Fat composition in infant formula contributes to the severity of necrotising enterocolitis.

Necrotising enterocolitis (NEC) is a devastating disease that typically affects formula-fed premature infants, suggesting that dietary components may influence disease pathogenesis. TAG are the major fat components of infant formula, and their digestion requires pancreatic lipases, which may be naturally deficient in premature neonates. We hypothesise that NEC develops partly from the accumulation of incompletely digested long-chain TAG-containing unsaturated fatty acids within the intestinal epithelial cells, leading to oxidative stress and enterocyte damage. We further hypothesise that the administration of a formula that contains reduced TAG ('pre-digested fat') that do not require lipase action may reduce NEC severity. To test these hypotheses, we induced NEC in neonatal mice using three different fat formulations, namely 'standard fat', 'pre-digested fat' or 'very low fat', and determined that mice fed 'standard fat' developed severe NEC, which was significantly reduced in mice fed 'pre-digested fat' or 'very low fat'. The expression level of the critical fat-digesting enzyme carboxyl ester lipase was significantly lower in the newborn compared with older pups, leading to impaired fat digestion. The accumulation of mal-digested fat resulted in the significant accumulation of fat droplets within the intestinal epithelium of the distal ileum, resulting in the generation of reactive oxygen species and intestinal inflammation. Strikingly, these changes were prevented in pups fed 'pre-digested fat' or 'very low fat' formulas. These findings suggest that nutritional formula containing a pre-digested fat system may overcome the natural lipase deficiency of the premature gut, and serve as a novel approach to prevent NEC.

Pulmonary Epithelial TLR4 Activation Leads to Lung Injury in Neonatal Necrotizing Enterocolitis.

We seek to define the mechanisms leading to the development of lung disease in the setting of neonatal necrotizing enterocolitis (NEC), a life-threatening gastrointestinal disease of premature infants characterized by the sudden onset of intestinal necrosis. NEC development in mice requires activation of the LPS receptor TLR4 on the intestinal epithelium, through its effects on modulating epithelial injury and repair. Although NEC-associated lung injury is more severe than the lung injury that occurs in premature infants without NEC, the mechanisms leading to its development remain unknown. In this study, we now show that TLR4 expression in the lung gradually increases during postnatal development, and that mice and humans with NEC-associated lung inflammation express higher levels of pulmonary TLR4 than do age-matched controls. NEC in wild-type newborn mice resulted in significant pulmonary injury that was prevented by deletion of TLR4 from the pulmonary epithelium, indicating a role for pulmonary TLR4 in lung injury development. Mechanistically, intestinal epithelial TLR4 activation induced high-mobility group box 1 release from the intestine, which activated pulmonary epithelial TLR4, leading to the induction of the neutrophil recruiting CXCL5 and the influx of proinflammatory neutrophils to the lung. Strikingly, the aerosolized administration of a novel carbohydrate TLR4 inhibitor prevented CXCL5 upregulation and blocked NEC-induced lung injury in mice. These findings illustrate the critical role of pulmonary TLR4 in the development of NEC-associated lung injury, and they suggest that inhibition of this innate immune receptor in the neonatal lung may prevent this devastating complication of NEC.

Valproic Acid Limits Pancreatic Recovery after Pancreatitis by Inhibiting Histone Deacetylases and Preventing Acinar Redifferentiation Programs.

The mechanisms by which drugs induce pancreatitis are unknown. A definite cause of pancreatitis is due to the antiepileptic drug valproic acid (VPA). On the basis of three crucial observations-that VPA inhibits histone deacetylases (HDACs), HDACs mediate pancreas development, and aspects of pancreas development are recapitulated during recovery of the pancreas after injury-we hypothesized that VPA does not cause injury on its own, but it predisposes patients to pancreatitis by inhibiting HDACs and provoking an imbalance in pancreatic recovery. In an experimental model of pancreatic injury, we found that VPA delayed recovery of the pancreas and reduced acinar cell proliferation. In addition, pancreatic expression of class I HDACs (which are the primary VPA targets) increased in the midphase of pancreatic recovery. VPA administration inhibited pancreatic HDAC activity and led to the persistence of acinar-to-ductal metaplastic complexes, with prolonged Sox9 expression and sustained ß-catenin nuclear activation, findings that characterize a delay in regenerative reprogramming. These effects were not observed with valpromide, an analog of VPA that lacks HDAC inhibition. This is the first report, to our knowledge, that VPA shifts the balance toward pancreatic injury and pancreatitis through HDAC inhibition. The work also identifies a new paradigm for therapies that could exploit epigenetic reprogramming to enhance pancreatic recovery and disorders of pancreatic injury.

Recurrent acute liver failure due to NBAS deficiency: phenotypic spectrum, disease mechanisms, and therapeutic concepts.

BACKGROUND: Acute liver failure (ALF) in infancy and childhood is a life-threatening emergency and in about 50% the etiology remains unknown. Recently biallelic mutations in NBAS were identified as a new molecular cause of ALF with onset in infancy, leading to recurrent acute liver failure (RALF). METHODS: The phenotype and medical history of 14 individuals with NBAS deficiency was studied in detail and functional studies were performed on patients' fibroblasts. RESULTS: The phenotypic spectrum of NBAS deficiency ranges from isolated RALF to a multisystemic disease with short stature, skeletal dysplasia, immunological abnormalities, optic atrophy, and normal motor and cognitive development resembling SOPH syndrome. Liver crises are triggered by febrile infections; they become less frequent with age but are not restricted to childhood. Complete recovery is typical, but ALF crises can be fatal. Antipyretic therapy and induction of anabolism including glucose and parenteral lipids effectively ameliorates the course of liver crises. Patients' fibroblasts showed an increased sensitivity to high temperature at protein and functional level and a disturbed tethering of vesicles, pointing at a defect of intracellular transport between the endoplasmic reticulum and Golgi. CONCLUSIONS: Mutations in NBAS cause a complex disease with a wide clinical spectrum ranging from isolated RALF to a multisystemic phenotype. Thermal susceptibility of the syntaxin 18 complex is the basis of fever dependency of ALF episodes. NBAS deficiency is the first disease related to a primary defect of retrograde transport. Identification of NBAS deficiency allows optimized therapy of liver crises and even prevention of further episodes.

The human milk oligosaccharide 2'-fucosyllactose attenuates the severity of experimental necrotising enterocolitis by enhancing mesenteric perfusion in the neonatal intestine.

Necrotising enterocolitis (NEC) is a common disease in premature infants characterised by intestinal ischaemia and necrosis. The only effective preventative strategy against NEC is the administration of breast milk, although the protective mechanisms remain unknown. We hypothesise that an abundant human milk oligosaccharide (HMO) in breast milk, 2'-fucosyllactose (2'FL), protects against NEC by enhancing intestinal mucosal blood flow, and we sought to determine the mechanisms underlying this protection. Administration of HMO-2'FL protected against NEC in neonatal wild-type mice, resulted in a decrease in pro-inflammatory markers and preserved the small intestinal mucosal architecture. These protective effects occurred via restoration of intestinal perfusion through up-regulation of the vasodilatory molecule endothelial nitric oxide synthase (eNOS), as administration of HMO-2'FL to eNOS-deficient mice or to mice that received eNOS inhibitors did not protect against NEC, and by 16S analysis HMO-2'FL affected the microbiota of the neonatal mouse gut, although these changes do not seem to be the primary mechanism of protection. Induction of eNOS by HMO-2'FL was also observed in cultured endothelial cells, providing a link between eNOS and HMO in the endothelium. These data demonstrate that HMO-2'FL protects against NEC in part through maintaining mesenteric perfusion via increased eNOS expression, and suggest that the 2'FL found in human milk may be mediating some of the protective benefits of breast milk in the clinical setting against NEC.

A continuous linear optimal transport approach for pattern analysis in image datasets.

We present a new approach to facilitate the application of the optimal transport metric to pattern recognition on image databases. The method is based on a linearized version of the optimal transport metric, which provides a linear embedding for the images. Hence, it enables shape and appearance modeling using linear geometric analysis techniques in the embedded space. In contrast to previous work, we use Monge's formulation of the optimal transport problem, which allows for reasonably fast computation of the linearized optimal transport embedding for large images. We demonstrate the application of the method to recover and visualize meaningful variations in a supervised-learning setting on several image datasets, including chromatin distribution in the nuclei of cells, galaxy morphologies, facial expressions, and bird species identification. We show that the new approach allows for high-resolution construction of modes of variations and discrimination and can enhance classification accuracy in a variety of image discrimination problems.

Toll-like receptor 4-mediated endoplasmic reticulum stress in intestinal crypts induces necrotizing enterocolitis.

The cellular cues that regulate the apoptosis of intestinal stem cells (ISCs) remain incompletely understood, yet may play a role in diseases characterized by ISC loss including necrotizing enterocolitis (NEC). Toll-like receptor-4 (TLR4) was recently found to be expressed on ISCs, where its activation leads to ISC apoptosis through mechanisms that remain incompletely explained. We now hypothesize that TLR4 induces endoplasmic reticulum (ER) stress within ISCs, leading to their apoptosis in NEC pathogenesis, and that high ER stress within the premature intestine predisposes to NEC development. Using transgenic mice and cultured enteroids, we now demonstrate that TLR4 induces ER stress within Lgr5 (leucine-rich repeat-containing G-protein-coupled receptor 5)-positive ISCs, resulting in crypt apoptosis. TLR4 signaling within crypts was required, because crypt ER stress and apoptosis occurred in TLR4(ΔIEC-OVER) mice expressing TLR4 only within intestinal crypts and epithelium, but not TLR4(ΔIEC) mice lacking intestinal TLR4. TLR4-mediated ER stress and apoptosis of ISCs required PERK (protein kinase-related PKR-like ER kinase), CHOP (C/EBP homologous protein), and MyD88 (myeloid differentiation primary response gene 88), but not ATF6 (activating transcription factor 6) or XBP1 (X-box-binding protein 1). Human and mouse NEC showed high crypt ER stress and apoptosis, whereas genetic inhibition of PERK or CHOP attenuated ER stress, crypt apoptosis, and NEC severity. Strikingly, using intragastric delivery into fetal mouse intestine, prevention of ER stress reduced TLR4-mediated ISC apoptosis and mucosal disruption. These findings identify a novel link between TLR4-induced ER stress and ISC apoptosis in NEC pathogenesis and suggest that increased ER stress within the premature bowel predisposes to NEC development.

Lactobacillus rhamnosus HN001 decreases the severity of necrotizing enterocolitis in neonatal mice and preterm piglets: evidence in mice for a role of TLR9.

Necrotizing enterocolitis (NEC) is the leading cause of death from gastrointestinal disease in premature infants and develops partly from an exaggerated intestinal epithelial immune response to indigenous microbes. There has been interest in administering probiotic bacteria to reduce NEC severity, yet concerns exist regarding infection risk. Mechanisms of probiotic activity in NEC are unknown although activation of the microbial DNA receptor Toll-like receptor-9 (TLR9) has been postulated. We now hypothesize that the Gram-positive bacterium Lactobacillus rhamnosus HN001 can attenuate NEC in small and large animal models, that its microbial DNA is sufficient for its protective effects, and that protection requires activation of the Toll-like receptor 9 (TLR9). We now show that oral administration of live or UV-inactivated Lactobacillus rhamnosus HN001 attenuates NEC severity in newborn mice and premature piglets, as manifest by reduced histology score, attenuation of mucosal cytokine response, and improved gross morphology. TLR9 was required for Lactobacillus rhamnosus-mediated protection against NEC in mice, as the selective decrease of TLR9 from the intestinal epithelium reversed its protective effects. Strikingly, DNA of Lactobacillus rhamnosus HN001 reduced the extent of proinflammatory signaling in cultured enterocytes and in samples of resected human ileum ex vivo, suggesting the therapeutic potential of this probiotic in clinical NEC. Taken together, these findings illustrate that Lactobacillus rhamnosus HN001 is an effective probiotic for NEC via activation of the innate immune receptor TLR9 and that Lactobacillus rhamnosus DNA is sufficient for its protective effects, potentially reducing concerns regarding the infectious risk of this novel therapeutic approach.

Neurofibromas with imaging characteristics resembling vascular anomalies.

OBJECTIVE: Although neurofibromas are rare, their initial clinical and imaging presentation can mimic those of vascular anomalies, particularly if the characteristic clinical features of neurofibromatosis are not present. The diagnostic challenges encountered in five cases of histologically proven neurofibromas, initially diagnosed as vascular anomalies, are reviewed and discussed. CONCLUSION: The clinical and imaging differences between neurofibromas and vascular anomalies are detailed with the histopathologic features to better understand why some neurofibromas are diagnosed as vascular anomalies.

Cancer diagnosis by nuclear morphometry using spatial information .

Methods for extracting quantitative information regarding nuclear morphology from histopathology images have been long used to aid pathologists in determining the degree of differentiation in numerous malignancies. Most methods currently in use, however, employ the naïve Bayes approach to classify a set of nuclear measurements extracted from one patient. Hence, the statistical dependency between the samples (nuclear measurements) is often not directly taken into account. Here we describe a method that makes use of statistical dependency between samples in thyroid tissue to improve patient classification accuracies with respect to standard naïve Bayes approaches. We report results in two sample diagnostic challenges.

Intestinal epithelial Toll-like receptor 4 regulates goblet cell development and is required for necrotizing enterocolitis in mice.

BACKGROUND & AIMS: Little is known about factors that regulate intestinal epithelial differentiation; microbial recognition receptors such as Toll-like receptor (TLR)4 might be involved. We investigated whether intestinal TLR4 regulates epithelial differentiation and is involved in development of necrotizing enterocolitis (NEC) of the immature intestine. METHODS: Mice with conditional disruption of TLR4 in the intestinal epithelium and TLR4 knockout (TLR4(-/-)) mice were generated by breeding TLR4(loxp/loxp) mice with villin-cre and Ella-cre, respectively. Enterocytes that did not express or overexpressed TLR4 were created by lentiviral or adenoviral transduction. Intestinal organoids were cultured on tissue matrices. Bile acids were measured by colorimetric assays, and microbial composition was determined by 16S pyrosequencing. NEC was induced in 7- to 10-day-old mice by induction of hypoxia twice daily for 4 days. RESULTS: TLR4(-/-) mice and mice with enterocyte-specific deletion of TLR4 were protected from NEC; epithelial differentiation into goblet cells was increased via suppressed Notch signaling in the small intestinal epithelium. TLR4 also regulates differentiation of goblet cells in intestinal organoid and enterocyte cell cultures; differentiation was increased on deletion of TLR4 and restored when TLR4 was expressed ectopically. TLR4 signaling via Notch was increased in intestinal tissue samples from patients with NEC, and numbers of goblet cells were reduced. 16S pyrosequencing revealed that wild-type and TLR4-deficient mice had similar microbial profiles; increased numbers of goblet cells were observed in mice given antibiotics. TLR4 deficiency reduced levels of luminal bile acids in vivo, and addition of bile acids to TLR4-deficient cell cultures prevented differentiation of goblet cells. CONCLUSIONS: TLR4 signaling and Notch are increased in intestinal tissues of patients with NEC and required for induction of NEC in mice. TLR4 prevents goblet cell differentiation, independently of the microbiota. Bile acids might initiate goblet cell development.

A flexible and robust approach for segmenting cell nuclei from 2D microscopy images using supervised learning and template matching.

We describe a new supervised learning-based template matching approach for segmenting cell nuclei from microscopy images. The method uses examples selected by a user for building a statistical model that captures the texture and shape variations of the nuclear structures from a given dataset to be segmented. Segmentation of subsequent, unlabeled, images is then performed by finding the model instance that best matches (in the normalized cross correlation sense) local neighborhood in the input image. We demonstrate the application of our method to segmenting nuclei from a variety of imaging modalities, and quantitatively compare our results to several other methods. Quantitative results using both simulated and real image data show that, while certain methods may work well for certain imaging modalities, our software is able to obtain high accuracy across several imaging modalities studied. Results also demonstrate that, relative to several existing methods, the template-based method we propose presents increased robustness in the sense of better handling variations in illumination, variations in texture from different imaging modalities, providing more smooth and accurate segmentation borders, as well as handling better cluttered nuclei.

A linear optimal transportation framework for quantifying and visualizing variations in sets of images.

Transportation-based metrics for comparing images have long been applied to analyze images, especially where one can interpret the pixel intensities (or derived quantities) as a distribution of 'mass' that can be transported without strict geometric constraints. Here we describe a new transportation-based framework for analyzing sets of images. More specifically, we describe a new transportation-related distance between pairs of images, which we denote as linear optimal transportation (LOT). The LOT can be used directly on pixel intensities, and is based on a linearized version of the Kantorovich-Wasserstein metric (an optimal transportation distance, as is the earth mover's distance). The new framework is especially well suited for computing all pairwise distances for a large database of images efficiently, and thus it can be used for pattern recognition in sets of images. In addition, the new LOT framework also allows for an isometric linear embedding, greatly facilitating the ability to visualize discriminant information in different classes of images. We demonstrate the application of the framework to several tasks such as discriminating nuclear chromatin patterns in cancer cells, decoding differences in facial expressions, galaxy morphologies, as well as sub cellular protein distributions.

Transport-based morphometry of nuclear structures of digital pathology images in cancers.

Alterations in nuclear morphology are useful adjuncts and even diagnostic tools used by pathologists in the diagnosis and grading of many tumors, particularly malignant tumors. Large datasets such as TCGA and the Human Protein Atlas, in combination with emerging machine learning and statistical modeling methods, such as feature extraction and deep learning techniques, can be used to extract meaningful knowledge from images of nuclei, particularly from cancerous tumors. Here we describe a new technique based on the mathematics of optimal transport for modeling the information content related to nuclear chromatin structure directly from imaging data. In contrast to other techniques, our method represents the entire information content of each nucleus relative to a template nucleus using a transport-based morphometry (TBM) framework. We demonstrate the model is robust to different staining patterns and imaging protocols, and can be used to discover meaningful and interpretable information within and across datasets and cancer types. In particular, we demonstrate morphological differences capable of distinguishing nuclear features along the spectrum from benign to malignant categories of tumors across different cancer tissue types, including tumors derived from liver parenchyma, thyroid gland, lung mesothelium, and skin epithelium. We believe these proof of concept calculations demonstrate that the TBM framework can provide the quantitative measurements necessary for performing meaningful comparisons across a wide range of datasets and cancer types that can potentially enable numerous cancer studies, technologies, and clinical applications and help elevate the role of nuclear morphometry into a more quantitative science. The source codes implementing our method is available at https://github.com/rohdelab/nuclear_morphometry.

Valproic acid-associated acute liver failure in children: case report and analysis of liver transplantation outcomes in the United States.

OBJECTIVE: To determine whether valproic acid (VPA)-associated acute liver failure (ALF; VPA-ALF) explains the poor outcomes after liver transplantation (LT) in children. STUDY DESIGN: Organ Procurement and Transplantation Network data of pediatric patients who underwent LT for VPA-ALF and ALF caused by other drugs (non-VPA-drug-induced acute liver failure [DIALF]) were analyzed. Pre- and post-transplant variables and post-LT survival were compared between VPA-ALF and non-VPA-DIALF. RESULTS: Seventeen children were transplanted for VPA-ALF. Of the 17 children, 82% died within 1 year of LT. Pre- and post-transplant parameters of VPA versus non-VPA-DIALF were comparable with two exceptions. The median alanine aminotransferase level at transplant was remarkably lower in VPA-ALF compared with non-VPA-DIALF (45 versus 1179 IU/L, P = .004). One-year survival probability was worse in VPA-ALF than non-VPA-DIALF (20% versus 69%, P < .0001). Median post-LT survival time for VPA-ALF was 2.8 months. CONCLUSION: Children who underwent LT for VPA-ALF had a significantly lower survival probability than children with non-VPA-DIALF. Current data suggest that VPA-ALF in children represents an "unmasking" of mitochondrial disease. VPA-ALF should be a contraindication for LT, even in the absence of a documented mitochondrial disease.

Reciprocal expression and signaling of TLR4 and TLR9 in the pathogenesis and treatment of necrotizing enterocolitis.

Necrotizing enterocolitis (NEC) is a common and often fatal inflammatory disorder affecting preterm infants that develops upon interaction of indigenous bacteria with the premature intestine. We now demonstrate that the developing mouse intestine shows reciprocal patterns of expression of TLR4 and TLR9, the receptor for bacterial DNA (CpG-DNA). Using a novel ultrasound-guided in utero injection system, we administered LPS directly into the stomachs of early and late gestation fetuses to induce TLR4 signaling and demonstrated that TLR4-mediated signaling within the developing intestine follows its expression pattern. Murine and human NEC were associated with increased intestinal TLR4 and decreased TLR9 expression, suggesting that reciprocal TLR4 and TLR9 signaling may occur in the pathogenesis of NEC. Enteral administration of adenovirus expressing mutant TLR4 to neonatal mice reduced the severity of NEC and increased TLR9 expression within the intestine. Activation of TLR9 with CpG-DNA inhibited LPS-mediated TLR4 signaling in enterocytes in a mechanism dependent upon the inhibitory molecule IRAK-M. Strikingly, TLR9 activation with CpG-DNA significantly reduced NEC severity, whereas TLR9-deficient mice exhibited increased NEC severity. Thus, the reciprocal nature of TLR4 and TLR9 signaling within the neonatal intestine plays a role in the development of NEC and provides novel therapeutic approaches to this disease.

Penalized Fisher Discriminant Analysis and Its Application to Image-Based Morphometry.

Image-based morphometry is an important area of pattern recognition research, with numerous applications in science and technology (including biology and medicine). Fisher Linear Discriminant Analysis (FLDA) techniques are often employed to elucidate and visualize important information that discriminates between two or more populations. We demonstrate that the direct application of FLDA can lead to undesirable errors in characterizing such information and that the reason for such errors is not necessarily the ill conditioning in the resulting generalized eigenvalue problem, as usually assumed. We show that the regularized eigenvalue decomposition often used is related to solving a modified FLDA criterion that includes a least-squares-type representation penalty, and derive the relationship explicitly. We demonstrate the concepts by applying this modified technique to several problems in image-based morphometry, and build discriminant representative models for different data sets.

Selected Topics in the Pathology of the Thyroid and Parathyroid Glands in Children and Adolescents.

The goals of this chapter in keeping with the overall general themes of this special edition will be (1) to highlight aspects of development of the thyroid and parathyroid glands with particular focus on the role and contribution of the neural crest (or not) and how this may impact on the pathology that is seen, (2) to emphasize those lesions particularly more commonly arising in the pediatric population that actually generate specimens that the surgical pathologist would encounter, and (3) highlight more in depth specific lesions associated with heritable syndromes or specific gene mutations since the heritable syndromes tends to manifest in the pediatric age group. In this light, the other interesting areas of pediatric thyroid disease including medical thyroid diseases, congenital hypothyroidism, anatomic variants and aberrations of development that lead to structural anomalies will not be emphasized here.

THE "-OMAS" and "-OPIAS": Targeted and Philosophical Considerations Regarding Hamartomas, Choristomas, Teratomas, Ectopias, and Heterotopias in Pediatric Otorhinolaryngologic Pathology.

The spectrum of "developmental" lesions that occur in the head and neck predominantly congenital in origin and arising at birth and/or discovered in childhood is broad and fascinating. These have been grouped into categories such as "ectopias", "heterotopias", "hamartomas", and "choristomas". On a philosophical and consequently systematic level, these lesions, mostly benign tumors seem to lack a true understanding of the pathogenetic foundation on which to base a more unified taxonomic designation. In this review, we will consider some of these select tumors as they represent syndromic associations (nasal chondromesenchymal hamartoma and DICER1 syndrome), the lingual choristoma from the perspective of its nomenclature and classification, lesions with ectopic meningothelial elements, and teratomas and the enigmatic "hairy polyp" in reference to a broader discussion of pathogenesis and pluripotent cells in the head and neck. A consistent thread will be how these lesions are designated with some final thoughts on future directions regarding the investigation of their pathogenesis and taxonomic nomenclature.

Deletion of AMPK minimizes graft-versus-host disease through an early impact on effector donor T cells.

Allogeneic hematopoietic stem cell transplantation is a viable treatment for multiple hematologic diseases, but its application is often limited by graft-versus-host disease (GVHD), where donor T cells attack host tissues in the skin, liver, and gastrointestinal tract. Here, we examined the role of the cellular energy sensor AMP kinase (AMPK) in alloreactive T cells during GVHD development. Early posttransplant, AMPK activity increased more than 15-fold in allogeneic T cells, and transplantation of T cells deficient in both AMPKα1 and AMPKα2 decreased GVHD severity in multiple disease models. Importantly, a lack of AMPK lessened GVHD without compromising antileukemia responses or impairing lymphopenia-driven immune reconstitution. Mechanistically, absence of AMPK decreased both CD4+ and CD8+ effector T cell numbers as early as day 3 posttransplant, while simultaneously increasing regulatory T cell (Treg) percentages. Improvements in GVHD resulted from cell-intrinsic perturbations in conventional effector T cells as depletion of donor Tregs had minimal impact on AMPK-related improvements. Together, these results highlight a specific role for AMPK in allogeneic effector T cells early posttransplant and suggest that AMPK inhibition may be an innovative approach to mitigate GVHD while preserving graft-versus-leukemia responses and maintaining robust immune reconstitution.