Hepatic Protein Research Articles

Regulation of carbohydrate metabolism and insulin secretion in diabetic rats following treatment with Hypericum perforatum L. hairy root aqueous methanol extract

Hairy root (HR) cultures of Hypericum perforatum L. have shown promise in controlling hyperglycemia, regulating blood lipid and enzyme profiles, and improving metabolic function in vivo. These benefits are mainly attributed to the phenolic acids, flavonoids, and xanthones present in HR extracts. However, the specific mechanism underlying these effects remains unclear. This study was performed to elucidate the biochemical and molecular mechanisms driving HR the antihyperglycemic and antidiabetic effects of HR extracts. HR extract (200 mg/kg body weight) was administered daily for 14 days to healthy rats and rats with streptozotocin-induced diabetes, with glibenclamide serving as a positive control. The phenolic composition of the HR extracts was confirmed through high-performance liquid chromatography/diode array detection/electrospray ionization mass spectrometry (HPLC/DAD/ESI-MSn ) analysis. The results showed that HR extract treatment increased the plasma insulin level and pancreatic poly (adenosine diphosphate-ribose) polymerase (PARP) activity in diabetic rats, thus normalizing blood glucose levels. Additionally, it reduced the activity of gluconeogenic enzymes, increased the activity of glycolytic enzymes, and normalized the glycogen content in the liver. HR extract-treated rats also exhibited increased hepatic adenosine monophosphate-activated protein kinase (AMPK) mRNA expression and a decreased protein kinase Cε (PKCε) concentration. In conclusion, HR extract demonstrated insulinotropic effects and effectively regulated hepatic carbohydrate metabolism in diabetic rats by modulating AMPK expression and the PKCε concentration. These findings suggest the potential use of HR extract as an herbal medicine for diabetes treatment and a source of antidiabetic drug development.

Alteration of Hepatic Cytochrome P450 Expression and Arachidonic Acid Metabolism by Arsenic Trioxide (ATO) in C57BL/6 Mice.

The success of arsenic trioxide (ATO) in acute promyelocytic leukemia has driven a plethora studies to investigate its efficacy in other malignancies. However, the inherent toxicity of ATO limits the expansion of its clinical applications. Such toxicity may be linked to ATO-induced metabolic derangements of endogenous substrates. Therefore, the primary objective of this study was to investigate the effectof ATO on the hepatic formation of arachidonic acid (AA) metabolites, hydroxyeicosatetraenoic acids (HETEs), as well as their most notable producing machinery, cytochrome P450 (CYP) enzymes. For this purpose, C57BL/6 mice were intraperitoneally injected with 8 mg/kg ATO for 6 and 24 h. Total RNA was extracted from harvested liver tissues for qPCR analysis of target genes. Hepatic microsomal proteins underwent incubation with AA, followed by identification/quantification of the produced HETEs. ATO downregulated Cyp2e1, while induced Cyp2j9 and most of Cyp4a and Cyp4f, and this has resulted in a significant increase in 17(S)-HETE and 18(R)-HETE, while significantly decreased 18(S)-HETE. Additionally, ATO induced Cyp4a10, Cyp4a14, Cyp4f13, Cyp4f16, and Cyp4f18, resulting in a significant elevation in 20-HETE formation. In conclusion, ATO altered hepatic AA metabolites formation through modulating the underlying network of CYP enzymes. Modifying the homeostatic production of bioactive AA metabolites, such as HETEs, may entail toxic events that can, at least partly, explain ATO-induced hepatotoxicity. Such modification can also compromise the overall body tolerability to ATO treatment in cancer patients.

Penthorum chinense Pursh extract ameliorates alcohol-related fatty liver disease in mice via the SIRT1/AMPK signaling axis

Penthorum chinense Pursh (P. chinense), a functional food, has been applied to protect the liver against alcohol-related fatty liver disease (ALD) for a long history in China. This study was designed to evaluate the ameliorative activity of the polyphenolic fraction in P. chinense (PGF) depending on the relief of ALD. The ALD mouse model was established by exposing the mice to a Lieber-DeCarli alcohol liquid diet. We found that PGF administration significantly ameliorated alcohol-induced liver injury, steatosis, oxidative stress, and inflammation in mice. Furthermore, alcohol-increased levels of the critical hepatic lipid synthesis proteins sterol regulatory element binding transcription factor (SREBP-1) and diacylglycerol o-acyltransferase 2 (DGAT2) were attenuated by PGF. Similarly, PGF inhibited the expression of the lipid transport protein very low-density lipoprotein receptor (VLDLR). Interestingly, PGF restored alcohol-inhibited expression of carnitine palmitoyltransferase 1 (CPT1) and peroxisome proliferator-activated receptor alpha (PPARα), essential fatty acid β-oxidation proteins. Mechanistic studies revealed that PGF protects against alcohol-induced hepatocyte injury and lipid deposition via the SIRT1/AMPK signaling pathway. In sum, this research clearly demonstrated the protective effects of PGF against ALD, which was mediated by activating SIRT1/AMPK pathways in hepatocytes. We provide a new theoretical basis for using P. chinense as a functional food in ALD.

Effects of the Addition of Trichoderma reesei Cellulase to Broiler Chicken Diets for a 21-Day Period.

The cellulose present in the cell wall of vegetables prevents the greater release of nutrients to the animal. Therefore, the use of the cellulase enzyme is a viable strategy as it is capable of breaking cellulose bonds, releasing nutrients such as glucose, increasing dietary energy, and thus improving the productive performance of birds. Trichoderma reesei is efficient in the production of cellulase, which is produced via submerged fermentation followed by purification, formulation, and drying. Therefore, an experiment was carried out using 240 male broilers of the Cobb-500® lineage to verify the effects resulting from the addition of powdered (500 g/t and 1000 g/t) and liquid (500 mL/t) cellulase over a period of 1 to 21 days. A completely randomized experimental design was used, consisting of four treatments with six replications and ten birds per replication that were housed in an experimental cage. It was observed that performance and digestibility results were significantly different with cellulase supplementation. Also, the relative weight of the large intestine in the period between one and seven days increased when cellulase was added at 1000 g/t. In the period of between eight and 14 days of life, the birds that consumed only the basal diet obtained higher levels of liver protein than those that received the treatments with the addition of the enzyme. However, 15 and 21 days, the consumed feed effect did not occur between thus, it is not conclusive whether hepatotoxicity occurs with the addition of cellulase. For the blood parameters, at 21 days, the diets with added cellulase were not significantly different regarding electrolytes. It was concluded that this cellulase produced by Trichoderma reesei can be included in the animals' diet.

Modulation of glycolipid metabolism in T2DM rats by Rubus irritans Focke extract: Insights from metabolic profiling and ERK/IRS-1 signaling pathway

Ethnopharmacological relevanceThe extracellular regulated protein kinase (ERK) plays a crucial role in the mitogen-activated protein kinase (MAPK) family, influencing apoptosis, proliferation, and differentiation. It connection to the insulin (INS) signaling cascade and the development of type 2 diabetes mellitus (T2DM) has been established. Rubus irritans Focke, an indispensable herb in Chinese Tibetan medicine for diabetes mellitus treatment, lacks a comprehensive understanding of its effects and pharmacological mechanisms in T2DM. Aim of the studyThis study aimed to elucidate the effects of Rubus irritans Focke extract (Rife) on a T2DM rat model, exploring its impact on glycemic and lipid metabolism, histopathological changes, and its potential targeting of the extracellular regulated protein kinase/insulin receptor substrate-1 (ERK/IRS-1) signaling pathway. Materials and methodsA T2DM rat model was induced by streptozotocin (STZ) injection (40 mg/kg) in high-fat diet-fed (HFD) male Wistar rats. Rife and metformin (Met) were administered for 4 weeks, and glycemic, lipid metabolism indices, and histopathological changes were assessed. Protein expression of ERK, IRS-1 in rat liver tissues was examined to evaluate the impact on the ERK/IRS-1 pathway. ResultsRife reducing hepatic ERK and IRS-1 protein expression in T2DM rats. Untargeted metabolomics identified 13 potential biomarkers and 4 differential metabolic pathways related to glycolipid metabolism disorders. ConclusionsRife demonstrated improved glycolipid metabolism in T2DM rats by inhibiting the ERK/IRS-1 related signaling pathway and influencing multiple metabolic pathways. This study provides valuable insights into the potential therapeutic mechanisms of Rife in the context of T2DM.

Copper exposure induces mitochondrial dysfunction and hepatotoxicity via the induction of oxidative stress and PERK/ATF4 -mediated endoplasmic reticulum stress

Copper is an essential trace element, and excessive exposure could result in hepatoxicity, however, the underlying molecular mechanisms remain incompletely understood. The present study is aimed to investigate the molecular mechanisms of copper sulfate (CuSO4) exposure-induced hepatoxicity both in vivo and in vitro. In vitro, HepG2 and L02 cells were exposed to various doses of CuSO4 for 24 h. Cell viability, ROS production, oxidative stress biomarkers, mitochondrial functions, ultrastructure, intracellular calcium (Ca2+) concentration, and the expression of proteins related to mitochondrial apoptosis and endoplasmic reticulum (ER) stress were assessed. In vivo, C57BL/6 mice were treated with CuSO4 at doses of 10 and 30 mg/kg BW/day and co-treated with 4-PBA at 100 mg/kg BW/day for 35 days. Subsequently, liver function, histopathological features, and protein expression were evaluated. Results found that exposure to CuSO4 at concentrations of 100–400 μM for 24 h significantly decreased the viabilities of HepG2 and L02 cells and it was in a dose-dependent manner. Additionally, CuSO4 exposure induced significant oxidative stress and mitochondrial dysfunction in HepG2 cells, which were partially ameliorated by the antioxidant N-acetylcysteine (NAC). Furthermore, CuSO4 exposure prominently triggered ER stress, as evidenced by the upregulation of GRP94, GRP78, phosphorylated forms of PERK and eIF2α, and CHOP proteins in livers of mice and HepG2 cells. NAC treatment significantly inhibited CuSO4 exposure -induced ER stress in HepG2 cells. Pharmacological inhibition of ER stress through co-treatment with 4-PBA and the PERK inhibitor GSK2606414, as well as genetic knockdown of ATF4, partially mitigated CuSO4-induced cytotoxicity in HepG2 cells by reducing mitochondrial dysfunction and inhibiting the mitochondrial apoptotic pathway. Moreover, 4-PBA treatment significantly attenuated CuSO4-induced caspase activation and hepatoxicity in mice. In conclusion, these results reveal that CuSO4-induced hepatotoxicity involves mitochondrial dysfunction and ER stress by activating oxidative stress induction and PERK/ATF4 pathway.

Exploring the relationship between dietary rapeseed meal, condensed tannin and growth, nutrive metabolism in largemouth bass (Micropterus salmoides)

This study evaluated the effects of rapeseed meal (RM) and condensed tannin (CT) on the growth and nutritional metabolism of largemouth bass (Micropterus salmoides), and speculated whether there was interaction between anti-nutritional factors in RM. Eight diets were formulated for 55-day feeding trial, six of which were practical diet groups, including control group (CG), five RM inclusive groups with graded RM levels of 5% (RM5), 10% (RM10), 15% (RM15), 20% (RM20) and 25% (RM25) partially replacing fishmeal, and the other two were semi-purified diets with 0.062% CT (CT5) ≈ the CT content in the RM5 and 0.31% CT (CT25) ≈ the CT content in the RM25. The results of the experiments on replacing fishmeal with RM demonstrated that with the increase of RM substitution level, the weight gain rate (WGR), survival rate (SR) and specific growth rate (SGR) of largemouth bass showed a downward trend. High levels of RM led to the inhibition of digestive enzyme activity in largemouth bass, as well as the manifestation of abnormal markers related to protein, fat, and carbohydrate metabolism in both plasma and liver. RM reduced muscle protein and fat content, although the effect was not statistically significant at 5% RM level. The results of CT concentration experiments showed that the addition of 0.31% CT significantly reduced WGR, SGR, SR, protein efficiency ratio (PER) and gastrointestinal digestive enzyme activity, and significantly increased the feed coefficient ratio (FCR), plasma ammonia (PA), plasma glucose (PG), and hepatic glycogen (HG) and liver triglyceride (TG). The addition of 0.062% CT significantly reduced PG and HG. Supplementation of CT significantly reduced muscle fat. Among the groups of CT supplementation and RM, CT5 exhibited superior WGR, PER and FCR compared to RM5, along with enhanced intestinal trypsin, amylase activity, liver glutamic pyruvic transaminase (GPT) activity and decreased content of HG and liver TG; CT25 exhibited decreased activities of gastric lipase (LPS), enteric trypsin and pepsin, and increased content of total cholesterol (T-CHO) and glucose in plasma in compariosn to RM25, accompanied with no significant change in growth performance. Therefore, largemouth bass exhibited tolerance to a 5% inclusion of RM in the feed without compromising their growth, digestion and metabolic functions. The incorporation of 5% RM, equivalent to a supplemental 0.062% CT, had regulatory effects on the digestion and metabolism of largemouth bass while improving feed utilization. There was an interaction among anti-nutritional factors in RM, with synergistic effect being evident at low RM levels, whereas antagonism more promient at high levels of RM.

YTHDF1 shapes immune-mediated hepatitis via regulating inflammatory cell recruitment and response

Severe immune responses regulate the various clinical hepatic injuries, including autoimmune hepatitis and acute viral hepatitis. N6-methyladenosine (m6A) modification is a crucial regulator of immunity and inflammation. However, the precise role of YTHDF1 in T cell-mediated hepatitis remains incompletely characterized. To address this, we utilized Concanavalin A (ConA)-induced mouse liver damage as an experimental model for T cell-mediated hepatitis. Our findings found that hepatic YTHDF1 protein rapidly decreased during ConA-induced hepatitis, and YTHDF1-deficient (Ythdf1−/−) mice showed more susceptibility to ConA-induced liver injury, along with an intensified inflammatory storm accompanied by aggravated hepatic inflammatory response via ERK and NF-κB pathways. Interestingly, hepatic-specific over-expression or deletion of YTHDF1 exhibited redundancy in ConA-induced liver injury. Validation in bone marrow chimeric mice confirmed the necessity of YTHDF1 in hematopoietic cells for controlling the response to ConA-induced hepatitis. Additionally, our data revealed that YTHDF1 deletion in macrophages exacerbated the inflammatory response induced by lipopolysaccharide. In summary, our study uncovered that YTHDF1 deficiency exacerbates the immune response in ConA-induced hepatitis by modulating the expression of inflammatory mediators, highlighting the potential of YTHDF1 as a therapeutic target for clinical hepatitis.

Smilax china L. polysaccharide prevents HFD induced-NAFLD by regulating hepatic fat metabolism and gut microbiota

BackgroundThe increasing incidence of nonalcoholic fatty liver disease(NAFLD) has urged the development of new therapeutics. NAFLD is intimately linked to gut microbiota due to the hepatic portal system, and utilizing natural polysaccharides as prebiotics has become a prospective strategy for preventing NAFLD. Smilax china L. polysaccharide (SCP) possesses excellent hepatoprotective and anti-inflammatory activity. However, its protective effects on NAFLD remains unclear. PurposeThe goal of this study was to explore the protective effects of SCP on high-fat diet (HFD)-induced NAFLD mice by regulating hepatic fat metabolism and gut microbiota. MethodsExtraction and isolation from Smilax china L. rhizome to obtain SCP. C57BL/6J mice were distributed to six groups: Control (normal chow diet), HFD-fed mice were assigned to HFD, simvastatin (SVT), and low-, medium-, high-doses of SCP for 12 weeks. The body, liver, and different adipose tissues weights were detected, and lipids in serum and liver were assessed. RT-PCR and Western blot were used to detect the hepatic fat metabolism-related genes and proteins. Gut microbiota of cecum contents was profiled through 16S rRNA gene sequencing. ResultsSCP effectively reversed HFD-induced increase weights of body, liver, and different adipose tissues. Lipid levels of serum and liver were also significantly reduced after SCP intervention. According to the results of RT-PCR and western blot analysis, SCP treatment up-regulated the genes and proteins related to lipolysis were up-regulated, while lipogenesis-related genes and proteins were down-regulated. Furthermore, the HFD-induced dysbiosis of intestinal microbiota was similarly repaired by SCP intervention, including enriching beneficial bacteria and depleting harmful bacteria. ConclusionSCP could effectively prevent HFD-induced NAFLD, might be considered as a prebiotic agent due to its excellent effects on altering hepatic fat metabolism and maintaining gut microbiota homeostasis.

Abstract 162: Post-transcriptional Regulation Of FGF21 In Obesity And Metabolism

Imbalances in energy intake and expenditure are the leading cause of the dramatic surge in obesity and cardiovascular disease affecting >60% of Americans. Increasing fat content in the average diet alters metabolism and contributes to disease. FGF21, a secreted hepatokine, modulates feeding behavior and energy expenditure (EE) primarily through signaling in brain and adipose tissue. FGF21 analogues are actively in clinical trials for several metabolic diseases, and it is therefore important to understand the various pathways that control FGF21. FGF21 is regulated by well-defined transcriptional programs, but its post-transcriptional regulation is less understood. Post-transcriptional regulation is mediated by RNA binding proteins (RBPs) that control mRNA stability. One family of such RBPs is the ZFP36 family proteins (ZFP36, ZFP36L1, ZFP36L2) that bind to AU-rich regions in the 3’UTR, leading to mRNA degradation. All 3 ZFP36 family proteins are redundant, therefore we knocked out Zfp36 , Zfp36l1 and Zfp36l2 in livers of adult mice. Fgf21 was the most upregulated gene following loss of ZFP36 family proteins in the liver. Consistent with elevated FGF21, these mice rapidly lost weight and fat mass while remaining metabolically healthy. Using metabolic cages, we observed that mice lacking hepatic ZFP36 family proteins had increased food intake and EE and decreased respiratory exchange ratio, indicating preferential fat burning. We then generated mice lacking the ZFP36 family proteins and Fgf21 which partially restored the metabolic phenotypes observed, leading to the hypothesis that ZFP36 family proteins post-transcriptionally regulate Fgf21 in controlling food intake and energy homeostasis. To determine if loss of ZFP36 family proteins protects against diet-induced obesity, we fed mice lacking the ZFP36 family proteins a high fat diet, however, they were entirely protected from diet induced obesity. The protection was partially reversed by additional deletion of Fgf21 . These findings demonstrate that FGF21 is regulated post-transcriptionally by the ZFP36 family proteins and loss of this mode of regulation has significant metabolic implications in diet-induced obesity and could lead to new therapeutic approaches to treat metabolic disease.

Assessing the ability of dihydrosterculic acid to activate PPARα

Peroxisome proliferator-activated receptor α (PPARα) is a ligand-activated transcription factor associated with the expression of specific target genes involved in fatty acid β-oxidation. Activation of PPARα could potentially be utilized to treat diseases associated with the accumulation of hepatic lipids such as non-alcoholic fatty liver disease (NAFLD), which currently impacts approximately 25% of the world’s population. At present, a class of synthetic ligands called fibrates can be prescribed to help manage NAFLD but are weak agonists for PPARα and therefore have limited effcacy. Consequently, alternative ligands for PPARα must be established and no natural endogenous ligands have been conclusively identified. This study expands on our previous work and aims to identify the direct effects of dihydrosterculic acid (DHSA), a cyclopropyl fatty acid found in cottonseed oil (CSO), on PPARα activity. To identify downstream effects of DHSA, male mice were placed on a CSO- (DHSA-containing) or isocaloric oil-enriched diet for 6 weeks and tissues were sent for genome-wide RNA sequencing analysis. CSO-fed mice demonstrated increased PPARα gene expression in the liver and concomitant decreased triglyceride in plasma (-75.1±11.5 milligrams/deciliter (mg/dL); p<0.01) and liver (-86.7±71.7 mg/gram (g) protein; p=0.1) and decreased free fatty acid in liver (-16.5±3.4 micromolar (μM)/g protein; p<0.01) relative to the oil-enriched control diet. Confirmational tissue analysis of liver samples show an increase in PPARα target gene expression in the DHSA containing groups compared to control diet groups, indicating that DHSA could potentially serve as an endogenous ligand for PPARα to increase its transcriptional activity to increase fat oxidation. To further test this hypothesis, female PPARα knockout mice were fed a DHSA containing diet or control diet for 4 weeks (n=8). Following the 4-week period, the chow-fed wild type females gained more weight (1.3±0.6 g; p<0.05) than the PPARα knockout mice. Otherwise, there were no significant differences in body weight or food intake between diets or genotypes. Additional in vitro studies show increased PPARα target gene expression following treatments with DHSA, specifically for CPT1a (2.7±0.2 fold increase; p<0.01), ACOX1 (1.3±0.1 fold increase; p<0.05), and ACADVL (1.6±0.1 fold increase, p<0.01), which encode key enzymes in lipid oxidation pathways. SCD1, the rate limiting enzyme in lipogenesis, was also downregulated (0.5±0.03 fold decrease, p<0.01) in response to a 24-hour treatment with DHSA indicating a reduction in endogenous lipogenesis. From these results, we believe that DHSA could function through a PPARα-dependent mechanism. Further analysis of the tissues collected from the PPARα knockout mouse model will aid in determining the mechanism of DHSA as well as identifying its potential for the treatment of excess lipid accumulation. This project is funded by Cotton Incorporated. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Effects of antagonism of thromboxane-prostanoid receptor on alcohol-associated liver disease

Backgrounds: Alcohol-associated liver disease (ALD) is one of the leading causes of liver diseases. Thromboxane-prostanoid receptor (TP-R) is widely expressed in the liver and is activated by thromboxane A2 (TXA2). A previous study reported an increase in TP-R mRNA level in liver after ethanol feeding. Preclinical studies have documented that TP-R antagonists exert a protective effect against cardiovascular and liver diseases. In particular, TP-R antagonist is reported to attenuate ALD in rodents. However, the mechanism(s) by which antagonizing TP-R attenuates ALD remains unclear. Methods: Herein, we used the chronic plus binge ethanol feeding model. Briefly, male, C57BL/6 wild-type mice (8-week) were fed a Lieber-Decarli ethanol (5% v/v) diet (ET, n = 10) or isocaloric control diet (CON, n = 6) for ten days followed by a single binge administration of ethanol or maltose-dextrin via oral gavage. A cohort of ethanol-fed mice received SQ 29,548, a TP-R antagonist (ET+SQ, n = 8) by oral administration. Results: Western blot analysis showed that the protein level of thromboxane A2 synthase 1 ( P<0.05), responsible for TXA2 production, was upregulated in the mouse liver upon ethanol exposure. Of note, ethanol feeding significantly increased plasma aspartate transferase (AST) level ( P<0.05), which was abolished by SQ administration ( P<0.05). Moreover, compared with CON mice, ET mice showed an increase in liver protein levels of pro-inflammatory markers, including TNFa ( P<0.05), cluster of differentiation 68 (CD68, P<0.001), and vascular cell adhesion protein 1 ( P<0.05). In addition, the pro-fibrogenic markers including collagen type 1 alpha 1 chain (COL1A1, P<0.05) and matrix metalloproteinase 9 (MMP9, P<0.01) were also increased in ET mice. However, these alterations were not found in ET+SQ mice compared to their controls. In particular, ET+SQ mice exhibited a marked reduction in hepatic protein levels of CD68 ( P<0.001), COL1A1 ( P<0.001), and MMP9 ( P<0.01) versus ET mice. Interestingly, we noted that ethanol consumption decreases the protein level of NADH:ubiquinone oxidoreductase subunit B8 (NDUFB8), a component of mitochondrial complex I, in mouse liver ( P<0.05), which was reversed by SQ administration( P<0.05). Conclusion: These findings suggest that blockade of TP-R activity attenuates ethanol-induced liver inflammation and fibrosis possibly through improving the function of mitochondrial complex I. This study was supported by the NIH-NIAAA P50 award-Alcohol Center of Research-Nebraska (P50AA030407-5130). This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Liver PKCε Heterozygous (LivPKCεfl/Δ) Female Mice Showed Better Glucose Tolerance than LivPKCεfl/Δ Male Mice When Fed on High Fat or High Fructose Diet for One Month

Introduction: Excess intake of high fat (HFD) and high fructose (HFrD) diets leads to liver fat accumulation that are linked with insulin resistance, glucose intolerance, hepatosteatosis and diet induced obesity. Activation of liver protein kinase Cε (PKCε) has been implicated as a key mediator of diet induced hepatic insulin resistance. However, this mechanism is not fully understood. Using PKCε knockout (PKCε-KO) mice, we aim to investigate the molecular mechanisms of how PKCε may interact with the insulin signaling cascade in both HFD and HFrD induced insulin resistance and obesity. Hypothesis: Studies have shown that PKCε−/− mice are protected from diet-induced insulin resistance and fasting-state glucose homeostasis was enhanced in PKC-KO mice. We hypothesize that hepatic PKCε deletion will protect against HFD and HFrD induced glucose intolerance in mice. Methods: In order to generate liver specific PKCε−/− mice, we crossed female PKCεfl/fl mice with male Alb Cre+/+ mice. At the time of this study, we have generated heterozygous (PKCεfl/Δ) mice which are being crossed to develop homozygous PKCε knockout (LivPKCεΔ/Δ) mice. Both Male and Female heterozygous (PKCεfl/Δ) mice were placed on three diets: Control (Cntrl, Research Diets Inc, RDI D12450J, Protein (20%), Carbs (70%, corn starch-50%, sucrose-maldextrin-20%, fructose-0%), Fat 10% (lard)), High Fructose (HFrD, RDI D02022704), Protein (20%), Carbs (70%, corn starch-50%, sucrose-maldextrin-20%, fructose-60%), Fat 10% (lard)) and High Fat (HFD, RDI D12492, Protein (20%), Carbs (20%, corn starch-0%, sucrose-maldextrin-20%, fructose-0%), Fat 60% (lard)). Within 1 month on the diets, the mice were genotyped and glucose tolerance tests (GTT, 2 g/kg body weight) were performed following 6 hours of starvation. Results: Our preliminary results showed that within one month, LivPKCεfl/Δ-HFD and LivPKCεfl/Δ-HFrD female mice displayed better glucose tolerance than male mice as seen by their lower glucose level during GTT. Interestingly, both LivPKCεfl/Δ male and LivPKCεfl/Δ female mice did not show significant differences in glucose tolerance when they were on control diet (LivPKCεfl/Δ-Cntrl) within the same duration. Conclusion: Conflicting evidence exists whether liver or adipose tissue specific deletion of PKCε protects mice from diet induced glucose intolerance. Studies in both humans and rodents have demonstrated that female subjects experience less endogenous glucose production and less hepatic insulin resistance than their male counterparts after high fat or high fructose challenge (PMID 23049932, PMID 18332156) and our preliminary findings are in line with these studies. Subsequent studies will use homozygous PKCε (LivPKCεΔ/Δ) mice and their littermates Wild-type mice for in-depth metabolic and molecular analysis. We would like to thank the National Science Foundation for award # NSF1931045 and the National Institutes of Health, award # 1R01DK126892-01A1. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Common snook (Centropomus undecimalis) juveniles maintained at high temperature and brackish water improve growth, osmoregulation, and antioxidant responses

The study evaluated the interactions between temperature (25 and 28 °C) and salinity (0, 15, and 32 ppt) on the growth performance, plasmatic ions, ATPases activities, and oxidative and antioxidant parameters in different tissues of common snook (Centropomus undecimalis) reared for 90 days by a 2 × 3 factorial design. The growth performance and feed conversion ratio were influenced by temperature and interaction between factors, with the best results observed in the combination between 28 °C and 15 ppt. There were effects of temperature and salinity (isolated or in interaction) for plasmatic ions, v-type H+-ATPase (VHA) and Na+/K+-ATPase (NKA) activities, and oxidative stress responses. Gill VHA (at 32 ppt) and NKA (at 0 and 15 ppt) activities were higher at 25 than at 28 °C. In the kidney, at 28 °C, the VHA (at 15 ppt) and NKA (at 32 ppt) activities were higher than at 25 °C. The VHA and NKA activities in the stomach at 28 °C and 32 ppt were higher than at the other treatments. In the intestine, the lowest activity of VHA at 25 and 28 °C occurred at 0 and 32 ppt, respectively, and the activity of NKA was highest in the interaction of 0 ppt and 25 °C. At 0 ppt, the plasmatic Na+ and Cl− values at both temperatures were decreased, and the plasmatic K+ values at 25 °C were increased. Liver thiobarbituric acid reactive substances and protein carbonyl content values were higher at the combinations of 15 ppt and 25 °C and 32 ppt and 28 °C than at the other interactions. Liver antioxidant responses (glutathione-S-transferase, glutathione peroxidase, non-protein thiols, and superoxide dismutase) were higher at 15 ppt than other salinities at both temperatures. In conclusion, the improved growth, osmoregulation, and antioxidant responses were found to be higher at 28 °C and 15 ppt.

Understanding the SARS-CoV-2–Human Liver Interactome Using a Comprehensive Analysis of the Individual Virus–Host Interactions

Many metabolic processes at the molecular level support both viral attack strategies and human defenses during COVID-19. This knowledge is of vital importance in the design of antiviral drugs. In this study, we extracted 18 articles (2021–2023) from PubMed reporting the discovery of hub nodes specific for the liver during COVID-19, identifying 142 hub nodes. They are highly connected proteins from which to obtain deep functional information on viral strategies when used as functional seeds. Therefore, we evaluated the functional and structural significance of each of them to endorse their reliable use as seeds. After filtering, the remaining 111 hubs were used to obtain by STRING an enriched interactome of 1111 nodes (13,494 interactions). It shows the viral strategy in the liver is to attack the entire cytoplasmic translational system, including ribosomes, to take control of protein biosynthesis. We used the SARS2-Human Proteome Interaction Database (33,791 interactions), designed by us with BioGRID data to implement a reverse engineering process that identified human proteins actively interacting with viral proteins. The results show 57% of human liver proteins are directly involved in COVID-19, a strong impairment of the ribosome and spliceosome, an antiviral defense mechanism against cellular stress of the p53 system, and, surprisingly, a viral capacity for multiple protein attacks against single human proteins that reveal underlying evolutionary–topological molecular mechanisms. Viral behavior over time suggests different molecular strategies for different organs.

Hepatocyte-Specific Fads1 Overexpression Attenuates Western Diet-Induced Metabolic Phenotypes in a Rat Model.

Fatty acid desaturase 1 (FADS1) is a rate-limiting enzyme in long-chain polyunsaturated fatty acid (LCPUFA) synthesis. Reduced activity of FADS1 was observed in metabolic dysfunction-associated steatotic liver disease (MASLD). The aim of this study was to determine whether adeno-associated virus serotype 8 (AAV8) mediated hepatocyte-specific overexpression of Fads1 (AAV8-Fads1) attenuates western diet-induced metabolic phenotypes in a rat model. Male weanling Sprague-Dawley rats were fed with a chow diet, or low-fat high-fructose (LFHFr) or high-fat high-fructose diet (HFHFr) ad libitum for 8 weeks. Metabolic phenotypes were evaluated at the endpoint. AAV8-Fads1 injection restored hepatic FADS1 protein levels in both LFHFr and HFHFr-fed rats. While AAV8-Fads1 injection led to improved glucose tolerance and insulin signaling in LFHFr-fed rats, it significantly reduced plasma triglyceride (by ~50%) and hepatic cholesterol levels (by ~25%) in HFHFr-fed rats. Hepatic lipidomics analysis showed that FADS1 activity was rescued by AAV8-FADS1 in HFHFr-fed rats, as shown by the restored arachidonic acid (AA)/dihomo-γ-linolenic acid (DGLA) ratio, and that was associated with reduced monounsaturated fatty acid (MUFA). Our data suggest that the beneficial role of AAV8-Fads1 is likely mediated by the inhibition of fatty acid re-esterification. FADS1 is a promising therapeutic target for MASLD in a diet-dependent manner.

Hepatic Enzymes and Proteins as Prognostic Factors in Liver Disease Management

Therefore, an article reviewing a hepatic enzyme/protein prognosis in liver disease studies is presented. Hepatocytic enzymes such as Alanine aminotransferase (ALT) and Aspartate aminotransferase (AST) being highly contributory indicators demonstrate mild to severe active liver inflammation and damage. Compared to this, it is worth noting that the so-called proteins which are referred to as albumin and alpha-fetoprotein (AFP) provide data on liver function and the severity of the disease, mainly in HCC (hepatocellular carcinoma). The review clarifies the role of these biomarkers in clinical practice attributing them to various indicators such as diagnosis, disease differentiation, treatment monitoring, and prognosis assessment. Difficulties, including variability, sensitivity problems, and complexity on the interpretation of results, are pointed out, showing the necessity of enhanced diagnosis tehnologies and personalized medicine strategies. The future of liver disease management concentrates on biomarker identification, better diagnostic equipment, embedding artificial intelligence, personized treatment, and involving authorities. The objective is to improve diagnostic accuracy, guid treatment outcomes, and apply ethics in genetic information. In summary, these pioneering technologies have the potential to make a substantial contribution to the level of liver disease treatment, provided that we address the connected problems, which will consequently result in better patients’ outcomes and quality of care.

Traditional Japanese medicine Kamikihito ameliorates sucrose preference, chronic inflammation and obesity induced by a high fat diet in middle-aged mice.

The high prevalence of obesity has become a pressing global public health problem and there exists a strong association between increased BMI and mortality at a BMI of 25 kg/m2 or higher. The prevalence of obesity is higher among middle-aged adults than among younger groups and the combination of aging and obesity exacerbate systemic inflammation. Increased inflammatory cytokines such as interleukin 6 and tumor necrosis factor alpha (TNFα) are hallmarks of obesity, and promote the secretion of hepatic C-reactive protein (CRP) which further induces systematic inflammation. The neuropeptide oxytocin has been shown to have anti-obesity and anti-inflammation effects, and also suppress sweet-tasting carbohydrate consumption in mammals. Previously, we have shown that the Japanese herbal medicine Kamikihito (KKT), which is used to treat neuropsychological stress disorders in Japan, functions as an oxytocin receptors agonist. In the present study, we further investigated the effect of KKT on body weight (BW), food intake, inflammation, and sweet preferences in middle-aged obese mice. KKT oral administration for 12 days decreased the expression of pro-inflammatory cytokines in the liver, and the plasma CRP and TNFα levels in obese mice. The effect of KKT administration was found to be different between male and female mice. In the absence of sucrose, KKT administration decreased food intake only in male mice. However, while having access to a 30% sucrose solution, both BW and food intake was decreased by KKT administration in male and female mice; but sucrose intake was decreased in female mice alone. In addition, KKT administration decreased sucrose intake in oxytocin deficient lean mice, but not in the WT lean mice. The present study demonstrates that KKT ameliorates chronic inflammation, which is strongly associated with aging and obesity, and decreases food intake in male mice as well as sucrose intake in female mice; in an oxytocin receptor dependent manner.

Repeated Injection of Very Small Superparamagnetic Iron Oxide Particles (VSOPs) in Murine Atherosclerosis: A Safety Study.

Citrate-coated electrostatically stabilized very small superparamagnetic iron oxide particles (VSOPs) have been successfully tested as magnetic resonance angiography (MRA) contrast agents and are promising tools for molecular imaging of atherosclerosis. Their repeated use in the background of pre-existing hyperlipidemia and atherosclerosis has not yet been studied. This study aimed to investigate the effect of multiple intravenous injections of VSOPs in atherosclerotic mice. Taurine-formulated VSOPs (VSOP-T) were repeatedly intravenously injected at 100 µmol Fe/kg in apolipoprotein E-deficient (ApoE KO) mice with diet-induced atherosclerosis. Angiographic imaging was carried out by in vivo MRI. Magnetic particle spectrometry was used to detect tissue VSOP content, and tissue iron content was quantified photometrically. Pathological changes in organs, atherosclerotic plaque development, and expression of hepatic iron-related proteins were evaluated. VSOP-T enabled the angiographic imaging of heart and blood vessels with a blood half-life of one hour. Repeated intravenous injection led to VSOP deposition and iron accumulation in the liver and spleen without affecting liver and spleen pathology, expression of hepatic iron metabolism proteins, serum lipids, or atherosclerotic lesion formation. Repeated injections of VSOP-T doses sufficient for MRA analyses had no significant effects on plaque burden, steatohepatitis, and iron homeostasis in atherosclerotic mice. These findings underscore the safety of VSOP-T and support its further development as a contrast agent and molecular imaging tool.

SiRNA-mediated reduction of a circulating protein in swine using lipid nanoparticles

Genetic manipulation of animal models is a fundamental research tool in biology and medicine but is challenging in large animals. In rodents, models can be readily developed by knocking out genes in embryonic stem cells or by knocking down genes through invivo delivery of nucleic acids. Swine are a preferred animal model for studying the cardiovascular and immune systems, but there are limited strategies for genetic manipulation. Lipid nanoparticles (LNPs) efficiently deliver small interfering RNA (siRNA) to knock down circulating proteins, but swine are sensitive to LNP-induced complement activation-related pseudoallergy (CARPA). We hypothesized that appropriately administering optimized siRNA-LNPs could knock down circulating levels of plasminogen, a blood protein synthesized in the liver. siRNA-LNPs against plasminogen (siPLG) reduced plasma plasminogen protein and hepatic plasminogen mRNA levels to below 5% of baseline values. Functional assays showed that reducing plasminogen levels modulated systemic blood coagulation. Clinical signs of CARPA were not observed, and occasional mild and transient hepatotoxicity was present in siPLG-treated animals at 5h post-infusion, which returned to baseline by 7days. These findings advance siRNA-LNPs in swine models, enabling genetic engineering of blood and hepatic proteins, which can likely expand to proteins in other tissues in the future.