d-Tetramethrin causes zebrafish hepatotoxicity by inducing oxidative stress and inhibiting cell proliferation.

d-Tetramethrin is one of the main components of mosquito control products, and is widely used for the control of dengue fever and insecticide production. Due to its widespread use, d-tetramethrin is a ubiquitous environmental pollutant and poses potential risks to human health. However, the effects of d-tetramethrin on liver morphology and function are not clearly established. In this study, we used zebrafish as an animal model to analyze the acute and chronic effects of d-tetramethrin exposure on the liver. We exposed zebrafish larvae and adults to different concentrations of d-tetramethrin and examined the impact of d-tetramethrin on lipid and glycogen metabolism, cellular properties, oxidative stress, cell proliferation, and apoptosis in the liver. We also analyzed transcriptional changes in genes related to apoptosis, inflammation, and cell proliferation using qPCR. Zebrafish exposed to d-tetramethrin exhibited severe liver damage, as evidenced by the presence of vacuoles and nuclear distortion in liver cells. The liver area in zebrafish larvae of the treatment group was significantly smaller than that of the control group. Significant lipid accumulation and decreased glycogen levels were observed in the livers of both zebrafish larvae and adults exposed to d-tetramethrin. Furthermore, d-tetramethrin exposure induced apoptosis and inflammation in zebrafish embryos. Additionally, d-tetramethrin caused liver damage, metabolic dysfunction, and impaired liver function. These results suggest that d-tetramethrin induces liver toxicity in zebrafish, by inducing oxidative stress and inhibiting cell proliferation.

Differential G Protein-Coupled Estrogen Receptor-1 Regulation of Counter-Regulatory Transmitter Marker and 5'-AMP-Activated Protein Kinase Expression in Ventrolateral versus Dorsomedial Ventromedial Hypothalamic Nucleus.

INTRODUCTION: The ventromedial hypothalamic nucleus (VMN) is an estrogen receptor (ER)-rich structure that regulates glucostasis. The role of nuclear but not membrane G protein-coupled ER-1 (GPER) in that function has been studied. METHODS: Gene silencing and laser-catapult microdissection/immunoblot tools were used to examine whether GPER regulates transmitter and energy sensor function in dorsomedial (VMNdm) and/or ventrolateral (VMNvl) VMN counter-regulatory nitrergic and γ-Aminobutyric acid (GABA) neurons. RESULTS: Intra-VMN GPER siRNA administration to euglycemic animals did not affect VMNdm or -vl nitrergic neuron nitric oxide synthase (nNOS), but upregulated (VMNdm) or lacked influence on (VMNvl) GABA nerve cell glutamate decarboxylase65/67 (GAD) protein. Insulin-induced hypoglycemia (IIH) caused GPER knockdown-reversible augmentation of nNOS, 5'-AMP-activated protein kinase (AMPK), and phospho-AMPK proteins in nitrergic neurons in both divisions. IIH had dissimilar effects on VMNvl (unchanged) versus VMNdm (increased) GABAergic neuron GAD levels, yet GPER knockdown affected these profiles. GPER siRNA prevented hypoglycemic upregulation of VMNvl and -dm GABA neuron AMPK without altering pAMPK expression. CONCLUSIONS: Outcomes infer that GPER exerts differential control of VMNdm versus -vl GABA transmission during glucostasis and is required for hypoglycemic upregulated nitrergic (VMNdm and -vl) and GABA (VMNdm) signaling. Glycogen metabolism is reported to regulate VMN nNOS and GAD proteins. Data show that GPER limits VMNvl glycogen phosphorylase (GP) protein expression and glycogen buildup during euglycemia but mediates hypoglycemic augmentation of VMNvl GP protein and glycogen content; VMNdm glycogen mass is refractory to GPER control. GPER regulation of VMNvl glycogen metabolism infers that this receptor may govern local counter-regulatory transmission in part by astrocyte metabolic coupling.

Optimal dietary zinc inclusion improved growth performance, serum antioxidant capacity, immune status, and liver lipid and glucose metabolism of largemouth bass (Micropterus salmoides).

This study was conducted to ascertain the effect of dietary Zn on growth and health status of juvenile largemouth bass (Micropterus salmoides). Six experimental diets with Zn level of 50.17, 56.74, 73.34, 86.03, 123.94, and 209.20 mg/kg, respectively were compounded using complex amino acid-chelated zinc, and were fed to juvenile fish (5.50 ± 0.10 g) for 70 d. The specific growth rate (SGR) varied with dietary Zn level in a quadratic model and peaked at the 73.34 mg/kg group, while the feeding rate exhibited an opposite trend (P < 0.05). The condition factor, hepatosomatic index and mesenteric fat index all exhibited a tendency similar with SGR (P < 0.05). Dietary Zn level affected serum total proteins, urea, triglycerides, and glucose (P < 0.05). Serum Zn and copper levels linearly increased with dietary Zn level, while serum iron and manganese showed the opposite trend. Serum superoxide dismutase (SOD) and total antioxidant capacity (T-AOC) increased with dietary Zn level and reached a plateau at 86.03 mg/kg. Serum complement component 3 (C3), IgM, and lysozyme also were enhanced by 73.34 mg/kg Zn. Body protein content increased with zinc level up to 73.34 mg/kg, and then remained steadily. As dietary Zn level increased, hepatic lipid level increased and then reached a plateau at 86.03 mg/kg group, while glycogen increased linearly. Moreover, gene expression related to lipid and glycogen metabolism from liver transcriptome further explained the liver lipid and glycogen variations. To conclude, a dietary Zn requirement of 76.99 mg/kg was suggested for juvenile largemouth bass to improve growth, antioxidant capacity, and immune status.

Artichoke (Cynara scolymus L.) water extract alleviates palmitate-induced insulin resistance in HepG2 hepatocytes via the activation of IRS1/PI3K/AKT/FoxO1 and GSK-3ß signaling pathway.

BACKGROUND: Artichoke (Cynara scolymus L.) is a typical element of a traditional Mediterranean diet and has potential health advantages for insulin resistance (IR) and type 2 diabetes mellitus (T2DM). This study aims to evaluate the effect and underlying mechanism of artichoke water extract (AWE) on palmitate (PA)-induced IR in human hepatocellular carcinoma (HepG2) cells. METHODS: The effect of AWE on cell viability was determined using CCK8 assay. Cellular glucose uptake, glucose consumption, glucose production, and glycogen content were assessed after AWE treatment. The gene expression and protein levels were examined by real-time polymerase chain reaction (qRT-PCR) and western blotting. RESULTS: The results showed that AWE dose-dependently increased cell viability in IR HepG2 cells (P < 0.01). AWE treatment significantly promoted glucose uptake and consumption, decreased glucose production, and increased the cellular glycogen content in IR HepG2 cells (P < 0.01). Mechanistically, AWE elevated the phosphorylation and total protein levels of major insulin signaling molecules in IR HepG2 cells, which resulted in a decrease in the expression of phosphoenolpyruvate carboxykinase (PEPCK) and glucose-6-phosphatase (G6Pase) and the inhibition of glycogen synthase (GS) phosphorylation in IR HepG2 cells. Furthermore, the protective effect of AWE on IR HepG2 cells might be ascribed to the inhibition of the endoplasmic reticulum (ER) stress. CONCLUSION: We conclude that AWE may improve glucose metabolism by regulating IRS1/PI3K/AKT/FoxO1 and GSK-3ß signaling associated with the inhibition of ER stress in IR HepG2 cells induced by PA.

[Mechanism of Gegen Qinlian Decoction in improving glucose metabolism in vitro and in vivo by alleviating hepatic endoplasmic reticulum stress].

This study investigated the mechanism of Gegen Qinlian Decoction(GQD) in improving glucose metabolism in vitro and in vivo by alleviating endoplasmic reticulum stress(ERS). Molecular docking was used to predict the binding affinity between the main effective plasma components of GQD and ERS-related targets. Liver tissue samples were obtained from normal rats, high-fat-induced diabetic rats, rats treated with metformin, and rats treated with GQD. RNA and protein were extracted. qPCR was used to measure the mRNA expression of ERS marker glucose-regulated protein 78(GRP78), and unfolded protein response(UPR) genes inositol requiring enzyme 1(Ire1), activating transcription factor 6(Atf6), Atf4, C/EBP-homologous protein(Chop), and caspase-12. Western blot was used to detect the protein expression of GRP78, IRE1, protein kinase R-like ER kinase(PERK), ATF6, X-box binding protein 1(XBP1), ATF4, CHOP, caspase-12, caspase-9, and caspase-3. The calcium ion content in liver tissues was determined by the colorimetric assay. The ERS-HepG2 cell model was established in vitro by inducing with tunicamycin for 6 hours, and 2.5%, 5%, and 10% GQD-containing serum were administered for 9 hours. The glucose oxidase method was used to measure extracellular glucose levels, flow cytometry to detect cell apoptosis, glycogen staining to measure cellular glycogen content, and immunofluorescence to detect the expression of GRP78. The intracellular calcium ion content was measured by the colorimetric assay. Whereas Western blot was used to detect GRP78 and ERS-induced IRE1, PERK, ATF6, and eukaryotic translation initiation factor 2α(eIF2α) phosphorylation. Additionally, the phosphorylation levels of insulin receptor substrate 1(IRS1), phosphatidylinositol 3-kinase regulatory subunit p85(PI3Kp85), and protein kinase B(Akt), which were involved in the insulin signaling pathway, were also measured. In addition, the phosphorylation levels of c-Jun N-terminal kinases(JNKs), which were involved in both the ERS and insulin signaling pathways, were measured by Western blot. Molecular docking results showed that GRP78, IRE1, PERK, ATF4, and various compounds such as baicalein, berberine, daidzein, jateorhizine, liquiritin, palmatine, puerarin and wogonoside had strong binding affinities, indicating that GQD might interfere with ERS-induced UPR. In vivo results showed that GQD down-regulated the mRNA transcription of Ire1, Atf6, Atf4, Grp78, caspase-12, and Chop in diabetic rats, and down-regulated GRP78, IRE1, PERK, as well as ERS-induced apoptotic factors ATF4 and CHOP, caspase-12, caspase-9, and caspase-3, while up-regulating XBP1 to enhance adaptive UPR. In addition, GQD increased the calcium ion content in liver tissues, which facilitated correct protein folding. In vitro results showed that GQD increased glucose consumption in ERS-induced HepG2 cells without significantly affecting cell viability, increased liver glycogen synthesis, down-regulated ATF6 and p-eIF2α(Ser51), and down-regulated IRE1, PERK, and GRP78, as well as p-IRS1(Ser312) and p-JNKs(Thr183/Tyr185), while up-regulating p-PI3Kp85(Tyr607) and p-Akt(Ser473). These findings suggested that GQD alleviates excessive ERS in the liver, reduces insulin resistance, and improves hepatic glucose metabolism in vivo and in vitro.

Sex-dependent endozepinergic regulation of ventromedial hypothalamic nucleus glucose counter-regulatory neuron aromatase protein expression in the adult rat.

The hypothalamic brain cell types that produce estradiol from testosterone remain unclear. Aromatase inhibition affects ventromedial hypothalamic nucleus (VMN) glucose-stimulatory nitric oxide (NO) and glucose-inhibitory γ-aminobutyric acid (GABA) transmission during insulin (INS)-induced hypoglycemia (IIH). Pure GABA and NO nerve cell samples acquired by laser-catapult-microdissection from consecutive rostro-caudal segments of the VMN were analyzed by Western blot to investigate whether regional subpopulations of each cell type contain machinery for neuro-estradiol synthesis. Astrocyte endozepinergic signaling governs brain steroidogenesis. Pharmacological tools were used here to determine if the glio-peptide octadecaneuropeptide (ODN) controls aromatase expression in GABA and NO neurons during eu- and/or hypoglycemia. Intracerebroventricular administration of the ODN G-protein coupled-receptor antagonist cyclo(1-8)[DLeu5]OP (LV-1075) decreased (male) or enhanced (female) VMN GABAergic neuron aromatase expression, but increased or reduced this profile in nitrergic neurons in a region-specific manner in each sex. IIH suppressed aromatase levels in GABA neurons located in the middle segment of the male VMN or distributed throughout this nucleus in the female. This inhibitory response was altered by the ODN isoactive surrogate octapeptide (OP) in female, but was refractory to OP in male. NO neuron aromatase protein in hypoglycemic male (middle and caudal VMN) and female (rostral and caudal VMN) rats, but was normalized in OP- plus INS-treated rats of both sexes. Results provide novel evidence that VMN glucose-regulatory neurons may produce neuro-estradiol, and that the astrocyte endozepine transmitter ODN may impose sex-specific control of baseline and/or hypoglycemic patterns of aromatase expression in distinct subsets of nitrergic and GABAergic neurons in this neural structure.

Platelet glycogenolysis is important for energy production and function.

Although the presence of glycogen in platelets was established in the 1960s, its importance to specific functions (i.e., activation, secretion, aggregation, and clot contraction) remains unclear. Patients with glycogen storage disease often present with increased bleeding and glycogen phosphorylase (GP) inhibitors, when used as treatments for diabetes, induce bleeding in preclinical studies suggesting some role for this form of glucose in hemostasis. In the present work, we examined how glycogen mobilization affects platelet function using GP inhibitors (CP316819 and CP91149) and a battery of ex vivo assays. Blocking GP activity increased glycogen levels in resting and thrombin-activated platelets and inhibited platelet secretion and clot contraction, with minimal effects on aggregation. Seahorse energy flux analysis and metabolite supplementation experiments suggested that glycogen is an important metabolic fuel whose role is affected by platelet activation and the availability of external glucose and other metabolic fuels. Our data shed light on the bleeding diathesis in glycogen storage disease patients and offer insights into the potential effects of hyperglycemia on platelets.

What did we know? Activated platelets transition from a low-energy-requiring, resting state to a high-energy-demanding state.Platelet glycogen is degraded upon activation.Glycogen storage disorders and glycogen phosphorylase inhibitors are associated with bleeding.What did we discover? Glycogen turnover occurs in resting platelets and its degradation is important for platelet functions.Glycogen phosphorylase inhibitors block secretion and clot contraction of which the latter can be reversed with alternative metabolic fuels.Glucose derived from glycogen may be routed through TCA/OxPhos versus aerobic glycolysis.What is the impact? Glycogen breakdown contributes to the high energy requirements of platelet function.Our work offers insights into potential energy sources in activated platelets.

Interleukin 6 acutely increases gluconeogenesis and decreases the suppressive effect of insulin on cAMP-stimulated glycogenolysis in rat liver.

Interleukin 6 (IL6) is an multifunctional cytokine that modulates several biological responses, including glucose metabolism. However, its acute effects on hepatic glucose release are still uncertain. The main purpose of this study was to investigate the effects of IL6 on gluconeogenesis from several glucose precursors (alanine, pyruvate and glutamine) and on the suppressive action of insulin on cAMP-stimulated glycogen catabolism in rat liver. IL6 effect on insulin peripheral sensitivity was also evaluated. IL6 was injected intravenously into rats and, 1 h later, gluconeogenesis and glycogenolysis were assessed in liver perfusion and peripheral insulin sensitivity by insulin tolerance test (ITT). IL6 intravenous injection increased hepatic glucose production from alanine, without changing pyruvate, lactate and urea production. IL6 injection also increased hepatic glucose production from pyruvate and glutamine. In addition, IL6 decreased the suppressive effect of insulin on cAMP-stimulated glucose and lactate production and glycogenolysis, without affecting pyruvate production. Furthermore, IL6 reduced the plasma glucose disappearance constant (kITT), an indicator of insulin resistance. In conclusion, IL6 acutely increased hepatic glucose release (gluconeogenesis and glycogenolysis) by a mechanism that likely involved the induction of insulin resistance in the liver, as evidenced by the reduced suppressive effect of insulin on cAMP-stimulated glycogen catabolism. In consistency, IL6 acutely induced peripheral insulin resistance.

Signal transduction of the insulin secretion induced by the chalcone analogue, (E)-3-(phenyl)-1-(3,4,5-trimethoxyphenyl)prop-2-en-1-one, and its role in glucose and lipid metabolism.

A chalcone analogue, (E)-3-(phenyl)-1-(3,4,5-trimethoxyphenyl)prop-2-en-1-one (DMU 101), was synthesized using classic base catalysis and Claisen-Schmidt condensation, and then screened for its antidiabetic properties. The compound's effects on glucose and lipid metabolism were assayed in rats that were treated acutely and for a short time to elucidate its mechanism of action, evaluating glucose tolerance and lactate dehydrogenase activity in response to chalcone analogue administration. The chalcone's in vitro and ex vivo effects on glycogen, glucose, lipid and lipolysis were also investigated, as well as the mechanism by which it induces 45Ca2+ influx-mediated insulin secretion. The analogue (10 mg/kg) diminished glycemia, without inducing acute cell damage, increased glycogen content in the skeletal muscle and reduced serum triacylglycerol and total cholesterol, but did not alter high-density lipoprotein or low-density lipoprotein. Chalcone (10 µM) stimulated glucose uptake in the soleus muscle and did not modulate in vitro or ex vivo lipolysis. This analogue also increased insulin secretion by triggering calcium influx and blocking ATP-sensitive K+ channels and voltage-dependent calcium channels. However, it also modulated stored calcium via sarco/endoplasmic reticulum calcium ATPase (SERCA) and ryanodine receptor (RYR) activity. These findings indicate that this chalcone may induce cellular repolarization via a mechanism mediated by calcium-dependent potassium channels.

Root-Extracted lignanamides from Limonium gmelinii (Willd.) Kuntze with a potential PTP1B inhibitory activity by regulating PI3K/AKT signaling pathway.

The phytochemical study of Limonium gmelinii roots resulted in the isolation of five lignanamides (1-5). Among them, limoniumins J, K, and M (1, 2, and 4) are undescribed compounds, limoniumin L (3) is a new naturally occurring lignanamide, and limoniumin B (5) is a known compound which showed PTP1B inhibition activity with an IC50 value of 5.05 ± 2.44 µM in our previous work. Spectroscopic data analysis, including 1D and 2D NMR and HRESIMS experiments, established the chemical structures of limoniumins J - M (1-4). Compounds 1-4 showed PTP1B inhibition activity, among which compound 3 showed the most potent PTP1B inhibition with an IC50 value of 2.07 ± 0.05 µM. Compounds 3 and 5 could significantly increase cellular glucose consumption and glucose uptake in L6 muscle cells and could synergize with insulin to promote glucose consumption and glucose uptake in a concentration-dependent manner. The treatment of compound 3 also promoted glycogen synthesis in skeletal muscle cells. Western blot analysis demonstrated that the good hypoglycemic effect of compounds 3 and 5 was achieved by activating PI3K/AKT signaling pathway to promote glucose consumption, glucose uptake, and glycogen synthesis. Furthermore, studies on molecular docking revealed the potent interactions between these bioactive substances and the PTP1B protein.

Euscaphic acid relieves fatigue by enhancing anti-oxidative and anti-inflammatory effects.

BACKGROUND: Oxidative stress and inflammation are involved in chronic fatigue. Euscaphic acid (EA) is an active compound of Eriobotrya japonica (Loquat) and has anti-oxidative effect. METHODS: The goal of present study is to prove whether EA could relieve fatigue through enhancing anti-oxidant and anti-inflammatory effects in in vitro/in vivo models. RESULTS: EA notably improved activity of superoxide dismutase (SOD) and catalase (CAT), while EA reduced levels of malondiadehyde (MDA) and inflammatory cytokines without cytotoxicity in H2O2-stimulated in myoblast cell line, C2C12 cells. EA significantly reduced levels of fatigue-causing factors such as lactate dehydrogenase (LDH) and creatin kinase (CK), while EA significantly incresed levels of anti-fatigue-related factor, glycogen compared to the H2O2-stimulated C2C12 cells. In treadmill stress test (TST), EA significantly enhanced activities of SOD and CAT as well as exhaustive time and decreased levels of MDA and inflammatory cytokines. After TST, levels of free fatty acid, citrate synthase, and muscle glycogen were notably enhanced by oral administration of EA, but EA decreased levels of lactate, LDH, cortisol, aspartate aminotransferase, alanine transaminase, CK, glucose, and blood urea nitrogen compared to the control group. Furthermore, in forced swimming test, EA significantly increased levels of anti-fatigue-related factors and decreased excessive accumulations of fatigue-causing factors. CONCLUSIONS: Therefore, the results indicate that potent anti-fatigue effect of EA can be achieved via the improvement of anti-oxidative and anti-inflammatory properties, and this study will provide scientific data for EA to be developed as a novel and efficient component in anti-fatigue health functional food.

Mechanism of Gegen Qinlian Decoction in improving glucose metabolism in vitro and in vivo by alleviating hepatic endoplasmic reticulum stress / 中国中药杂志

This study investigated the mechanism of Gegen Qinlian Decoction(GQD) in improving glucose metabolism in vitro and in vivo by alleviating endoplasmic reticulum stress(ERS). Molecular docking was used to predict the binding affinity between the main effective plasma components of GQD and ERS-related targets. Liver tissue samples were obtained from normal rats, high-fat-induced diabetic rats, rats treated with metformin, and rats treated with GQD. RNA and protein were extracted. qPCR was used to measure the mRNA expression of ERS marker glucose-regulated protein 78(GRP78), and unfolded protein response(UPR) genes inositol requiring enzyme 1(Ire1), activating transcription factor 6(Atf6), Atf4, C/EBP-homologous protein(Chop), and caspase-12. Western blot was used to detect the protein expression of GRP78, IRE1, protein kinase R-like ER kinase(PERK), ATF6, X-box binding protein 1(XBP1), ATF4, CHOP, caspase-12, caspase-9, and caspase-3. The calcium ion content in liver tissues was determined by the colorimetric assay. The ERS-HepG2 cell model was established in vitro by inducing with tunicamycin for 6 hours, and 2.5%, 5%, and 10% GQD-containing serum were administered for 9 hours. The glucose oxidase method was used to measure extracellular glucose levels, flow cytometry to detect cell apoptosis, glycogen staining to measure cellular glycogen content, and immunofluorescence to detect the expression of GRP78. The intracellular calcium ion content was measured by the colorimetric assay. Whereas Western blot was used to detect GRP78 and ERS-induced IRE1, PERK, ATF6, and eukaryotic translation initiation factor 2α(eIF2α) phosphorylation. Additionally, the phosphorylation levels of insulin receptor substrate 1(IRS1), phosphatidylinositol 3-kinase regulatory subunit p85(PI3Kp85), and protein kinase B(Akt), which were involved in the insulin signaling pathway, were also measured. In addition, the phosphorylation levels of c-Jun N-terminal kinases(JNKs), which were involved in both the ERS and insulin signaling pathways, were measured by Western blot. Molecular docking results showed that GRP78, IRE1, PERK, ATF4, and various compounds such as baicalein, berberine, daidzein, jateorhizine, liquiritin, palmatine, puerarin and wogonoside had strong binding affinities, indicating that GQD might interfere with ERS-induced UPR. In vivo results showed that GQD down-regulated the mRNA transcription of Ire1, Atf6, Atf4, Grp78, caspase-12, and Chop in diabetic rats, and down-regulated GRP78, IRE1, PERK, as well as ERS-induced apoptotic factors ATF4 and CHOP, caspase-12, caspase-9, and caspase-3, while up-regulating XBP1 to enhance adaptive UPR. In addition, GQD increased the calcium ion content in liver tissues, which facilitated correct protein folding. In vitro results showed that GQD increased glucose consumption in ERS-induced HepG2 cells without significantly affecting cell viability, increased liver glycogen synthesis, down-regulated ATF6 and p-eIF2α(Ser51), and down-regulated IRE1, PERK, and GRP78, as well as p-IRS1(Ser312) and p-JNKs(Thr183/Tyr185), while up-regulating p-PI3Kp85(Tyr607) and p-Akt(Ser473). These findings suggested that GQD alleviates excessive ERS in the liver, reduces insulin resistance, and improves hepatic glucose metabolism in vivo and in vitro.

Imperatorin Positively Regulates Melanogenesis through Signaling Pathways Involving PKA/CREB, ERK, AKT, and GSK3ß/ß-Catenin.

The present study investigated the melanogenic effects of imperatorin and isoimperatorin and the underlying mechanisms of imperatorin using a mouse melanoma B16F10 model. Interestingly, treatment with 25 µM of either imperatorin or isoimperatorin, despite their structural differences, did not produce differences in melanin content and intracellular tyrosinase activity. Imperatorin also activated the expression of melanogenic enzymes, such as tyrosinase (TYR) and tyrosinase-related proteins TYRP-1 and TYRP-2. Mechanistically, imperatorin increases melanin synthesis through the cyclic adenosine monophosphate (cAMP)-dependent protein kinase (PKA)/cAMP-responsive element-binding protein (CREB)-dependent upregulation of microphthalmia-associated transcription factor (MITF), which is a key transcription factor in melanogenesis. Furthermore, imperatorin exerted melanogenic effects by downregulating extracellular signal-regulated kinase (ERK) and upregulating phosphatidylinositol 3 kinase (PI3K)/protein kinase B (AKT)/glycogen synthesis kinase-3ß (GSK-3ß). Moreover, imperatorin increased the content of ß-catenin in the cell cytoplasm and nucleus by reducing the content of phosphorylated ß-catenin (p-ß-catenin). Finally, we tested the potential of imperatorin in topical application through primary human skin irritation tests. These tests were performed on the normal skin (upper back) of 31 volunteers to determine whether 25 or 50 µM of imperatorin had irritation or sensitization potential. During these tests, imperatorin did not induce any adverse reactions. Taken together, these findings suggest that the regulation of melanogenesis by imperatorin can be mediated by signaling pathways involving PKA/CREB, ERK, AKT, and GSK3ß/ß-catenin and that imperatorin could prevent the pathogenesis of pigmentation diseases when used as a topical agent.

CO2 narcosis induces a metabolic shift mediated via juvenile hormone in Bombus impatiens gynes.

Carbon dioxide (CO2) has pleiotropic effects on insect physiology and behavior. Although diverse, many impacts are related to changes in metabolism and reallocation of macronutrients. Here we examined the metabolic shift induced by CO2 and its regulation using Bombus impatiens. CO2 applied to bumble bee gynes induces bypass of diapause and transition into reproduction. We analyzed ovary activation and macronutrient amounts in four tissues/body parts (fat body, thorax, ovaries, and crop) at three timepoints following CO2 administration. To tease apart the effects of CO2 on reproduction and metabolism, we monitored the metabolic changes in gynes following ovary removal and CO2 narcosis. We also explored the role of juvenile hormone in mediating CO2 impact by feeding queens with a JH antagonist (Precocene). Gynes ovary activation was increased following CO2 treatment. Additionally, CO2-treated gynes showed lower lipid amount in the fat body and higher glycogen and protein amount in the ovary ten days after the treatment. CO2 treatment following ovary removal also resulted in decreased fat body lipids, suggesting that CO2 operates by inducing a metabolic shift independent of reproduction. Lastly, gynes fed with precocence did not show a metabolic shift following CO2, suggesting CO2 impact is mediated via juvenile hormone. Overall, these data suggest that CO2 induces transfer of macronutrients and utilization of stored reserved by accelerating metabolism. The proposed mechanism of CO2 may explain many of the pleiotropic effects of CO2 across species and can aid in understanding how this common anastatic influences insect physiology.

Sacubitril/valsartan combination enhanced cardiac glycophagy and prevented the progression of murine diabetic cardiomyopathy.

BACKGROUND: Diabetic cardiomyopathy (DCM) is linked to disturbance in cardiac glucose handling and increased cardiac glycogen storage. This study tested the potential role of sacubitril/valsartan on the progression of DCM in high fat diet (HFD)/streptozotocin (STZ)-induced type 2 diabetic rats compared to valsartan alone, including their effects on the cardiac glycophagy process. MATERIALS AND METHODS: Rats were fed on HFD for 6 weeks followed by single low-dose STZ (35 mg/kg). After confirming hyperglycemia, diabetic rats were continued on HFD and divided into three subgroups: Untreated-diabetic, Valsartan-treated diabetic and Sacubitril/valsartan-treated diabetic groups; in addition to a control group. Changes in ECG, blood glucose, serum insulin, lipid profile, and Homeostasis model of assessment of insulin resistance (HOMA-IR) were assessed and the degree of cardiac fibrosis was examined. Cardiac glycogen content and glycophagy process were evaluated. RESULTS: Sacubitril/valsartan administration to diabetic rats resulted in improvement of metabolic changes more than valsartan alone. Also, sacubitril/valsartan effectively prevented diabetes-associated cardiac hypertrophy, QTc prolongation, and fibrosis. Finally, cardiac glycogen concentrations in diabetic rats were decreased by sacubitril/valsartan combination, coupled with significant induction of glycophagy process in the diabetic rats' heart. CONCLUSION: Sacubitril/valsartan therapy provides a more favorable metabolic and cardioprotective response compared to valsartan alone in a rat model of DCM. These findings may be due to a direct cardioprotective impact of sacubitril/valsartan and secondary beneficial effects of improved hyperglycemia and dyslipidemia. In addition, these beneficial cardiac effects could be attributed to the induction of the glycophagy process and alleviating cardiac glycogen overload.

Does a Diet Rich in the Bacterium Rhodopirellula rubra Improve Daphnia magna Performance?

BACKGROUND: In the wild various organisms contribute to daphnids diet. This study, intendeds to evaluate the potential of the concentration of Rhodopirellula rubra as a single or supplementary food source for Daphnia magna. METHODS: Feeding assays were performed according to standard guidelines for chronic assays (21 days), and life-history parameters and several biomarkers (protein content, oxidative stress, energetic reserves and pigments) were measured. Five food regimens were conducted with 20 individual replicates (A - R. subcapitata; 0.2 - suspension of R. rubra at 0.2 arbitrary units (AU); 0.4 - suspension of R. rubra at 0.4 AU; 0.2+A - suspension of R. rubra at 0.2+alga; 0.2+A-suspension of R. rubra at 0.4 AU + alga). Additionally, the effects of three diets (A, 0.2, and 0.2+A) on the longevity of D. magna were assessed. RESULTS: The five diets showed a different C, N, and carotenoids composition, with an increase in the mixed diets. The results confirmed that the mixed diets improved D. magna life-history parameters. A decrease in glycogen, and the increase of haemoglobin, protein, and gluthione-S-transferase (GST) were observed. Furthermore, D. magna fed with bacterial single diets, presented worsen life history parameters and a decrease in the protein content. An induction of oxidative stress response (increased catalase and GST), and a significant decrease in lipid peroxidation and an accumulation of glycogen and carotenoids were observed. Overall, an increase in the amount of R. rubra provided to D. magna, from 0.2 AU to 0.4 AU, negatively impacted daphnid performance. No significant effects on Daphnia longevity (a 110-day assay) were observed among the three diets tested. However, a significant survival percentage and fertility (cumulative offspring is more than twice) was observed when D. magna was fed with the mixed diet. CONCLUSIONS: Results demonstrated that different diets provided a nutritional diversified food to the daphnids that induced differences in D. magna performance. The mixed diets proved to be beneficial (with increase in offspring) on D. magna performance, independently of the bacterial concentration tested. When in single diet, bacterial concentration is not nutritionally sufficient to raise D. magna even when in increased concentration.

ß-Hydroxyphosphocarnitine modifies fibrosis, steatosis and improves liver function in non-alcoholic steatohepatitis induced in rats.

BACKGROUND: Non-alcoholic steatohepatitis (NASH) is a chronic disease characterized by inflammation, steatosis, and liver fibrosis. The liver is particularly affected by alterations in lipid metabolism. Our aim was to evaluate the effect of ß-hydroxyphosphocarnitine (ß-HPC) on NASH induced in rats. METHODS: NASH was produced via the ad libitum daily chronic administration of a fructose solution (400 kcal) for 9 weeks, an oral dose of fat solution (16 kcal) for 7 weeks and a subcutaneous injection of CCl4 (30%) two times a week for 2 weeks to Wistar rats. To evaluate the effect of ß-HPC, a dose of 100 mg/kg was administered perorally for 4 weeks and its biochemical and hepatic effects on rats with NASH were analyzed. Serum levels of glucose, triglycerides, cholesterol, and liver enzymes were quantified. Histological changes were evaluated on slices stained with H&E, trichromic and PAS. Glycogen content was measured in liver samples. α-SMA and SREBP-1 immunopositive cells were identified in liver tissue. RESULTS: NASH was characterized by elevated triglycerides, elevated liver damage enzymes, and the presence of necrosis, inflammation, steatosis, and fibrosis. Significant amounts of glycogen were found, along with α-SMA positive cells in fibrosis areas. The over-expression of SREBP-1 in cytoplasm and nuclei was evident. Animals with NASH treated with ß-HPC showed a significant reduction in inflammation, necrosis, and glycogen content in the liver. A reduction in α-SMA and SREBP-1 immunopositive cells correlated with a significant reduction in the degree of fibrosis and steatosis found in liver tissue. ß-HPC reduced the levels of ALP and GGT, and significantly reduced triglyceride levels. Animals treated with ß-HPC did not show any alterations in liver enzyme function. CONCLUSIONS: Our research shows that ß-HPC can improve liver function and morphology in the case of NASH induced in rats, suggesting ß-HPC could be potentially used in the treatment of NASH.

[Activation of JNK induces apoptosis to autophagy conversion and enhances the survival of oxygen-glycogen deprived rat neurons].

Objective To investigate the efficacy and mechanism of c-Jun N-terminal kinase (JNK) in boosting survival of oxygen glucose deprivation (OGD) rat neurons. Methods The cortex neurons from fetal rats were primarily cultured to prepare a model of OGD neurons in vitro, and the characteristic endpoints were filtered to intervene with JNK inducer anisomycin (AN), respectively. The cells were randomly divided into control group, solvent control group (a same volume of solvent DMSO was added into the culture medium of the OGD neuron), AN group (OGD neurons were treated with JNK inducer AN for 5 hours at the end of OGD). After that, Western blotting and immunofluorescence cytometry were respectively performed to detect the protein expressions in OGD neurons, including beclin 1, microtubule-associated protein 1 light chain 3 (LC3), B cell lymphoma 2 (Bcl2), caspase-3, P62, ubiquitin, cathepsin B and lysosomal associated membrane protein 1 (LAMP1). The cell activity was evaluated by CCK-8 assay, and the axon length was measured by IPP software. Results Activation of JNK significantly promoted the expressions of beclin 1, LC3, and Bcl2, and markedly reduced the content of beclin 1-Bcl2 complex and attenuated the expressions of P62 and ubiquitin. Meanwhile, the expressions of cathepsin B and LAMP1 were not obviously altered. In this way, the survival rate of OGD neurons was improved. Conclusion Activation of JNK exerts a neuroprotective effect by facilitating dissociation of beclin 1-Bcl2 and inducing a switch from apoptosis to autophagy in OGD neurons.

ROS generation attenuates the anti-cancer effect of CPX on cervical cancer cells by inducing autophagy and inhibiting glycophagy.

Cervical cancer is one of the most common gynecological malignancies with poor prognosis due to constant chemoresistance and repeated relapse. Ciclopirox olamine (CPX), a synthetic antifungal agent, has recently been identified to be a promising anti-cancer candidate. However, the detailed mechanisms related to its anti-cancer effects remain unclear and need to be further elucidated. In this study, we found that CPX could induce proliferation inhibition in cervical cancer cells by targeting PARK7. Further results demonstrated that CPX could induce cytoprotective autophagy by downregulating the expression of PARK7 to activate PRKAA1 or by PARK7-independent accumulation of ROS to inhibit mTOR signaling. Meanwhile, CPX treatment increased the glycogen clustering and glycophagy in cervical cancer cells. The presence of N-acetyl-l-cysteine (NAC), a ROS scavenger, led to further clustering of glycogen in cells by reducing autophagy and enhancing glycophagy, which promoted CPX-induced inhibition of cervical cancer cell proliferation. Together, our study provides new insights into the molecular mechanisms of CPX in the anti-cancer therapy and opens new avenues for the glycophagy in cancer therapeutics.

ß-Carotene inhibits NF-κB and restrains diethylnitrosamine-induced hepatic inflammation in Wistar rats.

ß-Carotene exhibits antioxidant and hepatoprotective activities via a multitude of biochemical mechanisms. However, the action mechanism involved in antioxidant and anti-inflammatory effects of this carotene in chronic liver diseases is not fully understood. In the present investigation, we have attempted to outline a plausible mechanism of ß-carotene action against liver fibrosis in albino Wistar rats. To induce hepatic fibrosis, diethylnitrosamine (DEN) was administered in experimental rats for two weeks. DEN treated rats were divided into four groups, wherein each group comprised of five rats. ß-Carotene supplement attenuated DEN-induced elevation in LFT markers (P < 0.05); averted depletion of glycogen (24%, P < 0.05) and, increased nitrite (P < 0.05), hydroxyproline (~67%, P < 0.05) and collagen levels (~65%, P < 0.05). Confocal microscopy of tissue sections stained with picrosirius red revealed accrued collagen in DEN-administered group, which was found to be reduced by ß-carotene supplementation. Furthermore, ß-carotene decreased the expression of iNOS/NOS-2 and NF-κB, as revealed by immunohistochemistry and Western immunoblotting. Collectively, these results demonstrate that ß-carotene mitigates experimental liver fibrosis via inhibition of iNOS and NF-κB in-vivo. Thus, ß-carotene may be suggested as a possible nutraceutical to curb experimental liver fibrosis.