Effects Of BHBA Research Articles

Subclinical ketosis leads to lipid metabolism disorder by downregulating the expression of acetyl-coenzyme A acetyltransferase 2 in dairy cows

Ketosis is a metabolic disease that often occurs in dairy cows postpartum and is a result of disordered lipid metabolism. Acetyl-coenzyme A (CoA) acetyltransferase 2 (ACAT2) is important for balancing cholesterol and triglyceride (TG) metabolism; however, its role in subclinical ketotic dairy cows is unclear. This study aimed to explore the potential correlation between ACAT2 and lipid metabolism disorders in subclinical ketotic cows through in vitro and in vivo experiments. In the in vivo experiment, liver tissue and blood samples were collected from healthy cows (CON, n = 6, β-hydroxybutyric acid [BHBA] concentration <1.0 mM) and subclinical ketotic cows (subclinical ketosis [SCK], n = 6, BHBA concentration = 1.2-3.0 mM) to explore the effect of ACAT2 on lipid metabolism disorders in SCK cows. For the in vitro experiment, bovine hepatocytes (BHEC) were used as the model. The effects of BHBA on ACAT2 and lipid metabolism were investigated via BHBA concentration gradient experiments. Subsequently, the relation between ACAT2 and lipid metabolism disorder was explored by transfection with siRNA of ACAT2. Transcriptomics showed an upregulation of differentially expression genes during lipid metabolism and significantly lower ACAT2 mRNA levels in the SCK group. Compared with the CON group in vivo, the SCK group showed significantly higher expression levels of peroxisome proliferator-activated receptor γ (PPARγ) and sterol regulator element binding protein 1c (SREBP1c) and significantly lower expression levels of peroxisome proliferator-activated receptor α (PPARα), carnitine palmitoyl-transferase 1A (CPT1A), sterol regulatory element binding transcription factor 2 (SREBP2), and 3-hydroxy-3-methylglutaryl-CoA reductase (HMGCR). Moreover, the SCK group had a significantly higher liver TG content and significantly lower plasma total cholesterol (TC) and free cholesterol content. These results were indicative of TG and cholesterol metabolism disorders in the liver of dairy cows with SCK. Additionally, the SCK group showed an increased expression of perilipin-2 (PLIN2), decreased expression of apolipoprotein B, and decreased plasma concentration of very low-density lipoproteins (VLDL) and low-density lipoproteins cholesterol (LDL-C) by downregulating ACAT2, which indicated an accumulation of TG in liver. In vitro experiments showed that BHBA induced an increase in the TG content of BHEC, decreased content TC, increased expression of PPARγ and SREBP1c, and decreased expression of PPARα, CPT1A, SREBP2, and HMGCR. Additionally, BHBA increased the expression of PLIN2 in BHEC, decreased the expression and fluorescence intensity of ACAT2, and decreased the VLDL and LDL-C contents. Furthermore, silencing ACAT2 expression increased the TG content; decreased the TC, VLDL, and LDL-C contents; decreased the expression of HMGCR and SREBP2; and increased the expression of SREBP1c; but had no effect on the expression of PLIN2. These results suggest that ACAT2 downregulation in BHEC promotes TG accumulation and inhibits cholesterol synthesis, leading to TG and cholesterol metabolic disorders. In conclusion, ACAT2 downregulation in the SCK group inhibited cholesterol synthesis, increased TG synthesis, and reduced the contents of VLDL and LDL-C, eventually leading to disordered TG and cholesterol metabolism.

β-hydroxybutyrate Alleviates Learning and Memory Impairment Through the SIRT1 Pathway in D-Galactose-Injured Mice.

Learning and memory impairment is a common clinical symptom of aging and nervous system injuries, and seriously affects quality of life. Memory impairment is associated with increased oxidative stress (OS) and inflammatory response. β-hydroxybutyrate (BHBA) is a water-soluble endogenous small-molecule ketone body that easily crosses the blood-brain barrier and has shown neuroprotection activities. In this study, we investigated the effects and mechanisms of BHBA on D-galactose (D-gal)-induced memory impairment in mice by in vitro and in vivo experiments. BHBA was administered intragastrically to D-gal-injured C57BL/6 mice for 42 days. Water maze performance, the morphology of the hippocampus with Nissl staining, the ACh content, OS, and inflammation status were examined. To further investigate the mechanism, hippocampal neuronal cells (HT22) were treated with BHBA with or without the SIRT1 inhibitor or small interfering RNAs against sirt1 (si-SIRT1) before incubation with D-gal. BHBA significantly improved water maze performance; increased the ACh content, SOD activity, and SIRT1 expression; and decreased AChE and LDH activity, ROS, MDA, IL-1β, TNF-α contents, and NLRP3 expression. Further studies with the SIRT inhibitor or siRNAs against sirt1 reversed the above effects of BHBA. Collectively, BHBA inhibited hippocampal OS and the inflammation process to alleviate learning and memory impairment through activating the SIRT1 pathway in D-gal-injured mice, suggesting that BHBA could be a potential option for drug development of learning and memory impairment induced by nervous system injuries.

Effects of β-hydroxybutyricacid on the synthesis and assembly of very low-density lipoprotein in bovine hepatocytes invitro.

β-Hydroxybutyricacid (BHBA) is an important metabolite that involved in the development of ketosis and fatty liver in dairy cows. Dairy cows with fatty liver displayed high blood concentration of BHBA and very low-density lipoprotein (VLDL) assembly. The effects of BHBA on VLDL synthesis and assembly in hepatocytes of cows were unclear. In this study, bovine hepatocytes were cultured and treated with different concentrations of BHBA. We found that BHBA treatment upregulated the mRNA and protein levels of apolipoprotein B100 (ApoB 100), apolipoprotein E (ApoE) and microsomal triglyceride transfer protein (MTTP) and showed in a firstly increased and then decreased trend. Meanwhile, the mRNA and protein levels of LDLR showed in a reverse trend. Consequently, VLDL content was significantly increased in medium-dose BHBA treatment group, while decreased in high-dose group. These results indicate that the effects of BHBA on the VLDL synthesis showed in a dose-dependent manner that low levels of BHBA increase VLDL synthesis and high levels of BHBA decrease VLDL synthesis.

Effects of β-hydroxybutyrate and different calcium and potassium concentrations on the membrane potential and motility of abomasal smooth muscle cells in cattle

The left displacement of the abomasum (LDA) is a common disease in periparturient dairy cows. Plasma β-hydroxybutyrate (BHBA) levels above the reference range are regarded as risk factors for the occurrence of LDA. Additionally, hypokalemia and hypocalcemia have been observed in LDA cows. The aim of the present study was to characterize the membrane potential and the slow waves in abomasal smooth muscle cells by a microelectrode technique and to determine possible effects of BHBA and of various calcium and potassium concentrations on the membrane potential. Subsequently, the results obtained by the microelectrode technique were combined with in vitro motility experiments of abomasal smooth muscles. Strips of the abomasal circular smooth muscles were prepared and incubated in different buffer solutions. For the microelectrode technique, healthy bulls and cows that underwent surgery for LDA were sampled. These measurements showed a frequency of the slow waves between 3.5 and 10.9 per minute (for amplitudes ≥3mV) and between 0.6 and 4.5 per minute (for amplitudes ≥5mV). The frequency of contractions (1.8 to 3.1 per minute) were in the same order as the frequency of the slow waves with amplitudes ≥5mV. Blocking potassium conductance with barium chloride induced a depolarization of the basal membrane potential (from −43±2.9 to −37±4.1mV; mean ± standard error of the mean) without affecting the frequency or the height of the slow waves. The reduction in the potassium concentration from 5.4 to 2mmol/L resulted in a nominal decrease in the activity of contractions (from 22.2 to 18.6mN/min). The subsequent addition of 1mmol of KCl/L induced a nominal increase in contraction activity (from 18.6 to 25.7mN/min). An effect of BHBA (5mmol/L) could not be demonstrated, neither on the electric nor on the motility parameters. A simulated hypocalcemia (1.2mmol/L total, 0.9mmol/L ionized Ca) did not change slow waves and motility. In conclusion, changes in membrane potential ≥5mV correlated with contractions of abomasal muscles. Hypokalemic conditions may reduce abomasal contraction activity via an effect on the membrane potential. An exclusive increase in BHBA or a slight hypocalcemia (0.9mmol/L ionized Ca) had no effects on the motility of healthy abomasal smooth muscle cells. A prolonged or severe exposition of muscles to a combination of low Ca and high BHBA concentrations might nevertheless be able to affect abomasal motility.