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Background/Aims: Previously, we confirmed that liver-synthesized 5-HT rather than non-liver 5-HT, acting on the 5-HT₂ receptor (5-HT₂R), modulates lipid-induced excessive lipid synthesis (ELS). Here, we further revealed the effects of the hepatocellular 5-HT system in diabetes-related disorders. Methods: Studies were conducted in male ICR mice, human HepG2 cells, and primary mouse hepatocytes (PMHs) under gene or chemical inhibition of the 5-HT system, key lipid metabolism, and inflammation-related factors. Protein and messenger RNA expression and levels of the factors were determined via western blotting, reverse transcription PCR, and quantitative assay kits, respectively. Hepatic steatosis with inflammation and fibrosis, intracellular lipid droplet accumulation (LDA), and reactive oxygen species (ROS) location were determined via hematoxylin and eosin, Masson’s trichrome, Oil red O, and fluorescent-specific staining, respectively. Results: Palmitic acid induced the activation of the 5-HT system: the activation of 5-HT₂R, primarily 5-HT_2AR, in addition to upregulating monoamine oxidase A (MAO-A) expression and 5-HT synthesis, by activating the G protein/ phospholipase C pathway modulated PKCε activation, resulting in ELS with LDA; the activation of NF-κB, which mediates the generation of pro-inflammatory cytokines, was primarily due to ROS generation in the mitochondria induced by MAO-A–catalyzed 5-HT degradation, and secondarily due to the activation of PKCε. These effects of the 5-HT system were also detected in palmitic acid- or high glucose-treated PMHs and regulated multiple inflammatory signaling pathways. In diabetic mice, co-treatment with antagonists of both 5-HT synthesis and 5-HT₂R significantly abolished hepatic steatosis, inflammation, and fibrosis as well as hyperglycemia and dyslipidemia. Conclusion: Activation of the hepatocellular 5-HT system plays a crucial role in inducing diabetes-related hepatic dysfunction and is a potential therapeutic target.

Hepatic triglycerides production and adipose lipolysis are pivotal for long-term stress (LTS) or hyperglucocorticoidemia-induced insulin resistance. 5-hydroxytryptamine (5-HT) has been demonstrated to induce hepatic lipid metabolic abnormality by activating mammalian target of rapamycin (mTOR). In present study, we explored whether 5-HT is involved in LTS effects in liver using restraint stress-exposed rats and cultured primary rat hepatocytes and HepG2 cells. LTS with hyperglucocorticoidemia induced hepatic 5-HT synthetic increase with tryptophan hydroxylase 1 (Tph1) up-regulation, and 5-HT2 receptor (5-HT2R, including 5-HT2A, 2B receptor) up-regulation in liver and visceral adipose, as well as hepatic mTOR activation with triglycerides and VLDL overproduction with steatosis, and visceral adipose lipolytic increase with high blood free fatty acids (FFAs) level. 5-HT exposure exhibited LTS-like effects in both tissues, and both LTS and 5-HT effects could be abolished significantly by blocking 5-HT2R. In HepG2 cells dexamethasone or palmitate-induced mTOR activation with triglycerides and VLDL overproduction were accompanied by up-regulations of 5-HT synthesis and 5-HT2R, which were significantly abolished by gene silencing Tph1 or 5-HT2R and were almost fully abolished by co-silencing of both, especially on VLDL overproduction. Chemical inhibition of Tph1 or/and 5-HT2R in both hepatocytes exhibited similar abolishment with genetic inhibition on dexamethason-induced effects. 5-HT-stimulated effects in both hepatocytes were fully abolished by blocking 5-HT2R, while 5-HT itself also up-regulated 5-HT2R. In conclusion, up-regulated hepatic 5-HT synthesis and 5-HT2R induced by both glucocorticoid and FFAs are crucial for LTS-induced hepatic steatosis with VLDL overproduction, while 5-HT by acting on 5-HT2R mediates mTOR activation in liver.

Aim/IntroductionBoth glucocorticoids and 5‐hydroxytryptamine (5‐HT) have been shown to induce insulin resistance (IR) in hepatocytes and adipocytes. Here, we explore whether there is a correlation between them.Materials and MethodsExcept for the control group, male rats were exposed to dexamethasone treated with or without para‐chlorophenylalanine (pCPA), or carbidopa for 20 days. Except for the control group, buffalo rat liver 3A (BRL‐3A) cells were exposed to dexamethasone for 24 h, treated with or without pCPA, carbidopa, or clorgiline for 48 h, or exposed to 5‐HT treated with or without fluoxetine for 48 h. Whole‐body IR was determined by both glucose tolerance test and measurement of fasting blood glucose and insulin, whereas hepatocytes or adipocytes IR was determined by examining either hepatic gluconeogenesis, steatosis and glucose transporter 2 expression or lipolysis.ResultsDexamethasone‐induced whole‐body IR, liver and intraabdominal adipose IR were accompanied by upregulated expressions of tryptophan hydroxylase‐1 and aromatic amino acid decarboxylase with increased 5‐HT level in both tissues, which were attenuated significantly by pCPA, inhibiting tryptophan hydroxylase‐1, or carbidopa, inhibiting aromatic amino acid decarboxylase. [Correction added on 22 September 2015, after first online publication: ‘inhibiting aromatic amino acid decarboxylase’ was duplicated and has been replaced by ‘tryptophan hydroxylase‐1’.] In the BRL‐3A cells, dexamethasone‐induced IR was also accompanied by upregulated 5‐HT synthesis in dose‐ and time‐dependent manners, and was attenuated by pCPA or carbidopa, but exacerbated by clorgiline, inhibiting monoamine oxidase‐A to further increase 5‐HT level. Dexamethasone also enhanced 5‐HT 2A and 2B receptor expressions in both tissues and BRL‐3A cells. Additionally, blocking 5‐HT transporter with fluoxetine significantly suppressed 5‐HT‐induced IR in BRL‐3A cells.ConclusionEnhancement of 5‐HT synthesis in liver and intra‐abdominal adipose is an important reason for glucocorticoids‐induced IR.

Objectives. In the present study we try to elucidate the mechanism of Xiangqing anodyne spray (XQAS) effects on acute soft-tissue injury (STI). Methods. Acute STI model was established by hammer blow in the rat hind leg muscle. Within 8 hours, instantly after modeling and per 2-hour interval repeated topical applications with or without XQAS, CP or IH ethanol extracts spray (CPS and IHS) were performed, respectively; muscle swelling rate and inflammation-related biochemical parameters, muscle histological observation, and mRNA and protein expression were then examined. Results. XQAS dose-dependently suppressed STI-caused muscle swelling, proinflammatory mediator productions, and oxidative stress as well as severe pathological changes in the injured muscle tissue. Moreover, CPS mainly by blocking p38 activation while IHS majorly by blocking AKT activation led to cytoplastic IκBα degradation with NF-κB p65 translocated into the nucleus. There are synergistic effects between CP and IH components in the XQAS on preventing from acute STI with suppressing IκBα degradation, NF-κB p65 translocation, and subsequent inflammation and oxidative stress-related abnormality. Conclusion. Marked effects of XQAS on treating acute STI are ascribed to strong anti-inflammatory and antioxidative actions with a reasonable combination of CP active components, blocking p38-NF-κB pathway activated, and IH active components, blocking AKT-NF-κB pathway activated.