Pharmacologic inhibition of the renal sodium/glucose cotransporter-2 induces glycosuria and reduces glycemia. Given that SGLT2 inhibitors (SGLT2i) reduce mortality and cardiovascular risk in type 2 diabetes, improved understanding of molecular mechanisms mediating these metabolic effects is required. Treatment of obese but nondiabetic mice with the SGLT2i canagliflozin (CANA) reduces adiposity, improves glucose tolerance despite reduced plasma insulin, increases plasma ketones, and improves plasma lipid profiles. Utilizing an integrated transcriptomic-metabolomics approach, we demonstrate that CANA modulates key nutrient-sensing pathways, with activation of 5′ AMP-activated protein kinase (AMPK) and inhibition of mechanistic target of rapamycin (mTOR), independent of insulin or glucagon sensitivity or signaling. Moreover, CANA induces transcriptional reprogramming to activate catabolic pathways, increase fatty acid oxidation, reduce hepatic steatosis and diacylglycerol content, and increase hepatic and plasma levels of FGF21. Given that these phenotypes mirror the effects of FGF21 to promote lipid oxidation, ketogenesis, and reduction in adiposity, we hypothesized that FGF21 is required for CANA action. Using FGF21-null mice, we demonstrate that FGF21 is not required for SGLT2i-mediated induction of lipid oxidation and ketogenesis but is required for reduction in fat mass and activation of lipolysis. Taken together, these data demonstrate that SGLT2 inhibition triggers a fasting-like transcriptional and metabolic paradigm but requires FGF21 for reduction in adiposity.

Background: Nonalcoholic fatty liver disease (NAFLD) associates with cardiovascular disease independently of classic risk factors. Proprotein Convertase Subtilisin/Kexin type 9 (PCSK9) is secreted by hepatocytes and inhibits the uptake of low-density lipoproteins by targeting the receptor for degradation, and possibly lipogenesis. PCSK9 loss-of-function mutations and anti-PCKS9 drugs reduce LDL-cholesterol. Aim: To evaluate whether hepatic fat content is associated with circulating PCSK9. Materials and methods: In 201 consecutive patients biopsied for suspected nonalcoholic steatohepatitis, liver damage was quantified by NAFLD activity score, circulating PCSK9 by ELISA, and hepatic mRNA by qRT-PCR in a subset (n = 76). Results: Circulating PCSK9 was associated with steatosis grade (p = 0.0011), necroinflammation (p < 0.001), ballooning (p = 0.005), and fibrosis stage (p = 0.001). At multivariate analysis, PCSK9 was associated with steatosis grade (p = 0.012), older age and lower BMI, independently of sex, hyperglycemia, and fibrosis/inflammation. Circulating PCSK9 was associated with hepatic expression of SREBP-1c (p = 0.0002) and FAS (p = 0.03). PCSK9 mRNA levels were also correlated with steatosis severity (p = 0.04) and hepatic APOB (p < 0.001), SREBP-1c (p = 0.047) and FAS expression (p = 0.001). Conclusions: Circulating PCSK9 increases with hepatic fat accumulation and correlates with the severity of steatosis, independently of metabolic confounders and liver damage. Modulation of PCSK9 synthesis and release might be involved in NAFLD pathogenesis.

The suppressor of cytokine signaling (SOCS) proteins are negative regulators of the JAK/STAT pathway activated by pro-inflammatory cytokines, including the tumor necrosis factor-α (TNF-α). SOCS3 is also implicated in hypertriglyceridemia associated to insulin-resistance (IR). Proprotein Convertase Subtilisin Kexin Type 9 (PCSK9) levels are frequently found to be positively correlated to IR and plasma very low-density lipoprotein-triglycerides (VLDL-TG) concentrations. The present study aimed to investigate the possible role of TNF-α and JAK/STAT pathway on de novo lipogenesis and PCSK9 expression in HepG2 cells. TNF-α induced both SOCS3 and PCSK9 in a concentration-dependent manner. This effect was inhibited by transfection with siRNA anti-STAT3, suggesting the involvement of the JAK/STAT pathway. Retroviral overexpression of SOCS3 in HepG2 cells (HepG2^SOCS3) strongly inhibited STAT3 phosphorylation and induced PCSK9 mRNA and protein, with no effect on its promoter activity and mRNA stability. Consistently, siRNA anti-SOCS3 reduced PCSK9 mRNA levels while an opposite effect was observed with siRNA anti-STAT3. In addition, HepG2^SOCS3 express higher mRNA levels of key enzymes involved in the de novo lipogenesis, such as fatty-acid synthase (FAS), stearoyl-CoA desaturase, and apo-B. These responses were associated with significant increase of SCD-1 protein, activation of SREBP-1, accumulation of cellular TG, and secretion of apoB. HepG2^SOCS3 shows lower phosphorylation levels of IRS-1 Tyr⁸⁹⁶ and Akt Ser⁴⁷³ in response to insulin. Finally, insulin stimulation produced an additive effect with SOCS3 overexpression, further inducing PCSK9, SREBP-1, FAS and apoB mRNA. In conclusion, our data candidate PCSK9 as a gene involved in lipid metabolism regulated by pro-inflammatory cytokine TNF-α, in a SOCS3 dependent manner.