Serine synthesis via reversed SHMT2 activity drives glycine depletion and acetaminophen hepatotoxicity in MASLD.

Metabolic dysfunction-associated steatotic liver disease (MASLD) affects one-third of the global population. Understanding the metabolic pathways involved can provide insights into disease progression and treatment. Untargeted metabolomics of livers from mice with early-stage steatosis uncovered decreased methylated metabolites, suggesting altered one-carbon metabolism. The levels of glycine, a central component of one-carbon metabolism, were lower in mice with hepatic steatosis, consistent with clinical evidence. Stable-isotope tracing demonstrated that increased serine synthesis from glycine via reverse serine hydroxymethyltransferase (SHMT) is the underlying cause for decreased glycine in steatotic livers. Consequently, limited glycine availability in steatotic livers impaired glutathione synthesis under acetaminophen-induced oxidative stress, enhancing acute hepatotoxicity. Glycine supplementation or hepatocyte-specific ablation of the mitochondrial SHMT2 isoform in mice with hepatic steatosis mitigated acetaminophen-induced hepatotoxicity by supporting de novo glutathione synthesis. Thus, early metabolic changes in MASLD that limit glycine availability sensitize mice to xenobiotics even at the reversible stage of this disease.

The interplay between nonalcoholic fatty liver disease and atherosclerotic cardiovascular disease.

Therapeutic approaches that lower circulating low-density lipoprotein (LDL)-cholesterol significantly reduced the burden of cardiovascular disease over the last decades. However, the persistent rise in the obesity epidemic is beginning to reverse this decline. Alongside obesity, the incidence of nonalcoholic fatty liver disease (NAFLD) has substantially increased in the last three decades. Currently, approximately one third of world population is affected by NAFLD. Notably, the presence of NAFLD and particularly its more severe form, nonalcoholic steatohepatitis (NASH), serves as an independent risk factor for atherosclerotic cardiovascular disease (ASCVD), thus, raising interest in the relationship between these two diseases. Importantly, ASCVD is the major cause of death in patients with NASH independent of traditional risk factors. Nevertheless, the pathophysiology linking NAFLD/NASH with ASCVD remains poorly understood. While dyslipidemia is a common risk factor underlying both diseases, therapies that lower circulating LDL-cholesterol are largely ineffective against NASH. While there are no approved pharmacological therapies for NASH, some of the most advanced drug candidates exacerbate atherogenic dyslipidemia, raising concerns regarding their adverse cardiovascular consequences. In this review, we address current gaps in our understanding of the mechanisms linking NAFLD/NASH and ASCVD, explore strategies to simultaneously model these diseases, evaluate emerging biomarkers that may be useful to diagnose the presence of both diseases, and discuss investigational approaches and ongoing clinical trials that potentially target both diseases.

Sexual dimorphism in Louisiana pine snakes (Pituophis ruthveni).

The Louisiana pine snake, Pituophis ruthveni, is a cryptic, federally threatened snake species with several fragmented populations in Louisiana and Texas, USA. There are currently four captive breeding populations in zoos in the USA; however, little scientific data exists on their life history and anatomy. Accurate sex determination and identification of normal reproductive anatomy are an essential part of a veterinary exam and conservation programs. The authors had encountered various cases of sex misidentification in this species that were attributed to lack of lubrication of the sexing probes and enlarged musk glands. Anecdotal observation led to a hypothesis of sexual dimorphism based on body and tail shape. To test this hypothesis, we measured body length, tail length and width, and body to tail taper angle in 15 P. ruthveni (9 males and 6 females). We also obtained tail radiographs of all animals to document the presence of mineralized hemipenes. Significant dimorphism was identified in relative tail length, width, and taper angle; females consistently exhibited a more acute taper angle. Contrary to previous studies in other Pituophis species, a male-biased sexual size dimorphism was not identified. Mineralized hemipenes were confirmed in all males (a newly described trait in this species), and we found that the lateral view was consistently more reliable for identification of hemipenes compared to the ventrodorsal view. This information contributes to the scientific community's understanding of this species and is of use to biologists and veterinarians working toward conservation of this threatened species.

IL-1α Modulates IFN-γ-Induced Production of CXCL9/MIG during Herpes Simplex Virus Type-1 Corneal Infection.

PURPOSE: Investigating the modulation of neutrophil production of MIG and IP-10 during the inflammatory response to HSV-1 infection. MATERIALS AND METHODS: An ex vivo model of human corneal infection by HSV-1 was used for this study. This model permits the study of cytokine production by human corneal buttons in the presence, or absence, of gradient purified human neutrophils, under conditions of HSV-1 infection. All experimental samples were stimulated with a baseline concentration of recombinant human IFN-γ at 1 ng/mL. The relative levels of production for 12 pro-inflammatory mediators were screened using a multi-analyte ELISA assay. Neutrophil production of chemokines MIG and IP-10, under conditions of IFN-γ and/or HSV-1 stimulation were measured by quantitative ELISA. Lastly, antibody neutralization (goat IgG anti-human IL-1α, 2 µg/mL) of de novo production of IL-1α by corneal tissue was performed to investigated the effect on MIG and IP-10 production in the ex vivo model for HSV-1 infection. RESULTS: Four of the 12 pro-inflammatory mediators screened (IL-8, IL-6, IL-1α and IL-1ß) demonstrated elevated levels of production during corneal cell infection with HSV-1 and communication with neutrophils. Neutrophils were demonstrated to produce significant levels of both MIG and IP-10 under conditions of IFN-γ stimulation, and production of MIG was further upregulated by co-stimulation with IFN-γ and HSV-1. Neutralization of de novo IL-1α production in the model resulted in increased production of the chemokine production MIG but had no observable effect on IP-10 production. CONCLUSIONS: Our data provide evidence demonstrating the potential for expression patterns of MIG and IP-10 to be modulated by IL-1α, during the inflammatory response to HSV-1 corneal infection. Both corneal cells and neutrophils contribute to the production of T cell recruiting chemokines. However, IL-1α has the potential to upregulate MIG production by corneal cells while down-regulating MIG production by neutrophils.

Are vitamins A and E associated with persistent organic pollutants and fatty acids in the blubber of highly contaminated killer whales (Orcinus orca) from Greenland?

We quantified blubber concentrations of vitamins A (retinol) and E (α-tocopherol) and evaluated associations with persistent organic pollutants (ΣPOPs) in 14 highly-contaminated killer whales (Orcinus orca) sampled in Greenland from 2012 to 2014. We considered the influence of blubber depth, sex/age class and diet (based on biomass % of major fatty acids) in these relationships. Blubber concentrations of vitamin A averaged 34.1 ±â€¯4.7 µg g-1 wet weight (ww) and vitamin E averaged 35.6 ±â€¯4.4 µg g-1 ww. Although overall vitamin A concentrations did not vary between inner (closer to the muscle) and outer (closer to the skin) blubber layer or between sub-adults and adult females, concentrations in the outer layer of sub-adults were lower compared to the outer layer of adult females (p = 0.03). Outer layer may therefore reflect age accumulation of vitamin A, while in the more active inner layer, age effects might be masked by metabolic needs such as lactation. Neither diet nor ΣPOPs affected vitamin A variation, suggesting this vitamin is highly regulated in the body. Given the high exposures in these killer whales, vitamin A might not be a sensitive biomarker for POPs adverse effects. Vitamin E concentrations were significantly higher in inner compared to outer layer (p < 0.001), likely associated with blubber composition, suggesting that biopsies may not fully represent vitamin E concentrations in blubber. Age-accumulation of vitamin E also occurred with higher concentrations in adult females compared to sub-adults, independent of blubber depth (p < 0.01). Diet, ΣPOPs, and an interaction between these two variables significantly affected vitamin E variation in inner blubber, explaining 91% of this variation. The negative relationship between ΣPOPs (especially Σdichlorodiphenyltrichloroethane (DDT) and Σchlordanes in outer layers) and vitamin E was observed only in killer whales with a diet poorer in polyunsaturated fatty acids, suggested that killer whales feeding more consistently on marine mammals in Arctic environments over a fish-based diet, may be at higher risk of POP-induced disruption in vitamin E homeostasis. Considering diet is therefore important to understand the potential effects of elevated contaminant exposures on levels of certain essential nutrients, i.e., vitamin E, in killer whales.

Peripheral nerve blocks with sedation using propofol and alfentanil target-controlled infusion for hip fracture surgery: a review of 6 years in use.

BACKGROUND AND OBJECTIVES: Over the last 6 years, our center has introduced a novel technique combining peripheral nerve blocks (femoral and lateral femoral cutaneous nerves) with sedation using propofol with alfentanil target-controlled infusion for hip fracture surgery. The purpose of this review was to identify if adverse outcomes (of mortality and length of stay) were associated with its introduction compared to spinal or general anesthesia. METHODS: Retrospective data collection from hospital fracture database. Data were analyzed using Cox regression (adjusted for age, sex, and American Society of Anesthesiologists grade) to compare survival and length of stay data across the different anesthetic techniques used for hip fracture surgery. RESULTS: This technique was used in 472 (20%) of 2360 hip fractures. There was no significant difference between peripheral nerve blocks with propofol/alfentanil sedation/analgesia for mortality up to 120days (hazard ratio, 0.76; 95% confidence interval, 0.54-1.06; P=.11) and length of stay (hazard ratio, 1.03; 95% confidence interval, 0.91-1.17; P=.63) when compared to the other anesthetic techniques of spinal and general anesthesia. CONCLUSION: This novel technique does not appear to be associated with adverse mortality or length of stay after hip fracture surgery.

Depleting regulatory T cells with arginine-rich, cell-penetrating, peptide-conjugated morpholino oligomer targeting FOXP3 inhibits regulatory T-cell function.

CD4+CD25+regulatory T cells (T(reg)) impair anti-tumor and anti-viral immunity. As there are higher T(reg) levels in cancer patients compared with healthy individuals, there is considerable interest in eliminating them or altering their function as part of cancer or viral immunotherapy strategies. The scurfin transcriptional regulator encoded by the member of the forkhead winged helix protein family (FOXP3) is critical for maintaining the functions of T(reg). We hypothesized that targeting FOXP3 expression with a novel arginine-rich, cell-penetrating, peptide-conjugated phosphorodiamidate morpholino (PPMO) based antisense would eliminate T(reg) and enhance the induction of effector T-cell responses. We observed that the PPMO was taken up by activated T cells in vitro and could downregulate FOXP3 expression, which otherwise increases during antigen-specific T-cell activation. Generation of antigen-specific T cells in response to peptide stimulation was enhanced by pre-treatment of peripheral blood mononuclear cells with the FOXP3-targeted PPMO. In summary, modulation of T(reg) levels using the FOXP3 PPMO antisense-based genomic strategy has the potential to optimize immunotherapy strategies in cancer and viral immunotherapy.

Biochanin A, a naturally occurring inhibitor of fatty acid amide hydrolase.

BACKGROUND AND PURPOSE: Inhibitors of fatty acid amide hydrolase (FAAH), the enzyme responsible for the metabolism of the endogenous cannabinoid (CB) receptor ligand anandamide (AEA), are effective in a number of animal models of pain. Here, we investigated a series of isoflavones with respect to their abilities to inhibit FAAH. EXPERIMENTAL APPROACH: In vitro assays of FAAH activity and affinity for CB receptors were used to characterize key compounds. In vivo assays used were biochemical responses to formalin in anaesthetized mice and the 'tetrad' test for central CB receptor activation. KEY RESULTS: Of the compounds tested, biochanin A was adjudged to be the most promising. Biochanin A inhibited the hydrolysis of 0.5 microM AEA by mouse, rat and human FAAH with IC(50) values of 1.8, 1.4 and 2.4 microM respectively. The compound did not interact to any major extent with CB(1) or CB(2) receptors, nor with FAAH-2. In anaesthetized mice, URB597 (30 microg i.pl.) and biochanin A (100 microg i.pl.) both inhibited the spinal phosphorylation of extracellular signal-regulated kinase produced by the intraplantar injection of formalin. The effects of both compounds were significantly reduced by the CB(1) receptor antagonist/inverse agonist AM251 (30 microg i.pl.). Biochanin A (15 mg.kg(-1) i.v.) did not increase brain AEA concentrations, but produced a modest potentiation of the effects of 10 mg.kg(-1) i.v. AEA in the tetrad test. CONCLUSIONS AND IMPLICATIONS: It is concluded that biochanin A, in addition to its other biochemical properties, inhibits FAAH both in vitro and peripherally in vivo.

Composite tissue allotransplantation: current challenges.

Composite tissue allotransplantation (CTA) in the clinic is taking firm root. Success at hand, face, knee, trachea, and laryngeal transplantation has led to widespread interest and increasing application. Despite this, skepticism is common, particularly in the realm of reconstructive surgeons. The risks of immunosuppression remain a barrier to the advancement of the field, as these are perceived by many to be prohibitive. Significant progress in the field require the development of newer immunosuppressive agents with less toxicity and methods to achieve donor specific tolerance. This review focuses on the current state of CTA-both in the clinic and the laboratory. A thorough understanding of the immunology of CTA will allow the widespread application of this promising field.

Evaluation of cognitive assessment in stroke rehabilitation.

OBJECTIVE: To assess the effect of cognitive assessment on the functional outcome of stroke patients and quality of life for both patients and their carers. DESIGN: A multicentre, single-blind, randomized controlled trial. SETTING AND PARTICIPANTS: Two hundred and twenty-eight stroke patients were recruited from hospital wards in three UK centres. INTERVENTIONS: Patients were screened for cognitive impairment and randomly allocated to either routine care (116 patients) or routine care plus a detailed cognitive assessment (112 patients). MAIN OUTCOME MEASURES: Outcome was assessed three and six months after recruitment by an independent assessor blind to the intervention on Extended ADL, Cognitive Failures Questionnaire, General Health Questionnaire-28 for patients and carers and Carer Strain Index. RESULTS: There was no significant difference between the two groups in patients' functional outcome, perceived cognitive ability, level of psychological distress or satisfaction with care. There was a trend for the assessment group to have lower levels of carer strain (p = 0.06). CONCLUSIONS: The provision of information about cognitive assessment in stroke rehabilitation may decrease carer strain.

Signatures of hippocampal oxidative stress in aged spatial learning-impaired rodents.

Neurons and glia within the hippocampus of aged, spatial learning-impaired Long-Evans rats exhibit uniquely altered gene expression profiles, and we have postulated oxidative stress as the basis for this. To test this hypothesis we quantitated the extent of protein and nucleic acid oxidative damage, evaluated the status of mitochondrial DNA integrity, and examined several signaling entities and molecular indicators frequently associated with oxidative stress and gliosis. Immunoblotting demonstrated elevated heme oxygenase-1 in the aged-impaired hippocampus and immunocytochemistry suggested that heme oxygenase-1 is largely cytosolic and at least partly neuronal in nature. In the aged-impaired group, immunoreactivity to 8-hydroxy-2'-deoxyguanosine, an oxidative nucleic acid adduct, was found to be elevated in the dentate gyrus and in area CA1 of the hippocampal formation. Isolated mitochondrial DNA was found to be significantly damaged in the aged-impaired group. In the aged learning-impaired rats only, proteins in a 65-kDa band were found to contain excessive levels of carbonyl residues. Glial activation was examined by in situ hybridization histochemistry to tumor necrosis factor alpha and by immunocytochemistry with OX-6, which detects activated microglia. White matter in aged brains exhibited a modest up-regulation of tumor necrosis factor alpha mRNA and OX-6 immunoreactivity, but the hippocampal formation expressed tumor necrosis factor alpha mRNA equivalent to young animals and few OX-6-positive microglia. The mRNA for manganese-dependent superoxide dismutase, which is elevated in the aged hippocampus, was found preferentially expressed in neurons. We conclude that aged hippocampal neurons appear to be under oxidative stress and this is more severe in the learning-impaired subjects, suggesting a possible basis for age-induced cognitive decline.

The care of HIV-infected adults in rural areas of the United States.

OBJECTIVE: This study describes the population of HIV-infected adults receiving care in rural areas of the United States and compares HIV care received in rural and urban areas. METHODS: Interviews were conducted with a nationally representative sample of 367 HIV-infected adults receiving health care in rural areas and 2806 HIV-infected adults receiving health care in urban areas of the contiguous United States. RESULTS: We estimate that 4800 HIV-infected persons received medical care in rural areas during the first half of 1996. Patients in rural HIV care were more likely than patients in urban HIV care to receive care from providers seeing few (<10) HIV-infected patients (38% vs. 3%; p <.001). Rural care patients were less likely than urban care patients to have taken highly active antiretroviral agents (57% vs. 73%; p <.001) or Pneumocystis carinii pneumonia prophylactic medication when indicated (60% vs. 75%; p =.006). CONCLUSIONS: Few American adults received HIV care in rural areas of the United States. Our findings suggest ongoing disparities between urban and rural areas in access to high-quality HIV care.

Breast cancer action plan for Kentucky.

In 1998, Governor Paul E. Patton established the Kentucky Breast Cancer Task Force (KBCTF) to assess and make recommendations on the availability, accessibility, utilization, quality, and outcomes of breast cancer services across the spectrum of disease. Over a two-year period, the KBCTF reviewed the state of breast cancer control in Kentucky and made recommendations for reducing breast cancer morbidity and mortality. To achieve the provision of optimal breast cancer care, the KBCTF recommended universal adoption of professionally accepted guidelines for breast cancer treatment, pain management, and distress management. To better coordinate public education on breast cancer, the KBCTF recommended the development of a standardized packet of public education materials for dissemination through regional networks of "cancer control partners." KBCTF members also requested the Kentucky Cancer Program to investigate the feasibility of establishing a centralized mammography registry to gather more complete data on screening mammography programs.

Survival and plasticity of basal forebrain cholinergic systems in mice transgenic for presenilin-1 and amyloid precursor protein mutant genes.

The basalo-cortical cholinergic system was characterized in mice expressing mutant human genes for presenilin-1 (PS1), amyloid precursor protein (APP), and combined PS/APP. Dual immunocytochemistry for ChAT and A beta revealed swollen cholinergic processes within cortical plaques in both APP and PS/APP brains by 12 months, suggesting aberrant sprouting or redistribution of cholinergic processes in response to amyloid deposition. At 8 months, cortical and subcortical ChAT activity was normal (PS/APP) or elevated (PS, APP frontal cortex), while cholinergic cell counts (nBM/SI) and receptor binding were unchanged. ChAT mRNA was up-regulated in the nBM/SI of all three transgenic lines at 8 months. The data indicate that the basal forebrain cholinergic system does not degenerate in mice expressing AD-related transgenes, even in mice with extreme amyloid load. The

Induction of nitric-oxide synthase and activation of NF-kappaB by interleukin-12 p40 in microglial cells.

Interleukin-12 (IL-12) is composed of two different subunits, p40 and p35. Expression of p40 mRNA but not that of p35 mRNA in excessive amount in the central nervous system of patients with multiple sclerosis (MS) suggests that IL-12 p40 may have a role in the pathogenesis of the disease. However, the mode of action of p40 is completely unknown. Because nitric oxide produced from the induction of nitric-oxide synthase (iNOS) also plays a vital role in the pathophysiology of MS, the present study was undertaken to explore the role of p40 in the induction of NO production and the expression of iNOS in microglia. Both IL-12 and p40(2), the p40 homodimer, dose-dependently induced the production of NO in BV-2 microglial cells. This induction of NO production was accompanied by an induction of iNOS protein and mRNA. Induction of NO production by the expression of mouse p40 cDNA but not that of the mouse p35 cDNA suggests that the p40 but not the p35 subunit of IL-12 is involved in the expression of iNOS. In addition to BV-2 glial cells, p40(2) also induced the production of NO in mouse primary microglia and peritoneal macrophages. However, both IL-12 and p40(2) were unable to induce the production of NO in mouse primary astrocytes. Because activation of NF-kappaB is important for the expression of iNOS, we investigated the effect of p40(2) on the activation of NF-kappaB. Induction of the DNA binding as well as the transcriptional activity of NF-kappaB by p40(2) and inhibition of p40(2)-induced expression of iNOS by SN50, a cell-permeable peptide carrying the nuclear localization sequence of p50 NF-kappaB, but not by SN50M, a nonfunctional peptide mutant, suggests that p40(2) induces the expression of iNOS through the activation of NF-kappaB. This study delineates a novel role of IL-12 p40 in inducing the expression of iNOS in microglial cells, which may participate in the pathogenesis of neuroinflammatory diseases.

Visualization of muscarinic receptor-mediated phosphoinositide turnover in the hippocampus of young and aged, learning-impaired Long Evans rats.

Hippocampal receptor-mediated phosphoinositide (PI) turnover is severely blunted in aged rats that demonstrate cognitive deficits in the Morris water maze. To further examine the anatomical localization of this deficit, we examined the topography of muscarinic receptor-mediated PI turnover in young and aged-learning impaired rats by taking advantage of an autoradiographic method that visualizes PI turnover by measuring the diacylglycerol (DAG) branch of the PI turnover signal transduction system. Using this method, muscarinic cholinergic receptors were stimulated in hippocampal slices with agonist, and the receptor-mediated incorporation of [3H] cytidine into [3H]CDP-DAG was subsequently quantified in subregions of the hippocampus using film autoradiography. Our results show a significant decrease in basal incorporation of [ 3H]CDP-DAG in the subiculum and in the dentate gyrus in the aged rats. The muscarinic receptor-mediated [3H]CDP-DAG response was significantly blunted in the aged rats in subiculum, CA3, and CA1. In contrast, the receptor-mediated response was maintained in the dentate gyrus and hilus. These results indicate that the age-associated impairment in receptor-mediated PI turnover differs regionally, with a reduction in the subiculum and hippocampus proper that is pronounced relative to the hilus and dentate gyrus.

Who receives Ryan White CARE Act services? A demographic comparison of CARE act clients and the general AIDS population.

This study examines the extent to which health and social service providers funded by the Ryan White Comprehensive AIDS Resources Emergency (CARE) Act serve women, minorities, and other vulnerable populations emphasized by the legislation. Demographic characteristics of AIDS-diagnosed clients served by CARE Act-funded providers in four metropolitan areas and two states are compared with Centers for Disease Control and Prevention estimates of AIDS prevalence. Clients of CARE Act-funded providers tend to reflect the demographics of local HIV/AIDS epidemics. Where differences exist, CARE Act clients are more likely to be women and minorities and less likely to be injecting drug users. CARE Act-funded providers are effectively reaching most medically underserved populations.

Design, synthesis and binding at cloned muscarinic receptors of N-[5-(1'-substituted-acetoxymethyl)-3-oxadiazolyl] and N-[4-(1'-substituted-acetoxymethyl)-2-dioxolanyl] dialkyl amines.

Few muscarinic antagonists differentiate between the M4 and M2 muscarinic receptors. In a structure activity study, aimed at discovering leads for the development of a M4 muscarinic receptor-selective antagonist, we have synthesized and tested at cloned muscarinic receptors the binding of a group of dioxolane- or oxadiazole-dialkyl amines, and compared them to our compound 1, which contains the furan nucleus. Although none of these agents were particularly potent at M4 receptors (Kd values were typically 30-70 nM), furan derivatives (-)1 and (+)1 were significantly more potent at M4 receptors than at M2 receptors (approximately 3- and 4-fold, respectively). The dioxolane derivatives 12b and 12c were more than 10-fold selective for the M4 versus the M2 receptors, while the dioxolane derivative 12e was 15-fold more potent at M4 receptors than for M2 receptors. However, these agents bound to M3 receptors with potencies like that for the M4 receptor, so they are not M4-selective. The M4/M2 relative selectivities of some of our compounds are similar to the better hexahydrosiladifenidol derivatives, and may provide some important structural clues for the development of potent and selective M4 antagonists.

Prenatal HIV prevention practices in a low seroprevalence state.

The purpose of this study was to build understanding of prenatal HIV counseling and testing practices in low seroprevalence states. Responses from a 1998 population-based survey of Kentucky prenatal care providers (67% response; 312 analyzed) were compared with findings from patient focus groups. Sixty-two percent of clinicians said they routinely counsel prenatal patients with risk factors, but only 46% routinely counsel patients without risk factors. The proportions routinely offering HIV testing to patients with and without risk factors were 94% and 84%, respectively. Prenatal patients identified "fear of a positive test result" as the major barrier to test acceptance. This fear was fueled by lack of knowledge regarding the benefits of early detection. The study concludes that achieving universal prenatal HIV testing will require new strategies, such as the distribution of a standardized protocol, that address clinicians' concerns about "time burdens" without depriving patients of the opportunity to receive individualized counseling.