RESUMEN
BACKGROUND: The coronavirus disease of 2019 pandemic impacted all facets of health care in the United States, including the professional training for podiatry residents and students. In March of 2020, the Association of American Medical Colleges recommended pausing then modifying all clinical rotations. The podiatric community followed suit. In-person restrictions, cancellations of clerkships, limited clinical experiences, virtual didactic programs, and reduced surgical cases for students and residency programs occurred for many months during the ongoing pandemic. These adaptations impacted the ability of podiatric students to complete clinical rotations and clerkships, which are pivotal to their academic curriculum and residency program application and selection. METHODS: A survey was conducted by the Council of Teaching Hospitals (COTH) and sent out by the American Association of Colleges of Podiatric Medicine. The 2021 postinterview surveys were sent out to all participants in the 2021 Centralized Application Service for Podiatric Residencies Web application and match cycle, both programs and candidates. RESULTS: The COTH presents results and comments from the 2021 virtual interview experience and residency match. Data and anecdotal comments from the 2021 postinterview survey conducted by COTH, sent out by American Association of Colleges of Podiatric Medicine, are presented here. CONCLUSIONS: Results from the surveys of program directors and candidates show a preference by both groups for in-person interviews despite the personal time demands and increased costs associated with travel.
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COVID-19 , Internado y Residencia , Podiatría , Humanos , COVID-19/epidemiología , Curriculum , Hospitales de Enseñanza , Encuestas y CuestionariosRESUMEN
Ovarian cancer is the fifth-leading cause of cancer death among women. The dissemination of ovarian tumors and growth as spheroids accompanies late-stage disease. In cell culture, ovarian tumor cell spheroids can exhibit elevated resistance to environmental stressors, such as reactive oxygen species. Homeostatic balance of the antioxidant response is a protective mechanism that prevents anoikis, a form of programmed cell death. Signaling pathways activated by integrin receptors suppress anoikis. Rgnef (ARHGEF28/p190RhoGEF) is a guanine nucleotide exchange factor that is activated downstream of integrins. We find that Rgnef protein levels are elevated in late-stage serous ovarian cancer, high Rgnef mRNA levels are associated with decreased progression-free and overall survival, and genomic ARHGEF28 loss is associated with increased patient survival. Using transgenic and transplantable Rgnef knockout mouse models, we find that Rgnef is essential for supporting three-dimensional ovarian spheroid formation in vitro and tumor growth in mice. Using RNA-sequencing and bioinformatic analyses, we identify a conserved Rgnef-supported anti-oxidant gene signature including Gpx4, Nqo1, and Gsta4; common targets of the NF-kB transcription factor. Antioxidant treatment enhanced growth of Rgnef-knockout spheroids and Rgnef re-expression facilitated NF-κB-dependent tumorsphere survival. These studies reveal a new role for Rgnef in ovarian cancer to facilitate NF-κB-mediated gene expression protecting cells from oxidative stress.
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Factores de Intercambio de Guanina Nucleótido/fisiología , Neoplasias Ováricas/genética , Neoplasias Ováricas/patología , Estrés Oxidativo/genética , ras-GRF1/fisiología , Animales , Proliferación Celular/genética , Citoprotección/genética , Progresión de la Enfermedad , Femenino , Factores de Intercambio de Guanina Nucleótido/genética , Células HEK293 , Humanos , Ratones , Ratones Noqueados , FN-kappa B/metabolismo , Neoplasias Ováricas/metabolismo , Transducción de Señal/genética , Células Tumorales Cultivadas , ras-GRF1/genéticaRESUMEN
This practice memo, a collaborative effort between the Young Physicians' Program of the American Podiatric Medical Association (APMA) and the Young Surgeons Committee of the Society for Vascular Surgery (SVS), is intended to aid podiatrists and vascular surgeons in the early years of their respective careers, especially those involved in the care of patients with chronic wounds. During these formative years, learning how to successfully establish an inter-professional partnership is crucial in order to provide the best possible care to this important patient population.
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Evaluación de Resultado en la Atención de Salud , Práctica Asociada/organización & administración , Podiatría , Cirujanos/organización & administración , Procedimientos Quirúrgicos Vasculares/métodos , Prestación Integrada de Atención de Salud/organización & administración , Femenino , Humanos , Relaciones Interprofesionales , Masculino , Innovación Organizacional , Sociedades Médicas , Estados UnidosRESUMEN
This practice memo, a collaborative effort between the Young Physicians' Program of the American Podiatric Medical Association and the Young Surgeons Committee of the Society for Vascular Surgery, is intended to aid podiatrists and vascular surgeons in the early years of their respective careers, especially those involved in the care of patients with chronic wounds. During these formative years, learning how to successfully establish an interprofessional partnership is crucial to provide the best possible care to this important population of patients.
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Conducta Cooperativa , Prestación Integrada de Atención de Salud , Práctica Asociada , Grupo de Atención al Paciente , Podiatría , Cirujanos , Procedimientos Quirúrgicos Vasculares , Heridas y Lesiones/terapia , Enfermedad Crónica , Análisis Costo-Beneficio , Prestación Integrada de Atención de Salud/economía , Costos de la Atención en Salud , Humanos , Comunicación Interdisciplinaria , Práctica Asociada/economía , Grupo de Atención al Paciente/economía , Podiatría/economía , Cirujanos/economía , Resultado del Tratamiento , Procedimientos Quirúrgicos Vasculares/economía , Cicatrización de Heridas , Heridas y Lesiones/diagnóstico , Heridas y Lesiones/economía , Heridas y Lesiones/fisiopatologíaRESUMEN
OPINION STATEMENT: Despite an explosion in the number of options available for helping diabetic patients heal wounds, major amputation remains a critical issue for these persons. Since diabetes prematurely ages tissues and no organ system is immune to its presence, it makes inherent sense that multi-disciplinary team approaches to these patients is necessary to make significant strides forward. Here, we present literature from the fields of podiatric surgery/medicine, vascular and plastic surgery and introduce the successes that a multi-disciplinary limb salvage center can have on the lives and limbs of patients with diabetes.
RESUMEN
Eccrine syringofibroadenoma (ESFA) is a rare benign lesion of ductal and secretory differentiation exhibiting multiple cutaneous polymorphic presentations with an unknown etiology. We present a case of ESFA that uniquely exhibited large, thick, verrucous-like hyperplastic growths as well as superficial shiny mosaic plaques and deep ulcerations in three different anatomical locations in the same patient. The diagnosis of ESFA was confirmed histologically after biopsies were performed on all of the affected areas. In addition to a case report and literature review, we also present classification, clinical, and histologic aspects of ESFA.
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Adenoma de las Glándulas Sudoríparas/patología , Amputación Quirúrgica/métodos , Talón/patología , Neoplasias de las Glándulas Sudoríparas/patología , Neoplasias de las Glándulas Sudoríparas/cirugía , Adenoma de las Glándulas Sudoríparas/cirugía , Biopsia con Aguja , Estudios de Seguimiento , Humanos , Inmunohistoquímica , Extremidad Inferior/patología , Masculino , Persona de Mediana Edad , Invasividad Neoplásica/patología , Estadificación de Neoplasias , Medición de Riesgo , Resultado del TratamientoRESUMEN
Both vascular surgeons and podiatric physicians care for patients with diabetic foot ulcerations (DFUs), one of today's most challenging health-care populations in the United States. The prevalence of DFUs has steadily increased, along with the rising costs associated with care. Because of the numerous comorbidities affecting these patients, it is necessary to take a multidisciplinary approach in the management of these patients. Such efforts, primarily led by podiatric physicians and vascular surgeons, have been shown to effectively decrease major limb loss. Establishing an interprofessional partnership between vascular surgery and podiatric medicine can lead to an improvement in the delivery of care and outcomes of this vulnerable patient population.
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Pie Diabético/cirugía , Podiatría/métodos , Calidad de la Atención de Salud , Procedimientos Quirúrgicos Vasculares/métodos , Pie Diabético/diagnóstico , Pie Diabético/epidemiología , Femenino , Encuestas de Atención de la Salud , Humanos , Comunicación Interdisciplinaria , Relaciones Interprofesionales , Masculino , Práctica Asociada/organización & administración , Cirujanos/estadística & datos numéricos , Resultado del Tratamiento , Estados UnidosRESUMEN
INTRODUCTION: Focal adhesion kinase (FAK) controls cell growth and survival downstream of integrin-matrix receptors. Upon adhesion loss or FAK inhibition, FAK can translocate to the nucleus. The nucleolus is a non-membrane nuclear structure that regulates ribosome biogenesis and cell proliferation. Nucleostemin (NS), a nucleolar-localized protein, modulates cell cycle progression, stemness, and three-dimensional tumor spheroid formation. The signaling pathways that regulate NS levels in tumors remain undefined. METHODS: Human breast carcinoma cells were evaluated for growth in culture (adherent and anchorage-independent spheroid) and as orthotopic tumors. FAK signaling was evaluated by pharmacological FAK inhibitor addition (PF-271, IC50~0.1 µM) and by small hairpin RNA (shRNA) knockdown followed by re-expression of FAK wildtype (WT) or a kinase-dead (KD, K454R) FAK point mutant. Immunoblotting was used to evaluate FAK, NS, nucleolar phosphoprotein B23, and nucleolin levels. Total and phosphospecific antibody imunoblotting were used to detect changes in FAK, Akt kinase (Akt also known as protein kinase B), and 4E-binding protein 1 (4E-BP1) phosphorylation, a translation repressor protein and target of the mammalian target of rapamycin (mTOR) complex. Immunohistochemical, co-immunoprecipitation, and cellular fractionation analyses were used to evaluate FAK association with nucleoli. RESULTS: Pharmacological (0.1 µM PF-271) or genetic inhibition of FAK activity prevents MDA-MB-231 and 4T1L breast carcinoma growth as spheroids and as orthotopic tumors. FAK inhibition triggers proteasome-mediated decreased NS levels but no changes in other nucleolar proteins such as B23 (nucleophosmin) or nucleolin. Active FAK was associated with purified nucleoli of anchorage-independent cells and present within nucleoli of human invasive ductal carcinoma tumor samples. FAK co-immunoprecipitated with B23 that binds NS and a complex between FAK, NS, Akt, and mTOR was detected. Constitutively-active Akt kinase promoted tumor spheroid growth, stabilized NS levels, and promoted pS65 4E-BP1 phosphorylation in the presence of inhibited FAK. Rapamycin lowered NS levels and inhibited pS65 4E-BP1 phosphorylation in cells with activated Akt-mTOR signaling. CONCLUSIONS: FAK signaling occurs in the nucleolus, active FAK protects NS, and Akt-mTOR pathway regulates NS protein stability needed for breast carcinoma spheroid and tumor growth.
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Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Proteína-Tirosina Quinasas de Adhesión Focal/metabolismo , Proteínas de Unión al GTP/metabolismo , Proteínas Nucleares/metabolismo , Animales , Neoplasias de la Mama/genética , Ciclo Celular/genética , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Modelos Animales de Enfermedad , Activación Enzimática , Femenino , Proteína-Tirosina Quinasas de Adhesión Focal/antagonistas & inhibidores , Proteína-Tirosina Quinasas de Adhesión Focal/genética , Humanos , Ratones , Nucleofosmina , Inhibidores de Proteínas Quinasas/farmacología , Transporte de Proteínas , Proteínas Proto-Oncogénicas c-akt/metabolismo , Interferencia de ARN , ARN Mensajero/genética , ARN Interferente Pequeño/genética , Sirolimus/farmacología , Esferoides Celulares , Carga Tumoral/efectos de los fármacos , Carga Tumoral/genética , Células Tumorales Cultivadas , Ensayo de Tumor de Célula MadreRESUMEN
The guanine nucleotide exchange factor Rgnef (also known as ArhGEF28 or p190RhoGEF) promotes colon carcinoma cell motility and tumor progression via interaction with focal adhesion kinase (FAK). Mechanisms of Rgnef activation downstream of integrin or G protein-coupled receptors remain undefined. In the absence of a recognized G protein signaling homology domain in Rgnef, no proximal linkage to G proteins was known. Utilizing multiple methods, we have identified Rgnef as a new effector for Gα13 downstream of gastrin and the type 2 cholecystokinin receptor. In DLD-1 colon carcinoma cells depleted of Gα13, gastrin-induced FAK Tyr(P)-397 and paxillin Tyr(P)-31 phosphorylation were reduced. RhoA GTP binding and promoter activity were increased by Rgnef in combination with active Gα13. Rgnef co-immunoprecipitated with activated Gα13Q226L but not Gα12Q229L. The Rgnef C-terminal (CT, 1279-1582) region was sufficient for co-immunoprecipitation, and Rgnef-CT exogenous expression prevented Gα13-stimulated SRE activity. A domain at the C terminus of the protein close to the FAK binding domain is necessary to bind to Gα13. Point mutations of Rgnef-CT residues disrupt association with active Gα13 but not Gαq. These results show that Rgnef functions as an effector of Gα13 signaling and that this linkage may mediate FAK activation in DLD-1 colon carcinoma cells.
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Neoplasias del Colon/metabolismo , Subunidades alfa de la Proteína de Unión al GTP G12-G13/fisiología , Gastrinas/fisiología , Factores de Intercambio de Guanina Nucleótido/metabolismo , Línea Celular Tumoral , Neoplasias del Colon/patología , Proteína-Tirosina Quinasas de Adhesión Focal/metabolismo , Células HEK293 , Humanos , Paxillin/química , Paxillin/metabolismo , Fosforilación , Receptor de Colecistoquinina B/metabolismo , Factores de Intercambio de Guanina Nucleótido Rho/química , Factores de Intercambio de Guanina Nucleótido Rho/metabolismo , Tirosina/metabolismoRESUMEN
Ovarian cancer ascites fluid contains matrix proteins that can impact tumor growth via integrin receptor binding. In human ovarian tumor tissue arrays, we find that activation of the cytoplasmic focal adhesion (FAK) tyrosine kinase parallels increased tumor stage, ß5 integrin, and osteopontin matrix staining. Elevated osteopontin, ß5 integrin, and FAK mRNA levels are associated with decreased serous ovarian cancer patient survival. FAK remains active within ovarian cancer cells grown as spheroids, and anchorage-independent growth analyses of seven ovarian carcinoma cell lines identified sensitive (HEY, OVCAR8) and resistant (SKOV3-IP, OVCAR10) cells to 0.1 µmol/L FAK inhibitor (VS-4718, formerly PND-1186) treatment. VS-4718 promoted HEY and OVCAR8 G0-G1 cell-cycle arrest followed by cell death, whereas growth of SKOV3-IP and OVCAR10 cells was resistant to 1.0 µmol/L VS-4718. In HEY cells, genetic or pharmacological FAK inhibition prevented tumor growth in mice with corresponding reductions in ß5 integrin and osteopontin expression. ß5 knockdown reduced HEY cell growth in soft agar, tumor growth in mice, and both FAK Y397 phosphorylation and osteopontin expression in spheroids. FAK inhibitor-resistant (SKOV3-IP, OVCAR10) cells exhibited anchorage-independent Akt S473 phosphorylation, and expression of membrane-targeted and active Akt in sensitive cells (HEY, OVCAR8) increased growth but did not create a FAK inhibitor-resistant phenotype. These results link osteopontin, ß5 integrin, and FAK in promoting ovarian tumor progression. ß5 integrin expression may serve as a biomarker for serous ovarian carcinoma cells that possess active FAK signaling.
Asunto(s)
Quinasa 1 de Adhesión Focal/metabolismo , Cadenas beta de Integrinas/metabolismo , Neoplasias Quísticas, Mucinosas y Serosas/metabolismo , Neoplasias Ováricas/metabolismo , Aminopiridinas/farmacología , Animales , Antineoplásicos/farmacología , Adhesión Celular , Línea Celular Tumoral , Proliferación Celular , Resistencia a Antineoplásicos , Femenino , Quinasa 1 de Adhesión Focal/antagonistas & inhibidores , Quinasa 1 de Adhesión Focal/genética , Técnicas de Silenciamiento del Gen , Humanos , Estimación de Kaplan-Meier , Ratones Desnudos , Trasplante de Neoplasias , Neoplasias Quísticas, Mucinosas y Serosas/mortalidad , Osteopontina/metabolismo , Neoplasias Ováricas/mortalidad , Transducción de SeñalRESUMEN
OBJECTIVE: Focal adhesion kinase (FAK) is overexpressed in serous ovarian cancer. Loss of merlin, a product of the neurofibromatosis 2 tumor suppressor gene, is being evaluated as a biomarker for FAK inhibitor sensitivity in mesothelioma. Connections between merlin and FAK in ovarian cancer remain undefined. METHODS: Nine human and two murine ovarian cancer cell lines were analyzed for growth in the presence of a small molecule FAK inhibitor (PF-271, also termed VS-6062) from 0.1 to 1 µM for 72 h. Merlin was evaluated by immunoblotting and immunostaining of a human ovarian tumor tissue array. Growth of cells was analyzed in an orthotopic tumor model and evaluated in vitro after stable shRNA-mediated merlin knockdown. RESULTS: Greater than 50% inhibition of OVCAR8, HEY, and ID8-IP ovarian carcinoma cell growth occurred with 0.1 µM PF-271 in anchorage-independent (p<0.001) but not in adherent culture conditions. PF-271-mediated reduction in FAK Y397 phosphorylation occurred independently of growth inhibition. Suspended growth of OVCAR3, OVCAR10, IGROV1, IGROV1-IP, SKOV3, SKOV3-IP, A2780, and 5009-MOVCAR was not affected by 0.1 µM PF-271. Merlin expression did not correlate with serous ovarian tumor grade or stage. PF-271 (30 mg/kg, BID) did not inhibit 5009-MOVCAR tumor growth and merlin knockdown in SKOV3-IP and OVCAR10 cells did not alter suspended cell growth upon PF-271 addition. CONCLUSIONS: Differential responsiveness to FAK inhibitor treatment was observed. Intrinsic low merlin protein level correlated with PF-271-mediated anchorage-independent growth inhibition, but reduction in merlin expression did not induce sensitivity to FAK inhibition. Merlin levels may be useful for patient stratification in FAK inhibitor trials.
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Cistadenocarcinoma Seroso/tratamiento farmacológico , Quinasa 1 de Adhesión Focal/antagonistas & inhibidores , Neurofibromina 2/metabolismo , Neoplasias Ováricas/tratamiento farmacológico , Inhibidores de Proteínas Quinasas/farmacología , Animales , Biomarcadores de Tumor/metabolismo , Línea Celular Tumoral , Cistadenocarcinoma Seroso/enzimología , Cistadenocarcinoma Seroso/metabolismo , Femenino , Quinasa 1 de Adhesión Focal/metabolismo , Técnicas de Silenciamiento del Gen , Humanos , Ratones , Neurofibromina 2/genética , Neoplasias Ováricas/enzimología , Neoplasias Ováricas/metabolismoRESUMEN
Pharmacological focal adhesion kinase (FAK) inhibition prevents tumor growth and metastasis, via actions on both tumor and stromal cells. In this paper, we show that vascular endothelial cadherin (VEC) tyrosine (Y) 658 is a target of FAK in tumor-associated endothelial cells (ECs). Conditional kinase-dead FAK knockin within ECs inhibited recombinant vascular endothelial growth factor (VEGF-A) and tumor-induced VEC-Y658 phosphorylation in vivo. Adherence of VEGF-expressing tumor cells to ECs triggered FAK-dependent VEC-Y658 phosphorylation. Both FAK inhibition and VEC-Y658F mutation within ECs prevented VEGF-initiated paracellular permeability and tumor cell transmigration across EC barriers. In mice, EC FAK inhibition prevented VEGF-dependent tumor cell extravasation and melanoma dermal to lung metastasis without affecting primary tumor growth. As pharmacological c-Src or FAK inhibition prevents VEGF-stimulated c-Src and FAK translocation to EC adherens junctions, but FAK inhibition does not alter c-Src activation, our experiments identify EC FAK as a key intermediate between c-Src and the regulation of EC barrier function controlling tumor metastasis.
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Antígenos CD/metabolismo , Cadherinas/metabolismo , Proteína-Tirosina Quinasas de Adhesión Focal/fisiología , Animales , Antígenos CD/fisiología , Cadherinas/fisiología , Movimiento Celular , Proteína-Tirosina Quinasas de Adhesión Focal/metabolismo , Células HEK293 , Humanos , Ratones , Ratones Endogámicos BALB C , Metástasis de la Neoplasia , Fosforilación , Proteínas Proto-Oncogénicas pp60(c-src)/metabolismo , Transducción de Señal , Factor A de Crecimiento Endotelial Vascular/metabolismo , Factor A de Crecimiento Endotelial Vascular/fisiología , Receptor 2 de Factores de Crecimiento Endotelial Vascular/metabolismoRESUMEN
Rgnef (also known as p190RhoGEF or ARHGEF28) is a Rho guanine-nucleotide-exchange factor (GEF) that binds focal adhesion kinase (FAK). FAK is recruited to adhesions and activated by integrin receptors binding to matrix proteins, such as fibronectin (FN). Canonical models place Rgnef downstream of integrin-FAK signaling in regulating Rho GTPase activity and cell movement. Herein, we establish a new, upstream role for Rgnef in enhancing FAK localization to early peripheral adhesions and promoting FAK activation upon FN binding. Rgnef-null mouse embryo fibroblasts (MEFs) exhibit defects in adhesion formation, levels of FAK phosphotyrosine (pY)-397 and FAK localization to peripheral adhesions upon re-plating on FN. Rgnef re-expression rescues these defects, but requires Rgnef-FAK binding. A mutation in the Rgnef pleckstrin homology (PH) domain inhibits adhesion formation, FAK localization, and FAK-Y397 and paxillin-Y118 phosphorylation without disrupting the Rgnef-FAK interaction. A GEF-inactive Rgnef mutant rescues FAK-Y397 phosphorylation and early adhesion localization, but not paxillin-Y118 phosphorylation. This suggests that, downstream of FN binding, paxillin-pY118 requires Rgnef GEF activity through a mechanism distinct from adhesion formation and FAK activation. These results support a scaffolding role for Rgnef in FAK localization and activation at early adhesions in a PH-domain-dependent but GEF-activity-independent manner.
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Proteína-Tirosina Quinasas de Adhesión Focal/metabolismo , Integrina beta1/metabolismo , ras-GRF1/metabolismo , Secuencia de Aminoácidos , Animales , Adhesión Celular , Células Cultivadas , Activación Enzimática , Fibroblastos/citología , Fibroblastos/enzimología , Fibroblastos/metabolismo , Fibronectinas/metabolismo , Proteína-Tirosina Quinasas de Adhesión Focal/química , Proteína-Tirosina Quinasas de Adhesión Focal/genética , Integrina beta1/genética , Ratones , Ratones Noqueados , Datos de Secuencia Molecular , Paxillin/genética , Paxillin/metabolismo , Fosforilación , Unión Proteica , Estructura Terciaria de Proteína , Alineación de Secuencia , Transducción de Señal , ras-GRF1/química , ras-GRF1/genéticaRESUMEN
Recurrence and spread of ovarian cancer is the 5th leading cause of death for women in the United States. Focal adhesion kinase (FAK) is a cytoplasmic protein-tyrosine kinase located on chromosome 8q24.3 (gene is Ptk2), a site commonly amplified in serous ovarian cancer. Elevated FAK mRNA levels in serous ovarian carcinoma are associated with decreased (logrank P = 0.0007, hazard ratio 1.43) patient overall survival, but how FAK functions in tumor progression remains undefined. We have isolated aggressive ovarian carcinoma cells termed ID8-IP after intraperitoneal (IP) growth of murine ID8 cells in C57Bl6 mice. Upon orthotopic implantation within the peri-ovarian bursa space, ID8-IP cells exhibit greater tumor growth, local and distant metastasis, and elevated numbers of ascites-associated cells compared to parental ID8 cells. ID8-IP cells exhibit enhanced growth under non-adherent conditions with elevated FAK and c-Src tyrosine kinase activation compared to parental ID8 cells. In vitro, the small molecule FAK inhibitor (Pfizer, PF562,271, PF-271) at 0.1 uM selectively prevented anchorage-independent ID8-IP cell growth with the inhibition of FAK tyrosine (Y)397 but not c-Src Y416 phosphorylation. Oral PF-271 administration (30 mg/kg, twice daily) blocked FAK but not c-Src tyrosine phosphorylation in ID8-IP tumors. This was associated with decreased tumor size, prevention of peritoneal metastasis, reduced tumor-associated endothelial cell number, and increased tumor cell-associated apoptosis. FAK knockdown and re-expression assays showed that FAK activity selectively promoted anchorage-independent ID8-IP cell survival. These results support the continued evaluation of FAK inhibitors as a promising clinical treatment for ovarian cancer.
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División Celular/efectos de los fármacos , Progresión de la Enfermedad , Proteína-Tirosina Quinasas de Adhesión Focal/antagonistas & inhibidores , Neoplasias Ováricas/patología , Inhibidores de Proteínas Quinasas/farmacología , Femenino , Proteína-Tirosina Quinasas de Adhesión Focal/genética , Humanos , Neoplasias Ováricas/enzimología , ARN Mensajero/genéticaRESUMEN
Vascular cell adhesion molecule-1 (VCAM-1) plays important roles in development and inflammation. Tumor necrosis factor-α (TNF-α) and focal adhesion kinase (FAK) are key regulators of inflammatory and integrin-matrix signaling, respectively. Integrin costimulatory signals modulate inflammatory gene expression, but the important control points between these pathways remain unresolved. We report that pharmacological FAK inhibition prevented TNF-α-induced VCAM-1 expression within heart vessel-associated endothelial cells in vivo, and genetic or pharmacological FAK inhibition blocked VCAM-1 expression during development. FAK signaling facilitated TNF-α-induced, mitogen-activated protein kinase activation, and, surprisingly, FAK inhibition resulted in the loss of the GATA4 transcription factor required for TNF-α-induced VCAM-1 production. FAK inhibition also triggered FAK nuclear localization. In the nucleus, the FAK-FERM (band 4.1, ezrin, radixin, moesin homology) domain bound directly to GATA4 and enhanced its CHIP (C terminus of Hsp70-interacting protein) E3 ligase-dependent polyubiquitination and degradation. These studies reveal new developmental and anti-inflammatory roles for kinase-inhibited FAK in limiting VCAM-1 production via nuclear localization and promotion of GATA4 turnover.
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Núcleo Celular/metabolismo , Quinasa 1 de Adhesión Focal/metabolismo , Molécula 1 de Adhesión Celular Vascular/metabolismo , Transporte Activo de Núcleo Celular , Animales , Células Cultivadas , Embrión de Mamíferos/metabolismo , Activación Enzimática , Quinasa 1 de Adhesión Focal/genética , Factor de Transcripción GATA4/metabolismo , Regulación del Desarrollo de la Expresión Génica , Humanos , Ratones , Ratones Transgénicos , Factor de Necrosis Tumoral alfa/metabolismo , UbiquitinaciónRESUMEN
BACKGROUND: Cell migration is a highly regulated process that involves the formation and turnover of cell-matrix contact sites termed focal adhesions. Rho-family GTPases are molecular switches that regulate actin and focal adhesion dynamics in cells. Guanine nucleotide exchange factors (GEFs) activate Rho-family GTPases. Rgnef (p190RhoGEF) is a ubiquitous 190 kDa GEF implicated in the control of colon carcinoma and fibroblast cell motility. PRINCIPAL FINDINGS: Rgnef exon 24 floxed mice (Rgnef(flox)) were created and crossed with cytomegalovirus (CMV)-driven Cre recombinase transgenic mice to inactivate Rgnef expression in all tissues during early development. Heterozygous Rgnef(WT/flox) (Cre+) crosses yielded normal Mendelian ratios at embryonic day 13.5, but Rgnef(flox/flox) (Cre+) mice numbers at 3 weeks of age were significantly less than expected. Rgnef(flox/flox) (Cre+) (Rgnef-/-) embryos and primary mouse embryo fibroblasts (MEFs) were isolated and verified to lack Rgnef protein expression. When compared to wildtype (WT) littermate MEFs, loss of Rgnef significantly inhibited haptotaxis migration, wound closure motility, focal adhesion number, and RhoA GTPase activation after fibronectin-integrin stimulation. In WT MEFs, Rgnef activation occurs within 60 minutes upon fibronectin plating of cells associated with RhoA activation. Rgnef-/- MEF phenotypes were rescued by epitope-tagged Rgnef re-expression. CONCLUSIONS: Rgnef-/- MEF phenotypes were due to Rgnef loss and support an essential role for Rgnef in RhoA regulation downstream of integrins in control of cell migration.
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Movimiento Celular/fisiología , Adhesiones Focales/metabolismo , Factores de Intercambio de Guanina Nucleótido/genética , Cicatrización de Heridas/fisiología , ras-GRF1/genética , Proteína de Unión al GTP rhoA/metabolismo , Análisis de Varianza , Animales , Cartilla de ADN/genética , Embrión de Mamíferos/citología , Fibroblastos/metabolismo , Fibronectinas/metabolismo , Técnica del Anticuerpo Fluorescente , Genotipo , Factores de Intercambio de Guanina Nucleótido/metabolismo , Inmunohistoquímica , Integrinas/genética , Integrinas/metabolismo , Ratones , Ratones Noqueados , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , ras-GRF1/metabolismoRESUMEN
Mycetoma, also commonly referred to as Madura foot, is statistically rare in the United States. However, it is endemic to other parts of the world. It is a pseudotumor characterized by a triad of tumefaction, draining sinuses, and grains. Two types exist, with each caused by different groups of organisms that require different treatment approaches. Therefore, the exact diagnosis and culture of the organism is vital to successful treatment outcomes. Synovial sarcoma, in contrast, is a malignancy much more commonly seen in the United States. It is characterized by a well-circumscribed, often palpable, mass that is usually well delineated on magnetic resonance imaging. It has characteristic histologic and genetic features that help distinguish it from other soft tissue masses. We present a case of a soft tissue mass diagnosed in the United States. The patient had several clinical and radiographic features of synovial sarcoma but the histologic outcome was mycetoma. The case is followed by a review of the published data.
Asunto(s)
Dermatosis del Pie/microbiología , Micetoma/diagnóstico , Antifúngicos/uso terapéutico , Femenino , Dermatosis del Pie/diagnóstico , Dermatosis del Pie/terapia , Humanos , Itraconazol/uso terapéutico , Imagen por Resonancia Magnética , Malasia/etnología , Persona de Mediana Edad , Micetoma/terapia , Sarcoma Sinovial/diagnóstico , Neoplasias de los Tejidos Blandos/diagnósticoRESUMEN
Focal adhesion kinase (FAK) functions downstream of integrins and growth factor receptors to promote tumor cell motility and invasion. In colorectal cancer, FAK is activated by amidated gastrin, a protumorigenic hormone. However, it is unclear how FAK receives signals from the gastrin receptor or other G-protein-coupled receptors that can promote cell motility and invasion. The Rho guanine-nucleotide exchange factor p190RhoGEF (Rgnef) binds FAK and facilitates fibroblast focal adhesion formation on fibronectin. Here we report that Rgnef mRNA and protein expression are significantly increased during colorectal tumor progression. In human colon carcinoma cells, Rgnef forms a complex with FAK and upon gastrin stimulation, FAK translocates to newly-forming focal adhesions where it facilitates tyrosine phosphorylation of paxillin. short hairpin (shRNA)-mediated knockdown of Rgnef or FAK, or pharmacological inhibition of FAK activity, is sufficient to block gastrin-stimulated paxillin phosphorylation, cell motility, and invadopodia formation in a manner dependent upon upstream cholecystokinin-2 receptor expression. Overexpression of the C-terminal region of Rgnef (Rgnef-C, amino acid 1,279-1,582) but not Rgnef-CΔFAK (amino acid 1,302-1,582 lacking the FAK binding site) disrupted endogenous Rgnef-FAK interaction and prevented paxillin phosphorylation and cell motility stimulated by gastrin. Rgnef-C-expressing cells formed smaller, less invasive tumors with reduced tyrosine phosphorylation of paxillin upon orthotopic implantation, compared with Rgnef-CΔFAK-expressing cells. Our studies identify Rgnef as a novel regulator of colon carcinoma motility and invasion, and they show that a Rgnef-FAK linkage promotes colon carcinoma progression in vivo.
Asunto(s)
Neoplasias Colorrectales/metabolismo , Quinasa 1 de Adhesión Focal/metabolismo , Factores de Intercambio de Guanina Nucleótido/metabolismo , Secuencia de Aminoácidos , Animales , Células CACO-2 , Movimiento Celular/fisiología , Neoplasias Colorrectales/genética , Neoplasias Colorrectales/patología , Progresión de la Enfermedad , Activación Enzimática , Matriz Extracelular/metabolismo , Femenino , Quinasa 1 de Adhesión Focal/antagonistas & inhibidores , Gastrinas/metabolismo , Técnicas de Silenciamiento del Gen , Factores de Intercambio de Guanina Nucleótido/antagonistas & inhibidores , Factores de Intercambio de Guanina Nucleótido/biosíntesis , Factores de Intercambio de Guanina Nucleótido/genética , Células HCT116 , Humanos , Ratones , Ratones Desnudos , Datos de Secuencia Molecular , Paxillin/metabolismo , Fosforilación , Transducción de SeñalRESUMEN
The hypothalamus, pituitary, and gonads coordinate to direct the development and regulation of reproductive function in mammals. Control of the hypothalamic-pituitary-gonadal axis is dependent on correct migration of gonadotropin-releasing hormone (GnRH) neurons from the nasal placode to the hypothalamus, followed by proper synthesis and pulsatile secretion of GnRH, functions absent in patients with hypogonadal hypogonadism. In this study, we identify sine oculis-related homeobox 6 (Six6) as a novel factor necessary for proper targeting of GnRH expression to the limited population of GnRH neurons within the adult mouse hypothalamus and demonstrate that it is required for proper reproductive function in both male and female mice. Female Six6-null mice exhibit a striking decrease in fertility, failing to progress through the estrous cycle normally, show any signs of successful ovulation, or produce litters. Although basal gonadotropin production in these mice is relatively normal, analysis of GnRH expression reveals a dramatic decrease in total GnRH neuron numbers. We show that expression of Six6 is dramatically increased during GnRH neuronal maturation and that overexpression of Six6 induces GnRH transcription in neuronal cells. Finally, we demonstrate that this induction in GnRH expression is mediated via binding of Six6 to evolutionarily conserved ATTA sites located within the GnRH proximal promoter. Together, these data indicate that Six6 plays an important role in the regulation of GnRH expression and hypothalamic control of fertility.