Hepatic mRNA Research Articles

Abstract 4135443: A Phase 1, Single Ascending Dose Study to Evaluate ALN-TTRsc04, a Next-Generation RNA Interference Therapeutic, in Healthy Participants for Potential Treatment of Transthyretin Amyloidosis

Introduction: Transthyretin amyloidosis (ATTR) is a progressive, debilitating, fatal disease caused by deposition of misfolded toxic transthyretin (TTR) amyloid fibrils in multiple organs and tissues. Major ATTR manifestations include polyneuropathy and cardiomyopathy. Ongoing amyloid deposition drives rapid progression of ATTR, leading to morbidity and mortality. ALN-TTRsc04 is an investigational small interfering RNA that targets hepatic TTR mRNA to reduce circulating TTR protein levels. Hypothesis: ALN-TTRsc04 rapidly knocks down serum TTR and is well tolerated. Aims: Describe safety, pharmacokinetics, and pharmacodynamics of single ascending ALN-TTRsc04 doses. Methods: This ongoing, Phase 1, randomized, double-blind, placebo-controlled study (NCT05661916) enrolled healthy adults (18–65 yrs; 18.0–30 kg/m 2 ) sequentially included in 6 cohorts receiving subcutaneous single ascending doses of ALN-TTRsc04 (5, 25, 100, 300, 600, or 900 mg) or placebo (randomized 3:1). Urine and plasma concentrations were measured on Days 1–2 and 1–4, respectively; change from baseline (BL) in serum TTR and AEs will be assessed over 12 months. Results: Overall, 36 and 12 adults (of similar age/sex distribution) were randomized to 1 of the 6 ALN-TTRsc04 groups or placebo, respectively. ALN-TTRsc04 plasma concentrations declined below lower limit of quantification within 24–48 hrs; mean (SD) plasma half-life ranged from 4.5 (1.5) hrs (300 mg dose) to 7.6 (3.2) hrs (100 mg dose). ALN-TTRsc04 was minimally excreted by renal route (<20%) after 24 hrs. Rapid and sustained knockdown in serum TTR occurred following all ALN-TTRsc04 doses ( Figure ). Mean (SD) serum TTR was 236.2 (47.4) µg/mL and 290.1 (73.3) µg/mL at BL and 22.2 (1.8) µg/mL and 289.0 (59.1) µg/mL at Day 15, for ALN-TTRsc04 300 mg and placebo, respectively. Mean percent knockdown at Day 15 was 90.3% vs 1.8%, respectively. Robust mean serum TTR knockdown of >90% from BL was seen with ALN-TTRsc04 doses ≥300 mg at Day 29 and sustained until Day 90 (or Day 180 with 300 mg dose, 92.6% reduction). Majority of AEs across doses were mild; none were considered related to treatment. No injection-site reactions or safety signals, including liver-related signals, were identified. Conclusion: A single ALN-TTRsc04 dose led to a rapid and sustained knockdown in serum TTR; a reduction >90% from BL was sustained until Day 180 with the 300 mg dose. All ALN-TTRsc04 doses have been well tolerated to date; data through 12 months will be assessed.

E2F3a Activates Gadd45b Gene Expression Through PPARα-Mediated Transcriptional Regulation in Hepatic Cells

Growth arrest and DNA damage-inducible beta (GADD45b) plays a critical role in intracellular events such as cell growth and apoptosis. Although the functional study of GADD45b has been conducted, the mechanism for the transcriptional regulation of GADD45b is largely unknown. Due to the drastic induction of hepatic GADD45b mRNA by peroxisome proliferator-activated receptor alpha (PPARα) activation in wild-type mice, we investigated a key factor that affects the upregulation of GADD45b mRNA. Interestingly, we found that GADD45b-promoter luciferase activity was dramatically increased as the 5’-flanking regions were closer to the transcription start site (TSS) in Hepa1c1c7 cells. Additionally, we found two E2F (E2F-myc activator/cell cycle regulator) binding motifs in the region (-150/-1) of the GADD45b promoter. Notably, E2F3 mRNA was significantly increased only in wild-type mice treated with WY-14643, a PPARα agonist, followed by the induction of GADD45b mRNA. Furthermore, gene functional studies and Chromatin Immunoprecipitation (ChIP) assays revealed that E2F3 could regulate the GADD45b gene via the E2F binding motif. Thus, we suggest that E2F3 may play a key role in regulating the GADD45b gene as a positive transcription factor.

Liver-expressed antimicrobial peptide 2 is a hepatokine regulated by ghrelin, nutrients, and body weight.

Liver-expressed antimicrobial peptide 2 (LEAP2) is a peptide that counteracts the hunger hormone ghrelin-induced functions. Recently, we showed that vertical sleeve gastrectomy (VSG) did not alter the serum LEAP2 concentration in individuals with obesity. Here, we investigated the effects of VSG in both chow diet (CD)-fed and high-fat diet (HFD)-fed mice. In CD-fed mice, VSG increased plasma LEAP2 levels and hepatic Leap2 mRNA levels while decreasing body weight, blood glucose levels, and ghrelin levels. Intraperitoneal (ip) administration of ghrelin reversed these changes. These effects were found in both male and female mice. In contrast, VSG or weight loss in HFD-induced obese mice decreased LEAP2 levels. After fasting, the plasma LEAP2 concentration was in the following order: hepatic vein > abdominal aorta > portal vein. A high glucose concentration robustly increased the plasma LEAP2 concentration in the hepatic vein and abdominal aorta but not in the portal vein. In addition, corn oil or palmitate increased LEAP2 expression and secretion. The increase in LEAP2 levels after the meal tolerance test was delayed in the human subjects with diabetes. Our data suggest that various factors (metabolic, hormonal, and nutritional) regulate LEAP2, and the liver is the predominant site for the production and secretion of LEAP2. Furthermore, the interaction between ghrelin and LEAP2 is involved in the pathogenesis of obesity and diabetes.

Palygorskite improves growth performance and prevents liver damage in avian pathogenic Escherichia coli-challenged broiler chickens at an early age.

Avian pathogenic Escherichia coli (APEC) is a major bacterial infection that causes economic losses in the global poultry industry. Palygorskite (PAL) has been shown to enhance growth performance while improving antioxidative and anti-inflammatory properties of broilers. This study evaluated the protective effects of PAL on growth performance and liver function in broilers subjected to APEC challenge. A total of 320 one-day-old male Arbor Acres chicks were divided into four groups with eight replicates of ten birds each, based on a 2×2 factorial arrangement (basal diet or 5g/kg PAL-supplemented diet) and inoculation (bacterial culture medium or APEC). PAL increased body weight gain (BWG) prior to APEC challenge (P < 0.05). However, APEC caused losses in BWG, feed intake (FI), and feed efficiency, along with increased relative hepatic weight, hepatic pathology scores, and hepatic-cell apoptosis rate (P < 0.05). Compared to normal birds, APEC increased interleukin (IL)-1β, interferon-γ (IFN-γ), tumor necrosis factor-α (TNF-α), malondialdehyde (MDA), and nitric oxide (NO) levels, as well as lysozyme (LZM) and myeloperoxidase (MPO) activities, while decreasing total antioxidant capacity (T-AOC) and IL-10 levels, and total superoxide dismutase (T-SOD) and catalase (CAT) activities in both serum and liver, APEC also raised alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activities, but reduced total protein (TP), albumin (ALB), immunoglobulin (Ig) A, IgG, and IgM levels in serum (P < 0.05). Moreover, APEC increased hepatic mRNA level of IL-1β, IFN-γ, TNF-α, nuclear factor kappa B, and inducible nitric oxide synthase (iNOS), while inhibited mRNA level of IL-10 (P < 0.05). In contrast, PAL increased BWG and FI, alleviated hepatic-cell apoptosis rate during the challenge period (P < 0.05). An incorporation of PAL reduced triglyceride and NO contents, ALT and AST activities, while increasing TP, ALB, IL-10, IgG, and IgM levels in serum, enhancing serum T-SOD and CAT activities, elevating hepatic T-AOC and CAT activities, inhibiting hepatic MDA accumulation, and reducing IL-1β levels and LZM activity in both liver and serum (P < 0.05). An interactive effect was found for hepatic TNF-α and iNOS mRNA expression, in which PAL inhibited their mRNA expression in APEC-challenged birds (P < 0.05). Overall, PAL addition partially mitigated the negative impact of APEC challenge on growth performance and liver function of broiler chicks at an early age.

6503 A Randomized, Placebo-Controlled Phase 3 Study of Olezarsen In Patients With Familial Chylomicronemia Syndrome: The Balance Trial

Abstract Disclosure: E.S. Stroes: Advisory Board Member; Self; Amgen Inc, Sanofi, Novartis Pharmaceuticals, AstraZeneca, Novo Nordisk, Ionis Pharmaceuticals Inc., Merck, Daiichi Sankyo. Grant Recipient; Self; Ionis Pharmaceuticals Inc., Novo Nordisk, Novartis Pharmaceuticals, Sanofi. V. Alexander: Employee; Self; Ionis Pharmaceuticals Inc. E. Prokopczuk: Employee; Self; Ionis Pharmaceuticals Inc. R.A. Hegele: Consulting Fee; Self; Aegerion Pharmaceuticals, Ionis Pharmaceuticals Inc., Amgen Inc, Novartis Pharmaceuticals, Pfizer, Inc., Regeneron Pharmaceuticals, Sanofi, Ultragenyx, Acasti, and HLS Therapeutics. M. Arca: Grant Recipient; Self; Amgen Inc, Ionis Pharmaceuticals Inc., Novartis Pharmaceuticals, Pfizer, Inc., Regeneron Pharmaceuticals, Sanofi, Ultragenyx, Daiichi Sankyo, Alfasigma, Amryt, Sobi, and Viatris. C.M. Ballantyne: Consulting Fee; Self; 89Bio, Abbott Diagnostics, Alnylam Pharmaceuticals, Althera, Amarin, Amgen, Arrowhead, AstraZeneca, Denka Seiken, Esperion, Genentech, Gilead, Illumina, Ionis, Matinas BioPharma Inc, Merck, New Amster. Grant Recipient; Self; Abbott Diagnostic, Akcea, Amgen, Arrowhead, Esperion, Ionis, Merck, Novartis, Novo Nordisk, Regeneron, Roche Diagnostic, NIH, AHA, ADA. H. Soran: Grant Recipient; Self; Akcea Therapeutics, Alexion, Amryt, Kowa, MSD, NAPP, Novartis, Pfizer, Sanofi, Synageva, and Takeda. T.A. Prohaska: Employee; Self; Ionis Pharmaceuticals Inc. S. Xia: Employee; Self; Ionis Pharmaceuticals Inc. H.N. Ginsberg: Research Investigator; Self; Ionis Pharmaceuticals Inc. J.L. Witztum: Consulting Fee; Self; Ionis Pharmaceuticals Inc. S. Tsimikas: Consulting Fee; Self; Novartis. Employee; Self; Ionis Pharmaceuticals Inc.. Stock Owner; Self; Kleanathi, Oxitope. Background: Familial chylomicronemia syndrome (FCS) is a rare genetic disorder of lipoprotein lipase deficiency that results in severe hypertriglyceridemia and an increased risk of potentially life-threatening acute pancreatitis (AP). Decreasing high plasma levels of apolipoprotein C-III (apoC-III) reduces hypertriglyceridemia. Olezarsen is an investigational ligand-conjugated antisense oligonucleotide targeted to hepatic APOC3 mRNA to reduce triglycerides (TG). Methods: Balance is a Phase 3, multicenter, randomized, double-blind, placebo-controlled trial in 66 patients with genetically confirmed FCS and fasting TG ≥880 mg/dL at screening. Patients were advised to consume &amp;lt;20g fat/day and randomized 2:1 to olezarsen (50 mg or 80 mg) or placebo sc every 4 weeks for 53 weeks. The two primary endpoints were the placebo-adjusted % change in fasting TG from baseline to 6-months for (1) olezarsen 80 mg and (2) olezarsen 50 mg. Secondary endpoints included change in 12-month TG and apoC-III levels, and incidence of independently adjudicated AP events. Results: At baseline, TG levels were ∼2600 mg/dL and 47/66 (71%) of FCS patients had a history of AP. Placebo-corrected TG levels at 6 months were significantly reduced in olezarsen 80 mg (-43.5%, P=0.0009), and were numerically lower in the olezarsen 50 mg group (-22.4%, P=0.0775). Placebo-corrected TG levels at 12 months were reduced -59.4% in the olezarsen 80 mg group and -43.8% in the olezarsen 50 mg group. ApoC-III levels were reduced in both dose groups at 6 months (olezarsen 50 mg: -65.5%; olezarsen 80 mg: -73.7%) and 12 months (olezarsen 50 mg: -77.1%; olezarsen 80 mg: -81.3%). During 53 weeks of randomized treatment, 11 episodes of AP were observed in 23 patients on placebo versus 2 episodes in 43 olezarsen-treated patients (1 in 50 mg and 1 in 80 mg group). A favorable safety and tolerability profile was noted with more treatment emergent adverse events (TEAEs) and serious TEAEs in the placebo group. No serious TEAEs were related to study drug. Conclusion: In patients with FCS, olezarsen 80 mg significantly reduced TG levels and both olezarsen 50 and 80 mg groups demonstrated a marked reduction in AP events with a favorable safety and tolerability profile. Presentation: 6/1/2024

Arylacetamide deacetylase regulates hepatic iron homeostasis to protect against carbon tetrachloride-induced ferroptosis.

Arylacetamide deacetylase (AADAC) catalyzes the hydrolysis of small molecules containing ester and amide bonds. Recently, it has been reported that AADAC can suppress reactive oxygen species production in cancer cells. This study aimed to elucidate the possibility that AADAC protects against drug-induced liver injury accompanied by oxidative stress and to explore its molecular mechanisms. Intraperitoneal administration of carbon tetrachloride induced significantly more severe liver injury in Aadac knockout (KO) mice (plasma alanine aminotransferase level: 19,381 ± 10,578 U/L) than in wild-type (WT) mice (7219 ± 4729 U/L). More severe liver injury in Aadac KO mice was accompanied by higher hepatic malondialdehyde and antioxidant gene mRNA levels than those in WT mice. The increase in plasma alanine aminotransferase levels in Aadac KO mice was substantially suppressed by pretreatment with the ferroptosis inhibitors deferoxamine or ferrostatin-1, suggesting that Aadac deficiency increases susceptibility to ferroptosis. Immunoprecipitation followed by proteomic analysis revealed that AADAC interacts with ceruloplasmin (CP), which oxidizes ferrous iron to ferric iron. Hepatic CP activity was significantly lower in Aadac KO mice than that in WT mice, resulting in elevated hepatic ferrous iron levels in Aadac KO mice. Overexpression of human AADAC in Huh-7 cells significantly attenuated carbon tetrachloride-induced cytotoxicity by suppressing ferrous iron accumulation, suggesting that AADAC interacts with CP to suppress hepatic ferrous iron accumulation. The hepatoprotective role of Aadac in ferroptosis was also observed in mice with acetaminophen-induced liver injury. This study demonstrates a novel function of AADAC in protecting against ferroptosis induced by hepatotoxicants, carbon tetrachlorideand acetaminophen.

6826 Mevalonate Treatment Worsened Hypocholesterolemia During Prolonged Sepsis Without Affecting Adrenal And Muscle Function

Abstract Disclosure: L. De Bruyn: None. S. Derde: None. F. Van Beek: None. S. Vander Perre: None. I. Derese: None. L. Pauwels: None. G. Van den Berghe: None. L. Langouche: None. Introduction: Sepsis is hallmarked by an immediate and sustained reduction in total-, HDL- and LDL-cholesterol (-C), associated with illness severity and a potential contributor to the development of adrenal failure and muscle weakness. The underlying mechanisms that might be involved in sepsis-induced hypocholesterolemia are multifactorial and partly understood. We hypothesized that treatment with the cholesterol precursor mevalonate might improve cholesterol availability, thereby improving adrenal gland and skeletal muscle function. Our hypothesis was tested in a validated and clinically relevant mouse model of prolonged sepsis. Methods: In a catheterized mouse model of cecal ligation and puncture-induced, fluid resuscitated and antibiotics treated polymicrobial abdominal sepsis (5 d), septic mice received continuous infusion with either mevalonate (78 mg/d) or placebo solution, whereas healthy mice served as controls (N=50). Plasma total-C (HDL- + LDL-C), CORT and total bile acids were measured, in addition to ex vivo muscle force and adrenocortical cholesterol ester content. Gene expression markers of inflammation (Tnf-α, Il-6) were measured in the adrenal gland, as well as hepatic mRNA expression of cholesterol synthesis enzymes (Hmgcs1, Hmgcr, Fdft1). Metabolites of the mevalonate pathway (mevalonic acid, squalene, lanosterol, desmosterol and (7-dehydro) cholesterol) were quantified in the liver with LC-MS. Results: The sepsis-induced reduction in plasma total-C was further decreased in mevalonate-treated septic mice (26±3 mg/dL) as compared with placebo (53±3 mg/dL) and healthy controls (108±3 mg/dL) (p&amp;lt;0.0001). Metabolites of the mevalonate pathway were all increased in the liver after mevalonate treatment (p&amp;lt;0.01), whereas hepatic cholesterol content was decreased as compared with placebo (p&amp;lt;0.05). Also, gene expression markers of cholesterol synthesis enzymes were further decreased in the liver of mevalonate-treated septic mice as compared with placebo (p&amp;lt;0.05). In the context of the unexpected further reduction in plasma total-C, no additional effect on plasma CORT, total bile acids, and sepsis-induced loss of muscle force and depletion in adrenocortical cholesterol ester content was observed. In addition, the sepsis-induced rise in markers of adrenal inflammation (Tnf-α, Il-6) was not further affected by mevalonate treatment. Conclusion: Mevalonate treatment further reduced the sepsis-induced hypocholesterolemia in prolonged septic mice, potentially due to increased feedback inhibition on hepatic cholesterol biosynthesis, but without further affecting adrenal and muscle function. Other therapies such as substitution with bovine serum HDL-C are currently investigated to improve altered cholesterol availability during sepsis. Presentation: 6/2/2024

Stocking density affects growth, feed utilisation, metabolism, welfare and associated mRNA transcripts in liver and muscle of rainbow trout more pronouncedly than dietary fish meal inclusion level

In aquaculture, stocking density and diet composition are critical factors that influence the performance and welfare of the farmed animal. However, there is limited information on their interactive effects. In this context, applying a 2 × 2 factorial design, we conducted a 12-week feeding trial to evaluate the effect of stocking density (low, LSD: 3.3–15 kg m−3 and high, HSD: 5.8–25 kg m−3; initial-final values) and dietary fish meal content (high, HFM: 30 % and low, LFM: 15 %) on growth, feed utilisation, metabolic oxygen consumption, tissue indices, carcass composition, plasma metabolites, and transcriptional regulation of growth, metabolism and welfare-related biomarker mRNAs in liver and muscle of the juvenile rainbow trout. At the whole-animal level, the HSD group showed significantly higher routine metabolic rate and feed intake indicating higher energy demand; but growth and feed efficiency was lower in HSD than the LSD group. Morpho-anatomically, HSD fish had lower relative gut length and higher intra-peritoneal fat index than LSD fish. Lower plasma free amino acids and glucose levels reiterated high energy expenditure in HSD, and elevated albumin levels indicated potential oxidative stress in the HSD group. Correspondingly, at the transcriptional level, hepatic mRNA levels of antioxidative enzymes (sod, cat, gst, g6pd) and stress-related molecular chaperone hsp90 were up-regulated in HSD; while the transcript abundance of growth-axis markers (ghr, igf1 igf2) and nutrient-sensor mtor were decreased. In muscle, proteolytic pathway components fbx32 and murf1 were upregulated in HSD, and concurrently mtor was also upregulated suggesting higher protein turnover in this group. With respect to the effect of dietary fish meal inclusion levels, LFM showed decrease in growth, hepatic igf2 expression, and lower plasma triglycerides and free amino acid levels than HFM group. However, the sensory appearance of flesh was found to be slightly better in LFM. Significant diet-husbandry interactions were not observed phenotypically, but the mRNA abundance of hepatic markers (ampk1, hsp70, mhc1 and tnfa) linked to energy-sensing, stress and immune response were differentially regulated. Overall, the study shows that high stocking density has a greater degree of impact on fish growth and welfare than reduction in dietary fishmeal inclusion, with limited interactional effects.

Hepatic retinol dehydrogenase 11 dampens stress associated with the maintenance of cellular cholesterol levels

ObjectiveDysregulation of hepatic cholesterol metabolism can contribute to elevated circulating cholesterol levels, which is a significant risk factor for cardiovascular disease. Cholesterol homeostasis in mammalian cells is tightly regulated by an integrated network of transcriptional and post-transcriptional signalling pathways. Whilst prior studies have identified many of the central regulators of these pathways, the extended supporting networks remain to be fully elucidated. MethodsHere, we leveraged an integrated discovery platform, combining multi-omics data from 107 strains of mice to investigate these supporting networks. We identified retinol dehydrogenase 11 (RDH11; also known as SCALD) as a novel protein associated with cholesterol metabolism. Prior studies have suggested that RDH11 may be regulated by alterations in cellular cholesterol status, but its specific roles in this pathway are mostly unknown. ResultsHere, we show that mice fed a Western diet (high fat, high cholesterol) exhibited a significant reduction in hepatic Rdh11 mRNA expression. Conversely, mice treated with a statin (3-hydroxy-3-methyl-glutaryl-coenzyme A reductase (HMGCR) inhibitor) exhibited a 2-fold increase in hepatic Rdh11 mRNA expression. Studies in human and mouse hepatocytes demonstrated that RDH11 expression was regulated by altered cellular cholesterol conditions in a manner consistent with SREBP2 target genes HMGCR and LDLR. Modulation of RDH11 in vitro and in vivo demonstrated modulation of pathways associated with cholesterol metabolism, inflammation and cellular stress. Finally, RDH11 silencing in mouse liver was associated with a reduction in hepatic cardiolipin abundance and a concomitant reduction in the abundance of proteins of the mitochondrial electron transport chain. ConclusionTaken together, these findings suggest that RDH11 likely plays a role in protecting cells against the cellular toxicity that can arise as a by-product of endogenous cellular cholesterol synthesis.

Normalization of Fetal Cerebral and Hepatic Iron by Parental Iron Therapy to Pregnant Rats with Systemic Iron Deficiency without Anemia.

Iron (Fe) is a co-factor for enzymes of the developing brain necessitating sufficient supply. We investigated the effects of administering ferric derisomaltose/Fe isomaltoside (FDI) subcutaneously to Fe-deficient (ID) pregnant rats on cerebral and hepatic concentrations of essential metals and the expression of iron-relevant genes. Pregnant rats subjected to ID were injected with FDI on the day of mating (E0), 14 days into pregnancy (E14), or the day of birth (postnatal (P0)). The efficacy was evaluated by determination of cerebral and hepatic Fe, copper (Cu), and zinc (Zn) and gene expression of ferroportin, hepcidin, and ferritin H + L in pups on P0 and as adults on P70. Females fed an ID diet (5.2 mg/kg Fe) had offspring with significantly lower cerebral and hepatic Fe compared to female controls fed a standard diet (158 mg/kg Fe). Cerebral Cu increased irrespective of supplying a standard diet or administering FDI combined with the standard diet. Hepatic hepcidin mRNA was significantly lower following ID. Cerebral hepcidin mRNA was hardly detectable irrespective of iron status. In conclusion, administering FDI subcutaneously to ID pregnant rats on E0 normalizes fetal cerebral and hepatic Fe. When applied at later gestational ages, supplementation with additional Fe to the offspring is needed to normalize cerebral and hepatic Fe.

Parenteral nutrition results in peripheral ileal to hepatic circadian discordance in mice.

We have developed a mouse model of parenteral nutrition-associated liver disease (PNALD) in which parenteral nutrition (PN) infusion results in cholestatic liver injury. In the liver, the master circadian genes Arntl/Bmal drive rhythmic gene expression and regulate circadian expression of hepatic functions including bile acid synthesis. The aim of this study was to examine the effect of continuous PN on ileal and hepatic expression of circadian regulatory (CR) genes, farnesoid X receptor (FXR) signaling, and bile acid synthesis in mice. Wild-type mice were exposed to ad libitum Chow or continuous soy oil lipid emulsion-based PN infusion through a central venous catheter for 4 days (PN). Water was provided ad libitum, but no nutrients were provided enterally. On day 4, separate groups of Chow and PN-fed mice were euthanized every 6 h (7 AM, 1 PM, 7 PM, and 1 AM), and ileal, hepatic tissue and serum harvested. From tissue samples, the relative expression of circadian transcription factors and FXR signaling was assessed. Administration of 4-day PN increased hepatic injury, inflammatory cytokine expression, and gut permeability. In the ileum, PN activated FXR and induced expression of Fgf15 and Nr0b2. In the liver, expression of FXR-downstream targets was dysregulated. PN administrations impacted hepatic and ileal circadian transcription factor mRNA expression, which was discordant between the two organs. Dysregulation of circadian regulatory machinery is in part due to discordance of the gut-liver axis during PN. Pharmacological targeting of CR as a therapeutic strategy for PNALD thus deserves further investigation.NEW & NOTEWORTHY This study used a novel short-term model of parenteral nutrition (PN) that is translationally relevant. We find that short-term PN is sufficient to induce hepatic and ileal changes in circadian transcription factor expression and to prevent normal concordant coordination of circadian transcription factors between the ileum and liver. These data suggest that targeting circadian transcription may have some clinical benefit in patients receiving parenteral nutrition.

Study on the mechanism of Shenling Baizhu powder on the pathogenesis of pregnancy complicated with non-alcoholic fatty liver, based on PI3K/AKT/mTOR signal pathway.

This study investigates the effectiveness of Shenlin Baizhu powder in managing non-alcoholic fatty liver disease (NAFLD) during pregnancy and its mechanism through the PI3K/AKT/mTOR signaling pathway. Eight healthy male and 24 female Sprague-Dawley rats were used. After acclimatization, 6 female rats were fed normal chow, and 18 female rats were fed high-fat chow to induce NAFLD. After 8 weeks, female rats were mated with males to create a pregnant NAFLD model. The rats were divided into four groups: normal feeding, high-fat diet with saline, high-fat diet with 1.6 g/kg Shenlin Baizhu powder, and high-fat diet with 4.8 g/kg Shenlin Baizhu powder. Maternal body weight, serum and liver levels of aspartate aminotransferase (AST), alanine transaminase (ALT), triglyceride (TG), total cholesterol (TC), low density lipoprotein cholesterol (LDL-C), high density lipoprotein cholesterol (HDL-C), related inflammatory indexes interleukin-1 β (IL-1 β), tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) were measured. Liver tissue was examined using hematoxylin and oil red O staining, and protein expression related to the PI3K/AKT/mTOR pathway was assessed via Western blotting, immunohistochemistry and RT-PCR. Results showed significant weight gain and increases in ALT, AST, TG, TC, LDL-C, IL-1β, TNF-α, and IL-6, along with decreased HDL-C in NAFLD rats compared to controls. The high and low-dose Shenlin Baizhu powder groups exhibited improvements in body weight, liver histopathology, and reductions in serum TG, TC, LDL-C, ALT, AST, IL-1β, TNF-α, and IL-6, with increased HDL-C levels. Notably, the high-dose group showed greater efficacy in reducing hepatic fat accumulation, liver function markers, blood lipids, and inflammatory indexes, and decreased expression of hepatic PPARγ mRNA, SREBP1 mRNA, AKT mRNA, and related proteins. Shenlin Baizhu powder demonstrates potential in ameliorating high-fat diet-induced NAFLD in pregnant rats, likely through modulation of the PI3K/AKT/mTOR pathway, suggesting its therapeutic potential for gestational NAFLD.

197 Hepatic mRNA expression of innate and adaptive immune genes in beef steers with divergent residual body weight gain

Abstract Immune function has a pivotal role in dictating the overall health and productivity of cattle. In a proficient immune system, the liver assumes an integral function in detoxification and metabolic processes and contributes substantially to overall production and immunity. In this study, we evaluated the hepatic mRNA expression of genes involved in innate and adaptive immunity in crossbred beef steers with positive or negative residual body weight (BW) gain (RADG). Positive-RADG beef steers (n = 8; RADG = 0.76 kg/d) and negative-RADG beef steers (n = 8; RADG = -0.64 kg/d) were identified from a group of 108 growing crossbred beef steers (average BW = 286 ± 380kg) after a 56-d performance testing period. At the end of the 56-d period, liver tissue samples were collected from the beef steers for RNA extraction and cDNA synthesis. The mRNA expression of 84 genes involved in innate and adaptive immunity were analyzed using pathway-focused PCR-based arrays. The mRNA expression of genes with false discovery rate-adjusted P-values (FDR) ≤ 0.05 and absolute fold change (FC) ≥ 1.2 were determined to be differentially expressed. Out of the 84 genes analyzed, four genes [interleukin (IL)-2, MYD88, CD-80, NFkB-1] were differentially expressed and were all upregulated in positive compared with negative-RADG beef steers. Gene ontology pathway enrichment analysis of the differentially expressed genes revealed that pathways related to positive regulation of IL-17 production, cellular response to lipopolysaccharide, and positive regulation of lymphocyte activation were enriched in the positive-RADG steers (P ≤ 0.05). In conclusion, our study revealed that positive-RADG beef steers had increased expression of genes and pathways involved in pathogen recognition and initiation of responses to effectively fight off infections while mitigating excess inflammatory reactions compared with negative-RADG steers, which might explain, in part, the improved feed efficiency of these beef animals

Carthamus tinctorius L. (Safflower) Flower Extract Attenuates Hepatic Injury and Steatosis in a Rat Model of Type 2 Diabetes Mellitus via Nrf2-Dependent Hypoglycemic, Antioxidant, and Hypolipidemic Effects.

This study aimed to examine the hepatic and anti-steatotic protective effects of methanolic extract from Carthamus tinctorius (safflower) flowers (SFFE), using a rat model of type 2 diabetes mellitus (T2DM), and to examine the molecular mechanisms underlying these effects. Adult male Wistar rats were used for this study. First, T2DM was induced in some rats by feeding them a high-fat diet (HFD) for 4 weeks, followed by a single dose of streptozotocin (STZ) (35 mg/kg, i.p.). Experimental groups included the following five groups (n = 8 in each): control, control + SFFE, T2DM, T2DM + SFFE, and T2DM + SFFE + brusatol (an Nrf2 inhibitor, 2 mg/kg, i.p.). SFFE was administered at a concentration of 300 mg/kg, and all experiments concluded after 8 weeks. Treatments with SFFE significantly reduced fasting blood glucose levels, free fatty acids (FFAs), cholesterol, triglycerides, and low-density lipoprotein cholesterol in both the control and T2DM rats, but they failed to reduce fasting insulin levels in these groups. SFFE treatments also improved the liver structure and reduced hepatocyte vacuolization and hepatic levels of triglycerides and cholesterol in T2DM rats, in addition to increasing the hepatic mRNA levels of keap1 and the cytoplasmic levels and nuclear activities of Nrf2 in both the control and T2DM rats. SFFE also stimulated the expression levels of PPARα and CPT-1 but reduced the malondialdehyde (MDA), mRNA levels of SREBP1, fatty acid synthase, and acetyl CoA carboxylase in both the control and T2DM rats; meanwhile, it reduced hepatic mRNA and the nuclear activities of NF-κB and increased levels of glutathione, superoxide dismutase, and heme oxygenase-1 in the livers of both groups of treated rats. Furthermore, SFFE suppressed the levels of caspase-3, Bax, tumor necrosis factor-α, and interleukin-6 in the T2DM rats. Treatment with brusatol prevented all of these effects of SFFE. In conclusion, SFFE suppresses liver damage and hepatic steatosis in T2DM through Nrf2-dependent hypoglycemic, antioxidant, anti-inflammatory, and hypolipidemic effects.

Organosulfur Compounds, S-Allyl-L-Cysteine and S-Ethyl-L-Cysteine, Target PCSK-9/LDL-R-Axis to Ameliorate Cardiovascular, Hepatic, and Metabolic Changes in High Carbohydrate and High Fat Diet-Induced Metabolic Syndrome in Rats.

Metabolic syndrome (MetS) is an ever-evolving set of diseases that poses a serious health risk in many countries worldwide. Existing evidence illustrates that individuals with MetS have a 30%-40% higher chance of acquiring type 2 diabetes mellitus (T2DM), cardiovascular disease (CVD), or both. This study was undertaken to uncover the regulatory role of natural organosulfur compounds (OSCs), S-allyl-L-cysteine (SAC), and S-ethyl-L-cysteine (SEC), in targeting high carbohydrate high fat (HCHF)-diet-induced MetS-associated risk management. Our findings suggested that SAC and SEC ameliorated HCHF-diet-induced diabetic profiles, plasma lipid and lipoprotein level, liver function, oxidative-stress, inflammatory cytokines, and chemokines including monocyte chemoattractant protein-1 (MCP-1), lipid peroxidation, plasma proprotein convertase subtilisin/kexin type-9 (PCSK-9), and high-sensitivity C-reactive protein (hs-CRP). Moreover, the assessment of the hepatic mRNA expression of the key genes involved in cholesterol homeostasis depicted that SAC and SEC downregulated the PCSK-9 mRNA expression via targeting the expression of HNF-1α, a transcriptional activator of PCSK-9. On the other hand, the LDL-receptor (LDL-R) expression was upregulated through the activation of its transcriptional regulator sterol regulatory element binding protein-2 (SREBP-2). In addition, the activity and the mRNA expression of 3-hydroxy-3-methylglutaryl coenzyme-A reductases (HMG-R) and peroxisome proliferator-activated receptors (PPARs) were also improved by the treatment of SAC and SEC. We concluded that SAC and SEC can protect against MetS via improving the lipid and lipoprotein content, glycemic indices, hepatic function, targeting the inflammatory cascades, and oxidative imbalance, regulation of the mRNA expression of PCSK-9, LDL-R, SREBP-2, HNF-1α, PPARs, and inflammatory biomarkers.

Ontogeny of Hepatic Organic Cation Transporter-1 in Rat and Human.

The organic cation transporter (OCT)-1 mediates hepatic uptake of cationic endogenous compounds and xenobiotics. To date, limited information exists on how Oct1/OCT1 functionally develops with age in rat and human livers and how this would affect the pharmacokinetics of OCT substrates in children or juvenile animals. The functional ontogeny of rOct/hOCT was profiled in suspended rat (2-57 days old) and human hepatocytes (pediatric liver tissue donors: age 2-12 months) by determining uptake clearance of 4-[4-(dimethylamino)styryl]-N-methylpyridinium iodide (ASP+) as a known rOct/hOCT probe substrate. mRNA expression was determined in rat liver tissue corresponding to rat ages used in the functional studies, while hOCT1 mRNA expressions were determined in the same hepatocyte batches as those used for uptake studies. Maturation of rOct/hOCT activity and expression were evaluated by comparing values obtained at the various ages to the adult values. Relative to adult values (at 8 weeks), ASP+ uptake clearance in suspended rat hepatocytes aged 0, 1, 2, 3, 4, 5, and 6 weeks reached 26%, 29%, 33%, 37%, 72%, 63%, and 71%, respectively. Hepatic Oct1 mRNA expression was consistent with Oct activity (correlation coefficient of 0.92). In human hepatocytes, OCT1 activity was age dependent and also correlated with mRNA levels (correlation coefficient of 0.88). These data show that Oct1/OCT1 activities and expression mature gradually in rat/human liver, thereby mirroring the expression pattern of organic anion transporting polypeptide in rat. These high-resolution transporter ontogeny profiles will allow for more accurate prediction of the pharmacokinetics of OCT1/Oct1 substrates in pediatric populations and juvenile animals. SIGNIFICANCE STATEMENT: Organic cation transporter-1 (OCT1) represents a major drug uptake transporter in human liver. This study provides high-resolution data regarding the age-dependent function of OCT1 in the liver, based on in vitro experiments with rat and human hepatocytes obtained from donors between birth and adulthood. These ontogeny profiles will inform improved age-specific physiologically based pharmacokinetic models for OCT1 drug substrates in neonates, infants, children, and adults.

Phlorotannin Alleviates Liver Injury by Regulating Redox Balance, Apoptosis, and Ferroptosis of Broilers under Heat Stress.

Heat stress (HS) poses a great challenge to the poultry industry by inducing oxidative damage to the liver, endangering the health and production of broilers. As an important type of seaweed polyphenols, phlorotannin has been shown to have antioxidant properties. The present study evaluated the protective effects of dietary phlorotannin on HS-induced liver injury in broilers based on oxidative damage parameters. A total of 108 twenty-one days old male Arbor Acres plus (AA+) broilers were randomly divided into three groups: TN group (thermoneutral, 24 ± 1 °C, fed with basal diet), HS group (HS, 33 ± 1 °C for 8 h/day, fed with basal diet), and HS + phlorotannin group (HS + 600 mg/kg phlorotannin). Each group has six replicate cages with six birds per cage. The feeding experiment lasted 21 days. At the termination of the feeding experiment (42 days old), samples were collected for analysis of morphological and biochemical features. The results showed that HS decreased the liver index, serum albumin (ALB) content, hepatic antioxidant enzymes activities of catalase (CAT), total superoxide dismutase (T-SOD), glutathione S-transferase (GST), and glutathione peroxidase (GSH-Px) (p < 0.05), while increasing the hepatic histopathology score, apoptosis rate, and malondialdehyde (MDA) content (p < 0.05) in 42-day-old broilers. Compared with the HS group, dietary phlorotannin improved the activities of antioxidant enzymes (GST and GSH-Px) but decreased the histopathology score and apoptosis rate in the liver (p < 0.05). Moreover, HS down-regulated hepatic mRNA expression of CAT1, NQO1, HO-1, and SLC7A11 (p < 0.05), while up-regulated hepatic mRNA expression of Keap1, MafG, IκBα, NF-κB P65, IFN-γ, TFR1, ACSL4, Bax, and Caspase-9 (p < 0.05). Compared with HS group, dietary phlorotannin up-regulated hepatic mRNA expression of Nrf2, CAT1, MafF, GSTT1, NQO1, HO-1, GCLC, GPX1, TNF-α, Fpn1, and SLC7A11 (p < 0.05), while down-regulated hepatic mRNA expression of IκBα, Bax, Caspase-9, and TFR1 (p < 0.05). In conclusion, dietary supplementation of 600 mg/kg phlorotannin could alleviate HS-induced liver injury via regulating oxidative status, apoptosis, and ferroptosis in broilers; these roles of phlorotannin might be associated with the regulation of the Nrf2 signaling pathway.

Evaluation of Bisphenol S (BPS) toxicity on the reproductive system of Channa striatus: Insights for environmental risk assessment

Aquatic ecosystems face significant exposure to endocrine-disrupting chemicals (EDCs), which can mimic, block, or alter the synthesis of endogenous hormones. Bisphenol A (BPA), a widely known EDC, has been phased out from consumer products due to concerns about its potential impacts on human health. In its place, bisphenol S (BPS), an organic compound, has been increasingly used in the production of polycarbonate plastics, epoxy resins, thermal receipt papers, and currency. Vitellogenin (Vtg), a yolk precursor protein synthesized in the liver and present in oviparous fish, particularly males, serves as a pertinent biomarker for studying the effects of estrogenic EDCs on fish. This study aimed to assess the impact of BPS on reproductive parameters and hepatic vitellogenin expression in Channa striatus. The LC50 of BPS was determined to be 128.8 mg/L. Experimental groups included control and BPS-exposed fish, with sub-lethal concentrations of BPS (1 mg/L, 4 mg/L, and 12 mg/L) administered and effects monitored at seven- and twenty-one-day intervals. Significant decreases in gonadosomatic index (GSI), ova diameter, and fecundity were observed in BPS-exposed Channa striatus. Hepatic Vtg mRNA expression was downregulated in female and upregulated in male following BPS exposure. Serum hormone analysis confirmed the estrogenic activity of BPS. These findings underscore BPS's ability as an endocrine disruptor to interfere with hormone synthesis and disrupt spermatogenesis and oogenesis processes in Channa striatus. This research contributes to understanding the endocrine-disrupting effects of BPS on aquatic organisms, highlighting potential ecological implications and the need for continued monitoring and regulatory considerations.

FNDC4 reduces hepatocyte inflammatory cell death via AMPKα in metabolic dysfunction-associated steatotic liver disease

The molecular mediators responsible for the progression of metabolic dysfunction-associated steatotic liver disease (MASLD) to steatohepatitis (MASH) have not yet been completely disentangled. We sought to analyze whether FNDC4, an hepatokine and adipokine with anti-inflammatory properties, is involved in TNF-α-induced inflammatory cell death in patients with MASLD. Plasma FNDC4 (n=168) and hepatic FNDC4 and inflammatory cell death (n=65) were measured in samples from patients with severe obesity with available liver biopsy-proven MASLD diagnosis. The effect of FNDC4 on TNF-α-induced pyroptosis, apoptosis and necroptosis (PANoptosis) and mitochondrial dysfunction was studied invitro using human HepG2 hepatocytes. Compared with individuals with normal liver, patients with type 2 diabetes and MASLD exhibited decreased hepatic FNDC4 mRNA and protein levels, which were related to liver inflammation. An overexpression of TNF-α, its receptor TNF-R1 and factors involved in inflammatory cell death was also found in the liver of these patients. FNDC4-knockdown in HepG2 hepatocytes increased apoptotic cell death, while FNDC4 treatment blunted NLRP3 inflammasome-induced pyroptosis, apoptosis and necroptosis in TNF-α-stimulated hepatocytes. Moreover, FNDC4 improved TNF-α-induced hepatocyte mitochondrial dysfunction by enhancing mitochondrial DNA (mtDNA) copy number and OXPHOS complex subunits I, II, III and V protein expression. Mechanistically, AMP-activated protein kinase α (AMPKα) was required for the FNDC4-mediated inhibition of cell death and increase in mtDNA content. FNDC4 acts as a hepatocyte survival factor favouring mitochondrial homeostasis and decreasing inflammatory cell death via AMPKα. Collectively, our study identifies FNDC4 as an attractive target to prevent hepatocellular damage in patients with MASLD.

RNA interference therapy in cardiology: will new targets improve therapeutic goals?

The discovery of RNA interference in 1998 opened avenues for the manipulation of gene expression, leading to the development of small interfering RNA (siRNA) drugs. Patisiran, the first FDA-approved siRNA medication, targets hereditary transthyretin amyloidosis with polyneuropathy. Givosiran, lumasiran and nedosiran further expand siRNA applications in treating rare genetic diseases, demonstrating positive outcomes. In cardiology, inclisiran, approved for hypercholesterolaemia, showcases sustained reductions in LDL cholesterol levels. However, ongoing research aims to establish its impact on cardiovascular outcomes. Lipoprotein(a), an independent risk factor for atherosclerotic cardiovascular disease, has become a focus of siRNA therapies, precipitating the development of specific siRNA drugs like olpasiran, zerlasiran and lepodisiran, with promising reductions in lipoprotein(a) levels. Research to assess the effectiveness of these medications in reducing events is currently under way. Zodasiran and plozasiran address potential risk factors for cardiovascular diseases, targeting triglyceride-rich lipoproteins. Zilebesiran, which targets hepatic angiotensinogen mRNA, has demonstrated a dose-related reduction in serum angiotensinogen levels, thereby lowering blood pressure in patients with systemic arterial hypertension. The evolving siRNA methodology presents a promising future in cardiology, with ongoing studies assessing its effectiveness in various conditions. In the future, larger studies will provide insights into improvements in cardiovascular outcomes, long-term safety and broader applications in the general population. This review highlights the historical timeline of the development of siRNA-based drugs, their clinical indications, potential side-effects and future perspectives.