-G3. 3T3-L1 preadipocytes were differentiated in the presence of naked P-G3, microbead-conjugated P-G3 to prevent entering cells, or P-G3 beads in transwells to prevent the contact with cells. b, qPCR analyses of gene expression of cells in (a). Data were represented as mean ± s.e.m. (n = 4, 4, 4, 4). Statistical significance was calculated via 2-tailed Student’s t-test (treatment group vs vehicle group). c, The internalization of Cy5-labelled P-G3 into early endosome. Confocal images of Cy5-labelled P-G3 with early endosome marker in mature C3H10T1/2 adipocytes after 15 min or 1 hr of Cy5-P-G3 treatment. d, Colocalization of Cy5-P-G3 with lipid droplet, ER, and mitochondria in mature 3T3-L1 adipocytes after 24-hr of Cy5-P-G3 treatment. Representative data in c and d repeated twice with similar results. e, Representative gating strategy used in FACS analysis of lysosomal activity in Fig. 5b. f, Gene expression in C3H10T1/2 cells after indicated treatments from differentiation Day 4 to Day 9. Data were represented as mean ± s.e.m. (n = 4, 4). Statistical significance was calculated via 2-tailed Student’s t-test (treatment group vs vehicle group). g, P-G3 failed to affect NAD⁺ and NADH levels in mature adipocytes after 14-hr treatment. Data were represented as mean ± s.e.m. (n = 3, 3).