Ruscitto A, Morel MM, Shawber CJ, Reeve G, Lecholop MK, Bonthius D, Yao H, Embree MC
PMID: 32030828 | DOI: 10.1096/fj.201902287R
Temporomandibular joint osteoarthritis (TMJ OA) leads to permanent cartilage destruction, jaw dysfunction, and compromises the quality of life. However, the pathological mechanisms governing TMJ OA are poorly understood. Unlike appendicular articular cartilage, the TMJ has two distinct functions as the synovial joint of the craniofacial complex and also as the site for endochondral jaw bone growth. The established dogma of endochondral bone ossification is that hypertrophic chondrocytes undergo apoptosis, while invading vasculature with osteoprogenitors replace cartilage with bone. However, contemporary murine genetic studies support the direct differentiation of chondrocytes into osteoblasts and osteocytes in the TMJ. Here we sought to characterize putative vasculature and cartilage to bone transdifferentiation using healthy and diseased TMJ tissues from miniature pigs and humans. During endochondral ossification, the presence of fully formed vasculature expressing CD31+ endothelial cells and ?-SMA+ vascular smooth muscle cells were detected within all cellular zones in growing miniature pigs. Arterial, endothelial, venous, angiogenic, and mural cell markers were significantly upregulated in miniature pig TMJ tissues relative to donor matched knee meniscus fibrocartilage tissue. Upon surgically creating TMJ OA in miniature pigs, we discovered increased vasculature and putative chondrocyte to osteoblast transformation dually marked by COL2 and BSP or RUNX2 within the vascular bundles. Pathological human TMJ tissues also exhibited increased vasculature, while isolated diseased human TMJ cells exhibited marked increased in vasculature markers relative to control 293T cells. Our study provides evidence to suggest that the TMJ in higher order species are in fact vascularized. There have been no reports of cartilage to bone transdifferentiation or vasculature in human-relevant TMJ OA large animal models or in human TMJ tissues and cells. Therefore, these findings may potentially alter the clinical management of TMJ OA by defining new drugs that target angiogenesis or block the cartilage to bone transformation
Taieb, M;Ghannoum, D;Barré, L;Ouzzine, M;
PMID: 37296099 | DOI: 10.1038/s41419-023-05875-0
Genetic mutations in the Xylt1 gene are associated with Desbuquois dysplasia type II syndrome characterized by sever prenatal and postnatal short stature. However, the specific role of XylT-I in the growth plate is not completely understood. Here, we show that XylT-I is expressed and critical for the synthesis of proteoglycans in resting and proliferative but not in hypertrophic chondrocytes in the growth plate. We found that loss of XylT-I induces hypertrophic phenotype-like of chondrocytes associated with reduced interterritorial matrix. Mechanistically, deletion of XylT-I impairs the synthesis of long glycosaminoglycan chains leading to the formation of proteoglycans with shorter glycosaminoglycan chains. Histological and Second Harmonic Generation microscopy analysis revealed that deletion of XylT-I accelerated chondrocyte maturation and prevents chondrocytes columnar organization and arrangement in parallel of collagen fibers in the growth plate, suggesting that XylT-I controls chondrocyte maturation and matrix organization. Intriguingly, loss of XylT-I induced at embryonic stage E18.5 the migration of progenitor cells from the perichondrium next to the groove of Ranvier into the central part of epiphysis of E18.5 embryos. These cells characterized by higher expression of glycosaminoglycans exhibit circular organization then undergo hypertrophy and death creating a circular structure at the secondary ossification center location. Our study revealed an uncovered role of XylT-I in the synthesis of proteoglycans and provides evidence that the structure of glycosaminoglycan chains of proteoglycans controls chondrocyte maturation and matrix organization.
bioRxiv : the preprint server for biology
Wu, M;Zheng, W;Song, X;Bao, B;Wang, Y;Ramanan, D;Yang, D;Liu, R;Macbeth, JC;Do, EA;Andrade, WA;Yang, T;Cho, HS;Gazzaniga, FS;Ilves, M;Coronado, D;Thompson, C;Hang, S;Chiu, IM;Moffitt, JR;Hsiao, A;Mekalanos, JJ;Benoist, C;Kasper, DL;
PMID: 36778396 | DOI: 10.1101/2023.02.02.523770
Canonically, complement is a serum-based host defense system that protects against systemic microbial invasion. Little is known about the production and function of complement components on mucosal surfaces. Here we show gut complement component 3 (C3), central to complement function, is regulated by the composition of the microbiota in healthy humans and mice, leading to host-specific gut C3 levels. Stromal cells in intestinal lymphoid follicles (LFs) are the predominant source of intestinal C3. During enteric infection with Citrobacter rodentium or enterohemorrhagic Escherichia coli, luminal C3 levels increase significantly and are required for protection. C. rodentium is remarkably more invasive to the gut epithelium of C3-deficient mice than of wild-type mice. In the gut, C3-mediated phagocytosis of C. rodentium functions to clear pathogens. Our study reveals that variations in gut microbiota determine individuals†intestinal mucosal C3 levels, dominantly produced by LF stromal cells, which directly correlate with protection against enteric infection.Gut complement component 3 (C3) is induced by the microbiome in healthy humans and mice at a microbiota-specific level.Gut stromal cells located in intestinal lymphoid follicles are a major source of luminal C3 During enteric infections with Citrobacter rodentium or enterohemorrhagic Escherichia coli, gut luminal C3 levels increase and are required for protection. C. rodentium is significantly more invasive of the gut epithelium in C3-deficient mice when compared to WT mice. In the gut, C3-mediated opsonophagocytosis of C. rodentium functions to clear pathogens.
Garcia-Alonso, L;Lorenzi, V;Mazzeo, CI;Alves-Lopes, JP;Roberts, K;Sancho-Serra, C;Engelbert, J;Marečková, M;Gruhn, WH;Botting, RA;Li, T;Crespo, B;van Dongen, S;Kiselev, VY;Prigmore, E;Herbert, M;Moffett, A;Chédotal, A;Bayraktar, OA;Surani, A;Haniffa, M;Vento-Tormo, R;
PMID: 35794482 | DOI: 10.1038/s41586-022-04918-4
Gonadal development is a complex process that involves sex determination followed by divergent maturation into either testes or ovaries1. Historically, limited tissue accessibility, a lack of reliable in vitro models and critical differences between humans and mice have hampered our knowledge of human gonadogenesis, despite its importance in gonadal conditions and infertility. Here, we generated a comprehensive map of first- and second-trimester human gonads using a combination of single-cell and spatial transcriptomics, chromatin accessibility assays and fluorescent microscopy. We extracted human-specific regulatory programmes that control the development of germline and somatic cell lineages by profiling equivalent developmental stages in mice. In both species, we define the somatic cell states present at the time of sex specification, including the bipotent early supporting population that, in males, upregulates the testis-determining factor SRY and sPAX8s, a gonadal lineage located at the gonadal-mesonephric interface. In females, we resolve the cellular and molecular events that give rise to the first and second waves of granulosa cells that compartmentalize the developing ovary to modulate germ cell differentiation. In males, we identify human SIGLEC15+ and TREM2+ fetal testicular macrophages, which signal to somatic cells outside and inside the developing testis cords, respectively. This study provides a comprehensive spatiotemporal map of human and mouse gonadal differentiation, which can guide in vitro gonadogenesis.
Ma F, Ye H, He HH, Gerrin SJ, Chen S, Tanenbaum BA, Cai C, Sowalsky AG, He L, Wang H, Balk SP, Yuan X.
PMID: 27043282 | DOI: 10.1172/JCI78815.
The transcription factor SOX9 is critical for prostate development, and dysregulation of SOX9 is implicated in prostate cancer (PCa). However, the SOX9-dependent genes and pathways involved in both normal and neoplastic prostate epithelium are largely unknown. Here, we performed SOX9 ChIP sequencing analysis and transcriptome profiling of PCa cells and determined that SOX9 positively regulates multiple WNT pathway genes, including those encoding WNT receptors (frizzled [FZD] and lipoprotein receptor-related protein [LRP] family members) and the downstream β-catenin effector TCF4. Analyses of PCa xenografts and clinical samples both revealed an association between the expression of SOX9 and WNT pathway components in PCa. Finally, treatment of SOX9-expressing PCa cells with a WNT synthesis inhibitor (LGK974) reduced WNT pathway signaling in vitro and tumor growth in murine xenograft models. Together, our data indicate that SOX9 expression drives PCa by reactivating the WNT/β-catenin signaling that mediates ductal morphogenesis in fetal prostate and define a subgroup of patients who would benefit from WNT-targeted therapy.
King BC, Kulak K, Krus U, Rosberg R, Golec E, Wozniak K, Gomez MF, Zhang E, O'Connell DJ, Renström E, Blom AM.
PMID: - | DOI: 10.1016/j.cmet.2018.09.009
We show here that human pancreatic islets highly express C3, which is both secreted and present in the cytosol. Within isolated human islets, C3 expression correlates with type 2 diabetes (T2D) donor status, HbA1c, and inflammation. Islet C3 expression is also upregulated in several rodent diabetes models. C3 interacts with ATG16L1, which is essential for autophagy. Autophagy relieves cellular stresses faced by β cells during T2D and maintains cellular homeostasis. C3 knockout in clonal β cells impaired autophagy and led to increased apoptosis after exposure of cells to palmitic acid and IAPP. In the absence of C3, autophagosomes do not undergo fusion with lysosomes. Thus, C3 may be upregulated in islets during T2D as a cytoprotective factor against β cell dysfunction caused by impaired autophagy. Therefore, we revealed a previously undescribed intracellular function for C3, connecting the complement system directly to autophagy, with a broad potential importance in other diseases and cell types.
FASEB journal : official publication of the Federation of American Societies for Experimental Biology
Schaaf, CR;Polkoff, KM;Carter, A;Stewart, AS;Sheahan, B;Freund, J;Ginzel, J;Snyder, JC;Roper, J;Piedrahita, JA;Gonzalez, LM;
PMID: 37159340 | DOI: 10.1096/fj.202300223R
Intestinal epithelial stem cells (ISCs) are responsible for intestinal epithelial barrier renewal; thereby, ISCs play a critical role in intestinal pathophysiology research. While transgenic ISC reporter mice are available, advanced translational studies lack a large animal model. This study validates ISC isolation in a new porcine Leucine Rich Repeat Containing G Protein-Coupled Receptor 5 (LGR5) reporter line and demonstrates the use of these pigs as a novel colorectal cancer (CRC) model. We applied histology, immunofluorescence, fluorescence-activated cell sorting, flow cytometry, gene expression quantification, and 3D organoid cultures to whole tissue and single cells from the duodenum, jejunum, ileum, and colon of LGR5-H2B-GFP and wild-type pigs. Ileum and colon LGR5-H2B-GFP, healthy human, and murine biopsies were compared by mRNA fluorescent in situ hybridization (FISH). To model CRC, adenomatous polyposis coli (APC) mutation was induced by CRISPR/Cas9 editing in porcine LGR5-H2B-GFP colonoids. Crypt-base, green fluorescent protein (GFP) expressing cells co-localized with ISC biomarkers. LGR5-H2B-GFPhi cells had significantly higher LGR5 expression (p < .01) and enteroid forming efficiency (p < .0001) compared with LGR5-H2B-GFPmed/lo/neg cells. Using FISH, similar LGR5, OLFM4, HOPX, LYZ, and SOX9 expression was identified between human and LGR5-H2B-GFP pig crypt-base cells. LGR5-H2B-GFP/APCnull colonoids had cystic growth in WNT/R-spondin-depleted media and significantly upregulated WNT/β-catenin target gene expression (p < .05). LGR5+ ISCs are reproducibly isolated in LGR5-H2B-GFP pigs and used to model CRC in an organoid platform. The known anatomical and physiologic similarities between pig and human, and those shown by crypt-base FISH, underscore the significance of this novel LGR5-H2B-GFP pig to translational ISC research.
Verdile, N;Cardinaletti, G;Faccenda, F;Brevini, T;Gandolfi, F;Tibaldi, E;
| DOI: 10.1016/j.aquaculture.2022.739031
To develop more sustainable feed formulations, it is important to assess in detail their effect on gut function and health. We previously described the specific organization of the epithelial and stromal components of the intestinal stem cell niche (ISCN), in rainbow trout (RT) under actual farming conditions. In the present work, we used our previous observation, for performing a comparative analysis between a control diet (CF) and an experimental vegetable-based diet (CV) under a new perspective. We correlated diet-induced changes of the morphology and the absorptive capability of the RT mucosa with modifications of the ISCN. Histological analysis confirmed that CV diet caused a mucosa remodeling, characterized by the generation of accessory branches sprouting from the middle of the proximal intestine folds, determining a significant increase of the luminal surface. The newly-formed structures showed positivity for PepT1, Sglt-1, and Fabp2 indicating their active role in small molecule absorption. However, the cells lining the base of the new branches expressed both epithelial (sox9) and stromal (pdgfrα and foxl1) stem cell markers, rather than the expected markers of fully differentiated cells. Our results suggest that a nutritional challenge results in the formation of an ectopic ISNC at the middle of the intestinal folds that sustains the formation of functional collateral branches, presumably to compensate for the reduced intestinal absorption. Overall, these data highlight, for the first time, the plasticity of the ISCN and its possible role in compensating intestinal functions in response to challenging conditions.
EMBO J. 2016 Oct 17;35(20):2192-2212.
Suryo Rahmanto A, Savov V, Brunner A, Bolin S, Weishaupt H, Malyukova A, Rosén G, Čančer M, Hutter S, Sundström A, Kawauchi D, Jones DT, Spruck C, Taylor MD, Cho YJ, Pfister SM, Kool M, Korshunov A, Swartling FJ, Sangfelt O.
PMID: 27625374 | DOI: 10.15252/embj.201693889
SOX9 is a master transcription factor that regulates development and stem cell programs. However, its potential oncogenic activity and regulatory mechanisms that control SOX9 protein stability are poorly understood. Here, we show that SOX9 is a substrate of FBW7, a tumor suppressor, and a SCF (SKP1/CUL1/F-box)-type ubiquitin ligase. FBW7 recognizes a conserved degron surrounding threonine 236 (T236) in SOX9 that is phosphorylated by GSK3 kinase and consequently degraded by SCFFBW7α Failure to degrade SOX9 promotes migration, metastasis, and treatment resistance in medulloblastoma, one of the most common childhood brain tumors. FBW7 is either mutated or downregulated in medulloblastoma, and in cases where FBW7 mRNA levels are low, SOX9 protein is significantly elevated and this phenotype is associated with metastasis at diagnosis and poor patient outcome. Transcriptional profiling of medulloblastoma cells expressing a degradation-resistant SOX9 mutant reveals activation of pro-metastatic genes and genes linked to cisplatin resistance. Finally, we show that pharmacological inhibition of PI3K/AKT/mTOR pathway activity destabilizes SOX9 in a GSK3/FBW7-dependent manner, rendering medulloblastoma cells sensitive to cytostatic treatment.
Athwal VS, Pritchett J, Llewellyn J, Martin K, Camacho E, Raza SM, Phythian-Adams A, Birchall LJ, Mullan AF, Su K, Pearmain L, Dolman G, Zaitoun AM, Friedman SL, MacDonald A, Irving WL, Guha IN, Hanley NA, Piper Hanley K.
PMID: 29109128 | DOI: 10.15252/emmm.201707860
Fibrosis and organ failure is a common endpoint for many chronic liver diseases. Much is known about the upstream inflammatory mechanisms provoking fibrosis and downstream potential for tissue remodeling. However, less is known about the transcriptional regulation in vivo governing fibrotic matrix deposition by liver myofibroblasts. This gap in understanding has hampered molecular predictions of disease severity and clinical progression and restricted targets for antifibrotic drug development. In this study, we show the prevalence of SOX9 in biopsies from patients with chronic liver disease correlated with fibrosis severity and accurately predicted disease progression toward cirrhosis. Inactivation of Sox9 in mice protected against both parenchymal and biliary fibrosis, and improved liver function and ameliorated chronic inflammation. SOX9 was downstream of mechanosignaling factor, YAP1. These data demonstrate a role for SOX9 in liver fibrosis and open the way for the transcription factor and its dependent pathways as new diagnostic, prognostic, and therapeutic targets in patients with liver fibrosis.
Wiedemann, J;Billi, A;Bocci, F;Kashgari, G;Xing, E;Tsoi, L;Meller, L;Swindell, W;Wasikowski, R;Xing, X;Ma, F;Gharaee-Kermani, M;Kahlenberg, J;Harms, P;Maverakis, E;Nie, Q;Gudjonsson, J;Andersen, B;
| DOI: 10.1016/j.celrep.2023.111994
Palmoplantar skin is structurally and functionally unique, but the transcriptional programs driving this specialization are unclear. Here, we use bulk and single-cell RNA sequencing of human palm, sole, and hip skin to describe the distinguishing characteristics of palmoplantar and non-palmoplantar skin while also uncovering differences between palmar and plantar sites. Our approach reveals an altered immune environment in palmoplantar skin, with downregulation of diverse immunological processes and decreased immune cell populations. Further, we identify specific fibroblast populations that appear to orchestrate key differences in cell-cell communication in palm, sole, and hip. Dedicated keratinocyte analysis highlights major differences in basal cell fraction among the three sites and demonstrates the existence of two spinous keratinocyte populations constituting parallel, site-selective epidermal differentiation trajectories. In summary, this deep characterization of highly adapted palmoplantar skin contributes key insights into the fundamental biology of human skin and provides a valuable data resource for further investigation.
Human Adult Fibroblast-like Synoviocytes and Articular Chondrocytes Exhibit Prominent Overlap in Their Transcriptomic Signatures
Jones, K;Angelozzi, M;Gangishetti, U;Haseeb, A;de Charleroy, C;Lefebvre, V;Bhattaram, P;
PMID: 33931959 | DOI: 10.1002/acr2.11255
Fibroblast-like synoviocytes (FLS) and articular chondrocytes (AC) derive from a common pool of embryonic precursor cells. They are currently believed to engage in largely distinct differentiation programs to build synovium and articular cartilage and maintain healthy tissues throughout life. We tested this hypothesis by deeply characterizing and comparing their transcriptomic attributes. We profiled the transcriptomes of freshly isolated AC, synovium, primary FLS, and dermal fibroblasts from healthy adult humans using bulk RNA sequencing assays and downloaded published single-cell RNA sequencing data from freshly isolated human FLS. We integrated all data to define cell-specific signatures and validated findings with quantitative reverse transcription PCR of human samples and RNA hybridization of mouse joint sections. We identified 212 AC and 168 FLS markers on the basis of exclusive or enriched expression in either cell and 294 AC/FLS markers on the basis of similar expression in both cells. AC markers included joint-specific and pan-cartilaginous genes. FLS and AC/FLS markers featured 37 and 55 joint-specific genes, respectively, and 131 and 239 pan-fibroblastic genes, respectively. These signatures included many previously unrecognized markers with potentially important joint-specific roles. AC/FLS markers overlapped in their expression patterns among all FLS and AC subpopulations, suggesting that they fulfill joint-specific properties in all, rather than in discrete, AC and FLS subpopulations. This study broadens knowledge and identifies a prominent overlap of the human adult AC and FLS transcriptomic signatures. It also provides data resources to help further decipher mechanisms underlying joint homeostasis and degeneration and to improve the quality control of tissues engineered for regenerative treatments.
