Neuregulin 4 mediates the metabolic benefits of mild cold exposure by promoting beige fat thermogenesis

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Abstract

Interorgan crosstalk via secreted hormones and metabolites is a fundamental aspect of mammalian metabolic physiology. Beyond the highly specialized endocrine cells, peripheral tissues are emerging as an important source of metabolic hormones that influence energy and nutrient metabolism and contribute to disease pathogenesis. Neuregulin 4 (Nrg4) is a fat-derived hormone that protects mice from nonalcoholic steatohepatitis (NASH) and NASH-associated liver cancer by shaping hepatic lipid metabolism and the liver immune microenvironment. Despite its enriched expression in brown fat, whether NRG4 plays a role in thermogenic response and mediates the metabolic benefits of cold exposure remain unexplored. Here we show that Nrg4 expression in inguinal white adipose tissue (iWAT) is highly responsive to chronic cold exposure. Nrg4 deficiency impairs beige fat induction and renders mice more susceptible to diet-induced metabolic disorders under mild cold conditions. Using mice with adipocyte and hepatocyte-specific Nrg4 deletion, we reveal that adipose tissue-derived NRG4, but not hepatic NRG4, is essential for beige fat induction following cold acclimation. Furthermore, treatment with recombinant NRG4-Fc fusion protein promotes beige fat induction in iWAT and improves metabolic health in diet-induced obese mice. These findings highlight a critical role of NRG4 in mediating beige fat induction and preserving metabolic health under mild cold conditions.

Authors

Zhimin Chen, Peng Zhang, Tongyu Liu, Xiaoxue Qiu, Siming Li, Jiandie D. Lin

Hypoxia-activated prodrug and anti-angiogenic therapy cooperatively treat pancreatic cancer but elicit immunosuppressive G-MDSC infiltration

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Abstract

We previously showed that ablation of tumor hypoxia can sensitize tumors to immune checkpoint blockade (ICB). Here, we used a Kras+/G12DTP53+/R172HPdx1-Cre (KPC) derived model of pancreatic adenocarcinoma (PDAC) to examine the tumor response and adaptive resistance mechanisms involved in response to two established methods of hypoxia-reducing therapy: the hypoxia-activated prodrug TH-302 and vascular endothelial growth factor receptor 2 (VEGFR-2) blockade. The combination of both modalities normalized tumor vasculature, increased DNA damage and cell death, and delayed tumor growth. In contrast to prior cancer models, the combination did not alleviate overall tissue hypoxia or sensitize these KPC tumors to ICB therapy despite qualitative improvements to the CD8 T cell response. Bulk-tumor RNA sequencing, flow cytometry, and adoptive myeloid cell transfer suggested that treated tumor cells increased their capacity to recruit granulocytic myeloid derived suppressor cells (G-MDSC) through CCL9 secretion. Blockade of the CCL9-CCR1 axis could limit G-MDSC migration, and depletion of Ly6G-positive cells could sensitize tumors to the combination of TH-302 and anti-VEGFR-2 with ICB. Together, these data suggest that pancreatic tumors modulate G-MDSC migration as an adaptive response to vascular normalization, and that these immunosuppressive myeloid cells act in a setting of persistent hypoxia to maintain adaptive immune resistance.

Authors

Arthur Liu, Seth T. Gammon, Federica Pisaneschi, Akash Boda, Casey R. Ager, David Piwnica-Worms, David S. Hong, Michael A. Curran

Spatial transcriptomics identifies candidate stromal drivers of benign prostatic hyperplasia

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Abstract

Benign prostatic hyperplasia (BPH) is the nodular proliferation of the prostate transition zone in older men, leading to urinary storage and voiding problems that can be recalcitrant to therapy. Decades ago, John McNeal proposed that BPH originates with the “reawakening” of embryonic inductive activity by adult prostate stroma, which spurs new ductal proliferation and branching morphogenesis. Here, by laser microdissection and transcriptional profiling of the BPH stroma adjacent to hyperplastic branching ducts, we identified secreted factors likely mediating stromal induction of prostate glandular epithelium and coinciding processes. The top stromal factors were Insulin Like Growth Factor 1 (IGF1) and C-X-C Motif Chemokine Ligand 13 (CXCL13), which we confirmed by RNA in situ hybridization to be co-expressed in BPH fibroblasts, along with their cognate receptors (IGF1R and CXCR5) on adjacent epithelium. In contrast, IGF1 but not CXCL13 was expressed in human embryonic prostate stroma. Finally, we demonstrated that IGF1 is necessary for the generation of BPH-1 cell spheroids and patient-derived BPH cell organoids in three-dimensional culture. Our findings partially support historic speculations on the etiology of BPH, and provide what we believe to be new molecular targets for rational therapies directed against the underlying processes driving BPH.

Authors

Anna S. Pollack, Christian A. Kunder, Noah Brazer, Zhewei Shen, Sushama Varma, Robert B. West, Gerald R. Cunha, Laurence S. Baskin, James D. Brooks, Jonathan R. Pollack

Disruption of CFAP418 interaction with lipids causes widespread abnormal membrane-associated cellular processes in retinal degenerations

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Abstract

Syndromic ciliopathies and retinal degenerations are large heterogeneous groups of genetic diseases. Pathogenic variants in the CFAP418 gene may cause both disorders, and its protein sequence is evolutionarily conserved. However, the disease mechanism underlying CFAP418 mutations has not been explored. Here, we apply quantitative lipidomic, proteomic, and phosphoproteomic profiling and affinity purification coupled with mass spectrometry to address the molecular function of CFAP418 in retinas. We show that CFAP418 protein binds to lipid metabolism precursor phosphatidic acid (PA) and mitochondrion-specific lipid cardiolipin but does not form a tight and static complex with proteins. Loss of Cfap418 in mice disturbs membrane lipid homeostasis and membrane-protein association, which subsequently causes mitochondrial defects and membrane remodeling abnormalities across multiple vesicular trafficking pathways in photoreceptors, especially the endosomal sorting complexes required for transport (ESCRT) pathway. Ablation of Cfap418 also increases the activity of PA-binding protein kinase Cα in the retina. Overall, our results indicate that membrane lipid imbalance is a pathological mechanism underlying syndromic ciliopathies and retinal degenerations, which is associated with other known causative genes of these diseases.

Authors

Anna M. Clark, Dongmei Yu, Grace Neiswanger, Daniel Zhu, Junhuang Zou, J. Alan Maschek, Thomas Burgoyne, Jun Yang

MYC-driven increases in mitochondrial DNA copy number occur early and persist throughout prostatic cancer progression

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Abstract

Increased mitochondrial function may render some cancers vulnerable to mitochondrial inhibitors. Since mitochondrial function is regulated partly by mitochondrial DNA copy number (mtDNAcn), accurate measurements of mtDNAcn could help reveal which cancers are driven by increased mitochondrial function and may be candidates for mitochondrial inhibition. However, prior studies have employed bulk macrodissections that fail to account for cell type-specific or tumor cell heterogeneity in mtDNAcn. These studies have often produced unclear results, particularly in prostate cancer. Herein, we developed a multiplex in situ method to spatially quantify cell type specific mtDNAcn. We show that mtDNAcn is increased in luminal cells of high-grade prostatic intraepithelial neoplasia (HGPIN), is increased in prostatic adenocarcinomas (PCa), and is further elevated in metastatic castration-resistant prostate cancer. Increased PCa mtDNAcn was validated by two orthogonal methods and is accompanied by increases in mtRNAs and enzymatic activity. Mechanistically, MYC inhibition in prostate cancer cells decreases mtDNA replication and expression of several mtDNA replication genes, and MYC activation in the mouse prostate leads to increased mtDNA levels in the neoplastic prostate cells. Our in situ approach also revealed elevated mtDNAcn in precancerous lesions of the pancreas and colon/rectum, demonstrating generalization across cancer types using clinical tissue samples.

Authors

Jiayu Chen, Qizhi Zheng, Jessica L. Hicks, Levent Trabzonlu, Busra Ozbek, Tracy Jones, Ajay M. Vaghasia, Tatianna C. Larman, Rulin Wang, Mark C. Markowski, Samuel R. Denmeade, Kenneth J. Pienta, Ralph H. Hruban, Emmanuel S. Antonarakis, Anuj Gupta, Chi V. Dang, Srinivasan Yegnasubramanian, Angelo M. De Marzo

DLL4-Notch3-WNT5B axis mediates bi-directional pro-metastatic crosstalk between melanoma and lymphatic endothelial cells

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Abstract

Despite strong indications that melanoma interaction with lymphatic vessels actively promotes melanoma progression, the molecular mechanisms are not yet completely understood. To characterize molecular factors of this crosstalk we established human primary lymphatic endothelial cell (LEC) co-cultures with human melanoma cell lines. Here, we show that co-culture with melanoma cells induced transcriptomic changes in LECs and led to multiple alterations in their function. WNT5B, a paracrine signaling molecule upregulated in melanoma cells upon LEC interaction, was found contributing to the functional changes in LECs. Moreover, WNT5B transcription was regulated by Notch3 in melanoma cells following the co-culture with LECs, and Notch3 and WNT5B were coexpressed in melanoma patient primary tumor and metastasis samples. Moreover, melanoma cells derived from LEC co-culture escaped efficiently from the primary site to the proximal tumor draining lymph nodes, which was impaired upon WNT5B depletion. This supported the role of WNT5B in promoting the metastatic potential of melanoma cells through its effects on LECs. Finally, DLL4, a Notch ligand expressed in LECs, was identified as an upstream inducer of the Notch3-WNT5B axis in melanoma. This study elucidated WNT5B as a key molecular factor mediating bi-directional crosstalk between melanoma cells and lymphatic endothelium and promoting melanoma metastasis.

Authors

Sanni Alve, Silvia Gramolelli, Joonas Jukonen, Susanna Juteau, Anne Pink, Atte A. Manninen, Satu Hänninen, Elisa Monto, Madeleine H. Lackman, Olli Carpén, Pipsa Saharinen, Sinem Karaman, Kari Vaahtomeri, Päivi M. Ojala

Efficient pulmonary lymphatic drainage is necessary for inflammation resolution in ARDS

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Abstract

The lymphatic vasculature is the natural pathway for the resolution of inflammation, while the role of pulmonary lymphatic drainage function in sepsis-induced acute respiratory distress syndrome (ARDS) remains poorly characterized. In this study, Indocyanine green (ICG)-Near Infrared (NIR) lymphatic living imaging was performed to examine pulmonary lymphatic drainage function in septic mice models. We found that the pulmonary lymphatic drainage was impaired owing to the damaged lymphatic structure in sepsis-induced ARDS. Moreover, prior lymphatic defects by blocking vascular endothelial growth factor receptor-3 (VEGFR3), worsened sepsis-induced lymphatic dysfunction and inflammation. The post-treatment of vascular endothelial growth factor-C (Cys156Ser) (VEGF-C156S), a ligand of VEGFR3, ameliorated lymphatic drainage through rejuvenating lymphatics to reduce the pulmonary edema and promote pulmonary macrophages and neutrophils to drain to pretracheal lymph nodes (pLNs). Meanwhile, VEGF-C156S post-treatment reversed sepsis-inhibited C-C motif chemokine ligand 21 (CCL21), which co-localizes with the pulmonary lymphatic vessels. Furthermore, the advantages of VEGF-C156S on the drainage of inflammatory cells and edema fluid were abolished by blocking VEGFR3 or CCL21. These results suggest that efficient pulmonary lymphatic drainage is necessary for inflammation resolution in ARDS. Our findings offer a novel therapeutic approach to sepsis-induced ARDS by promoting lymphatic drainage function.

Authors

Pu-hong Zhang, Wen-wu Zhang, Shun-shun Wang, Cheng-hua Wu, Yang-dong Ding, Xin-yi Wu, Fang Gao Smith, Yu Hao, Sheng-wei Jin

Ghrelin-responsive mediobasal hypothalamic neurons mediate exercise-associated food intake and exercise endurance

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Abstract

Previous studies have implicated the orexigenic hormone ghrelin as a mediator of exercise endurance and the feeding response post-exercise. Specifically, plasma ghrelin levels nearly double in mice when they are submitted to an hour-long bout of high-intensity interval exercise (HIIE) using treadmills. Also, GHSR (ghrelin receptor)-null mice exhibit decreased food intake following HIIE and a diminished running distance (time until exhaustion) during a longer, step-wise exercise endurance protocol. To investigate whether ghrelin-responsive mediobasal hypothalamus (MBH) neurons mediate these effects, we stereotaxically delivered the inhibitory DREADD virus AAV2-hSyn-DIO-hM4(Gi)-mCherry to the MBH of Ghsr-IRES-Cre mice, which express Cre-recombinase directed by the Ghsr promoter. We found that chemogenetic inhibition of GHSR-expressing MBH neurons [upon delivery of clozapine-N-oxide (CNO)] 1) suppressed food intake following HIIE by 31.3%, 2) reduced maximum running distance by 20.7%-22.7% and raised blood glucose and blood lactate levels by 18.4%-51.5% and 24.6%-39.2%, respectively, during an exercise endurance protocol, 3) reduced food intake following ghrelin administration by 57.2%, but 4) did not affect glucose tolerance. Further, HIIE increased MBH Ghsr expression. These results indicate that activation of ghrelin-responsive MBH neurons is required for the normal feeding response to HIIE and the usual amount of running exhibited during an exercise endurance protocol.

Authors

Omprakash Singh, Sean B. Ogden, Salil Varshney, Kripa Shankar, Deepali Gupta, Subhojit Paul, Sherri Osborne-Lawrence, Corine P. Richard, Nathan P. Metzger, Connor Lawrence, Luis León-Mercado, Jeffrey M. Zigman

FGF23 directly inhibits osteoprogenitor differentiation in Dmp1 knockout mice

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Abstract

Fibroblast growth factor 23 (FGF23) is a phosphate (Pi)-regulating hormone produced by bone. Hereditary hypophosphatemic disorders are associated with FGF23 excess, impaired skeletal growth and osteomalacia. Blocking FGF23 became an effective therapeutic strategy in X-linked hypophosphatemia, but testing remains limited in autosomal recessive hypophosphatemic rickets (ARHR). This study investigates the effects of Pi repletion and bone specific deletion of Fgf23 on bone and mineral metabolism in the Dmp1 knockout (Dmp1KO) mouse model of ARHR.At 12 weeks, Dmp1KO mice showed increased serum FGF23 and PTH levels, hypophosphatemia, impaired growth, rickets and osteomalacia. Six weeks of dietary Pi supplementation exacerbated FGF23 production, hyperparathyroidism, renal Pi excretion and osteomalacia. In contrast, osteocyte-specific deletion of Fgf23 resulted in a partial correction of FGF23 excess, which was sufficient to fully restore serum Pi levels, but only partially corrected the bone phenotype. In vitro, we show that FGF23 directly impairs osteoprogenitors differentiation and that DMP1 deficiency contributes to impaired mineralization independently of FGF23 or Pi levels. In conclusion, FGF23-induced hypophosphatemia is only partially responsible for the bone defects observed in Dmp1KO mice. Our data suggest that combined DMP1 repletion and FGF23 blockade could effectively correct ARHR-associated mineral and bone disorders.

Authors

Guillaume Courbon, Dominik Kentrup, Jane Joy Thomas, Xueyan Wang, Hao-Hsuan Tsai, Jadeah J. Spindler, John Von Drasek, Laura Mazudie Ndjonko, Marta Martinez-Calle, Sana Lynch, Lauriane Hivert, Xiaofang Wang, Wenhan Chang, Jian Q. Feng, Valentin David, Aline Martin

TMTC4 is a hair-cell-specific human deafness gene

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Abstract

Transmembrane and tetratricopeptide repeat 4 (Tmtc4) is a recently described novel deafness gene in mice. Tmtc4-knockout mice have rapidly progressive postnatal hearing loss due to overactivation of the unfolded protein response (UPR); however, the cellular basis and human relevance of Tmtc4-associated hearing loss in the cochlea was not heretofore appreciated. We created a hair-cell-specific conditional knockout mouse that phenocopies the constitutive knockout with postnatal onset deafness, demonstrating that Tmtc4 is a hair-cell specific deafness gene. Furthermore, we identified a human family in which Tmtc4 variants segregate with adult-onset progressive hearing loss. Lymphoblastoid cells derived from multiple affected and unaffected family members, as well as human embryonic kidney cells engineered to harbor each of the variants, demonstrated that the human Tmtc4 variants confer hypersensitivity of the UPR towards apoptosis. These findings provide evidence that TMTC4 is a deafness gene in humans and further implicate the UPR in progressive hearing loss.

Authors

Jiang Li, Byung Yoon Choi, Yasmin Eltawil, Noura Ismail Mohamad, Yesai Park, Ian R. Matthews, Jin-Hee Han, Bong Jik Kim, Elliott H. Sherr, Dylan K. Chan