Lactate transport inhibition therapeutically reprograms fibroblast metabolism in experimental pulmonary fibrosis.

Myofibroblast differentiation, essential for driving extracellular matrix synthesis in pulmonary fibrosis, requires increased glycolysis. While glycolytic cells must export lactate, the contributions of lactate transporters to myofibroblast differentiation are unknown. In this study, we investigated how MCT1 and MCT4, key lactate transporters, influence myofibroblast differentiation and experimental pulmonary fibrosis. Our findings reveal that inhibiting MCT1 or MCT4 reduces TGFß-stimulated pulmonary myofibroblast differentiation in vitro and decreases bleomycin-induced pulmonary fibrosis in vivo. Through comprehensive metabolic analyses, including bioenergetics, stable isotope tracing, metabolomics, and imaging mass spectrometry in both cells and mice, we demonstrate that inhibiting lactate transport enhances oxidative phosphorylation, reduces reactive oxygen species production, and diminishes glucose metabolite incorporation into fibrotic lung regions. Furthermore, we introduce VB253, a novel MCT4 inhibitor, which ameliorates pulmonary fibrosis in both young and aged mice, with comparable efficacy to established antifibrotic therapies. These results underscore the necessity of lactate transport for myofibroblast differentiation, identify MCT1 and MCT4 as promising pharmacologic targets in pulmonary fibrosis, and support further evaluation of lactate transport inhibitors for patients for whom limited therapeutic options currently exist.

JAK2 R683S Mutation Resulting in Dual Diagnoses of Chronic Eosinophilic Leukemia and Myelodysplastic/Myeloproliferative Overlap Syndrome.

A 66-year-old male presented with hypereosinophilia, thrombocytosis, extensive thrombosis refractory to direct oral anticoagulant therapy, and evidence of end-organ damage, including rash, splenic infarcts, and pulmonary infiltrates. Bone marrow biopsy revealed myeloid malignancy consistent with both chronic eosinophilic leukemia and myelodysplastic/myeloproliferative neoplasms (MDS/MPN) with SF3B1 mutation and thrombocytosis. Next-generation sequencing of the patient's eosinophils and neutrophil compartments revealed pathologic variants in EZH2 and SF3B1 in addition to a noncanonical JAK2 R683S mutation that has not been previously described in myeloproliferative disorders or other chronic myeloid neoplasms. These mutations were not present in the patient's lymphoid cell fraction, suggesting that the hematopoietic malignancy arose in a myeloid-committed progenitor cell. Based on this case and previous work from our group, we propose that noncanonical JAK2 mutations may permit signal transduction that biases toward eosinophilic differentiation in chronic myeloid neoplasms. Although the patient's blood counts initially responded to ruxolitinib and hydroxyurea, the response was not durable. Early referral for allogenic bone marrow transplant appears necessary to prevent long-term complications and disease progression in myeloid neoplasms with clonal hypereosinophilia driven by noncanonical JAK2 mutations.

The impact of antibiotic allergy labels on antibiotic exposure, clinical outcomes, and healthcare costs: A systematic review.

OBJECTIVE: A growing body of evidence suggests that antibiotic allergy labels as documented in medical records are a risk factor for poor clinical outcomes. In this systematic review, we aimed to determine how antibiotic allergy labels influence 3 domains: antibiotic use and exposure, clinical outcomes, and healthcare-related costs. DESIGN: We performed a systematic review to identify studies reporting outcomes in patients with antibiotic allergy labels compared to nonallergic counterparts. The search included PubMed, EMBASE, Cochrane CENTRAL, EBSCO, Cochrane Database of Abstracts of Reviews of Effects and Web of Science. Two reviewers independently screened studies for inclusion and abstracted data. Studies were graded using the Newcastle-Ottawa quality assessment scale. Study outcomes included antibiotic use, clinical outcomes, and economic outcomes. RESULTS: In total, 41 studies met our criteria for inclusion. These studies varied in medical specialty, patient population, healthcare delivery system, and design, but most were conducted among adults age >18 years (85%) in the inpatient setting (82.5%). Among 34 studies examining antibiotic exposure, 32 (94%) found that patients with antibiotic allergy labels received more broad-spectrum antibiotics. Moreover, 31 studies examined clinical outcomes such as length of hospitalization, ICU admission, hospital readmission, multidrug-resistant or opportunistic infection, or mortality, and 27 (87%) found that allergy-labeled patients had at least 1 negative outcome. Of 9 studies examining healthcare costs, 7 (78%) found that allergy-labeled patients incurred significantly higher drug or hospital-related costs. CONCLUSIONS: Antibiotic allergy labels have negative effects on antibiotic use, clinical outcomes, and economic outcomes in a variety of clinical settings and populations.

Regulation of Tumor Initiation by the Mitochondrial Pyruvate Carrier.

Although metabolic adaptations have been demonstrated to be essential for tumor cell proliferation, the metabolic underpinnings of tumor initiation are poorly understood. We found that the earliest stages of colorectal cancer (CRC) initiation are marked by a glycolytic metabolic signature, including downregulation of the mitochondrial pyruvate carrier (MPC), which couples glycolysis and glucose oxidation through mitochondrial pyruvate import. Genetic studies in Drosophila suggest that this downregulation is required because hyperplasia caused by loss of the Apc or Notch tumor suppressors in intestinal stem cells can be completely blocked by MPC overexpression. Moreover, in two distinct CRC mouse models, loss of Mpc1 prior to a tumorigenic stimulus doubled the frequency of adenoma formation and produced higher grade tumors. MPC loss was associated with a glycolytic metabolic phenotype and increased expression of stem cell markers. These data suggest that changes in cellular pyruvate metabolism are necessary and sufficient to promote cancer initiation.

Prevention and Reversion of Pancreatic Tumorigenesis through a Differentiation-Based Mechanism.

Activating mutations in Kras are nearly ubiquitous in human pancreatic cancer and initiate precancerous pancreatic intraepithelial neoplasia (PanINs) when induced in mouse acinar cells. PanINs normally take months to form but are accelerated by deletion of acinar cell differentiation factors such as Ptf1a, suggesting that loss of cell identity is rate limiting for pancreatic tumor initiation. Using a genetic mouse model that allows for independent control of oncogenic Kras and Ptf1a expression, we demonstrate that sustained Ptf1a is sufficient to prevent Kras-driven tumorigenesis, even in the presence of tumor-promoting inflammation. Furthermore, reintroducing Ptf1a into established PanINs reverts them to quiescent acinar cells in vivo. Similarly, Ptf1a re-expression in human pancreatic cancer cells inhibits their growth and colony-forming ability. Our results suggest that reactivation of an endogenous differentiation program can prevent and reverse oncogene-driven transformation in cells harboring tumor-driving mutations, introducing a potential paradigm for solid tumor prevention and treatment.

Economic Burden of Home Antimicrobial Therapy: OPAT Versus Oral Therapy.

BACKGROUND: There is increasing evidence that outpatient parenteral antimicrobial therapy (OPAT) is overused for children and that outcomes with oral therapy are equivalent. Our objective was to compare economic burden between OPAT and oral therapy, accounting for direct and indirect costs and caregiver quality of life (QoL). METHODS: We conducted a prospective cohort study of caregivers for children after hospitalization who were treated with prolonged antimicrobial therapy. We collected data about missed work and school and time spent administering therapy. Caregivers completed the Pediatric Quality of Life Inventory to assess QoL. Clinical information included length of stay, treatment indication, and type of therapy (OPAT versus oral therapy). Direct medical costs were obtained by using a microcosting system and accounted for medication, supplies, and home-nursing visits. The primary cost outcome was the mean daily cost of therapy. Multivariable models were developed to adjust for potential confounders. RESULTS: Two hundred and twelve caregivers completed surveys: 123 (58%) for oral therapy and 89 (42%) for OPAT. Caregivers administering OPAT reported more missed work, missed school for their children, time with daily medication administration (90 vs 6 minutes; P < .01) and lower QoL scores (77.8 vs 68.9) than caregivers administering oral therapy. The mean daily cost was $65 (95% confidence interval: $51-$78) for OPAT and $7 (95% confidence interval: $4-$9) for oral therapy. Relative differences in cost and QoL between groups did not change after model adjustment. CONCLUSIONS: The overall burden of OPAT is substantially higher than that of oral therapy, including higher direct and indirect costs and greater impact on caregiver QoL. These findings strongly support efforts to use oral therapy in place of OPAT when clinically appropriate.

Outpatient Parenteral Antimicrobial Therapy in Young Infants.

We examined clinical outcomes for 53 young infants (<3 months of age) treated with outpatient parenteral antimicrobial therapy after discharge from a freestanding children's hospital. None of the patients experienced treatment failure or disease progression; 9% of them experienced a catheter-related complication, but this percentage is not different than that for older children.

EWS/FLI is a Master Regulator of Metabolic Reprogramming in Ewing Sarcoma.

Ewing sarcoma is a bone malignancy driven by a translocation event resulting in the fusion protein EWS/FLI1 (EF). EF functions as an aberrant and oncogenic transcription factor that misregulates the expression of thousands of genes. Previous work has focused principally on determining important transcriptional targets of EF, as well as characterizing important regulatory partnerships in EF-dependent transcriptional programs. Less is known, however, about EF-dependent metabolic changes or their role in Ewing sarcoma biology. Therefore, the metabolic effects of silencing EF in Ewing sarcoma cells were determined. Metabolomic analyses revealed distinct separation of metabolic profiles in EF-knockdown versus control-knockdown cells. Mitochondrial stress tests demonstrated that knockdown of EF increased respiratory as well as glycolytic functions. Enzymes and metabolites in several metabolic pathways were altered, including de novo serine synthesis and elements of one-carbon metabolism. Furthermore, phosphoglycerate dehydrogenase (PHGDH) was found to be highly expressed in Ewing sarcoma and correlated with worse patient survival. PHGDH knockdown or pharmacologic inhibition in vitro caused impaired proliferation and cell death. Interestingly, PHGDH modulation also led to elevated histone expression and methylation. These studies demonstrate that the translocation-derived fusion protein EF is a master regulator of metabolic reprogramming in Ewing sarcoma, diverting metabolites toward biosynthesis. As such, these data suggest that the metabolic aberrations induced by EF are important contributors to the oncogenic biology of these tumors.Implications: This previously unexplored role of EWS/FLI1-driven metabolic changes expands the understanding of Ewing sarcoma biology, and has potential to significantly inform development of therapeutic strategies. Mol Cancer Res; 15(11); 1517-30. ©2017 AACR.

Differentiation and Inflammation: 'Best Enemies' in Gastrointestinal Carcinogenesis.

While recent studies demonstrate that cancer can arise from mutant stem cells, this hypothesis does not explain why tissues without defined stem cell populations are susceptible to inflammation-driven tumorigenesis. We propose that chronic inflammatory diseases, such as colitis and pancreatitis, predispose to gastrointestinal (GI) adenocarcinoma by reprogramming differentiated cells. Focusing on colon and pancreas, we discuss recently discovered connections between inflammation and loss of cell differentiation, and propose that dysregulation of cell fate may be a novel rate-limiting step of tumorigenesis. We review studies identifying differentiation mechanisms that limit tumor initiation and that, upon reactivation, can prevent or revert the cancer cell transformed phenotype. Together, these findings suggest that differentiation-targeted treatments hold promise as a therapeutic strategy in GI cancer.

The acinar differentiation determinant PTF1A inhibits initiation of pancreatic ductal adenocarcinoma.

Understanding the initiation and progression of pancreatic ductal adenocarcinoma (PDAC) may provide therapeutic strategies for this deadly disease. Recently, we and others made the surprising finding that PDAC and its preinvasive precursors, pancreatic intraepithelial neoplasia (PanIN), arise via reprogramming of mature acinar cells. We therefore hypothesized that the master regulator of acinar differentiation, PTF1A, could play a central role in suppressing PDAC initiation. In this study, we demonstrate that PTF1A expression is lost in both mouse and human PanINs, and that this downregulation is functionally imperative in mice for acinar reprogramming by oncogenic KRAS. Loss of Ptf1a alone is sufficient to induce acinar-to-ductal metaplasia, potentiate inflammation, and induce a KRAS-permissive, PDAC-like gene expression profile. As a result, Ptf1a-deficient acinar cells are dramatically sensitized to KRAS transformation, and reduced Ptf1a greatly accelerates development of invasive PDAC. Together, these data indicate that cell differentiation regulators constitute a new tumor suppressive mechanism in the pancreas.

SOCS3 is an endogenous inhibitor of pathologic angiogenesis.

Inflammatory cytokines and growth factors drive angiogenesis independently; however, their integrated role in pathologic and physiologic angiogenesis is not fully understood. Suppressor of cytokine signaling-3 (SOCS3) is an inducible negative feedback regulator of inflammation and growth factor signaling. In the present study, we show that SOCS3 curbs pathologic angiogenesis. Using a Cre/Lox system, we deleted SOCS3 in vessels and studied developmental and pathologic angiogenesis in murine models of oxygen-induced retinopathy and cancer. Conditional loss of SOCS3 leads to increased pathologic neovascularization, resulting in pronounced retinopathy and increased tumor size. In contrast, physiologic vascularization is not regulated by SOCS3. In vitro, SOCS3 knockdown increases proliferation and sprouting of endothelial cells costimulated with IGF-1 and TNFα via reduced feedback inhibition of the STAT3 and mTOR pathways. These results identify SOCS3 as a pivotal endogenous feedback inhibitor of pathologic angiogenesis and a potential therapeutic target acting at the converging crossroads of growth factor- and cytokine-induced vessel growth.

Retinal expression of Wnt-pathway mediated genes in low-density lipoprotein receptor-related protein 5 (Lrp5) knockout mice.

Mutations in low-density lipoprotein receptor-related protein 5 (Lrp5) impair retinal angiogenesis in patients with familial exudative vitreoretinopathy (FEVR), a rare type of blinding vascular eye disease. The defective retinal vasculature phenotype in human FEVR patients is recapitulated in Lrp5 knockout (Lrp5(-/-)) mouse with delayed and incomplete development of retinal vessels. In this study we examined gene expression changes in the developing Lrp5(-/-) mouse retina to gain insight into the molecular mechanisms that underlie the pathology of FEVR in humans. Gene expression levels were assessed with an Illumina microarray on total RNA from Lrp5(-/-) and WT retinas isolated on postnatal day (P) 8. Regulated genes were confirmed using RT-qPCR analysis. Consistent with a role in vascular development, we identified expression changes in genes involved in cell-cell adhesion, blood vessel morphogenesis and membrane transport in Lrp5(-/-) retina compared to WT retina. In particular, tight junction protein claudin5 and amino acid transporter slc38a5 are both highly down-regulated in Lrp5(-/-) retina. Similarly, several Wnt ligands including Wnt7b show decreased expression levels. Plasmalemma vesicle associated protein (plvap), an endothelial permeability marker, in contrast, is up-regulated consistent with increased permeability in Lrp5(-/-) retinas. Together these data suggest that Lrp5 regulates multiple groups of genes that influence retinal angiogenesis and may contribute to the pathogenesis of FEVR.

Wnt signaling mediates pathological vascular growth in proliferative retinopathy.

BACKGROUND: Ischemic proliferative retinopathy, characterized by pathological retinal neovascularization, is a major cause of blindness in working-age adults and children. Defining the molecular pathways distinguishing pathological neovascularization from normal vessels is critical to controlling these blinding diseases with targeted therapy. Because mutations in Wnt signaling cause defective retinal vasculature in humans with some characteristics of the pathological vessels in retinopathy, we investigated the potential role of Wnt signaling in pathological retinal vascular growth in proliferative retinopathy. METHODS AND RESULTS: In this study, we show that Wnt receptors (Frizzled4 and low-density lipoprotein receptor-related protein5 [Lrp5]) and activity are significantly increased in pathological neovascularization in a mouse model of oxygen-induced proliferative retinopathy. Loss of Wnt coreceptor Lrp5 and downstream signaling molecule dishevelled2 significantly decreases the formation of pathological retinal neovascularization in retinopathy. Loss of Lrp5 also affects retinal angiogenesis during development and formation of the blood-retinal barrier, which is linked to significant downregulation of tight junction protein claudin5 in Lrp5(-/-) vessels. Blocking claudin5 significantly suppresses Wnt pathway-driven endothelial cell sprouting in vitro and developmental and pathological vascular growth in retinopathy in vivo. CONCLUSIONS: These results demonstrate an important role of Wnt signaling in pathological vascular development in retinopathy and show a novel function of Cln5 in promoting angiogenesis.

Resveratrol inhibits pathologic retinal neovascularization in Vldlr(-/-) mice.

PURPOSE: Macular telangiectasia (MacTel) is a vision-threatening retinal disease with unknown pathogenesis and no approved treatment. Very low-density lipoprotein receptor mutant mice (Vldlr(-/-)) exhibit critical features of MacTel such as retinal neovascularization and photoreceptor degeneration. In this study, the authors evaluate the therapeutic potential of resveratrol, a plant polyphenol, in Vldlr(-/-) mice as a model for MacTel. METHODS: Vldlr(-/-) and wild-type mice at postnatal day (P) 21 to P60 or P10 to P30 were treated orally with resveratrol. The number of neovascular lesions was evaluated on retinal flatmounts, and resveratrol effects on endothelial cells were assessed by Western blot for phosphorylated ERK1/2, aortic ring, and migration assays. Vegf and Gfap expression was evaluated in laser-capture microdissected retinal layers of angiogenic lesions and nonlesion areas from Vldlr(-/-) and wild-type retinas. RESULTS: From P15 onward, Vldlr(-/-) retinas develop vascular lesions associated with the local upregulation of Vegf in photoreceptors and Gfap in the inner retina. Oral resveratrol reduces lesion formation when administered either before or after disease onset. The reduction of vascular lesions in resveratrol-treated Vldlr(-/-) mice is associated with the suppression of retinal Vegf transcription. Resveratrol also reduces endothelial ERK1/2 signaling as well as the migration and proliferation of endothelial cells. Furthermore, a trend toward increased rhodopsin mRNA in Vldlr(-/-) retinas is observed. CONCLUSIONS: Oral administration of resveratrol is protective against retinal neovascular lesions in Vldlr(-/-) mice by inhibiting Vegf expression and angiogenic activation of retinal endothelial cells. These results suggest that resveratrol might be a safe and effective intervention for treating patients with MacTel.

5-Lipoxygenase metabolite 4-HDHA is a mediator of the antiangiogenic effect of ω-3 polyunsaturated fatty acids.

Lipid signaling is dysregulated in many diseases with vascular pathology, including cancer, diabetic retinopathy, retinopathy of prematurity, and age-related macular degeneration. We have previously demonstrated that diets enriched in ω-3 polyunsaturated fatty acids (PUFAs) effectively reduce pathological retinal neovascularization in a mouse model of oxygen-induced retinopathy, in part through metabolic products that suppress microglial-derived tumor necrosis factor-α. To better understand the protective effects of ω-3 PUFAs, we examined the relative importance of major lipid metabolic pathways and their products in contributing to this effect. ω-3 PUFA diets were fed to four lines of mice deficient in each key lipid-processing enzyme (cyclooxygenase 1 or 2, or lipoxygenase 5 or 12/15), retinopathy was induced by oxygen exposure; only loss of 5-lipoxygenase (5-LOX) abrogated the protection against retinopathy of dietary ω-3 PUFAs. This protective effect was due to 5-LOX oxidation of the ω-3 PUFA lipid docosahexaenoic acid to 4-hydroxy-docosahexaenoic acid (4-HDHA). 4-HDHA directly inhibited endothelial cell proliferation and sprouting angiogenesis via peroxisome proliferator-activated receptor γ (PPARγ), independent of 4-HDHA's anti-inflammatory effects. Our study suggests that ω-3 PUFAs may be profitably used as an alternative or supplement to current anti-vascular endothelial growth factor (VEGF) treatment for proliferative retinopathy and points to the therapeutic potential of ω-3 PUFAs and metabolites in other diseases of vasoproliferation. It also suggests that cyclooxygenase inhibitors such as aspirin and ibuprofen (but not lipoxygenase inhibitors such as zileuton) might be used without losing the beneficial effect of dietary ω-3 PUFA.

Postnatal weight gain modifies severity and functional outcome of oxygen-induced proliferative retinopathy.

In clinical studies, postnatal weight gain is strongly associated with retinopathy of prematurity (ROP). However, animal studies are needed to investigate the pathophysiological mechanisms of how postnatal weight gain affects the severity of ROP. In the present study, we identify nutritional supply as one potent parameter that affects the extent of retinopathy in mice with identical birth weights and the same genetic background. Wild-type pups with poor postnatal nutrition and poor weight gain (PWG) exhibit a remarkably prolonged phase of retinopathy compared to medium weight gain or extensive weight gain pups. A high (r(2) = 0.83) parabolic association between postnatal weight gain and oxygen-induced retinopathy severity is observed, as is a significantly prolonged phase of proliferative retinopathy in PWG pups (20 days) compared with extensive weight gain pups (6 days). The extended retinopathy is concomitant with prolonged overexpression of retinal vascular endothelial growth factor in PWG pups. Importantly, PWG pups show low serum levels of nonfasting glucose, insulin, and insulin-like growth factor-1 as well as high levels of ghrelin in the early postoxygen-induced retinopathy phase, a combination indicative of poor metabolic supply. These differences translate into visual deficits in adult PWG mice, as demonstrated by impaired bipolar and proximal neuronal function. Together, these results provide evidence for a pathophysiological correlation between poor postnatal nutritional supply, slow weight gain, prolonged retinal vascular endothelial growth factor overexpression, protracted retinopathy, and reduced final visual outcome.

Short communication: PPAR gamma mediates a direct antiangiogenic effect of omega 3-PUFAs in proliferative retinopathy.

RATIONALE: Omega3 long-chain polyunsaturated fatty acids (omega3-PUFAs) are powerful modulators of angiogenesis. However, little is known about the mechanisms governing omega3-PUFA-dependent attenuation of angiogenesis. OBJECTIVE: This study aims to identify a major mechanism by which omega3-PUFAs attenuate retinal neovascularization. METHODS AND RESULTS: Administering omega3-PUFAs exclusively during the neovascular stage of the mouse model of oxygen-induced retinopathy induces a direct neovascularization reduction of more than 40% without altering vasoobliteration or the regrowth of normal vessels. Cotreatment with an inhibitor of peroxisome proliferator-activated receptor (PPAR)gamma almost completely abrogates this effect. Inhibition of PPARgamma also reverses the omega3-PUFA-induced reduction of retinal tumor necrosis factor-alpha, intercellular adhesion molecule-1, vascular cell adhesion molecule-1, endothelial selectin, and angiopoietin 2 but not vascular endothelial growth factor. CONCLUSIONS: These results identify a direct, PPARgamma-mediated effect of omega3-PUFAs on retinal neovascularization formation and retinal angiogenic activation that is independent of vascular endothelial growth factor.

The mouse retina as an angiogenesis model.

The mouse retina has been used extensively over the past decades to study both physiologic and pathologic angiogenesis. Over time, various mouse retina models have evolved into well-characterized and robust tools for in vivo angiogenesis research. This article is a review of the angiogenic development of the mouse retina and a discussion of some of the most widely used vascular disease models. From the multitude of studies performed in the mouse retina, a selection of representative works is discussed in more detail regarding their role in advancing the understanding of both the ocular and general mechanisms of angiogenesis.

Loss of psbS expression reduces vegetative growth, reproductive output, and light-limited, but not light-saturated, photosynthesis in Arabidopsis thaliana (Brassicaceae) grown in temperate light environments.

Plants protect themselves against the deleterious effects of high light intensities by inducing a mechanism ubiquitous among plants known as energy dissipation, which safely converts excess light to heat before it can lead to the formation of free radicals. Mutants possessing a deletion of the psbS gene, such as the npq4 mutant, cannot perform energy dissipation and thus offer an opportunity to assess the importance of this process to plant function. In a temperate light environment, greenhouse-grown npq4 mutants of Arabidopsis thaliana had smaller rosette diameters and leaf numbers. The reduction in size observed in npq4 plants was associated with fewer floral stalks, fewer fruits, lower whole-plant and individual seed masses, and lower germination rates. In the field, npq4 mutants developed fewer fruits. After a controlled exposure to high light stress, both PSII efficiency and CO(2) assimilation were more significantly compromised in npq4 mutants at low light intensities, but not at high light intensities. Thus, the protective nature of energy dissipation manifests in light environments that include periods of high light, which predispose plants to PSII photoinactivation, and periods of low light, when PSII photoinactivation decreases the rate of photosynthesis.