The minichromosome maintenance complex drives esophageal basal zone hyperplasia
Rochman et al. report proteomics of eosinophilic esophagitis, which identifies the minichromosome maintenance complex as a driver of esophageal basal zone hyperplasia. The cover image is an artistic rendering of minichromosome maintenance proteins in esophageal tissue. Image credit: Chris Woods, Mark Rochman, and Marc Rothenberg.
-
Technical Advances
×
Abstract
IgG4-related disease (IgG4-RD) is a systemic autoimmune disease with unclear pathogenesis. We performed single-cell RNA-seq and surface proteome analyses on 61,379 PBMCs from 9 treatment-naive IgG4-RD patients and 7 age- and sex-matched healthy controls. Integrative analyses were performed for altered gene expression in IgG4-RD, and flow cytometry and immunofluorescence were used for validation. We observed expansion of plasmablasts with enhanced protein processing and activation, which correlated with the number of involved organs in IgG4-RD. Increased proportions of CD4+ cytotoxic T lymphocytes (CTLs), CD8+ CTLs-GNLY (granulysin), and γδT cells with enhanced chemotaxis and cytotoxicity but with suppressed inhibitory receptors characterize IgG4-RD. Prominent infiltration of lymphocytes with distinct compositions were found in different organs of IgG4-RD patients. Transcription factors (TFs), including PRDM1/XBP1 and RUNX3, were upregulated in IgG4-RD, promoting the differentiation of plasmablasts and CTLs, respectively. Monocytes in IgG4-RD have stronger expression of genes related to cell adhesion and chemotaxis, which may give rise to profibrotic macrophages in lesions. The gene activation pattern in peripheral immune cells indicated activation of multiple interaction pathways between cell types, in part through chemokines or growth factors and their receptors. Specific upregulation of TFs and expansion of plasmablasts and CTLs may be involved in the pathogenesis of IgG4-RD, and each of these populations are candidate targets for therapeutic interventions in this disease.
Authors
Chenyang Lu, Shasha Li, Pingying Qing, Qiuping Zhang, Xing Ji, Zhigang Tang, Chunyan Chen, Tong Wu, Yidan Hu, Yi Zhao, Xiaohui Zhang, Qi He, David A. Fox, Chunyu Tan, Yubin Luo, Yi Liu
×Abstract
Femoral atherosclerotic plaques are less inflammatory than carotid plaques histologically, but limited cell-level data exist regarding comparative immune landscapes and polarization at these sites. We investigated intraplaque leukocyte phenotypes and transcriptional polarization in 49 patients undergoing femoral (n = 23) or carotid (n = 26) endarterectomy using single-cell RNA-Seq (scRNA-Seq; n = 13), flow cytometry (n = 24), and IHC (n = 12). Comparative scRNA-Seq of CD45+-selected leukocytes from femoral (n = 9; 35,265 cells) and carotid (n = 4; 30,655 cells) plaque revealed distinct transcriptional profiles. Inflammatory foam cell–like macrophages and monocytes comprised higher proportions of myeloid cells in carotid plaques, whereas noninflammatory foam cell–like macrophages and LYVE1-overexpressing macrophages comprised higher proportions of myeloid cells in femoral plaque (P < 0.001 for all). A significant comparative excess of CCR2+ macrophages in carotid versus plaque was observed by flow cytometry in a separate validation cohort. B cells were more prevalent and exhibited a comparatively antiinflammatory profile in femoral plaque, whereas cytotoxic CD8+ T cells were more prevalent in carotid plaque. In conclusion, human femoral plaques exhibit distinct macrophage phenotypic and transcriptional profiles as well as diminished CD8+ T cell populations compared with human carotid plaques
Authors
Joshua Slysz, Arjun Sinha, Matthew DeBerge, Shalini Singh, Harris Avgousti, Inhyeok Lee, Kristofor Glinton, Reina Nagasaka, Prarthana Dalal, Shaina Alexandria, Ching Man Wai, Ricardo Tellez, Mariavittoria Vescovo, Ashwin Sunderraj, Xinkun Wang, Matthew Schipma, Ryan Sisk, Rishab Gulati, Jenifer Vallejo, Ryosuke Saigusa, Donald M. Lloyd-Jones, Jon Lomasney, Samuel Weinberg, Karen Ho, Klaus Ley, Chiara Giannarelli, Edward B. Thorp, Matthew J. Feinstein
-
Research Articles
×
Abstract
Nonalcoholic fatty liver disease (NAFLD) and type 2 diabetes are interacting comorbidities of obesity, and increased hepatic de novo lipogenesis (DNL), driven by hyperinsulinemia and carbohydrate overload, contributes to their pathogenesis. Fatty acid synthase (FASN), a key enzyme of hepatic DNL, is upregulated in association with insulin resistance. However, the therapeutic potential of targeting FASN in hepatocytes for obesity-associated metabolic diseases is unknown. Here, we show that hepatic FASN deficiency differentially affects NAFLD and diabetes depending on the etiology of obesity. Hepatocyte-specific ablation of FASN ameliorated NAFLD and diabetes in melanocortin 4 receptor–deficient mice but not in mice with diet-induced obesity. In leptin-deficient mice, FASN ablation alleviated hepatic steatosis and improved glucose tolerance but exacerbated fed hyperglycemia and liver dysfunction. The beneficial effects of hepatic FASN deficiency on NAFLD and glucose metabolism were associated with suppression of DNL and attenuation of gluconeogenesis and fatty acid oxidation, respectively. The exacerbation of fed hyperglycemia by FASN ablation in leptin-deficient mice appeared attributable to impairment of hepatic glucose uptake triggered by glycogen accumulation and citrate-mediated inhibition of glycolysis. Further investigation of the therapeutic potential of hepatic FASN inhibition for NAFLD and diabetes in humans should thus consider the etiology of obesity.
Authors
Toshiya Matsukawa, Takashi Yagi, Tohru Uchida, Mashito Sakai, Masaru Mitsushima, Takao Naganuma, Hiroyuki Yano, Yuka Inaba, Hiroshi Inoue, Keisuke Yanagida, Masaaki Uematsu, Kazuki Nakao, Harumi Nakao, Atsu Aiba, Yoji Nagashima, Tetsuya Kubota, Naoto Kubota, Yoshihiko Izumida, Naoya Yahagi, Hiroyuki Unoki-Kubota, Yasushi Kaburagi, Shun-ichiro Asahara, Yoshiaki Kido, Hideo Shindou, Michiko Itoh, Yoshihiro Ogawa, Shiro Minami, Yasuo Terauchi, Kazuyuki Tobe, Kohjiro Ueki, Masato Kasuga, Michihiro Matsumoto
×Abstract
Pulmonary hypertension (PH) is a life-threatening disease characterized by a progressive narrowing of pulmonary arterioles. Although VEGF is highly expressed in lung of patients with PH and in animal PH models, the involvement of angiogenesis remains elusive. To clarify the pathophysiological function of angiogenesis in PH, we compared the angiogenic response in hypoxia (Hx) and SU5416 (a VEGFR2 inhibitor) plus Hx (SuHx) mouse PH models using 3D imaging. The 3D imaging analysis revealed an angiogenic response in the lung of the Hx-PH, but not of the severer SuHx-PH model. Selective VEGFR2 inhibition with cabozantinib plus Hx in mice also suppressed angiogenic response and exacerbated Hx-PH to the same extent as SuHx. Expression of endothelial proliferator-activated receptor γ coactivator 1α (PGC-1α) increased along with angiogenesis in lung of Hx-PH but not SuHx mice. In pulmonary endothelial cell–specific Ppargc1a-KO mice, the Hx-induced angiogenesis was suppressed, and PH was exacerbated along with increased oxidative stress, cellular senescence, and DNA damage. By contrast, treatment with baicalin, a flavonoid enhancing PGC-1α activity in endothelial cells, ameliorated Hx-PH with increased Vegfa expression and angiogenesis. Pulmonary endothelial PGC-1α–mediated angiogenesis is essential for adaptive responses to Hx and might represent a potential therapeutic target for PH.
Authors
Takayuki Fujiwara, Norifumi Takeda, Hironori Hara, Satoshi Ishii, Genri Numata, Hiroyuki Tokiwa, Manami Katoh, Sonoko Maemura, Takaaki Suzuki, Hiroshi Takiguchi, Tomonobu Yanase, Yoshiaki Kubota, Seitaro Nomura, Masaru Hatano, Kazutaka Ueda, Mutsuo Harada, Haruhiro Toko, Eiki Takimoto, Hiroshi Akazawa, Hiroyuki Morita, Satoshi Nishimura, Issei Komuro
×Abstract
X-linked hypophosphatemia (XLH) is characterized by high serum fibroblast growth factor 23 (FGF23) levels, resulting in impaired 1,25-dihydroxyvitamin D3 (1,25D) production. Adults with XLH develop a painful mineralization of the tendon-bone attachment site (enthesis), called enthesopathy. Treatment of mice with XLH (Hyp) with 1,25D or an anti–FGF23 Ab, both of which increase 1,25D signaling, prevents enthesopathy. Therefore, we undertook studies to determine a role for impaired 1,25D action in enthesopathy development. Entheses from mice lacking vitamin D 1α-hydroxylase (Cyp27b1) (C–/–) had a similar enthesopathy to Hyp mice, whereas deletion of Fgf23 in Hyp mice prevented enthesopathy, and deletion of both Cyp27b1 and Fgf23 in mice resulted in enthesopathy, demonstrating that the impaired 1,25D action due to high FGF23 levels underlies XLH enthesopathy development. Like Hyp mice, enthesopathy in C–/– mice was observed by P14 and was prevented, but not reversed, with 1,25D therapy. Deletion of the vitamin D receptor in scleraxis-expressing cells resulted in enthesopathy, indicating that 1,25D acted directly on enthesis cells to regulate enthesopathy development. These results show that 1,25D signaling was necessary for normal postnatal enthesis maturation and played a role in XLH enthesopathy development. Optimizing 1,25D replacement in pediatric patients with XLH is necessary to prevent enthesopathy.
Authors
Rakshya Rana, Jiana T. Baker, Melissa Sorsby, Supriya Jagga, Shreya Venkat, Shaza Almardini, Eva S. Liu
×Abstract
The intricate interplay between maternal immune response to SARS-CoV-2 and the transfer of protective factors to the fetus remains unclear. By analyzing mother-neonate dyads from second and third trimester SARS-CoV-2 infections, our study shows that neutralizing antibodies (NAbs) are infrequently detected in cord blood. We uncovered that this is due to impaired IgG-NAb placental transfer in symptomatic infection and to the predominance of maternal SARS-CoV-2 NAbs of the IgA and IgM isotypes, which are prevented from crossing the placenta. Crucially, the balance between maternal antiviral response and transplacental transfer of IgG-NAbs appears to hinge on IL-6 and IL-10 produced in response to SARS-CoV-2 infection. In addition, asymptomatic maternal infection was associated with expansion of anti–SARS-CoV-2 IgM and NK cell frequency. Our findings identify a protective role for IgA/IgM-NAbs in gestational SARS-CoV-2 infection and open the possibility that the maternal immune response to SARS-CoV-2 infection might benefit the neonate in 2 ways, first by skewing maternal immune response toward immediate viral clearance, and second by endowing the neonate with protective mechanisms to curtail horizontal viral transmission in the critical postnatal period, via the priming of IgA/IgM-NAbs to be transferred by the breast milk and via NK cell expansion in the neonate.
Authors
Juliana Gonçalves, Magda Melro, Marta Alenquer, Catarina Araújo, Júlia Castro-Neves, Daniela Amaral-Silva, Filipe Ferreira, José S. Ramalho, Nádia Charepe, Fátima Serrano, Carlos Pontinha, Maria João Amorim, Helena Soares
×Abstract
Pathogenic mutations in mitochondrial (mt) tRNA genes that compromise oxidative phosphorylation (OXPHOS) exhibit heteroplasmy and cause a range of multisyndromic conditions. Although mitochondrial disease patients are known to suffer from abnormal immune responses, how heteroplasmic mtDNA mutations affect the immune system at the molecular level is largely unknown. Here, in mice carrying pathogenic C5024T in mt-tRNAAla and in patients with mitochondrial encephalomyopathy, lactic acidosis, stroke-like episodes (MELAS) syndrome carrying A3243G in mt-tRNALeu, we found memory T and B cells to have lower pathogenic mtDNA mutation burdens than their antigen-inexperienced naive counterparts, including after vaccination. Pathogenic burden reduction was less pronounced in myeloid compared with lymphoid lineages, despite C5024T compromising macrophage OXPHOS capacity. Rapid dilution of the C5024T mutation in T and B cell cultures could be induced by antigen receptor–triggered proliferation and was accelerated by metabolic stress conditions. Furthermore, we found C5024T to dysregulate CD8+ T cell metabolic remodeling and IFN-γ production after activation. Together, our data illustrate that the generation of memory lymphocytes shapes the mtDNA landscape, wherein pathogenic variants dysregulate the immune response.
Authors
Jingdian Zhang, Camilla Koolmeister, Jinming Han, Roberta Filograna, Leo Hanke, Monika Àdori, Daniel J. Sheward, Sina Teifel, Shreekara Gopalakrishna, Qiuya Shao, Yong Liu, Keying Zhu, Robert A. Harris, Gerald McInerney, Ben Murrell, Mike Aoun, Liselotte Bäckdahl, Rikard Holmdahl, Marcin Pekalski, Anna Wedell, Martin Engvall, Anna Wredenberg, Gunilla B. Karlsson Hedestam, Xaquin Castro Dopico, Joanna Rorbach
×Abstract
The identity and origin of the stem/progenitor cells for adult joint cartilage repair remain unknown, impeding therapeutic development. Simulating the common therapeutic modality for cartilage repair in humans, i.e., full-thickness microfracture joint surgery, we combined the mouse full-thickness injury model with lineage tracing and identified a distinct skeletal progenitor cell type enabling long-term (beyond 7 days after injury) articular cartilage repair in vivo. Deriving from a population with active Prg4 expression in adulthood while lacking aggrecan expression, these progenitors proliferate, differentiate to express aggrecan and type II collagen, and predominate in long-term articular cartilage wounds, where they represent the principal repair progenitors in situ under native repair conditions without cellular transplantation. They originate outside the adult bone marrow or superficial zone articular cartilage. These findings have implications for skeletal biology and regenerative medicine for joint injury repair.
Authors
Mei Massengale, Justin L. Massengale, Catherine R. Benson, Ninib Baryawno, Toshihiko Oki, Matthew L. Steinhauser, Alissa Wang, Deepak Balani, Luke S. Oh, Mark A. Randolph, Thomas J. Gill III, Henry M. Kronenberg, David T. Scadden
×Abstract
DNASE1L3, an enzyme highly expressed in DCs, is functionally important for regulating autoimmune responses to self-DNA and chromatin. Deficiency of DNASE1L3 leads to development of autoimmune diseases in both humans and mice. However, despite the well-established causal relationship between DNASE1L3 and immunity, little is known about the involvement of DNASE1L3 in regulation of antitumor immunity, the foundation of modern antitumor immunotherapy. In this study, we identify DNASE1L3 as a potentially new regulator of antitumor immunity and a tumor suppressor in colon cancer. In humans, DNASE1L3 is downregulated in tumor-infiltrating DCs, and this downregulation is associated with poor patient prognosis and reduced tumor immune cell infiltration in many cancer types. In mice, Dnase1l3 deficiency in the tumor microenvironment enhances tumor formation and growth in several colon cancer models. Notably, the increased tumor formation and growth in Dnase1l3-deficient mice are associated with impaired antitumor immunity, as evidenced by a substantial reduction of cytotoxic T cells and a unique subset of DCs. Consistently, Dnase1l3-deficient DCs directly modulate cytotoxic T cells in vitro. To our knowledge, our study unveils a previously unknown link between DNASE1L3 and antitumor immunity and further suggests that restoration of DNASE1L3 activity may represent a potential therapeutic approach for anticancer therapy.
Authors
Wenling Li, Hideki Nakano, Wei Fan, Yuanyuan Li, Payel Sil, Keiko Nakano, Fei Zhao, Peer W. Karmaus, Sara A. Grimm, Min Shi, Xin Xu, Ryushin Mizuta, Daisuke Kitamura, Yisong Wan, Michael B. Fessler, Donald N. Cook, Igor Shats, Xiaoling Li, Leping Li
×Abstract
Pediatric cardiomyopathy (CM) represents a group of rare, severe disorders that affect the myocardium. To date, the etiology and mechanisms underlying pediatric CM are incompletely understood, hampering accurate diagnosis and individualized therapy development. Here, we identified biallelic variants in the highly conserved flightless-I (FLII) gene in 3 families with idiopathic, early-onset dilated CM. We demonstrated that patient-specific FLII variants, when brought into the zebrafish genome using CRISPR/Cas9 genome editing, resulted in the manifestation of key aspects of morphological and functional abnormalities of the heart, as observed in our patients. Importantly, using these genetic animal models, complemented with in-depth loss-of-function studies, we provided insights into the function of Flii during ventricular chamber morphogenesis in vivo, including myofibril organization and cardiomyocyte cell adhesion, as well as trabeculation. In addition, we identified Flii function to be important for the regulation of Notch and Hippo signaling, crucial pathways associated with cardiac morphogenesis and function. Taken together, our data provide experimental evidence for a role for FLII in the pathogenesis of pediatric CM and report biallelic variants as a genetic cause of pediatric CM.
Authors
Claudine W.B. Ruijmbeek, Filomena Housley, Hafiza Idrees, Michael P. Housley, Jenny Pestel, Leonie Keller, Jason K.H. Lai, Herma C. van der Linde, Rob Willemsen, Janett Piesker, Zuhair N. Al-Hassnan, Abdulrahman Almesned, Michiel Dalinghaus, Lisa M. van den Bersselaar, Marjon A. van Slegtenhorst, Federico Tessadori, Jeroen Bakkers, Tjakko J. van Ham, Didier Y.R. Stainier, Judith M.A. Verhagen, Sven Reischauer
×Abstract
A T50I substitution in the K-Ras interswitch domain causes Noonan syndrome and emerged as a third-site mutation that restored the in vivo transforming activity and constitutive MAPK pathway activation by an attenuated KrasG12D,E37G oncogene in a mouse leukemia model. Biochemical and crystallographic data suggested that K-RasT50I increases MAPK signal output through a non-GTPase mechanism, potentially by promoting asymmetric Ras:Ras interactions between T50 and E162. We generated a “switchable” system in which K-Ras mutant proteins expressed at physiologic levels supplant the fms like tyrosine kinase 3 (FLT3) dependency of MOLM-13 leukemia cells lacking endogenous KRAS and used this system to interrogate single or compound G12D, T50I, D154Q, and E162L mutations. These studies support a key role for the asymmetric lateral assembly of K-Ras in a plasma membrane–distal orientation that promotes the formation of active Ras:Raf complexes in a membrane-proximal conformation. Disease-causing mutations such as T50I are a valuable starting point for illuminating normal Ras function, elucidating mechanisms of disease, and identifying potential therapeutic opportunities for Rasopathy disorders and cancer.
Authors
Pan-Yu Chen, Benjamin J. Huang, Max Harris, Christopher Boone, Weijie Wang, Heidi Carias, Brian Mesiona, Daniela Mavrici, Amanda C. Kohler, Gideon Bollag, Chao Zhang, Ying Zhang, Kevin Shannon
×Abstract
Nitric oxide (NO) is a critical signaling molecule that has been implicated in the pathogenesis of neurocognitive diseases. Both excessive and insufficient NO production have been linked to pathology. Previously, we have shown that argininosuccinate lyase deficiency (ASLD) is a novel model system to investigate cell-autonomous, nitric oxide synthase–dependent NO deficiency. Humans with ASLD are at increased risk for developing hyperammonemia due to a block in ureagenesis. However, natural history studies have shown that individuals with ASLD have multisystem disease including neurocognitive deficits that can be independent of ammonia. Here, using ASLD as a model of NO deficiency, we investigated the effects of NO on brain endothelial cells in vitro and the blood-brain barrier (BBB) in vivo. Knockdown of ASL in human brain microvascular endothelial cells (HBMECs) led to decreased transendothelial electrical resistance, indicative of increased cell permeability. Mechanistically, treatment with an NO donor or inhibition of Claudin-1 improved barrier integrity in ASL-deficient HBMECs. Furthermore, in vivo assessment of a hypomorphic mouse model of ASLD showed increased BBB leakage, which was partially rescued by NO supplementation. Our results suggest that ASL-mediated NO synthesis is required for proper maintenance of brain microvascular endothelial cell functions as well as BBB integrity.
Authors
Jordan Kho, Urszula Polak, Ming-Ming Jiang, John D. Odom, Jill V. Hunter, Saima M. Ali, Lindsay C. Burrage, Sandesh C.S. Nagamani, Robia G. Pautler, Hannah P. Thompson, Akihiko Urayama, Zixue Jin, Brendan Lee
×Abstract
Metabolic mechanisms underlying the heterogeneity of major adverse cardiovascular (CV) event (MACE) risk in individuals with type 2 diabetes mellitus (T2D) remain unclear. We hypothesized that circulating metabolites reflecting mitochondrial dysfunction predict incident MACE in T2D. Targeted mass-spectrometry profiling of 60 metabolites was performed on baseline plasma samples from the Trial Evaluating Cardiovascular Outcomes with Sitagliptin (TECOS; discovery cohort) and Exenatide Study of Cardiovascular Event Lowering (EXSCEL; validation cohort) biomarker substudy cohorts. A principal components analysis metabolite factor comprising medium-chain acylcarnitines (MCACs) was associated with MACE in TECOS and validated in EXSCEL, with higher levels associated with higher MACE risk. Meta-analysis showed that long-chain acylcarnitines (LCACs) and dicarboxylacylcarnitines were also associated with MACE. Metabolites remained associated with MACE in multivariate models and favorably changed with exenatide therapy. A third cohort (Cardiac Catheterization Genetics [CATHGEN]) with T2D was assessed to determine whether these metabolites improved discriminative capability of multivariate models for MACE. Nine metabolites (MCACs and LCACs and 1 dicarboxylacylcarnitine) were associated with time to MACE in the CATHGEN cohort. Addition of these metabolites to clinical models minimally improved the discriminative capability for MACE but did significantly down reclassify risk. Thus, metabolites reporting on dysregulated mitochondrial fatty acid oxidation are present in higher levels in individuals with T2D who experience subsequent MACE. These biomarkers may improve CV risk prediction models, be therapy responsive, and highlight emerging risk mechanisms.
Authors
Jessica A. Regan, Robert J. Mentz, Maggie Nguyen, Jennifer B. Green, Lauren K. Truby, Olga Ilkayeva, Christopher B. Newgard, John B. Buse, Harald Sourij, C. David Sjöström, Naveed Sattar, Robert W. McGarrah, Yinggan Zheng, Darren K. McGuire, Eberhard Standl, Paul Armstrong, Eric D. Peterson, Adrian F. Hernandez, Rury R. Holman, Svati H. Shah
×Abstract
Intact fibroblast growth factor 23 (iFGF23) is a phosphaturic hormone that is cleaved by furin into N-terminal and C-terminal fragments. Several studies have implicated vitamin D in regulating furin in infections. Thus, we investigated the effect of 1,25-dihydroxyvitamin D3 [1,25(OH)2D] and the vitamin D receptor (VDR) on furin-mediated iFGF23 cleavage. Mice lacking VDR (Vdr–/–) had a 25-fold increase in iFGF23 cleavage, with increased furin levels and activity compared with wild-type (WT) littermates. Inhibition of furin activity blocked the increase in iFGF23 cleavage in Vdr–/– animals and in a Vdr-knockdown osteocyte OCY454 cell line. Chromatin immunoprecipitation revealed VDR binding to DNA upstream of the Furin gene, with more transcription in the absence of VDR. In WT mice, furin inhibition reduced iFGF23 cleavage, increased iFGF23, and reduced serum phosphate levels. Similarly, 1,25(OH)2D reduced furin activity, decreased iFGF23 cleavage, and increased total FGF23. In a post hoc analysis of a randomized clinical trial, we found that ergocalciferol treatment, which increased serum 1,25(OH)2D, significantly decreased serum furin activity and iFGF23 cleavage, compared with placebo. Thus, 1,25(OH)2D inhibits iFGF23 cleavage via VDR-mediated suppression of Furin expression, thereby providing a mechanism by which vitamin D can augment phosphaturic iFGF23 levels.
Authors
Han Xie, Isinsu Bastepe, Wen Zhou, Birol Ay, Zara Ceraj, Ignacio A. Portales-Castillo, Eva S. Liu, Sherri-Ann M. Burnett-Bowie, Harald Jüppner, Eugene P. Rhee, Murat Bastepe, Petra Simic
×Abstract
The growth of skeletal muscle relies on a delicate equilibrium between protein synthesis and degradation; however, how proteostasis is managed in the endoplasmic reticulum (ER) is largely unknown. Here, we report that the SEL1L-HRD1 ER-associated degradation (ERAD) complex, the primary molecular machinery that degrades misfolded proteins in the ER, is vital to maintain postnatal muscle growth and systemic energy balance. Myocyte-specific SEL1L deletion blunts the hypertrophic phase of muscle growth, resulting in a net zero gain of muscle mass during this developmental period and a 30% reduction in overall body growth. In addition, myocyte-specific SEL1L deletion triggered a systemic reprogramming of metabolism characterized by improved glucose sensitivity, enhanced beigeing of adipocytes, and resistance to diet-induced obesity. These effects were partially mediated by the upregulation of the myokine FGF21. These findings highlight the pivotal role of SEL1L-HRD1 ERAD activity in skeletal myocytes for postnatal muscle growth, and its physiological integration in maintaining whole-body energy balance.
Authors
Benedict Abdon, Yusheng Liang, Débora da Luz Scheffer, Mauricio Torres, Neha Shrestha, Rachel B. Reinert, You Lu, Brent Pederson, Amara Bugarin-Lapuz, Sander Kersten, Ling Qi
×Abstract
Osteoarthritis (OA) is the most common joint disorder, and disease-modifying OA drugs (DMOADs) represent a major need in OA management. Krüppel-like factor 4 (KLF4) is a central transcription factor upregulating regenerative and protective functions in joint tissues. This study was aimed to identify small molecules activating KLF4 expression and to determine functions and mechanisms of the hit compounds. High-throughput screening (HTS) with 11,948 clinical-stage compounds was performed using a reporter cell line detecting endogenous KLF4 activation. Eighteen compounds were identified through the HTS and confirmed in a secondary screen. After testing in SW1353 chondrosarcoma cells and human chondrocytes, mocetinostat — a class I selective histone deacetylase (HDAC) inhibitor — had the best profile of biological activities. Mocetinostat upregulated cartilage signature genes in human chondrocytes, meniscal cells, and BM-derived mesenchymal stem cells, and it downregulated hypertrophic, inflammatory, and catabolic genes in those cells and synoviocytes. I.p. administration of mocetinostat into mice reduced severity of OA-associated changes and improved pain behaviors. Global gene expression and proteomics analyses revealed that regenerative and protective effects of mocetinostat were dependent on peroxisome proliferator-activated receptor γ coactivator 1-α. These findings show therapeutic and protective activities of mocetinostat against OA, qualifying it as a candidate to be used as a DMOAD.
Authors
Manabu Kawata, Daniel B. McClatchy, Jolene K. Diedrich, Merissa Olmer, Kristen A. Johnson, John R. Yates, Martin K. Lotz
×Abstract
The proportions and phenotypes of immune cell subsets in peripheral blood undergo continual and dramatic remodeling throughout the human life span, which complicates efforts to identify disease-associated immune signatures in type 1 diabetes (T1D). We conducted cross-sectional flow cytometric immune profiling on peripheral blood from 826 individuals (stage 3 T1D, their first-degree relatives, those with ≥2 islet autoantibodies, and autoantibody-negative unaffected controls). We constructed an immune age predictive model in unaffected participants and observed accelerated immune aging in T1D. We used generalized additive models for location, shape, and scale to obtain age-corrected data for flow cytometry and complete blood count readouts, which can be visualized in our interactive portal (ImmScape); 46 parameters were significantly associated with age only, 25 with T1D only, and 23 with both age and T1D. Phenotypes associated with accelerated immunological aging in T1D included increased CXCR3+ and programmed cell death 1–positive (PD-1+) frequencies in naive and memory T cell subsets, despite reduced PD-1 expression levels on memory T cells. Phenotypes associated with T1D after age correction were predictive of T1D status. Our findings demonstrate advanced immune aging in T1D and highlight disease-associated phenotypes for biomarker monitoring and therapeutic interventions.
Authors
Melanie R. Shapiro, Xiaoru Dong, Daniel J. Perry, James M. McNichols, Puchong Thirawatananond, Amanda L. Posgai, Leeana D. Peters, Keshav Motwani, Richard S. Musca, Andrew Muir, Patrick Concannon, Laura M. Jacobsen, Clayton E. Mathews, Clive H. Wasserfall, Michael J. Haller, Desmond A. Schatz, Mark A. Atkinson, Maigan A. Brusko, Rhonda Bacher, Todd M. Brusko
×Abstract
Inadequate adaption to mechanical forces, including blood pressure, contributes to development of arterial aneurysms. Recent studies have pointed to a mechanoprotective role of YAP and TAZ in vascular smooth muscle cells (SMCs). Here, we identified reduced expression of YAP1 in human aortic aneurysms. Vascular SMC–specific knockouts (KOs) of YAP/TAZ were thus generated using the integrin α8–Cre (Itga8-Cre) mouse model (i8-YT-KO). i8-YT-KO mice spontaneously developed aneurysms in the abdominal aorta within 2 weeks of KO induction and in smaller arteries at later times. The vascular specificity of Itga8-Cre circumvented gastrointestinal effects. Aortic aneurysms were characterized by elastin disarray, SMC apoptosis, and accumulation of proteoglycans and immune cell populations. RNA sequencing, proteomics, and myography demonstrated decreased contractile differentiation of SMCs and impaired vascular contractility. This associated with partial loss of myocardin expression, reduced blood pressure, and edema. Mediators in the inflammatory cGAS/STING pathway were increased. A sizeable increase in SOX9, along with several direct target genes, including aggrecan (Acan), contributed to proteoglycan accumulation. This was the earliest detectable change, occurring 3 days after KO induction and before the proinflammatory transition. In conclusion, Itga8-Cre deletion of YAP and TAZ represents a rapid and spontaneous aneurysm model that recapitulates features of human abdominal aortic aneurysms.
Authors
Marycarmen Arévalo Martínez, Olivia Ritsvall, Joakim Armstrong Bastrup, Selvi Celik, Gabriel Jakobsson, Fatima Daoud, Christopher Winqvist, Anders Aspberg, Catarina Rippe, Lars Maegdefessel, Alexandru Schiopu, Thomas A. Jepps, Johan Holmberg, Karl Swärd, Sebastian Albinsson
×Abstract
Maternal decidual CD8+ T cells must integrate the antithetical demands of providing immunity to infection while maintaining immune tolerance for fetal and placental antigens. Human decidual CD8+ T cells were shown to be highly differentiated memory T cells with mixed signatures of dysfunction, activation, and effector function. However, no information is present on how specificity for microbial or fetal antigens relates to their function or dysfunction. In addition, a key question, whether decidual CD8+ T cells include unique tissue-resident memory T cells (Trm) or also effector memory T cell (Tem) types shared with peripheral blood populations, is unknown. Here, high-dimensional flow cytometry of decidual and blood CD8+ T cells identified 2 Tem populations shared in blood and decidua and 9 functionally distinct Trm clusters uniquely found in decidua. Interestingly, fetus- and virus-specific decidual CD8+ Trm cells had similar features of inhibition and cytotoxicity, with no significant differences in their expression of activation, inhibitory, and cytotoxic molecules, suggesting that not all fetus-specific CD8+ T cell responses are suppressed at the maternal-fetal interface. Understanding how decidual CD8+ T cell specificity relates to their function and tissue residency is crucial in advancing understanding of their contribution to placental inflammation and control of congenital infections.
Authors
Shweta Mahajan, Aria Alexander, Zachary Koenig, Nicholas Saba, Nina Prasanphanich, David A. Hildeman, Claire A. Chougnet, Emily DeFranco, Sandra Andorf, Tamara Tilburgs
×Abstract
Eosinophilic esophagitis (EoE) is a chronic gastrointestinal disorder characterized by food antigen–driven eosinophilic inflammation and hyperproliferation of esophageal mucosa. By utilizing a large-scale, proteomic screen of esophageal biopsies, we aimed to uncover molecular drivers of the disease. Proteomic analysis by liquid chromatography–tandem mass spectrometry identified 402 differentially expressed proteins (DEPs) that correlated with the EoE transcriptome. Immune cell–related proteins were among the most highly upregulated DEPs in EoE compared with controls, whereas proteins linked to epithelial differentiation were primarily downregulated. Notably, in the inflamed esophageal tissue, all 6 subunits of the minichromosome maintenance (MCM) complex, a DNA helicase essential for genomic DNA replication, were significantly upregulated at the gene and protein levels. Furthermore, treating esophageal epithelial cells with a known inhibitor of the MCM complex (ciprofloxacin) blocked esophageal epithelial proliferation. In a murine model of EoE driven by overexpression of IL-13, ciprofloxacin treatment decreased basal zone thickness and reduced dilated intercellular spaces by blocking the transition of epithelial cells through the S-phase of the cell cycle. Collectively, a broad-spectrum proteomic screen has identified the involvement of the MCM complex in EoE and has highlighted MCM inhibitors as potential therapeutic agents for the disease.
Authors
Mark Rochman, Yrina Rochman, Julie M. Caldwell, Lydia E. Mack, John A. Besse, Nathan P. Manes, Sung Hwan Yoon, Tetsuo Shoda, Aleksandra Nita-Lazar, Marc E. Rothenberg
