Skeletal muscle is a major determinant of systemic metabolic homeostasis that plays a critical role in glucose metabolism and insulin sensitivity. By contrast, despite being a major user of fatty acids, and evidence that muscular disorders can lead to abnormal lipid deposition (e.g., nonalcoholic fatty liver disease in myopathies), our understanding of skeletal muscle regulation of systemic lipid homeostasis is not well understood. Here we show that skeletal muscle Krüppel-like factor 15 (KLF15) coordinates pathways central to systemic lipid homeostasis under basal conditions and in response to nutrient overload. Mice with skeletal muscle–specific KLF15 deletion demonstrated (a) reduced expression of key targets involved in lipid uptake, mitochondrial transport, and utilization, (b) elevated circulating lipids, (c) insulin resistance/glucose intolerance, and (d) increased lipid deposition in white adipose tissue and liver. Strikingly, a diet rich in short-chain fatty acids bypassed these defects in lipid flux and ameliorated aspects of metabolic dysregulation. Together, these findings establish skeletal muscle control of lipid flux as critical to systemic lipid homeostasis and metabolic health.
Liyan Fan, David R. Sweet, Domenick A. Prosdocimo, Vinesh Vinayachandran, Ernest R. Chan, Rongli Zhang, Olga Ilkayeva, Yuan Lu, Komal S. Keerthy, Chloe E. Booth, Christopher B. Newgard, Mukesh K. Jain
The effect of SARS-CoV-2 infection on the pathophysiology of the placenta and its impact on pregnancy outcome has not yet been fully elucidated. Here, we present a comprehensive clinical, morphological, and molecular analysis of placental tissues from pregnant women with and without SARS-CoV-2 infection. SARS-CoV-2 could be detected in half of placental tissues from SARS-CoV-2-positive women. The presence of the virus was not associated with any distinctive pathological, maternal or neonatal outcome features. SARS-CoV-2 tissue load was low in all but one patient which exhibited severe placental damage leading to neonatal neurological manifestations. The placental transcriptional response induced by high viral load of SARS-CoV-2 showed an immunopathology phenotype similar to autopsy lung tissues from patients with severe COVID-19. This finding contrasted with the lack of inflammatory response in placental tissues from SARS-CoV-2-positive women with low viral tissue load and from SARS-CoV-2-negative women. Importantly, no evidence of vertical transmission of SARS-CoV-2 was found in any newborns, suggesting that the placenta may be an effective maternal-neonatal barrier against the virus even in the presence of severe infection. Our observations suggest that severe placental damage induced by the virus may be detrimental for the neonate independently of vertical transmission.
Fulvia Milena Cribiù, Roberta Erra, Lorenza Pugni, Carlota Rubio-Perez, Lidia Alonso, Sara Simonetti, Giorgio A. Croci, Garazi Serna, Andrea Ronchi, Carlo Pietrasanta, Giovanna Lunghi, Anna Maria Fagnani, Maria Piñana, Matthias S. Matter, Alexandar Tzankov, Luigi Terracciano, Andres Anton, Enrico Ferrazzi, Stefano Ferrero, Enrico Iurlaro, Joan Seoane, Paolo Nuciforo
Medulloblastoma is an aggressive pediatric brain tumor that can be driven by misactivation of the Hedgehog (HH) pathway. CDK6 is a critical effector of oncogenic Hedgehog signaling, but attempts to target the Hedgehog pathway in medulloblastoma have been encumbered by resistance to single-agent molecular therapy. We identified resistance mechanisms to CDK6 inhibition in HH-associated medulloblastoma by performing orthogonal CRISPR and CRISPR interference screens in medulloblastoma cells treated with a CDK4/6 inhibitor, and RNA-sequencing of a mouse model of HH-associated medulloblastoma with genetic deletion of Cdk6. Our concordant in vitro and in vivo data revealed decreased ribosomal protein expression underlies resistance to CDK6 inhibition in HH-associated medulloblastoma, leading to endoplasmic reticular (ER) stress and activation of the unfolded protein response (UPR). These pathways increased the activity of enzymes producing Smoothened-activating sterol lipids that sustained oncogenic HH signaling in medulloblastoma despite cell cycle attenuation. Consistently, we demonstrated concurrent genetic deletion or pharmacological inhibition of CDK6 and HSD11ß2, an enzyme producing Smoothened-activating lipids, additively blocked cancer growth in multiple mouse genetic models of HH-associated medulloblastoma. Our data reveal a resistance pathway to CDK4/6 inhibition and a combination therapy to treat the most common malignant brain tumor in children that we believe are novel.
Vikas Daggubati, Jordan Hochstetler, Anirudh Bommireddy, Abrar Choudhury, Alexis Leigh Krup, Pervinder K. Choksi, Pakteema Tong, Amy Li, Libin Xu, Jeremy F. Reiter, David R. Raleigh
Innate lymphoid cells (ILCs) are enriched at barrier surfaces, including the gastrointestinal tract. While most studies have focused on the balance between pathogenic group 1 ILCs (ILC1s) and protective ILC3s in maintaining gut homeostasis and during chronic intestinal inflammation, such as Crohn’s disease (CD), less is known regarding ILC2s. Using an established murine model of CD-like ileitis, i.e., SAMP1/YitFc (SAMP) strain, we showed that ILC2s, compared to ILC1s and ILC3s, were increased within draining mesenteric lymph nodes and ilea of SAMP vs. AKR (parental control) mice early, during the onset of disease. Gut-derived ILC2s from Crohn’s patients vs. healthy controls were also increased and expand, similar to ILC1s, in greater proportion compared to ILC3s. Importantly, we report that the intracellular bacterial-sensing protein, nucleotide-binding oligomerization domaining-containing protein-2, encoded by NOD2, the first and strongest susceptibility gene identified for CD, promoted ILC2 expansion, which was dramatically reduced in SAMP lacking NOD2 and SAMP raised under germ-free conditions. Furthermore, these effects occurred through a mechanism involving the IL-33/ST2 ligand-receptor pair. Collectively, our results indicate a functional link between NOD2 and ILC2s, regulated by the IL-33/ST2 axis, that mechanistically may contribute to early events leading to CD pathogenesis.
Carlo De Salvo, Kristine-Ann Buela, Brecht Creyns, Daniele Corridoni, Nitish Rana, Hannah L. Wargo, Chiara Cominelli, Peter G. Delaney, Fabio Cominelli, Alexander Rodriguez-Palacios, Séverine Vermeire, Theresa T. Pizarro
Gene editing holds the potential to correct mutations and cure devastating genetic disorders. The technology has not yet proven efficacious for therapeutic use in central nervous system (CNS) diseases with ubiquitous neuronal defects. Angelman syndrome (AS), a severe neurodevelopmental disorder, is caused by a lack of maternal expression of the UBE3A gene. Due to genomic imprinting, only neurons are affected. One therapeutic approach focuses on the intact paternal UBE3A copy in AS patients that is silenced by an antisense transcript (UBE3A-ATS). We show here that gene editing of Ube3a-ATS in the mouse brain results in the formation of base pair insertions/deletions (indels) in neurons and the subsequent unsilencing of the paternal Ube3a allele in neurons, which partially corrects the behavior phenotype of a murine AS model. This study provides compelling evidence to further investigate editing of the homologous region of the human UBE3A-ATS, since this may provide a lasting therapeutic effect for AS patients.
Ralf S. Schmid, Xuefeng Deng, Priyalakshmi Panikker, Msema Msackyi, Camilo Breton, James M. Wilson
The emergence of drug-resistant fungi has prompted an urgent threat alert from the Centers for Disease Control. Biofilm assembly by these pathogens further impairs effective therapy. We recently identifed an antifungal, turbinmicin, that inhibits the fungal vesicle-mediated trafficking pathway and demonstrates broad-spectrum activity against planktonically growing fungi. During biofilm growth, vesicles with unique features play a critical role in the delivery of the biofilm extracellular matrix components. As these components are largely responsible for the drug resistance associated with biofilm growth, we explored the utility of turbinmicin in the biofilm setting. We found that turbinmicin disrupts extracellular vesicle delivery during biofilm growth, and this impairs the subsequent assembly of the biofilm matrix. We demonstrated that elimination of the extracellular matrix renders the drug-resistant biofilm communities susceptible to fungal killing by turbinmicin. Furthermore, the addition of turbinmicin to otherwise ineffective antifungal therapy potentiated the activity of these drugs. The underlying role of vesicles explains this dramatic activity and was supported by phenotype reversal with the addition of exogenous biofilm extracellular vesicles. This striking capacity to cripple biofilm assembly mechanisms reveals a new approach to eradicating biofilms and sheds light on turbinmicin as a promising anti-biofilm drug.
Miao Zhao, Fan Zhang, Robert Zarnowski, Kenneth J. Barns, Ryley Jones, Jen L. Fossen, Hiram Sanchez, Scott R. Rajski, Anjon Audhya, Tim S. Bugni, David R. Andes
Propranolol, a pleiotropic β-adrenergic blocker, was anecdotally reported to reduce cerebral cavernous malformations (CCM) in humans. However, propranolol has neither been rigorously evaluated in animal models nor was its mechanism of action in CCM defined. We report that propranolol or its S(-) enantiomer dramatically reduced embryonic venous cavernomas in ccm2 mosaic zebrafish, whereas R-(+)-propranolol, lacking β-antagonism, had no effect. Silencing of β1, but not β2, adrenergic receptor mimicked the beneficial effects of propranolol in a zebrafish CCM model as did a β1-selective antagonist, metoprolol. Thus, propranolol ameliorates cavernous malformations by β1 adrenergic antagonism in zebrafish. Oral propranolol significantly reduced lesion burden in two chronic murine models of the exceptionally aggressive Pdcd10/Ccm3 form of CCM. Propranolol or other β1-selective antagonists may be beneficial in CCM disease.
Wenqing Li, Robert Shenkar, Matthew R. Detter, Thomas Moore, Christian R. Benavides, Rhonda Lightle, Romuald Girard, Nicholas Hobson, Ying Cao, Yan Li, Erin Griffin, Carol Gallione, Joseph M. Zabramski, Mark H. Ginsberg, Douglas A. Marchuk, Issam A. Awad
The SARS-CoV-2 novel coronavirus global pandemic (COVID-19) has led to millions of cases and hundreds of thousands of deaths globally. While older adults appear at high risk for severe disease, hospitalizations and deaths due to SARS-CoV-2 among children have been relatively rare. Integrating single-cell RNA sequencing (scRNA-seq) of developing mouse lung with temporally-resolved immunofluorescence in mouse and human lung tissue, we found expression of SARS-CoV-2 Spike protein primer TMPRSS2 was highest in ciliated cells and type I alveolar epithelial cells (AT1), and TMPRSS2 expression increased with aging in mice and humans. Analysis of autopsy tissue from fatal COVID-19 cases detected SARS-CoV-2 RNA most frequently in ciliated and secretory cells in airway epithelium and AT1 cells in peripheral lung. SARS-CoV-2 RNA was highly colocalized in cells expressing TMPRSS2. Together, these data demonstrate the cellular spectrum infected by SARS-CoV-2 in lung epithelium and suggest that developmental regulation of TMPRSS2 may underlie the relative protection of infants and children from severe respiratory illness.
Bryce A. Schuler, A. Christian Habermann, Erin J. Plosa, Chase J. Taylor, Christopher Jetter, Nicholas M. Negretti, Meghan E. Kapp, John T. Benjamin, Peter Gulleman, David S. Nichols, Lior Z. Braunstein, Alice Hackett, Michael Koval, Susan H. Guttentag, Timothy S. Blackwell, Steven A. Webber, Nicholas E. Banovich, Jonathan A. Kropski, Jennifer M. S. Sucre
Diabetes mellitus (DM) is a risk factor for cancer development. However, the role of DM induced hyperglycemic stress (HG) in the development of blood cancer is poorly understood, largely due to lack of appropriate animal models. Epidemiologic studies show that individuals with DM are more likely to possess higher rate of mutations in genes found in pre-leukemic stem and progenitor cells (pre-LHSC/Ps) including in the epigenetic regulator TET2. TET2-mutant pre-LHSC/Ps require additional hits to evolve into a full-blown leukemia and/or aggressive myeloproliferative neoplasm (MPN). Cell intrinsic mutations have been shown to cooperate with Tet2 to promote leukemic transformation. However, the role of extrinsic factors is poorly understood. Utilizing a novel mouse model bearing haploinsufficiency of Tet2, to mimic the human pre-LHSC/P condition and HG stress, in the form of an Ins2Akita/+ mutation, which induces HG and Type-1 DM, we show that the compound mutant mice develop a lethal form of MPN and/or acute myeloid leukemia (AML). RNAseq revealed that this is in part due to upregulation of pro-inflammatory pathways, thereby generating a feedforward loop, including the expression of an anti-apoptotic lncRNA Morrbid. Loss of Morrbid in the compound mutants rescues the lethality and mitigates the development of MPN/AML. Our results describe a novel mouse model for age-dependent AML/MPN and suggest that HG stress acts as an environmental driver for myeloid neoplasm, which could be effectively prevented by reducing the expression of inflammation-related lncRNA Morrbid.
Zhigang Cai, Xiaoyu Lu, Chi Zhang, Sai Nelanuthala, Fabiola Aguilera, Abigail Hadley, Baskar Ramdas, Fang Fang, Kenneth P. Nephew, Jonathan J. Kotzin, Adam Williams, Jorge Henao-Mejia, Laura S. Haneline, Reuben Kapur
A considerable fraction of B cells recognize SARS-CoV-2 with germline-encoded elements of their B cell receptor resulting in the production of neutralizing and non-neutralizing antibodies. We found that antibody sequences from different discovery cohorts shared biochemical properties and could be retrieved across validation cohorts confirming the stereotyped character of this naive response in COVID-19. While neutralizing antibody sequences were found independently of disease severity in line with serological data, individual non-neutralizing antibody sequences were associated with fatal clinical courses suggesting detrimental effects of these antibodies. We mined 200 immune repertoires of healthy individuals and 500 of patients with blood or solid cancers - all acquired prior to the pandemic - for SARS-CoV-2 antibody sequences. While the largely unmutated B cell rearrangements occurred in a substantial fraction of immune repertoires from young and healthy individuals, these sequences were less likely found in individuals over 60 years of age and in cancer. This reflects B cell repertoire restriction in aging and cancer and may to a certain extent explain the different clinical COVID-19 courses observed in these risk groups. Future studies will have to address if this stereotyped B cell response to SARS-CoV-2 emerging from unmutated antibody rearrangements will create long-lived memory.
Lisa Paschold, Donjete Simnica, Edith Willscher, Maria J.G.T. Vehreschild, Jochen Dutzmann, Daniel G. Sedding, Christoph Schultheiß, Mascha Binder
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