Denisovan, modern human and mouse TNFAIP3 alleles tune A20 phosphorylation and immunity.

Resisting and tolerating microbes are alternative strategies to survive infection, but little is known about the evolutionary mechanisms controlling this balance. Here genomic analyses of anatomically modern humans, extinct Denisovan hominins and mice revealed a TNFAIP3 allelic series with alterations in the encoded immune response inhibitor A20. Each TNFAIP3 allele encoded substitutions at non-catalytic residues of the ubiquitin protease OTU domain that diminished IκB kinase-dependent phosphorylation and activation of A20. Two TNFAIP3 alleles encoding A20 proteins with partial phosphorylation deficits seemed to be beneficial by increasing immunity without causing spontaneous inflammatory disease: A20 T108A;I207L, originating in Denisovans and introgressed in modern humans throughout Oceania, and A20 I325N, from an N-ethyl-N-nitrosourea (ENU)-mutagenized mouse strain. By contrast, a rare human TNFAIP3 allele encoding an A20 protein with 95% loss of phosphorylation, C243Y, caused spontaneous inflammatory disease in humans and mice. Analysis of the partial-phosphorylation A20 I325N allele in mice revealed diminished tolerance of bacterial lipopolysaccharide and poxvirus inoculation as tradeoffs for enhanced immunity.

A nonredundant role for T cell-derived interleukin 22 in antibacterial defense of colonic crypts.

Interleukin (IL)-22 is central to immune defense at barrier sites. We examined the contributions of innate lymphoid cell (ILC) and T cell-derived IL-22 during Citrobacter rodentium (C.r) infection using mice that both report Il22 expression and allow lineage-specific deletion. ILC-derived IL-22 activated STAT3 in C.r-colonized surface intestinal epithelial cells (IECs) but only temporally restrained bacterial growth. T cell-derived IL-22 induced a more robust and extensive activation of STAT3 in IECs, including IECs lining colonic crypts, and T cell-specific deficiency of IL-22 led to pathogen invasion of the crypts and increased mortality. This reflected a requirement for T cell-derived IL-22 for the expression of a host-protective transcriptomic program that included AMPs, neutrophil-recruiting chemokines, and mucin-related molecules, and it restricted IFNγ-induced proinflammatory genes. Our findings demonstrate spatiotemporal differences in the production and action of IL-22 by ILCs and T cells during infection and reveal an indispensable role for IL-22-producing T cells in the protection of the intestinal crypts.

Distal colonocytes targeted by C. rodentium recruit T-cell help for barrier defence.

Interleukin 22 (IL-22) has a non-redundant role in immune defence of the intestinal barrier1-3. T cells, but not innate lymphoid cells, have an indispensable role in sustaining the IL-22 signalling that is required for the protection of colonic crypts against invasion during infection by the enteropathogen Citrobacter rodentium4 (Cr). However, the intestinal epithelial cell (IEC) subsets targeted by T cell-derived IL-22, and how T cell-derived IL-22 sustains activation in IECs, remain undefined. Here we identify a subset of absorptive IECs in the mid-distal colon that are specifically targeted by Cr and are differentially responsive to IL-22 signalling. Major histocompatibility complex class II (MHCII) expression by these colonocytes was required to elicit sustained IL-22 signalling from Cr-specific T cells, which was required to restrain Cr invasion. Our findings explain the basis for the regionalization of the host response to Cr and demonstrate that epithelial cells must elicit MHCII-dependent help from IL-22-producing T cells to orchestrate immune protection in the intestine.

Insulin-Like Growth Factors Are Key Regulators of T Helper 17 Regulatory T Cell Balance in Autoimmunity.

Appropriate balance of T helper 17 (Th17) and regulatory T (Treg) cells maintains immune tolerance and host defense. Disruption of Th17-Treg cell balance is implicated in a number of immune-mediated diseases, many of which display dysregulation of the insulin-like growth factor (IGF) system. Here, we show that, among effector T cell subsets, Th17 and Treg cells selectively expressed multiple components of the IGF system. Signaling through IGF receptor (IGF1R) activated the protein kinase B-mammalian target of rapamycin (AKT-mTOR) pathway, increased aerobic glycolysis, favored Th17 cell differentiation over that of Treg cells, and promoted a heightened pro-inflammatory gene expression signature. Group 3 innate lymphoid cells (ILC3s), but not ILC1s or ILC2s, were similarly responsive to IGF signaling. Mice with deficiency of IGF1R targeted to T cells failed to fully develop disease in the experimental autoimmune encephalomyelitis (EAE) model of multiple sclerosis. Thus, the IGF system represents a previously unappreciated pathway by which type 3 immunity is modulated and immune-mediated pathogenesis controlled.

BioCanRx Summit for Cancer Immunotherapy 2022 Proceedings.

From 19 to 21 November 2022, BioCanRx held its first post-pandemic in-person Summit for Cancer Immunotherapy in Montreal, Canada. The meeting was well attended by patients, trainees, researchers, clinicians, and industry professionals, who came together to discuss the current state and future of biotherapeutics for cancer in Canada and beyond. Three plenaries, three keynote speakers, a lively debate, and panel discussions, together with poster sessions and a social event, made the event memorable and productive. The current state of cellular therapies, cellular engineering, clinical trials, and the role of the cancer microbiome were discussed in plenary session, and the patient voice was welcomed and present throughout the meeting, in large part due to the Learning Institute, a BioCanRx initiative to include patient partners in research. In this meeting review, we highlight the platform presentations, keynote speakers, debate combatants, panellists, and the patient perspective on the annual meeting.

Feeding under fire: Relations between parental stress hormones and controlling feeding behaviors.

Under stress, parents tend to use more controlling feeding behaviors toward their children (Berge et al., 2017; Doan et al., 2022; Loth et al., 2016). However, the majority of prior work focuses on subjective reports of stress, and there is a dearth of research examining parental physiological stress and its impact on feeding behaviors. In the current study, we examined how parental physiological stress reactivity would influence their feeding behaviors under mild stress in a lab-based setting. Parents (n = 83, 50 % females) and their children (59% female, Mage = 42 months, SD = 4.48) participated. Stress was induced using the Trier Social Stress Test in the laboratory (Kirshbaum et al., 1993). Salivary samples were collected at 4 time points during the visit to index stress reactivity and later assayed for cortisol and DHEA. Parent-child interactions during the anticipatory period of the stress test were observationally coded for parent use of controlling feeding behaviors. To examine whether parent stress physiology predicts their feeding behaviors, we ran a Poisson regression using income, parent ethnicity, parent sex (mom/dad), time of day, and DHEA/cortisol ratio as predictors of controlling feeding behavior. Latinx parents used less controlling feeding behaviors, b = -0.323, p = 0.041 than non-Latinx parents. Parents with a higher DHEA/Cortisol ratio were less likely to use controlling feeding behaviors, b = -0.231, p = 0.008. To our knowledge, this is the first study to show that for both mothers and fathers, DHEA relative to cortisol has a protective role in controlling feeding practices, and lends support to the role of acute stress reactivity in predicting behavioral outcomes.

RELA governs a network of islet-specific metabolic genes necessary for beta cell function.

AIMS/HYPOTHESIS: NF-κB activation unites metabolic and inflammatory responses in many diseases yet less is known about the role that NF-κB plays in normal metabolism. In this study we investigated how RELA impacts the beta cell transcriptional landscape and provides network control over glucoregulation. METHODS: We generated novel mouse lines harbouring beta cell-specific deletion of either the Rela gene, encoding the canonical NF-κB transcription factor p65 (ßp65KO mice), or the Ikbkg gene, encoding the NF-κB essential modulator NEMO (ßNEMOKO mice), as well as ßA20Tg mice that carry beta cell-specific and forced transgenic expression of the NF-κB-negative regulator gene Tnfaip3, which encodes the A20 protein. Mouse studies were complemented by bioinformatics analysis of human islet chromatin accessibility (assay for transposase-accessible chromatin with sequencing [ATAC-seq]), promoter capture Hi-C (pcHi-C) and p65 binding (chromatin immunoprecipitation-sequencing [ChIP-seq]) data to investigate genome-wide control of the human beta cell metabolic programme. RESULTS: Rela deficiency resulted in complete loss of stimulus-dependent inflammatory gene upregulation, consistent with its known role in governing inflammation. However, Rela deletion also rendered mice glucose intolerant because of functional loss of insulin secretion. Glucose intolerance was intrinsic to beta cells as ßp65KO islets failed to secrete insulin ex vivo in response to a glucose challenge and were unable to restore metabolic control when transplanted into secondary chemical-induced hyperglycaemic recipients. Maintenance of glucose tolerance required Rela but was independent of classical NF-κB inflammatory cascades, as blocking NF-κB signalling in vivo by beta cell knockout of Ikbkg (NEMO), or beta cell overexpression of Tnfaip3 (A20), did not cause severe glucose intolerance. Thus, basal p65 activity has an essential and islet-intrinsic role in maintaining normal glucose homeostasis. Genome-wide bioinformatic mapping revealed the presence of p65 binding sites in the promoter regions of specific metabolic genes and in the majority of islet enhancer hubs (~70% of ~1300 hubs), which are responsible for shaping beta cell type-specific gene expression programmes. Indeed, the islet-specific metabolic genes Slc2a2, Capn9 and Pfkm identified within the large network of islet enhancer hub genes showed dysregulated expression in ßp65KO islets. CONCLUSIONS/INTERPRETATION: These data demonstrate an unappreciated role for RELA as a regulator of islet-specific transcriptional programmes necessary for the maintenance of healthy glucose metabolism. These findings have clinical implications for the use of anti-inflammatories, which influence NF-κB activation and are associated with diabetes.

Investigating dual inhibition of ACC and CD36 for the treatment of nonalcoholic fatty liver disease in mice.

Nonalcoholic fatty liver disease (NAFLD) is the most common chronic liver disease worldwide. Dysregulation in hepatic lipid metabolism, including increased fatty acid uptake and de novo lipogenesis (DNL), is a hallmark of NAFLD. Here, we investigated dual inhibition of the fatty acid transporter fatty acid translocase (FAT/CD36), and acetyl-CoA carboxylase (ACC), the rate-limiting enzyme in DNL, for the treatment of NAFLD in mice. Mice with hepatic CD36 deletion (Cd36LKO) and wild-type littermates were fed a high-fat diet for 12 wk and treated daily with either oral administration of an ACC inhibitor (GS-834356, Gilead Sciences; ACCi) or vehicle for 8 wk. Neither CD36 deletion or ACC inhibition impacted body composition, energy expenditure, or glucose tolerance. Cd36LKO mice had elevated fasting plasma insulin, suggesting mild insulin resistance. Whole body fatty acid oxidation was significantly decreased in Cd36LKO mice. Liver triglyceride content was significantly reduced in mice treated with ACCi; however, CD36 deletion caused an unexpected increase in liver triglycerides. This was associated with upregulation of genes and proteins of DNL, including ACC, and decreased liver triglyceride secretion ex vivo. Overall, these data confirm the therapeutic utility of ACC inhibition for steatosis resolution but indicate that inhibition of CD36 is not an effective treatment for NAFLD in mice.NEW & NOTEWORTHY Dysregulation of hepatic lipid metabolism is a hallmark of nonalcoholic fatty liver disease. Here, we show that dual inhibition of the de novo lipogenesis enzyme, ACC, and hepatic deletion of the fatty acid transporter, CD36, was ineffective for the treatment of NAFLD in mice. This was due to a paradoxical increase in liver triglycerides with CD36 deletion resulting from decreased hepatic triglyceride secretion and increased lipogenic gene expression.

The impact of the cytoplasmic ubiquitin ligase TNFAIP3 gene variation on transcription factor NF-κB activation in acute kidney injury.

Nuclear factor κB (NF-κB) activation is a deleterious molecular mechanism that drives acute kidney injury (AKI) and manifests in transplanted kidneys as delayed graft function. The TNFAIP3 gene encodes A20, a cytoplasmic ubiquitin ligase and a master negative regulator of the NF- κB signaling pathway. Common population-specific TNFAIP3 coding variants that reduce A20's enzyme function and increase NF- κB activation have been linked to heightened protective immunity and autoimmune disease, but have not been investigated in AKI. Here, we functionally identified a series of unique human TNFAIP3 coding variants linked to the autoimmune genome-wide association studies single nucleotide polymorphisms of F127C; namely F127C;R22Q, F127C;G281E, F127C;W448C and F127C;N449K that reduce A20's anti-inflammatory function in an NF- κB reporter assay. To investigate the impact of TNFAIP3 hypomorphic coding variants in AKI we tested a mouse Tnfaip3 hypomorph in a model of ischemia reperfusion injury (IRI). The mouse Tnfaip3 coding variant I325N increases NF- κB activation without overt inflammatory disease, providing an immune boost as I325N mice exhibit enhanced innate immunity to a bacterial challenge. Surprisingly, despite exhibiting increased intra-kidney NF- κB activation with inflammation in IRI, the kidney of I325N mice was protected. The I325N variant influenced the outcome of IRI by changing the dynamic expression of multiple cytoprotective mechanisms, particularly by increasing NF- κB-dependent anti-apoptotic factors BCL-2, BCL-XL, c-FLIP and A20, altering the active redox state of the kidney with a reduction of superoxide levels and the enzyme super oxide dismutase-1, and enhancing cellular protective mechanisms including increased Foxp3+ T cells. Thus, TNFAIP3 gene variants represent a kidney and population-specific molecular factor that can dictate the course of IRI.

Negative emodiversity is associated with emotional eating in adolescents: An examination of emotion dynamics in daily life.

INTRODUCTION: Emotional eating is a mental health concern, common in adolescents, that develops as a result of their tendency to use high-energy food to regulate their fluctuating emotions. Due to their highly fluctuating emotional life, adolescents tend to have unique within-person profiles of emotional experiences that change across moments and days, often lost in global assessments of emotions. Hence, it is imperative to examine individual differences in dynamics of emotions, as experienced in daily life, in relation to emotional eating in adolescents. METHODS: In an Ecological Momentary Assessment study, we examined individual differences in three within-person dynamic characteristics (baseline levels, intraindividual variability, and emodiversity) of emotions in 158 dominantly Hispanic adolescents in the United States, aged 14-17 years old, predicting trait-level emotional eating. RESULTS: Results indicated that higher negative emodiversity, baselines, and variability in stress were predictive of emotional eating in adolescents. When all considered together, negative emodiversity (i.e., variety of the types of negative emotions experienced in one's daily life) remained the only significant predictor of emotional eating. CONCLUSIONS: This study affirms the importance of diversity in emotional experiences in relation to emotional eating, particularly in daily contexts of adolescents' lives. Additionally, the study emphasizes the importance of distinguishing between diversity (i.e., variety in types) in positive versus negative emotional experiences with regard to emotional eating. By taking into account the ecological validity of adolescents' daily lives and individual differences in dynamical changes in emotions, we are taking a step forward by shedding light on how the dynamics of negative emotions-in terms of within-person baselines, variability, and diversity-might be related to general levels of emotional eating in adolescents.

A Quasi-Experimental Evaluation of Single Trough-Based Area Under the Curve Guided Dosing on the Incidence of Vancomycin Associated Nephrotoxicity in Veteran Patients.

Background: Two common dosing strategies for vancomycin are trough-based and area under the curve (AUC)-based dosing. Objective: To compare the incidence of nephrotoxicity in trough-based dosing group with the single trough-based AUC dosing at the Salem VA Medical Center. Methods: This retrospective study included patients who received trough-based dosing of vancomycin between January 1, 2017, and January 1, 2019 (preimplementation group) and AUC-based dosing (postimplementation) between October 1, 2019, and October 1, 2021, at the Salem VA Medical Center. The primary outcome was nephrotoxicity at 96 hours, 7 days, and entire hospital length of stay (LOS). Secondary outcomes included 30-day readmission and all-cause mortality rates, cumulative doses at 24, 48, and 72 hours, and percentage of patients considered at goal (AUC 400-600 or trough between 10 and 20 mg/L). Propensity score (PS) matching was utilized to adjust for confounding. Results: After PS matching 100 patients were included in preimplementation and 95 patients in the postimplementation group. The average study patient was a 68-year-old white male. There was significant reduction in the risk of nephrotoxicity in postimplementation cohort at 96 hours (adjusted (a)HR: 0.28, 95% CI (0.12-0.66); 7 days (aHR: 0.39, 95% CI (0.18-0.85); and entire hospital LOS (aHR: 0.46, 95% CI (0.22-0.95). Secondary outcomes showed no difference between the groups except significantly higher proportion of patients were considered at therapeutic goal in the postimplementation cohort compared with pre-implementation cohort. Conclusion: This hypothesis generating study shows that AUC-based dosing calculated using single trough concentration may result in reduced rate of nephrotoxicity than trough-based dosing.

Profile and Predictors of Blood Tumor Mutational Burden in Advanced Hepatocellular Carcinoma.

Advanced hepatocellular carcinoma (HCC) is responsive to immune checkpoint inhibitors, but there are currently no known biomarkers to predict treatment benefit. Blood TMB (bTMB) estimation via circulating tumor DNA (ctDNA) profiling can provide a convenient means to estimate HCC TMB. Here we provide the first landscape of bTMB in advanced HCC using a commercially available next-generation sequencing assay, show that it is approximately three times as high as matched tissue TMB, and show that bTMB correlates with NAFLD cirrhosis etiology and the presence of genomic alterations in HTERT and TP53. These results lay the foundation for subsequent studies evaluating bTMB as an immune therapy predictive biomarker in HCC.

Urinary metabotype of severe asthma evidences decreased carnitine metabolism independent of oral corticosteroid treatment in the U-BIOPRED study.

INTRODUCTION: Asthma is a heterogeneous disease with poorly defined phenotypes. Patients with severe asthma often receive multiple treatments including oral corticosteroids (OCS). Treatment may modify the observed metabotype, rendering it challenging to investigate underlying disease mechanisms. Here, we aimed to identify dysregulated metabolic processes in relation to asthma severity and medication. METHODS: Baseline urine was collected prospectively from healthy participants (n=100), patients with mild-to-moderate asthma (n=87) and patients with severe asthma (n=418) in the cross-sectional U-BIOPRED cohort; 12-18-month longitudinal samples were collected from patients with severe asthma (n=305). Metabolomics data were acquired using high-resolution mass spectrometry and analysed using univariate and multivariate methods. RESULTS: A total of 90 metabolites were identified, with 40 significantly altered (p<0.05, false discovery rate <0.05) in severe asthma and 23 by OCS use. Multivariate modelling showed that observed metabotypes in healthy participants and patients with mild-to-moderate asthma differed significantly from those in patients with severe asthma (p=2.6×10-20), OCS-treated asthmatic patients differed significantly from non-treated patients (p=9.5×10-4), and longitudinal metabotypes demonstrated temporal stability. Carnitine levels evidenced the strongest OCS-independent decrease in severe asthma. Reduced carnitine levels were associated with mitochondrial dysfunction via decreases in pathway enrichment scores of fatty acid metabolism and reduced expression of the carnitine transporter SLC22A5 in sputum and bronchial brushings. CONCLUSIONS: This is the first large-scale study to delineate disease- and OCS-associated metabolic differences in asthma. The widespread associations with different therapies upon the observed metabotypes demonstrate the need to evaluate potential modulating effects on a treatment- and metabolite-specific basis. Altered carnitine metabolism is a potentially actionable therapeutic target that is independent of OCS treatment, highlighting the role of mitochondrial dysfunction in severe asthma.

Providing metabolomics education and training: pedagogy and considerations.

BACKGROUND: Metabolomics is a highly multidisciplinary and non-standardised research field. Metabolomics researchers must possess and apply extensive cross-disciplinary content knowledge, subjective experience-based judgement, and the associated diverse skill sets. Accordingly, appropriate educational and training initiatives are important in developing this knowledge and skills base in the metabolomics community. For these initiatives to be successful, they must consider both pedagogical best practice and metabolomics-specific contextual challenges. AIM OF REVIEW: The aim of this review is to provide consolidated pedagogical guidance for educators and trainers in metabolomics educational and training programmes. KEY SCIENTIFIC CONCEPTS OF REVIEW: In this review, we discuss the principles of pedagogical best practice as they relate to metabolomics. We then discuss the challenges and considerations in developing and delivering education and training in metabolomics. Finally, we present examples from our own teaching practice to illustrate how pedagogical best practice can be integrated into metabolomics education and training programmes.

Stressed out and fed up: The effect of stress on maternal feeding behaviors and the moderating role of executive function.

Stress is associated with a range of unhealthy eating habits, yet few studies have examined how stress may influence the intergenerational transmission of eating habits from parents to their children. Specifically, there is a lack of data regarding the role of stress on feeding practices. Moreover, most work investigating the associations between parental stress and their feeding behaviors has been correlational, limiting our understanding of causality. In the current study, we used an experimental design, induced high and low stress in mothers using a standard laboratory stressor, and observed mother-child interactions during a snack break. We also examined the potential role of maternal executive functioning (EF) for buffering the effects of stress on maternal feeding behaviors. Levels of maternal stress were manipulated with the Trier Social Stress Task (TSST) in a community sample (N = 80 dyads, Child Mage = 41.89 months, female = 43). We measured maternal EF with a series of computerized tasks. Maternal feeding behaviors were coded for controlling behaviors, which included pressuring and restricting behaviors. Results indicate a main effect of stress on controlling feeding behaviors, such that mothers in the high-stress condition exhibited higher levels of controlling behaviors. The effect of stress on controlling feeding behaviors was ameliorated among mothers with higher levels of EF after controlling for child age and income. Results provide causal evidence for the role of stress on feeding behaviors and suggest EF as a factor to be considered in the treatment and prevention of diet-related illnesses.

A mother's touch: Preschool-aged children are regulated by positive maternal touch.

Positive maternal touch plays an important role in the development of children's physiological regulation and cognitive development in infancy, as well as the development of sociality in early childhood. However, few studies have looked beyond infancy to consider the possible continuing impact of positive maternal touch on child stress reactivity during early childhood. A diverse community sample of mothers (N = 114, Mage  = 33.52 years, SD = 5.33) and their preschool-aged children (Mage  = 41.68 months, SD = 4.67; 49.1% female) participated in the study. Basic demographics were reported by mothers. We coded maternal touch behaviors during an emotionally charged laboratory conversation task and assessed children's physiological reactivity to stressful laboratory tasks with salivary cortisol. Results reveal a significant negative association between positive maternal touch and child salivary cortisol reactivity. In addition, family income, adjusted for family size, and child sex were significantly associated with child cortisol stress reactivity. Findings are discussed in terms of persistent downregulating effects of positive maternal touch on child stress reactivity, as well as possible links of stress reactivity with family income, a proxy for economic stress, and child sex.

The impact of Covid-19 on education equity: A view from Barbados and Jamaica.

The outbreak of Covid-19 worldwide has presented an unprecedented challenge for the equity-in-education agenda, especially in developing countries of the Global South (e.g., the English-speaking Caribbean). This article examines the impact school closures have had in Jamaica and Barbados, and highlights the emerging disparities the global pandemic has had on education. The central organizing questions are as follows: Who was affected by school closures in Barbados and Jamaica? How did the Ministries of Education (MOEs) support curriculum and instruction during the pandemic? What challenges does Covid-19 present for MOEs? What are the implications for education after Covid-19? School closure data suggest a gender disparity, with more males than females out of school due to Covid-19 from preprimary to secondary school in Barbados and Jamaica. MOEs in the region responded to school closures primarily by increasing access to technology to facilitate remote learning. Some of the challenges with continuing education for students during Covid-19 were due to a lack of infrastructure and amenities to support remote learning. Implications for education post-Covid-19 are considered.

Perilipin 5 S155 phosphorylation by PKA is required for the control of hepatic lipid metabolism and glycemic control.

Perilipin 5 (PLIN5) is a lipid-droplet-associated protein that coordinates intracellular lipolysis in highly oxidative tissues and is thought to regulate lipid metabolism in response to phosphorylation by protein kinase A (PKA). We sought to identify PKA phosphorylation sites in PLIN5 and assess their functional relevance in cultured cells and the livers of mice. We detected phosphorylation on S155 and identified S155 as a functionally important site for lipid metabolism. Expression of phosphorylation-defective PLIN5 S155A in Plin5 null cells resulted in decreased rates of lipolysis and triglyceride-derived fatty acid oxidation. FLIM-FRET analysis of protein-protein interactions showed that PLIN5 S155 phosphorylation regulates PLIN5 interaction with adipose triglyceride lipase at the lipid droplet, but not with α-ß hydrolase domain-containing 5. Re-expression of PLIN5 S155A in the liver of Plin5 liver-specific null mice reduced lipolysis compared with wild-type PLIN5 re-expression, but was not associated with other changes in hepatic lipid metabolism. Furthermore, glycemic control was impaired in mice with expression of PLIN5 S155A compared with mice expressing PLIN5. Together, these studies demonstrate that PLIN5 S155 is required for PKA-mediated lipolysis and builds on the body of evidence demonstrating a critical role for PLIN5 in coordinating lipid and glucose metabolism.

GIP receptor deletion in mice confers resistance to high-fat diet-induced obesity via alterations in energy expenditure and adipose tissue lipid metabolism.

Glucose-dependent insulinotropic polypeptide (GIP) is best known as an incretin hormone that is secreted from K-cells of the proximal intestine, but evidence also implicates a role for GIP in regulating lipid metabolism and adiposity. It is well-established that GIP receptor knockout (GIPR KO) mice are resistant to diet-induced obesity; however, the factors mediating this effect remain unresolved. Accordingly, we aimed to elucidate the mechanisms leading to adiposity resistance in GIPR KO mice with a focus on whole-body energy balance and lipid metabolism in adipose tissues. Studies were conducted in age-matched male GIPR KO and wild-type (WT) mice fed a high-fat diet for 10 weeks. GIPR KO mice gained less body weight and fat mass compared to WT littermates, and this was associated with increased energy expenditure but no differences in food intake or fecal energy loss. Upon an oral lipid challenge, fatty acid storage in inguinal adipose tissue was significantly increased in GIPR KO compared with WT mice. This was not related to differential expression of lipoprotein lipase in adipose tissue. Adipose tissue lipolysis was increased in GIPR KO compared with WT mice, particularly following ß-adrenergic stimulation, and could explain why GIPR KO mice gain less adipose tissue despite increased rates of fatty acid storage in inguinal adipose tissue. Taken together, these results suggest that the GIPR is required for normal maintenance of body weight and adipose tissue mass by regulating energy expenditure and lipolysis.NEW & NOTEWORTHY GIPR KO mice fed a high-fat diet have reduced adiposity despite transporting more ingested lipids into adipose tissue. This can be partly explained by accelerated adipose tissue lipolysis and increased energy expenditure in GIPR KO mice. These new insights rationalize targeting the GIPR as part of a weight management strategy in obesity.

Selection of a novel AAV2/TNFAIP3 vector for local suppression of islet xenograft inflammation.

BACKGROUND: Neonatal porcine islets (NPIs) can restore glucose control in mice, pigs, and non-human primates, representing a potential abundant alternative islet supply for clinical beta cell replacement therapy. However, NPIs are vulnerable to inflammatory insults that could be overcome with genetic modifications. Here, we demonstrate in a series of proof-of-concept experiments the potential of the cytoplasmic ubiquitin-editing protein A20, encoded by the TNFAIP3 gene, as an NPI cytoprotective gene. METHODS: We forced A20 expression in NPI grafts using a recombinant adenovirus 5 (Ad5) vector and looked for impact on TNF-stimulated NF-κB activation and NPI graft function. As adeno-associated vectors (AAV) are clinically preferred vectors but exhibit poor transduction efficacy in NPIs, we next screened a series of AAV serotypes under different transduction protocols for their ability achieve high transduction efficiency and suppress NPI inflammation without impacting NPI maturation. RESULTS: Forcing the expression of A20 in NPI with Ad5 vector blocked NF-κB activation by inhibiting IκBα phosphorylation and degradation, and reduced the induction of pro-inflammatory genes Cxcl10 and Icam1. A20-expressing NPIs also exhibited superior functional capacity when transplanted into diabetic immunodeficient recipient mice, evidenced by a more rapid return to euglycemia and improved GTT compared to unmodified NPI grafts. We found AAV2 combined with a 14-day culture period maximized NPI transduction efficiency (>70% transduction rate), and suppressed NF-κB-dependent gene expression without adverse impact upon NPI maturation. CONCLUSION: We report a new protocol that allows for high-efficiency genetic modification of NPIs, which can be utilized to introduce candidate genes without the need for germline engineering. This approach would be suitable for preclinical and clinical testing of beneficial molecules. We also report for the first time that A20 is cytoprotective for NPI, such that A20 gene therapy could aid the clinical development of NPIs for beta cell replacement.