Protective Effects of Hepatocyte Stress Defenders, Nrf1 and Nrf2, against MASLD Progression.

Progression of metabolic dysfunction-associated steatites liver disease (MASLD) to steatohepatitis (MASH) is driven by stress-inducing lipids that promote liver inflammation and fibrosis, and MASH can lead to cirrhosis and hepatocellular carcinoma. Previously, we showed coordinated defenses regulated by transcription factors, nuclear factor erythroid 2-related factor-1 (Nrf1) and -2 (Nrf2), protect against hepatic lipid stress. Here, we investigated protective effects of hepatocyte Nrf1 and Nrf2 against MASH-linked liver fibrosis and tumorigenesis. Male and female mice with flox alleles for genes encoding Nrf1 (Nfe2l1), Nrf2 (Nfe2l2), or both were fed a MASH-inducing diet enriched with high fat, fructose, and cholesterol (HFFC) or a control diet for 24-52 weeks. During this period, hepatocyte Nrf1, Nrf2, or combined deficiency for ~7 days, ~7 weeks, and ~35 weeks was induced by administering mice hepatocyte-targeting adeno-associated virus (AAV) expressing Cre recombinase. The effects on MASH, markers of liver fibrosis and proliferation, and liver tumorigenesis were compared to control mice receiving AAV-expressing green fluorescent protein. Also, to assess the impact of Nrf1 and Nrf2 induction on liver fibrosis, HFFC diet-fed C57bl/6J mice received weekly injections of carbon tetrachloride, and from week 16 to 24, mice were treated with the Nrf2-activating drug bardoxolone, hepatocyte overexpression of human NRF1 (hNRF1), or both, and these groups were compared to control. Compared to the control diet, 24-week feeding with the HFFC diet increased bodyweight as well as liver weight, steatosis, and inflammation. It also increased hepatocyte proliferation and a marker of liver damage, p62. Hepatocyte Nrf1 and combined deficiency increased liver steatosis in control diet-fed but not HFFC diet-fed mice, and increased liver inflammation under both diet conditions. Hepatocyte Nrf1 deficiency also increased hepatocyte proliferation, whereas combined deficiency did not, and this also occurred for p62 level in control diet-fed conditions. In 52-week HFFC diet-fed mice, 35 weeks of hepatocyte Nrf1 deficiency, but not combined deficiency, resulted in more liver tumors in male mice, but not in female mice. In contrast, hepatocyte Nrf2 deficiency had no effect on any of these parameters. However, in the 15-week CCL4-exposed and 24-week HFFC diet-fed mice, Nrf2 induction with bardoxolone reduced liver steatosis, inflammation, fibrosis, and proliferation. Induction of hepatic Nrf1 activity with hNRF1 enhanced the effect of bardoxolone on steatosis and may have stimulated liver progenitor cells. Physiologic Nrf1 delays MASLD progression, Nrf2 induction alleviates MASH, and combined enhancement synergistically protects against steatosis and may facilitate liver repair.

Irciniaplysins A-D: New Psammaplysin Derivatives from Philippine Marine Sponge Ircinia sp.

Four new psammaplysin derivatives (1-4) with fatty acyl substituents, designated irciniaplysins A-D, and three known psammaplysins (5-7) were isolated from a marine sponge Ircinia sp. Their structures were elucidated using extensive spectroscopic analyses. The positions of the double bonds and the branch points of the fatty acyl side chains were determined by GC-MS analysis of their fatty acid methyl ester (FAME) derivatives. Irciniaplysins A (1) and B (2) contained an unusual long-chain fatty acyl substituent with a 5,9-diene unit. The isolated compounds were evaluated for their cytotoxic activity against the human colorectal carcinoma (HCT 116) cells, however, none of these compounds showed significant activity.

Sterol O-acyltransferase (SOAT/ACAT) activity is required to form cholesterol crystals in hepatocyte lipid droplets.

OBJECTIVE: Excess cholesterol storage can induce the formation of cholesterol crystals in hepatocyte lipid droplets. Such crystals distinguish metabolic dysfunction associated steatohepatitis (MASH) from simple steatosis and may underlie its pathogenesis by causing cell damage that triggers liver inflammation. The mechanism linking cholesterol excess to its crystallization in lipid droplets is unclear. As cholesteryl esters localize to and accumulate in lipid droplets more readily than unesterified free cholesterol, we investigated whether cholesterol esterification by sterol O-acyltransferase (SOAT), also known as acyl co-A cholesterol acyltransferase (ACAT), is required for hepatocyte lipid droplet crystal formation. METHOD: Cholesterol crystals were measured in cholesterol loaded Hep3B hepatocytes, RAW264.7 macrophages, and mouse liver using polarizing light microscopy. We examined the effect of blocking SOAT activity on crystal formation and compared these results to features of cholesterol metabolism and the progression to intracellular crystal deposits. RESULTS: Cholesterol loading of Hep3B cells caused robust levels of lipid droplet localized crystal formation in a dose- and time-dependent manner. Co-treatment with SOAT inhibitors and genetic ablation of SOAT1 blocked crystal formation. SOAT inhibitor also blocked crystal formation in low density lipoprotein (LDL) treated Hep3B cells, acetylated LDL treated RAW 264.7 macrophages, and in the liver of mice genetically predisposed to hepatic cholesterol overload and in mice with cholesterol enriched diet-induced MASH. CONCLUSION: SOAT1-mediated esterification may underlie cholesterol crystals associated with MASH by concentrating it in lipid droplets. These findings imply that inhibiting hepatocyte SOAT1 may be able to alleviate cholesterol associated MASH. Moreover, that either a lipid droplet localized cholesteryl ester hydrolase is required for cholesterol crystal formation, or the crystals are composed of cholesteryl ester.

The complex polyploid genome architecture of sugarcane.

Sugarcane, the world's most harvested crop by tonnage, has shaped global history, trade and geopolitics, and is currently responsible for 80% of sugar production worldwide1. While traditional sugarcane breeding methods have effectively generated cultivars adapted to new environments and pathogens, sugar yield improvements have recently plateaued2. The cessation of yield gains may be due to limited genetic diversity within breeding populations, long breeding cycles and the complexity of its genome, the latter preventing breeders from taking advantage of the recent explosion of whole-genome sequencing that has benefited many other crops. Thus, modern sugarcane hybrids are the last remaining major crop without a reference-quality genome. Here we take a major step towards advancing sugarcane biotechnology by generating a polyploid reference genome for R570, a typical modern cultivar derived from interspecific hybridization between the domesticated species (Saccharum officinarum) and the wild species (Saccharum spontaneum). In contrast to the existing single haplotype ('monoploid') representation of R570, our 8.7 billion base assembly contains a complete representation of unique DNA sequences across the approximately 12 chromosome copies in this polyploid genome. Using this highly contiguous genome assembly, we filled a previously unsized gap within an R570 physical genetic map to describe the likely causal genes underlying the single-copy Bru1 brown rust resistance locus. This polyploid genome assembly with fine-grain descriptions of genome architecture and molecular targets for biotechnology will help accelerate molecular and transgenic breeding and adaptation of sugarcane to future environmental conditions.

HDL functionality is dependent on hepatocyte stress defense factors Nrf1 and Nrf2.

High density lipoproteins (HDL) promote homeostasis and counteract stressful tissue damage that underlie cardiovascular and other diseases by mediating reverse cholesterol transport, reducing inflammation, and abrogating oxidative damage. However, metabolically stressful conditions associated with atherosclerosis can impair these effects. Hepatocytes play a major role in the genesis and maturation of circulating HDL, and liver stress elicits marked regulatory changes to circulating HDL abundance and composition, which affect its functionality. The mechanisms linking liver stress to HDL function are incompletely understood. In this study, we sought to determine whether stress defending transcription factors nuclear factor erythroid 2 related factor-1 (Nrf1) and -2 (Nrf2) promote hepatocyte production of functional HDL. Using genetically engineered mice briefly fed a mild metabolically stressful diet, we investigated the effect of hepatocyte-specific deletion of Nrf1, Nrf2, or both on circulating HDL cholesterol, protein composition, and function. Combined deletion, but not single gene deletion, reduced HDL cholesterol and apolipoprotein A1 levels as well as the capacity of HDL to accept cholesterol undergoing efflux from cultured macrophages and to counteract tumor necrosis factor α-induced inflammatory effect on cultured endothelial cells. This coincided with substantial alteration to the HDL proteome, which correlated with liver gene expression profiles of corresponding proteins. Thus, our findings show complementary actions by hepatocyte Nrf1 and Nrf2 play a role in shaping HDL abundance and composition to promote production of functionally viable HDL. Consequently, our study illuminates the possibility that enhancing stress defense programming in the liver may improve atheroprotective and perhaps other health promoting actions of HDL.

Euglycemia is affected by stress defense factor hepatocyte NRF1, but not NRF2.

Hepatocyte stress signaling has been established to alter glucose metabolism and impair systemic glucose homeostasis. In contrast, the role of stress defenses in the control of glucose homeostasis is less understood. Nuclear factor erythroid 2 related factor-1 (NRF1) and -2 (NRF2) are transcription factors that promote stress defense and can exert hepatocyte stress defense programming via complementary gene regulation. To identify whether there are independent or complementary roles of these factors in hepatocytes on glucose homeostasis, we investigated the effect of adult-onset, hepatocyte-specific deletion of NRF1, NRF2, or both on glycemia in mice fed 1-3 weeks with a mildly stressful diet enriched with fat, fructose, and cholesterol. Compared to respective control, NRF1 deficiency and combined deficiency reduced glycemia, in some cases resulting in hypoglycemia, whereas there was no effect of NRF2 deficiency. However, reduced glycemia in NRF1 deficiency did not occur in the leptin-deficient mouse model of obesity and diabetes, suggesting hepatocyte NRF1 support defenses that counteract hypoglycemia but does not promote hyperglycemia. Consistent with this, NRF1 deficiency was associated with reduced liver glycogen and glycogen synthase expression as well as marked alteration to circulating level of glycemia-influencing hormones, growth hormone and insulin-like growth factor-1 (IGF1). Overall, we identify a role for hepatocyte NRF1 in modulating glucose homeostasis, which may be linked to liver glycogen storage and the growth hormone/IGF1 axis.

Complementary gene regulation by NRF1 and NRF2 protects against hepatic cholesterol overload.

Hepatic cholesterol overload promotes steatohepatitis. Insufficient understanding of liver stress defense impedes therapy development. Here, we elucidate the role of stress defense transcription factors, nuclear factor erythroid 2 related factor-1 (NRF1) and -2 (NRF2), in counteracting cholesterol-linked liver stress. Using a diet that increases liver cholesterol storage, expression profiles and phenotypes of liver from mice with hepatocyte deficiency of NRF1, NRF2, or both are compared with controls, and chromatin immunoprecipitation sequencing is undertaken to identify target genes. Results show NRF1 and NRF2 co-regulate genes that eliminate cholesterol and mitigate inflammation and oxidative damage. Combined deficiency, but not deficiency of either alone, results in severe steatohepatitis, hepatic cholesterol overload and crystallization, altered bile acid metabolism, and decreased biliary cholesterol. Moreover, therapeutic effects of NRF2-activating drug bardoxolone require NRF1 and are supplemented by NRF1 overexpression. Thus, we discover complementary gene programming by NRF1 and NRF2 that counteract cholesterol-associated fatty liver disease progression.

Neural responses to pup calls and pup odors in California mouse fathers and virgin males.

The onset of mammalian maternal care is associated with plasticity in neural processing of infant-related sensory stimuli; however, little is known about sensory plasticity associated with fatherhood. We quantified behavioral and neural responses of virgin males and new fathers to olfactory and auditory stimuli from young, unfamiliar pups in the biparental California mouse (Peromyscus californicus). Each male was exposed for 10 min to one of four combinations of a chemosensory stimulus (pup-scented or unscented cotton [control]) and an auditory stimulus (pup vocalizations or white noise [control]). Behavior did not differ between fathers and virgins during exposure to sensory stimuli or during the following hour; however, males in both groups were more active both during and after exposure to pup-related stimuli compared to control stimuli. Fathers had lower expression of Fos in the main olfactory bulbs (MOB) but higher expression in the medial preoptic area (MPOA) and bed nucleus of the stria terminalis medial division, ventral part (STMV) compared to virgins. Lastly, males had higher Fos expression in MPOA when exposed to pup odor compared to control stimuli, and when exposed to pup odor and pup calls compared to pup calls only or control stimuli. These findings suggest that the onset of fatherhood alters activity of MOB, MPOA and STMV and that pup odors and vocalizations have additive or synergistic effects on males' behavior and MPOA activation.

Immunometabolic factors contributing to obesity-linked hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is a major public health concern that is promoted by obesity and associated liver complications. Onset and progression of HCC in obesity is a multifactorial process involving complex interactions between the metabolic and immune system, in which chronic liver damage resulting from metabolic and inflammatory insults trigger carcinogenesis-promoting gene mutations and tumor metabolism. Moreover, cell growth and proliferation of the cancerous cell, after initiation, requires interactions between various immunological and metabolic pathways that provide stress defense of the cancer cell as well as strategic cell death escape mechanisms. The heterogenic nature of HCC in addition to the various metabolic risk factors underlying HCC development have led researchers to focus on examining metabolic pathways that may contribute to HCC development. In obesity-linked HCC, oncogene-induced modifications and metabolic pathways have been identified to support anabolic demands of the growing HCC cells and combat the concomitant cell stress, coinciding with altered utilization of signaling pathways and metabolic fuels involved in glucose metabolism, macromolecule synthesis, stress defense, and redox homeostasis. In this review, we discuss metabolic insults that can underlie the transition from steatosis to steatohepatitis and from steatohepatitis to HCC as well as aberrantly regulated immunometabolic pathways that enable cancer cells to survive and proliferate in the tumor microenvironment. We also discuss therapeutic modalities targeted at HCC prevention and regression. A full understanding of HCC-associated immunometabolic changes in obesity may contribute to clinical treatments that effectively target cancer metabolism.

Modeling the conversion between specific IgE test platforms for nut allergens in children and adolescents.

BACKGROUND: Multiplex tests allow for measurement of allergen-specific IgE responses to multiple extracts and molecular allergens and have several advantages for large cohort studies. Due to significant methodological differences, test systems are difficult to integrate in meta-analyses/systematic reviews since there is a lack of datasets with direct comparison. We aimed to create models for statistical integration of allergen-specific IgE to peanut/tree nut allergens from three IgE test platforms. METHODS: Plasma from Canadian and Austrian children/adolescents with peanut/tree nut sensitization and a cohort of sensitized, high-risk, pre-school asthmatics (total n = 166) were measured with three R&D multiplex IgE test platforms: Allergy Explorer version 1 (ALEX) (Macro Array Dx), MeDALL-chip (Mechanisms of Development of Allergy) (Thermo Fisher), and EUROLINE (EUROIMMUN). Skin prick test (n = 51) and ImmunoCAP (Thermo Fisher) (n = 62) results for extracts were available in a subset. Regression models (Multivariate Adaptive Regression Splines, local polynomial regression) were applied if >30% of samples were positive to the allergen. Intra-test correlations between PR-10 and nsLTP allergens were assessed. RESULTS: Using two regression methods, we demonstrated the ability to model allergen-specific relationships with acceptable measures of fit (r2  = 94%-56%) for peanut and tree nut sIgE testing at the extract and molecular-level, in order from highest to lowest: Ara h 2, Ara h 6, Jug r 1, Ana o 3, Ara h 1, Jug r 2, and Cor a 9. CONCLUSION: Our models support the notion that quantitative conversion is possible between sIgE multiplex platforms for extracts and molecular allergens and may provide options to aggregate data for future meta-analysis.

Mir142 loss unlocks IDH2R140-dependent leukemogenesis through antagonistic regulation of HOX genes.

AML is a genetically heterogeneous disease and understanding how different co-occurring mutations cooperate to drive leukemogenesis will be crucial for improving diagnostic and therapeutic options for patients. MIR142 mutations have been recurrently detected in IDH-mutated AML samples. Here, we have used a mouse model to investigate the interaction between these two mutations and demonstrate a striking synergy between Mir142 loss-of-function and IDH2R140Q, with only recipients of double mutant cells succumbing to leukemia. Transcriptomic analysis of the non-leukemic single and leukemic double mutant progenitors, isolated from these mice, suggested a novel mechanism of cooperation whereby Mir142 loss-of-function counteracts aberrant silencing of Hoxa cluster genes by IDH2R140Q. Our analysis suggests that IDH2R140Q is an incoherent oncogene, with both positive and negative impacts on leukemogenesis, which requires the action of cooperating mutations to alleviate repression of Hoxa genes in order to advance to leukemia. This model, therefore, provides a compelling rationale for understanding how different mutations cooperate to drive leukemogenesis and the context-dependent effects of oncogenic mutations.

Persistent ventilation inhomogeneity after an acute exacerbation in preschool children with recurrent wheezing.

BACKGROUND: Preschool children with recurrent wheezing suffer high morbidity. It is unclear whether objective measures of asthma control, such as pulmonary function tests (PFTs), provide additional information to the clinical assessment. METHODS: We recruited children between 3 and 6 years old, with a history of recurrent wheezing in the preceding year and treated for acute wheezing exacerbation in the emergency department (ED) into an observational cohort study. Children attended two outpatient visits: the first study visit within five days of discharge from the ED and the second study visit 12 weeks after the ED visit. We performed standardized symptom score (test for respiratory and asthma control in kids (TRACK)), multiple breath washout (MBW), spirometry, and clinical assessment at both visits. RESULTS: Seventy-four children, mean (standard deviation (SD)) age of 4.32 years (0.84), attended both visits. Paired FEV0.75 and lung clearance index (LCI) measurements at both time points were obtained in 37 and 34 subjects, respectively. Feasibility for all tests improved at visit 2 and was not age-dependent. At the second study visit, a third had controlled asthma based on the TRACK score, and the mean lung clearance index (LCI) improved from 9.86 to 8.31 (P = .003); however, 46% had an LCI in the abnormal range. FEV0.75 z-score improved from -1.66 to -1.17 (P = .05) but remained in the abnormal range in 24%. LCI was abnormal in more than half of the children with "well-controlled" asthma based on the TRACK score. There was no correlation between PFT measures and TRACK scores at either visit. CONCLUSIONS: Lung clearance index demonstrates a persistent deficit post-exacerbation in a large proportion of preschoolers with recurrent wheezing, highlighting that symptom scores alone may not suffice for monitoring these children.

Effects of an entertaining, culturally targeted narrative and an appealing expert interview on the colorectal screening intentions of African American women.

Universal screening for colorectal cancer (CRC) is recommended for individuals 50-75 years of age, but screening uptake is suboptimal and African Americans have suffered persistent racial disparities in CRC incidence and deaths. We compared a culturally tailored fictional narrative and an engaging expert interview on the ability to increase intentions to be screened for CRC among African American women. In a post-only experiment, women (N = 442) in face-to-face listening groups in African American churches heard audio recordings of either a narrative or an expert interview. Questionnaires were completed immediately afterward and 30 days later. Women who heard narratives reported stronger intentions to be screened with a home stool blood test than women who heard the interview; the effect lasted at least 30 days. Culturally tailored, fictional narratives appear to be an effective persuasive strategy for reducing racial disparities in CRC outcomes.

Author Correction: Osmotic modulation of chromatin impacts on efficiency and kinetics of cell fate modulation.

A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has been fixed in the paper.

Osmotic modulation of chromatin impacts on efficiency and kinetics of cell fate modulation.

Chromatin structure is a major regulator of transcription and gene expression. Herein we explore the use of osmotic modulation to modify the chromatin structure and reprogram gene expression. In this study we use the extracellular osmotic pressure as a chromatin structure and transcriptional modulator. Hyposmotic modulation promotes chromatin loosening and induces changes in RNA polymerase II (Pol II) activity. The chromatin decondensation opens space for higher amounts of DNA engaged RNA Pol II. Hyposmotic modulation constitutes an alternative route to manipulate cell fate decisions. This technology was tested in model protocols of induced pluripotency and transdifferentiation in cells growing in suspension and adherent to substrates, CD34+ umbilical-cord-blood (UCB), fibroblasts and B-cells. The efficiency and kinetics of these cell fate modulation processes were improved by transient hyposmotic modulation of the cell environment.

Perturbed adipose tissue hydrogen peroxide metabolism in centrally obese men: Association with insulin resistance.

OBJECTIVE: Although adipose tissue hydrogen peroxide (H2O2) and its metabolizing enzymes have been linked to obesity and insulin resistance in animal studies, this relation remains to be evaluated in humans. METHODS: Non-diabetic men (N = 43, median age, 49 (37, 54 y)) undergoing abdominal surgeries were studied. Participants were classified by body mass index (BMI) into normal-weight (N = 19), or overweight/obese (Ow/Ob; BMI ≥25; N = 24). Centrally obese men were identified by waist-height ratio ≥0.5. H2O2 and activities of superoxide dismutase, catalase and glutathione peroxidase enzymes were assayed in subcutaneous fat samples, and visceral fat (available from N = 33), and their associations with anthropometric parameters, fasting serum lipids, and the homeostasis model of insulin resistance (HOMA-IR) were tested using correlations and multivariate linear regression. RESULTS: H2O2 concentrations and catalase activity were increased in visceral fat from Ow/Ob men, compared to normal-weight subjects (+32%, P = 0.038 and +51%, P = 0.043 respectively). Centrally obese subjects had >2-fold higher superoxide dismutase activity (P = 0.005), 46% higher H2O2 (P = 0.028), and 89% higher catalase activity (P = 0.009) in visceral fat, compared to lean subjects. Central obesity did not alter these markers in subcutaneous fat, apart from a 50% increase in catalase, and did not affect glutathione peroxidase in either fat depot. H2O2 in visceral fat positively correlated with insulin resistance (r = 0.40, P = 0.032). Catalase activity in visceral fat was an independent determinant of HOMA-IR, explaining ~18% of the variance (ß = 0.42, P = 0.016), after adjustment for age and BMI. CONCLUSION: These findings suggest that adipose tissue catalase shows compensatory up-regulation in response to obesity-induced H2O2 accumulation, and that perturbed H2O2 metabolism in visceral fat is linked to insulin resistance in obese humans.

Host associations and beta diversity of fungal endophyte communities in New Guinea rainforest trees.

Processes shaping the distribution of foliar fungal endophyte species remain poorly understood. Despite increasing evidence that these cryptic fungal symbionts of plants mediate interactions with pathogens and herbivores, there remain basic questions regarding the extent to which dispersal limitation and host specificity might shape fungal endophyte community composition in rainforests. To assess the relative importance of spatial pattern and host specificity, we isolated fungi from a sample of mapped trees in lowland Papua New Guinea. Sequences of the internal transcribed spacer (ITS) region were obtained for 2079 fungal endophytes from three sites and clustered into molecular operational taxonomic units (MOTUs) at 95% similarity. Multivariate analyses suggest that host affinity plays a significant role in structuring endophyte community composition whereas there was no evidence of endophyte spatial pattern at the scale of tens to hundreds of metres. Differences in endophyte communities between sampled trees were weakly correlated with variation in foliar traits but not with tree species relatedness. The dominance of relatively few generalist endophytes and the presence of a large number of rare MOTUs was a consistent observation at three sites separated by hundreds of kilometres and regional turnover was low. Host specificity appears to play a relatively weak but more important role than dispersal limitation in shaping the distribution of fungal endophyte communities in New Guinea forests. Our results suggest that in the absence of strong ecological gradients and host turnover, beta diversity of endophyte communities could be low in large areas of contiguous forest.