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1.
Am J Pathol ; 194(1): 135-149, 2024 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-37918800

RESUMEN

Osteophytes in osteoarthritis (OA) joints contribute to restriction of joint movement, joint pain, and OA progression, but little is known about osteophyte regulators. Examination of gene expression related to cartilage extracellular matrix, endochondral ossification, and growth factor signaling in articular cartilage and osteophytes obtained from OA knee joints showed that several genes such as COL1A1, VCAN, BGLAP, BMP8B, RUNX2, and SOST were overexpressed in osteophytes compared with articular cartilage. Ratios of mesenchymal stem/progenitor cells, which were characterized by co-expression of CD105 and CD166, were significantly higher in osteophytic cells than articular cells. A three-dimensional culture method for cartilage and osteophyte cells was developed by modification of cultures of self-assembled spheroid cell organoids (spheroids). These spheroids cultured in the media for mesenchymal stem cells containing transforming growth factor-ß3 showed characteristic morphologies and gene expression profiles of articular cartilage and osteophytes, respectively. The effects of IL-1ß, tumor necrosis factor-α, and IL-6 on the spheroids of articular and osteophytic cells were studied. To the best of our knowledge, they provide the first evidence that IL-6 suppresses the spheroid size of osteophytic cells by inducing apoptosis and reducing extracellular matrix molecules. These data show that IL-6 is the suppressor of osteophyte growth and suggest that IL-6 expression and/or activity are implicated in the regulation of osteophyte formation in pathologic joints.


Asunto(s)
Cartílago Articular , Osteoartritis de la Rodilla , Osteoartritis , Osteofito , Humanos , Cartílago Articular/patología , Condrocitos/metabolismo , Péptidos y Proteínas de Señalización Intercelular/metabolismo , Interleucina-6/metabolismo , Articulación de la Rodilla/patología , Osteoartritis/patología , Osteoartritis de la Rodilla/metabolismo , Osteofito/genética , Osteofito/metabolismo , Osteofito/patología
2.
Lab Invest ; 102(1): 102-111, 2022 01.
Artículo en Inglés | MEDLINE | ID: mdl-34718343

RESUMEN

The ADAMTS (a disintegrin and metalloproteinase with thrombospondin motifs) family includes nine members with aggrecan-degrading activity, i.e., ADAMTS1, 4, 5, 8, 9, 15, 16, 18, and 20. However, their systematic expression profile in knee osteoarthritis (OA) synovium and effects of cytokines and growth factors on the expression in OA synovial fibroblasts remain elusive. In this study, expression of all nine aggrecanolytic ADAMTS species was assessed by quantitative real-time PCR in OA and control normal synovial tissues. OA synovial fibroblasts were treated with interleukin-1α (IL-1α), IL-1ß, tumor necrosis factor-α (TNF-α), transforming growth factor-ß (TGF-ß), vascular endothelial growth factor165, and heparin-binding epidermal growth factor, and analyzed for the expression of the ADAMTS species. The signaling pathways and inhibition of ADAMTS4 expression by high-molecular-weight hyaluronan, adalimumab, tocilizumab, and signaling molecule inhibitors were studied. ADAMTS1, 4, 5, 9, and 16 were expressed in OA synovium, but only ADAMTS4 expression was significantly higher in OA as compared to normal synovium. IL-1α, TNF-α, and TGF-ß markedly increased ADAMTS4 expression, while their effects were minimal for the other ADAMTS species. ADAMTS4 was synergistically upregulated by treatment with IL-1α and TNF-α, IL-1α and TGF-ß, or IL-1α, TNF-α and TGF-ß. The signaling molecules' inhibitors demonstrated that IL-1α-induced ADAMTS4 expression is predominantly through TGF-ß-associated kinase 1 (TAK1), and the TNF-α-stimulated expression is via TAK1 and nuclear factor-κB (NF-κB). The TGF-ß-promoted expression was through the activin receptor-like kinase 5 (ALK5)/Smad2/3, TAK1, and non-TAK1 pathways. Adalimumab blocked TNF-α-stimulated expression. ADAMTS4 expression co-stimulated with IL-1α, TNF-α and TGF-ß was abolished by treatment with adalimumab, TAK1 inhibitor, and ALK5/Smad2/3 inhibitor. These data demonstrate marked and synergistic upregulation of ADAMTS4 by IL-1α, TNF-α and TGF-ß in OA synovial fibroblasts, and suggest that concurrent therapy with an anti-TNF-α drug and inhibitor(s) may be useful for prevention against aggrecan degradation in OA.


Asunto(s)
Proteína ADAMTS4/genética , Citocinas/farmacología , Fibroblastos/efectos de los fármacos , Osteoartritis de la Rodilla/metabolismo , Membrana Sinovial/metabolismo , Regulación hacia Arriba/efectos de los fármacos , Proteína ADAMTS4/metabolismo , Células Cultivadas , Sinergismo Farmacológico , Fibroblastos/metabolismo , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Interleucina-1/farmacología , Isoenzimas/genética , Isoenzimas/metabolismo , Quinasas Quinasa Quinasa PAM/genética , Quinasas Quinasa Quinasa PAM/metabolismo , FN-kappa B/metabolismo , Transducción de Señal/efectos de los fármacos , Membrana Sinovial/citología , Factor de Crecimiento Transformador beta/farmacología , Inhibidores del Factor de Necrosis Tumoral/farmacología , Factor de Necrosis Tumoral alfa/farmacología , Factor A de Crecimiento Endotelial Vascular/farmacología
3.
Am J Pathol ; 191(11): 1986-1998, 2021 11.
Artículo en Inglés | MEDLINE | ID: mdl-34390681

RESUMEN

Hyaluronan (HA)-binding protein involved in HA depolymerization (HYBID) is involved in cartilage destruction via HA depolymerization in human knee osteoarthritis. However, the role of HYBID in the progression of osteoarthritis remain elusive. This study sought to examine whether genetic depletion of Hybid could suppress surgically induced osteoarthritis of mouse knee joints. In osteoarthritis induced by medial collateral ligament transection with meniscus removal, articular cartilage destruction and osteophyte formation at the medial femoral-tibial joint were significantly inhibited in Hybid-deficient (Hybid-/-) mice compared with wild-type mice. Hybid was highly produced by synovial cells and articular chondrocytes in the osteoarthritis joints of wild-type mice. IL-1ß, IL-6, and tumor necrosis factor-α were up-regulated in the osteoarthritis joint tissues of both wild-type and Hybid-/- mice. Vascular density at the synovial and periosteal junction was significantly reduced in Hybid-/- mice compared with wild-type mice. High-molecular-weight HA accumulated in osteoarthritis joint tissues of Hybid-/- mice. Injections of high-molecular-weight HA to knee joints attenuated the cartilage destruction and osteophyte formation in wild-type mouse osteoarthritis group. Inhibition of cartilage destruction and osteophyte formation in Hybid-/- mice was also observed in destabilization of the medial meniscus model. These data are the first to demonstrate that cartilage destruction and osteophyte formation are suppressed in Hybid-/- mice and suggest that Hybid-mediated HA depolymerization is implicated for the progression of mechanically-induced knee osteoarthritis.


Asunto(s)
Ácido Hialurónico/metabolismo , Osteoartritis de la Rodilla/metabolismo , Osteoartritis de la Rodilla/patología , Animales , Cartílago Articular/metabolismo , Cartílago Articular/patología , Modelos Animales de Enfermedad , Ratones
4.
J Biol Chem ; 295(45): 15328-15341, 2020 11 06.
Artículo en Inglés | MEDLINE | ID: mdl-32868297

RESUMEN

Dental enamel, the hardest tissue in the human body, is derived from dental epithelial cell ameloblast-secreted enamel matrices. Enamel mineralization occurs in a strictly synchronized manner along with ameloblast maturation in association with ion transport and pH balance, and any disruption of these processes results in enamel hypomineralization. G protein-coupled receptors (GPCRs) function as transducers of external signals by activating associated G proteins and regulate cellular physiology. Tissue-specific GPCRs play important roles in organ development, although their activities in tooth development remain poorly understood. The present results show that the adhesion GPCR Gpr115 (Adgrf4) is highly and preferentially expressed in mature ameloblasts and plays a crucial role during enamel mineralization. To investigate the in vivo function of Gpr115, knockout (Gpr115-KO) mice were created and found to develop hypomineralized enamel, with a larger acidic area because of the dysregulation of ion composition. Transcriptomic analysis also revealed that deletion of Gpr115 disrupted pH homeostasis and ion transport processes in enamel formation. In addition, in vitro analyses using the dental epithelial cell line cervical loop-derived dental epithelial (CLDE) cell demonstrated that Gpr115 is indispensable for the expression of carbonic anhydrase 6 (Car6), which has a critical role in enamel mineralization. Furthermore, an acidic condition induced Car6 expression under the regulation of Gpr115 in CLDE cells. Thus, we concluded that Gpr115 plays an important role in enamel mineralization via regulation of Car6 expression in ameloblasts. The present findings indicate a novel function of Gpr115 in ectodermal organ development and clarify the molecular mechanism of enamel formation.


Asunto(s)
Ameloblastos/metabolismo , Esmalte Dental/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Animales , Células Cultivadas , Ratones , Ratones Noqueados , Ratas , Receptores Acoplados a Proteínas G/deficiencia , Receptores Acoplados a Proteínas G/genética
5.
Am J Pathol ; 190(5): 1046-1058, 2020 05.
Artículo en Inglés | MEDLINE | ID: mdl-32084364

RESUMEN

Cell migration-inducing hyaluronidase 1 (CEMIP), also known as hyaluronan (HA)-binding protein involved in HA depolymerization (HYBID), plays a role in HA degradation. CEMIP2, also known as transmembrane protein 2 (TMEM2), possessing a sequence similarity with HYBID, is reported as a hyaluronidase in mice. However, the expression of these molecules in osteoarthritic synovium and their involvement in HA degradation in synovial fluid (SF) from patients with knee osteoarthritis remain elusive. This study examined their expression in synovial tissue and the relationship with molecular weight of HA in SF in knee osteoarthritis patients. Quantification of mRNA demonstrated that HYBID expression is significantly (5.5-fold) higher in osteoarthritic synovium than in normal control synovium, whereas TMEM2 expression level is similar between the two groups. By immunohistochemistry, HYBID was localized mainly to CD68-negative and fibroblast-specific protein 1-positive synovial lining cells and sublining fibroblasts in osteoarthritic synovium. The mRNA expression levels of HYBID, but not TMEM2, in osteoarthritic synovium positively correlated with distribution of lower-molecular-weight HA with below 1000 kDa in SF. HA-degrading activity in osteoarthritic synovial fibroblasts was abrogated by siRNA-mediated knockdown of HYBID. Among the 12 factors examined, IL-6 significantly up-regulated the HYBID expression and HA-degrading activity in osteoarthritic synovial fibroblasts. These data suggest that HYBID overexpressed by IL-6-stimulated synovial fibroblasts is implicated in HA degradation in osteoarthritic synovium.


Asunto(s)
Fibroblastos/metabolismo , Ácido Hialurónico/metabolismo , Hialuronoglucosaminidasa/metabolismo , Proteínas de la Membrana/metabolismo , Osteoartritis de la Rodilla/metabolismo , Anciano , Femenino , Humanos , Masculino , Osteoartritis de la Rodilla/patología , Membrana Sinovial/metabolismo , Membrana Sinovial/patología
6.
J Biol Chem ; 294(10): 3406-3418, 2019 03 08.
Artículo en Inglés | MEDLINE | ID: mdl-30504223

RESUMEN

The development of ectodermal organs, such as teeth, requires epithelial-mesenchymal interactions. Basic helix-loop-helix (bHLH) transcription factors regulate various aspects of tissue development, and we have previously identified a bHLH transcription factor, AmeloD, from a tooth germ cDNA library. Here, we provide both in vitro and in vivo evidence that AmeloD is important in tooth development. We created AmeloD-knockout (KO) mice to identify the in vivo functions of AmeloD that are critical for tooth morphogenesis. We found that AmeloD-KO mice developed enamel hypoplasia and small teeth because of increased expression of E-cadherin in inner enamel epithelial (IEE) cells, and it may cause inhibition of the cell migration. We used the CLDE dental epithelial cell line to conduct further mechanistic analyses to determine whether AmeloD overexpression in CLDE cells suppresses E-cadherin expression and promotes cell migration. Knockout of epiprofin (Epfn), another transcription factor required for tooth morphogenesis and development, and analysis of AmeloD expression and deletion revealed that AmeloD also contributed to multiple tooth formation in Epfn-KO mice by promoting the invasion of dental epithelial cells into the mesenchymal region. Thus, AmeloD appears to play an important role in tooth morphogenesis by modulating E-cadherin and dental epithelial-mesenchymal interactions. These findings provide detailed insights into the mechanism of ectodermal organ development.


Asunto(s)
Movimiento Celular , Células Epiteliales/citología , Diente/citología , Factores Generales de Transcripción/metabolismo , Secuencia de Aminoácidos , Animales , Cadherinas/metabolismo , Línea Celular , Proliferación Celular , Células Epiteliales/metabolismo , Regulación de la Expresión Génica , Técnicas de Inactivación de Genes , Ratones , Diente/metabolismo
7.
Biochem Biophys Res Commun ; 523(1): 171-176, 2020 02 26.
Artículo en Inglés | MEDLINE | ID: mdl-31839217

RESUMEN

Cell adhesion between oligodendrocytes and neuronal axons is a critical step for myelination that enables the rapid propagation of action potential in the central nervous system. Here, we show that the transmembrane protein teneurin-4 plays a role in the cell adhesion required for the differentiation of oligodendrocytes. We found that teneurin-4 formed molecular complexes with all of the four teneurin family members and promoted cell-cell adhesion. Oligodendrocyte lineage cells attached to the recombinant extracellular domain of all the teneurins and formed well-branched cell processes. In an axon-mimicking nanofibers assay, nanofibers coated with the recombinant teneurin-4 extracellular domain increased the differentiation of oligodendrocytes. Our results show that teneurin-4 binds to all teneurins through their extracellular domain, which facilitates the oligodendrocyte-axon adhesion, and promotes oligodendrocyte differentiation via its homophilic interaction.


Asunto(s)
Adhesión Celular , Diferenciación Celular , Espacio Extracelular/metabolismo , Proteínas de la Membrana/química , Proteínas de la Membrana/metabolismo , Oligodendroglía/citología , Animales , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Oligodendroglía/metabolismo , Dominios Proteicos , Ratas , Ratas Wistar
8.
Microvasc Res ; 129: 103986, 2020 05.
Artículo en Inglés | MEDLINE | ID: mdl-32017943

RESUMEN

Wet age-related macular degeneration (AMD) and diabetic retinopathy are the leading causes of blindness through increased angiogenesis. Although VEGF-neutralizing proteins provide benefit, inconsistent responses indicate a need for new therapies. We previously identified the Fibulin-7 C-terminal fragment (Fbln7-C) as an angiogenesis inhibitor in vitro. Here we show that Fbln7-C inhibits neovascularization in vivo, in both a model of wet AMD involving choroidal neovascularization (CNV) and diabetic retinopathy involving oxygen-induced ischemic retinopathy. Furthermore, a short peptide sequence from Fbln7-C is responsible for the anti-angiogenic properties of Fbln7-C. Our work suggests Fbln7-C as a therapeutic candidate for wet AMD and ischemic retinopathy.


Asunto(s)
Inhibidores de la Angiogénesis/farmacología , Proteínas de Unión al Calcio/farmacología , Coroides/irrigación sanguínea , Neovascularización Coroidal/prevención & control , Fragmentos de Péptidos/farmacología , Neovascularización Retiniana/prevención & control , Vasos Retinianos/efectos de los fármacos , Degeneración Macular Húmeda/prevención & control , Animales , Proteínas de Unión al Calcio/síntesis química , Proteínas de Unión al Calcio/genética , Neovascularización Coroidal/genética , Neovascularización Coroidal/metabolismo , Neovascularización Coroidal/patología , Modelos Animales de Enfermedad , Femenino , Ratones Endogámicos C57BL , Fragmentos de Péptidos/síntesis química , Neovascularización Retiniana/genética , Neovascularización Retiniana/metabolismo , Neovascularización Retiniana/patología , Vasos Retinianos/metabolismo , Vasos Retinianos/patología , Degeneración Macular Húmeda/genética , Degeneración Macular Húmeda/metabolismo , Degeneración Macular Húmeda/patología
9.
Int J Cancer ; 145(8): 2157-2169, 2019 10 15.
Artículo en Inglés | MEDLINE | ID: mdl-30924128

RESUMEN

Glioblastoma (GBM) is pathologically characterized by highly malignant neoplastic cells, focal necrosis and aberrant blood vessels composed of disorganized endothelial cells and pericytes. The recent cancer microarray database revealed upregulation of fibulin-7 (Fbln7), a member of the fibulin family, but provided no information on the tissue localization or biological function. In the present study, we demonstrated that Fbln7 is markedly overexpressed by the GBM tissue among astrocytic tumors, and immunolocalized mainly to endothelial cells and pericytes of the glomeruloid and hypertrophied microvessels. The production of Fbln7 by endothelial cells and pericytes was confirmed in cultured human umbilical vein endothelial cells (HUVEC) and human brain vascular pericytes (HBVP) and vascular endothelial growth factor (VEGF) stimulated the Fbln7 expression in HUVEC. Fbln7 bound to angiopoietin-1, but not angiopoietin-2 or Tie2 receptor, through interaction between the N-terminal portions of Fbln7 and angiopoietin-1, and it blocked phosphorylation of Tie2 receptor in HUVEC. In a coculture assay using HUVEC and HBVP, multilayered and irregular-shaped tube-like structures of HUVEC were induced by treatment with a high concentration of VEGF. This was accompanied by Fbln7 overproduction by HUVEC and angiopoietin-1 expression by HBVP. The production of aberrant VEGF-induced tube-like structures was attenuated by treatment with antibody or synthetic peptides specific to the Fbln7 N-terminal domain or knockdown of Fbln7. These data demonstrate that Fbln7 is overexpressed by endothelial cells and pericytes of the abnormal microvessels in GBM, and suggest that Fbln7 may contribute to the aberrant vessel formation by modulation of the angiopoietin-1/angiopoietin-2-Tie2 axis.


Asunto(s)
Angiopoyetina 1/genética , Neoplasias Encefálicas/genética , Proteínas de Unión al Calcio/genética , Glioblastoma/genética , Neovascularización Patológica/genética , Angiopoyetina 1/metabolismo , Neoplasias Encefálicas/irrigación sanguínea , Neoplasias Encefálicas/metabolismo , Proteínas de Unión al Calcio/metabolismo , Células Cultivadas , Técnicas de Cocultivo , Regulación Neoplásica de la Expresión Génica , Glioblastoma/irrigación sanguínea , Glioblastoma/metabolismo , Células Endoteliales de la Vena Umbilical Humana/efectos de los fármacos , Células Endoteliales de la Vena Umbilical Humana/metabolismo , Células Endoteliales de la Vena Umbilical Humana/fisiología , Humanos , Neovascularización Patológica/metabolismo , Neovascularización Fisiológica/efectos de los fármacos , Neovascularización Fisiológica/genética , Pericitos/citología , Pericitos/efectos de los fármacos , Pericitos/metabolismo , Unión Proteica , Factor A de Crecimiento Endotelial Vascular/farmacología
10.
FASEB J ; 32(9): 4889-4898, 2018 09.
Artículo en Inglés | MEDLINE | ID: mdl-29634368

RESUMEN

Fibulin-7 (Fbln7) has been identified as the latest member of the fibulin family of secreted glycoproteins in developing teeth, functioning as a cell adhesion molecule and interacting with other matrix proteins, receptors, and growth factors. More recently, we have shown that the C-terminal Fbln7 fragment (Fbln7-C) has antiangiogenic activity in vitro. Fbln7 is also expressed in immune-privileged tissues, such as eye and placenta, but its functional significance is unknown. In the current study, we show that human monocytes adhere to both full-length Fbln7 (Fbln7-FL) and Fbln7-C, in part, via integrins α5ß1 and α2ß1. Morphologic studies and surface expression analyses of CD14, mannose receptor (CD206), major histocompatibility complex II, and CD11b receptors revealed that both Fbln7-FL and Fbln7-C inhibit M-CSF-induced monocyte differentiation. Fbln7-C had significantly greater negative effects on cell spreading and stress fiber formation, including the production of IL-6 and metalloproteinase-1/-9 compared with Fbln7-FL. Furthermore, in an LPS-induced systemic inflammation model, Fbln7-C and Fbln7-FL reduced the infiltration of immune cells, such as neutrophils and macrophages, to the inflamed peritoneum. Thus, these results suggest that Fbln7 and Fbln7-C could modulate the activity of immune cells and have therapeutic potential for inflammatory diseases.-Sarangi, P. P., Chakraborty, P., Dash, S. P., Ikeuchi, T., de Vega, S., Ambatipudi, K., Wahl, L., Yamada, Y. Cell adhesion protein fibulin-7 and its C-terminal fragment negatively regulate monocyte and macrophage migration and functions in vitro and in vivo.


Asunto(s)
Moléculas de Adhesión Celular/metabolismo , Adhesión Celular/fisiología , Macrófagos/metabolismo , Monocitos/metabolismo , Animales , Proteínas de Unión al Calcio/metabolismo , Adhesión Celular/efectos de los fármacos , Movimiento Celular/efectos de los fármacos , Movimiento Celular/fisiología , Células Cultivadas , Humanos , Lectinas Tipo C/metabolismo , Lipopolisacáridos/farmacología , Macrófagos/efectos de los fármacos , Receptor de Manosa , Lectinas de Unión a Manosa/metabolismo , Ratones Endogámicos C57BL , Monocitos/efectos de los fármacos , Neutrófilos/metabolismo , Receptores de Superficie Celular/metabolismo
11.
Biopolymers ; 106(2): 184-195, 2016 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-26491858

RESUMEN

Despite the research done on pathological angiogenesis, there is still a need for the development of new therapies against angiogenesis-related diseases. Fibulin-7 (Fbln7) is a member of the extracellular matrix fibulin protein family. The Fbln7 C-terminal fragment, Fbln7-C, binds to endothelial cells and inhibits their tube formation in culture. In this study, we screened 12 synthetic peptides, covering the fibulin-globular domain of Fbln7-C, to identify active sites for endothelial cell adhesion and in vitro antiangiogenic activity. Three peptides, fc10, fc11, and fc12, promoted Human Umbilical Vein Endothelial Cells (HUVECs) adhesion, and the morphology of HUVECs on fc10 was similar to that on Fbln7-C. EDTA and the anti-integrin ß1 function-blocking antibody inhibited HUVECs adhesion to both fc10 and fc12, and heparin inhibited HUVECs adhesion to both fc11 and fc12. fc10 and fc11 inhibited HUVECs tube formation. Our results suggest that three peptides from Fbln7-C are biologically active for endothelial cell adhesion and disrupt the tube formation, suggesting a potential therapeutic use of these peptides for angiogenesis-related diseases. © 2015 Wiley Periodicals, Inc. Biopolymers (Pept Sci) 106: 184-195, 2016.

12.
Am J Pathol ; 184(6): 1683-94, 2014 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-24720953

RESUMEN

Laminin α1 (LAMA1), a subunit of the laminin-111 basement membrane component, has been implicated in various biological functions in vivo and in vitro. Although LAMA1 is present in kidney, its roles in the kidney are unknown because of early embryonic lethality. Herein, we used a viable conditional knockout mouse model with a deletion of Lama1 in the epiblast lineage (Lama1(CKO)) to study the role of LAMA1 in kidney development and function. Adult Lama1(CKO) mice developed focal glomerulosclerosis and proteinuria with age. In addition, mesangial cell proliferation was increased, and the mesangial matrix, which normally contains laminin-111, was greatly expanded. In vitro, mesangial cells from Lama1(CKO) mice exhibited significantly increased proliferation compared with those from controls. This increased proliferation was inhibited by the addition of exogenous LAMA1-containing laminin-111, but not by laminin-211 or laminin-511, suggesting a specific role for LAMA1 in regulating mesangial cell behavior. Moreover, the absence of LAMA1 increased transforming growth factor (TGF)-ß1-induced Smad2 phosphorylation, and inhibitors of TGF-ß1 receptor I kinase blocked Smad2 phosphorylation in both control and Lama1(CKO) mesangial cells, indicating that the increased Smad2 phosphorylation occurred in the absence of LAMA1 via the TGF-ß1 receptor. These findings suggest that LAMA1 plays a critical role in kidney function and kidney aging by regulating the mesangial cell population and mesangial matrix deposition through TGF-ß/Smad signaling.


Asunto(s)
Envejecimiento/metabolismo , Proliferación Celular , Matriz Extracelular/metabolismo , Mesangio Glomerular/metabolismo , Laminina/metabolismo , Envejecimiento/genética , Envejecimiento/patología , Animales , Matriz Extracelular/genética , Matriz Extracelular/patología , Mesangio Glomerular/patología , Glomerulonefritis/genética , Glomerulonefritis/metabolismo , Glomerulonefritis/patología , Laminina/genética , Ratones , Ratones Noqueados , Fosforilación/genética , Proteinuria/genética , Proteinuria/metabolismo , Proteinuria/patología , Transducción de Señal/genética , Proteína Smad2/genética , Proteína Smad2/metabolismo , Factor de Crecimiento Transformador beta1/genética , Factor de Crecimiento Transformador beta1/metabolismo
13.
FASEB J ; 28(3): 1386-97, 2014 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-24344332

RESUMEN

Teneurin-4 (Ten-4), a transmembrane protein, is highly expressed in the central nervous system; however, its cellular and molecular function in neuronal differentiation remains unknown. In this study, we aimed to elucidate the function of Ten-4 in neurite outgrowth. Ten-4 expression was induced during neurite outgrowth of the neuroblastoma cell line Neuro-2a. Ten-4 protein was localized at the neurite growth cones. Knockdown of Ten-4 expression in Neuro-2a cells decreased the formation of the filopodia-like protrusions and the length of individual neurites. Conversely, overexpression of Ten-4 promoted filopodia-like protrusion formation. In addition, knockdown and overexpression of Ten-4 reduced and elevated the activation of focal adhesion kinase (FAK) and Rho-family small GTPases, Cdc42 and Rac1, key molecules for the membranous protrusion formation downstream of FAK, respectively. Inhibition of the activation of FAK and neural Wiskott-Aldrich syndrome protein (N-WASP), which is a downstream regulator of FAK and Cdc42, blocked protrusion formation by Ten-4 overexpression. Further, Ten-4 colocalized with phosphorylated FAK in the filopodia-like protrusion regions. Together, our findings show that Ten-4 is a novel positive regulator of cellular protrusion formation and neurite outgrowth through the FAK signaling pathway.


Asunto(s)
Proteína-Tirosina Quinasas de Adhesión Focal/metabolismo , Proteínas de la Membrana/fisiología , Neuritas , Transducción de Señal , Animales , Secuencia de Bases , Cartilla de ADN , Ratones , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa
14.
Arch Biochem Biophys ; 545: 148-53, 2014 Mar 01.
Artículo en Inglés | MEDLINE | ID: mdl-24480309

RESUMEN

We have previously demonstrated that fibulin-7 (Fbln7) is expressed in teeth by pre-odontoblast and odontoblast cells, localized in the basement membrane and dentin matrices, and is an adhesion molecule for dental mesenchyme cells and odontoblasts. Fbln7 is also expressed in blood vessels by endothelial cells. In this report, we show that a recombinant C-terminal Fbln7 fragment (Fbln7-C) bound to Human Umbilical Vein Endothelial Cells (HUVECs) but did not promote cell spreading and actin stress fiber formation. Fbln7-C binding to HUVECs induced integrin clustering at cell adhesion sites with other focal adhesion molecules, and sustained activation of FAK, p130Cas, and Rac1. In addition, RhoA activation was inhibited, thereby preventing HUVEC spreading. As endothelial cell spreading is an important step for angiogenesis, we examined the effect of Fbln7-C on angiogenesis using in vitro assays for endothelial cell tube formation and vessel sprouting from aortic rings. We found that Fbln7-C inhibited the HUVEC tube formation and the vessel sprouting in aortic ring assays. Our findings suggest potential anti-angiogenic activity of the Fbln7 C-terminal region.


Asunto(s)
Proteínas de Unión al Calcio/metabolismo , Células Endoteliales de la Vena Umbilical Humana/metabolismo , Neovascularización Fisiológica , Animales , Proteínas de Unión al Calcio/química , Adhesión Celular , Proteína Sustrato Asociada a CrK/metabolismo , Activación Enzimática , Adhesiones Focales/metabolismo , Adhesiones Focales/ultraestructura , Células Endoteliales de la Vena Umbilical Humana/citología , Humanos , Integrinas/metabolismo , Ratones , Fosforilación , Unión Proteica , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Fibras de Estrés/metabolismo , Proteína de Unión al GTP rac1/metabolismo , Proteína de Unión al GTP rhoA/metabolismo
15.
Int J Dev Biol ; 68(1): 19-24, 2024.
Artículo en Inglés | MEDLINE | ID: mdl-38591690

RESUMEN

Tooth formation is a process tightly regulated by reciprocal interactions between epithelial and mesenchymal tissues. These epithelial-mesenchyme interactions regulate the expression of target genes via transcription factors. Among the regulatory elements governing this process, Epiprofin/Sp6 is a zinc finger transcription factor which is expressed in the embryonic dental epithelium and in differentiating pre-odontoblasts. Epiprofin knockout (Epfn-/-) mice present severe dental abnormalities, such as supernumerary teeth and enamel hypoplasia. Here, we describe dentin defects in molars and incisors of Epfn-/- mice. We observed that in the absence of Epfn, markers of early odontoblast differentiation, such as alkaline phosphatase activity, Dsp/Dpp expression, and Collagen Type I deposition, are downregulated. In addition, the expression of tight and gap junction proteins was severely impaired in the predontoblastic cell layer of developing Epfn-/- molars. Altogether, our data shows that Epfn is crucial for the proper differentiation of dental mesenchymal cells towards functional odontoblasts and subsequent dentin-matrix deposition.


Asunto(s)
Displasia de la Dentina , Odontoblastos , Ratones , Animales , Odontoblastos/metabolismo , Displasia de la Dentina/metabolismo , Diferenciación Celular , Odontogénesis , Factores de Transcripción/genética , Factores de Transcripción/metabolismo
16.
Structure ; 2024 Oct 03.
Artículo en Inglés | MEDLINE | ID: mdl-39395422

RESUMEN

Pathogenic bacteria must secure the uptake of nutritional metals such as iron for their growth, making their import systems attractive targets for the development of new antimicrobial modalities. In the pathogenic bacterium Streptococcus pyogenes, the iron uptake system FtsABCD transports iron encapsulated by siderophores of the hydroxamate class. However, the inability of S. pyogenes to produce these metabolites makes the biological and clinical relevance of this route unresolved. Herein, we demonstrated that the periplasmic binding protein FtsB recognizes not only the hydroxamate siderophore ferrichrome, as previously documented, but also ferrioxamine E (FOE), ferrioxamine B (FOB), and bisucaberin (BIS), each of them with high affinity (nM level). Up to seven aromatic residues in the binding pocket accommodate the variable backbones of the different siderophores through CH-π interactions, explaining ligand promiscuity. Collectively, our observations revealed how S. pyogenes exploits the diverse xenosiderophores produced by other microorganisms as iron sources to secure this precious nutrient.

17.
J Neurosci ; 32(34): 11586-99, 2012 Aug 22.
Artículo en Inglés | MEDLINE | ID: mdl-22915103

RESUMEN

Myelination is essential for proper functioning of the CNS. In this study, we have identified a mouse mutation, designated furue, which causes tremors and hypomyelination in the CNS, particularly in the spinal cord, but not in the sciatic nerve of the PNS. In the spinal cord of the furue mice, myelination of small-diameter axons was dramatically reduced, and differentiation of oligodendrocytes, the myelin-forming cells in the CNS, was inhibited. We subsequently found that the furue mutation was associated with a transgene insertion into the teneurin-4 (Ten-4, Ten-m4/Odz4) gene, encoding a transmembrane protein of unknown function. Ten-4 was strongly expressed in the spinal cord of wild-type mice and was induced during normal oligodendrocyte differentiation. In contrast, in the furue mice, the expression of Ten-4 was absent. Differentiation and cellular process formation of oligodendrocytes were inhibited in primary cell culture from the furue mice. Cell differentiation and process formation were also inhibited in the oligodendrocyte progenitor cell line CG-4 after suppression of Ten-4 expression by shRNA. Furthermore, Ten-4 positively regulated focal adhesion kinase, an essential signaling molecule for oligodendrocyte process formation and myelination of small-diameter axons. These findings suggest that Ten-4 is a novel regulator of oligodendrocyte differentiation and that it plays a critical role in the myelination of small-diameter axons in the CNS.


Asunto(s)
Axones/metabolismo , Diferenciación Celular/genética , Sistema Nervioso Central , Enfermedades Desmielinizantes/genética , Proteínas Nucleares/deficiencia , Oligodendroglía/citología , 2',3'-Nucleótido Cíclico 3'-Fosfodiesterasa/genética , 2',3'-Nucleótido Cíclico 3'-Fosfodiesterasa/metabolismo , Proteína de la Poliposis Adenomatosa del Colon/metabolismo , Factores de Edad , Animales , Animales Recién Nacidos , Antígenos/metabolismo , Axones/patología , Axones/ultraestructura , Encéfalo/citología , Tamaño de la Célula , Sistema Nervioso Central/metabolismo , Sistema Nervioso Central/patología , Sistema Nervioso Central/fisiopatología , Quinasa 1 de Adhesión Focal/genética , Quinasa 1 de Adhesión Focal/metabolismo , Galactosilceramidasa/metabolismo , Regulación del Desarrollo de la Expresión Génica/genética , Humanos , Proteínas de la Membrana , Ratones , Ratones Endogámicos C57BL , Ratones Mutantes , Ratones Transgénicos , Microscopía Electrónica de Transmisión , Proteína Básica de Mielina/metabolismo , Neuroglía/fisiología , Proteínas Nucleares/genética , Organogénesis , Proteoglicanos/metabolismo , ARN Mensajero/metabolismo , ARN Interferente Pequeño/genética , Receptores de Interleucina-2/genética , Receptores de Interleucina-2/metabolismo , Transfección
18.
Sci Rep ; 12(1): 17242, 2022 10 14.
Artículo en Inglés | MEDLINE | ID: mdl-36241903

RESUMEN

Destruction of articular cartilage in osteoarthritis (OA) is initiated by depletion of the hyaluronan (HA)-aggrecan network, followed by degradation of the collagen fibrils. Previously, we reported the implications of HA-binding protein involved in HA depolymerization (HYBID), alias cell migration-inducing protein (CEMIP) and KIAA1199, for HA degradation. However, transmembrane protein 2 (TMEM2), which is ~ 50% homologous to HYBID, was discovered as another hyaluronidase, but their expression and regulation by OA chondrocytes remain elusive. Here we report that the absolute mRNA copy numbers of HYBID are significantly (7.1-fold) higher in OA cartilage than normal cartilage, whereas TMEM2 levels are not different between the groups. HA-degrading activity of cultured OA chondrocytes disappeared by siRNA-mediated knockdown of HYBID, but not TMEM2. HYBID expression was significantly up-regulated by treatment with interleukin-6 (IL-6) or tumor necrosis factor-α (TNF-α) and additively increased by the combined treatment. No significant changes in the TMEM2 expression were seen by the factors examined. IL-1α remarkably enhanced IL-6 production and increased HYBID expression when soluble IL-6 receptor was supplemented. These results demonstrate that in stark contrast to the constitutive expression of TMEM2 and its negligible HA-degrading activity, HYBID is overexpressed in OA cartilage and up-regulated by IL-6 and TNF-α in OA chondrocytes.


Asunto(s)
Cartílago Articular , Osteoartritis , Humanos , Agrecanos/metabolismo , Cartílago Articular/patología , Células Cultivadas , Condrocitos/metabolismo , Colágeno/metabolismo , Ácido Hialurónico/metabolismo , Hialuronoglucosaminidasa/genética , Hialuronoglucosaminidasa/metabolismo , Interleucina-6/metabolismo , Osteoartritis/patología , Receptores de Interleucina-6/metabolismo , ARN Mensajero/metabolismo , ARN Interferente Pequeño/metabolismo , Factor de Necrosis Tumoral alfa/metabolismo , Factor de Necrosis Tumoral alfa/farmacología
19.
Dev Biol ; 337(2): 363-74, 2010 Jan 15.
Artículo en Inglés | MEDLINE | ID: mdl-19913006

RESUMEN

The formation and maintenance of the apical ectodermal ridge (AER) is critical for the outgrowth and patterning of the vertebrate limb. In the present work, we have investigated the role of Epiprofin (Epfn/Sp6), a member of the SP/KLF transcription factor family that is expressed in the limb ectoderm and the AER, during limb development. Epfn mutant mice have a defective autopod that shows mesoaxial syndactyly in the forelimb and synostosis (bony fusion) in the hindlimb and partial bidorsal digital tips. Epfn mutants also show a defect in the maturation of the AER that appears flat and broad, with a double ridge phenotype. By genetic analysis, we also show that Epfn is controlled by WNT/b-CATENIN signaling in the limb ectoderm. Since the less severe phenotypes of the conditional removal of b-catenin in the limb ectoderm strongly resemble the limb phenotype of Epfn mutants, we propose that EPFN very likely functions as a modulator of WNT signaling in the limb ectoderm.


Asunto(s)
Extremidades/embriología , Factores de Transcripción de Tipo Kruppel/metabolismo , Dedos de Zinc , Animales , Tipificación del Cuerpo/genética , Proteína Morfogenética Ósea 4/metabolismo , Muerte Celular , Proliferación Celular , Ectodermo/embriología , Ectodermo/metabolismo , Ectodermo/patología , Embrión de Mamíferos/metabolismo , Embrión de Mamíferos/patología , Extremidades/patología , Factor 8 de Crecimiento de Fibroblastos/metabolismo , Regulación del Desarrollo de la Expresión Génica , Factores de Transcripción de Tipo Kruppel/deficiencia , Factores de Transcripción de Tipo Kruppel/genética , Ratones , Mutación/genética , Fenotipo , Transducción de Señal , Proteínas Wnt/metabolismo , beta Catenina/metabolismo
20.
J Biol Chem ; 285(24): 18948-58, 2010 Jun 11.
Artículo en Inglés | MEDLINE | ID: mdl-20404334

RESUMEN

Pannexin 3 (Panx3) is a new member of the gap junction pannexin family, but its expression profiles and physiological function are not yet clear. We demonstrate in this study that Panx3 is expressed in cartilage and regulates chondrocyte proliferation and differentiation. Panx3 mRNA was expressed in the prehypertrophic zone in the developing growth plate and was induced during the differentiation of chondrogenic ATDC5 and N1511 cells. Panx3-transfected ATDC5 and N1511 cells promoted chondrogenic differentiation, but the suppression of endogenous Panx3 inhibited differentiation of ATDC5 cells and primary chondrocytes. Panx3-transfected ATDC5 cells reduced parathyroid hormone-induced cell proliferation and promoted the release of ATP into the extracellular space, possibly by action of Panx3 as a hemichannel. Panx3 expression in ATDC5 cells reduced intracellular cAMP levels and the activation of cAMP-response element-binding, a protein kinase A downstream effector. These Panx3 activities were blocked by anti-Panx3 antibody. Our results suggest that Panx3 functions to switch the chondrocyte cell fate from proliferation to differentiation by regulating the intracellular ATP/cAMP levels.


Asunto(s)
Adenosina Trifosfato/metabolismo , Condrocitos/citología , Conexinas/metabolismo , AMP Cíclico/metabolismo , Regulación de la Expresión Génica , Animales , Diferenciación Celular , Proliferación Celular , Condrocitos/metabolismo , Proteínas Quinasas Dependientes de AMP Cíclico/metabolismo , Humanos , Ratones , Modelos Biológicos , ARN Mensajero/metabolismo , Elementos de Respuesta
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