3-Hydroxybutyrate Ameliorates the Progression of Diabetic Nephropathy

Abstract

Studies report beneficial effects of 3-hydroxybutyrate (3-OHB) on the treatment of type 2 diabetes and obesity, but the effects of 3-OHB on diabetic nephropathy have not been elucidated. This study was designed to investigate the efficacy and mechanism of 3-OHB against progression of diabetic nephropathy (DN). Mice (db/db) were fed normal chow, high-fat, or ketogenic diets (KD) containing precursors of 3-OHB. Hyperglycemia was determined based on random glucose level (≥250 mg/dL). Fasting blood glucose and body weights were measured once a week. Twenty four-hour urine albumin to creatinine ratio was determined 5 weeks after the differential diet. Energy expenditure was measured 9 weeks after the differential diet. Body weights were significantly lower in the KD group than those in other groups, but no significant differences in fasting blood glucose levels among three groups were observed. Urine albumin to creatinine ratio and serum blood urea nitrogen (BUN) to creatinine ratio in the KD group were significantly lower than in other groups. Histologic and quantitative analysis of mesangial area suggested that KD delayed the progression of DN phenotype in db/db mice. Metabolic cage analysis also revealed that KD increased energy expenditure in db/db mice. In vitro studies with proximal tubular cells revealed that 3-OHB stimulated autophagic flux. 3-OHB increased LC3 I to LC3 II ratio, phosphorylation of AMPK, beclin, p62 degradation, and NRF2 expression. Moreover, we found that 3-OHB attenuated high glucose-induced reactive oxygen species (ROS) levels in proximal tubular cells. In vivo study also confirmed increased LC3 and decreased ROS levels in the kidney of KD mice. In summary, this study shows in both in vivo and in vitro models that 3-OHB delays the progression of DN by augmenting autophagy and inhibiting oxidative stress.

Critical Reanalysis of the Mechanisms Underlying the Cardiorenal Benefits of SGLT2 Inhibitors and Reaffirmation of the Nutrient Deprivation Signaling/Autophagy Hypothesis

“…26,37 Ketone Body-Induced Nutrient Deprivation Signaling and Autophagic Flux SGLT2 inhibitors induce a state of starvation mimicry characterized by ketogenesis 46,51 and the delivery of ketone bodies to organs can exert a direct effect to increase AMPK and decrease mTOR phosphorylation, to increase the expression of PGC-1α, and to promote autophagic flux. 57,[234][235][236][237][238] Potentially acting through these mediators, The following refer to specialized cell lines or mutant mice: AML-12 hepatocytes, db/db mice, H9c2 cardiomyocytes, HK2 proximal renal tubular cells, HL-1 cardiomyocytes, LLC-PK1 proximal tubular cells, LO2 hepatocytes, RAW264.7 macrophages, and THP-1 monocytes. AMPK indicates adenosine monophosphate-activated protein kinase; Atg5, autophagy related gene 5; BNIP3, B-cell lymphoma 2 interacting protein 3; FUNDC1, FUN14 domain containing 1; LAMP-1, lysosomalassociated membrane protein 1; LC3, microtubule-associated protein 1A/1B-light chain 3; SGLT2, sodium-glucose cotransporter 2; SGLT2 p62, sequestosome 1 (ubiquitin-binding protein); SGLT2i, sodium-glucose cotransporter 2 inhibitor; SIRT3, sirtuin-3; TLR9, Toll-like receptor 9; and Ulk,Unc-51-like kinase.…”

“…AMPK indicates adenosine monophosphate-activated protein kinase; Atg5, autophagy related gene 5; BNIP3, B-cell lymphoma 2 interacting protein 3; FUNDC1, FUN14 domain containing 1; LAMP-1, lysosomalassociated membrane protein 1; LC3, microtubule-associated protein 1A/1B-light chain 3; SGLT2, sodium-glucose cotransporter 2; SGLT2 p62, sequestosome 1 (ubiquitin-binding protein); SGLT2i, sodium-glucose cotransporter 2 inhibitor; SIRT3, sirtuin-3; TLR9, Toll-like receptor 9; and Ulk,Unc-51-like kinase. 57,[234][235][236][237][238][239][240] Ketone body supplementation inhibits phenylephrine-induced hypertrophy in experiments 238 and interruption of ketogenesis negates the benefits of SGLT2 inhibitors on the kidney. 57 These observations suggest that the ketonemia seen after SGLT2 inhibitors use may exert cardioprotective and nephroprotective effects as a result of a direct action to stimulate nutrient deprivation signaling rather than because of an ability to serve as an efficient fuel for the generation of ATP.…”

“…SGLT2 inhibitors induce a state of starvation mimicry characterized by ketogenesis 46,51 and the delivery of ketone bodies to organs can exert a direct effect to increase AMPK and decrease mTOR phosphorylation, to increase the expression of PGC-1α, and to promote autophagic flux. 57,234–238 Potentially acting through these mediators, ketonemia has been shown to reduce oxidative stress, mute cellular inflammation, enhance mitochondrial biogenesis, and preserve cardiac and renal function after diverse stresses. 57,234–240 Ketone body supplementation inhibits phenylephrine-induced hypertrophy in experiments 238 and interruption of ketogenesis negates the benefits of SGLT2 inhibitors on the kidney.…”

“…57,234–238 Potentially acting through these mediators, ketonemia has been shown to reduce oxidative stress, mute cellular inflammation, enhance mitochondrial biogenesis, and preserve cardiac and renal function after diverse stresses. 57,234–240 Ketone body supplementation inhibits phenylephrine-induced hypertrophy in experiments 238 and interruption of ketogenesis negates the benefits of SGLT2 inhibitors on the kidney. 57 These observations suggest that the ketonemia seen after SGLT2 inhibitors use may exert cardioprotective and nephroprotective effects as a result of a direct action to stimulate nutrient deprivation signaling rather than because of an ability to serve as an efficient fuel for the generation of ATP.…”

Oxidative stress, inflammation, dysfunctional redox homeostasis and autophagy cause age-associated diseases

“…Phosphorylated PGC-1α activates Nrf1/2 that resulting in the expression of mitochondrial transcription factor A (mtTFA), which stimulates replication and transcription of mtDNA, leading to mitochondrial biogenesis and mitochondrial OP process to satisfy the energy demand [174,175]. Additionally, exercising muscle also releases myokines, including muscle-derived IL-6 and irisin [176,177], as well as β-hydroxybutyrate, a ketone metabolite [178,179], all of which synergistically suppress inflammation in muscle as well as in other organs, thus helping to alleviate AADs, such as in liver (NAFLD, liver fibrosis, HCC), adipose tissue and pancreas (insulin resistance, diabetes), blood vessels (AS, hypertension, and CVD) and brain (AD and PD) [180,181]. Mechanistically, β-hydroxybutyrate can activate AMPK and Nrf2 as well as inhibit inflammation by suppressing inflammasome formation and endoplasmic reticulum stress [179,180].…”