Fig 1: Effects of Salusin-α overexpression or interference on the levels of AdipoR2, APPRα, ApoA5 and SREBP-1c. (A) HepG2 cells were transfected with pHAGE and pHAGE-Salusin-α. Two groups of HepG2 cells infected with pHAGE-Salusin-α, after 24 h of culture, one of the pHAGE-Salusin-α groups was treated with Tha for 24 h, whereas the other groups were continued to cultivate and remained unchanged. WB analysis detected the changes in the levels of AdipoR2, PPARα, ApoA5 and SREBP-1c proteins. (B) HepG2 cells were transfected with shMock and shSalusin-α#1. Two groups of HepG2 cells infected with shSalusin-α#1, then after 24 h, one of the shSalusin-α#1 groups was treated with PBA for 24 h, whereas the other groups were continued to cultivate and remain unchanged. WB analysis detected the changes in the protein levels of AdipoR2, PPARα, ApoA5 and SREBP-1c. The data are presented as the mean ± SD. *P<0.05, **P<0.01 and ***P<0.001 by t-test. NC, negative control; shMock, a meaningless RNA as a control for shSalusin-α; shSalusin-α#1, pLKO.1-shSalusin-α#1, representing interference salusin-α; pHAGE-Salusin-α, representing overexpression salusin-α; Tha, thapsigargin; PBA, 4-phenyl butyric acid; AdipoR2, adiponectin receptor 2; PPARα, peroxisome proliferator-activated receptor-α; ApoA5, apolipoprotein A5; SREBP-1c, sterol regulatory element-binding transcription factor 1; sh-, short hairpin.
Fig 2: Overexpression of salusin-α upregulates AdipoR2 and activates the PPARα/ApoA5/SREBP-1c pathway. Based on the present study, the following hypothesis is proposed: Salusin-α and the interference sequences were transported into cells by lentivirus, and after a series of modification, the expression level of salusin-α was changed. Salusin-α can influence AdipoR2, and its overexpression leads to upregulation of the AdipoR2 expression level, thereby upregulating the expression of PPARα in hepatocytes, whereas PPARα upregulates the expression of ApoA5 and inhibits the expression of SREBP-1c, ultimately promoting fatty acid oxidation and reducing lipid accumulation in hepatocytes. There are numerous possibilities that could account for how salusin-α interacts with AdipoR2: It may be that salusin-α directly binds with AdipoR2 and activates it, or salusin-α indirectly acts on AdipoR2 through adiponectin, and so on; these aspects need to be confirmed by subsequent research. The gray arrow represents the mechanism process of overexpression lentivirus transporting salusin-α and regulating salusin-α expression. The red arrow represents the mechanism process that interferes with the lentivirus to transport the interfering sequence of salusin-α and thus regulates salusin-α. shsalusin-α, Interference sequence of salusin-α. PPARα, peroxisome proliferator-activated receptor-α; ApoA5, apolipoprotein A5; SREBP-1c, sterol regulatory element-binding transcription factor 1; AdipoR2, adiponectin receptor 2; sh-, short hairpin.
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