RESUMEN
The major populations at risk for developing pressure ulcers are older adults who have multiple risk factors that increase their vulnerability, people who are critically ill and those with spinal cord injury/disease. The reported prevalence of pressure ulcers in the United States is 2.5 million. However, this estimate is derived from acute care facilities and does not include people who are living at home or in nursing facilities. Despite the implementation of hospital and facility-based preventive measures, the incidence of pressure ulcers has not decreased in decades. In addition to the burden of pain, infection and death, it is estimated that hospital-acquired pressure ulcers cost the health system $26.8 billion annually with over 50% of the cost attributed to treating Stage 3 and 4 pressure injuries. Thus, it is critical to examine the literature and develop guidelines that will improve the outcomes of this complex and costly condition. This guideline update is a compendium of the best available evidence for the treatment of Pressure Ulcers published since the last update in 2015 and includes a new section based on changing demographics entitled 'Palliative wound care for seriously ill patients with pressure ulcers'. The overall goal of the Wound Healing Society Guideline project is to present clear, concise and commercial free guidelines that clinicians can use to guide care, that researchers can use to develop studies that will improve treatment and that both clinicians and researchers can use to understand the gaps in our knowledge base.
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Úlcera por Presión , Humanos , Anciano , Úlcera por Presión/epidemiología , Úlcera por Presión/terapia , Úlcera por Presión/etiología , Cicatrización de Heridas , Factores de Riesgo , PrevalenciaRESUMEN
Dental enamel formation is coordinated by ameloblast differentiation, production of enamel matrix proteins, and crystal growth. The factors regulating ameloblast differentiation are not fully understood. Here we show that the high mobility group N (HMGN) nucleosomal binding proteins modulate the rate of ameloblast differentiation and enamel formation. We found that HMGN1 and HMGN2 proteins are downregulated during mouse ameloblast differentiation. Genetically altered mice lacking HMGN1 and HMGN2 proteins show faster ameloblast differentiation and a higher rate of enamel deposition in mice molars and incisors. In vitro differentiation of induced pluripotent stem cells to dental epithelium cells showed that HMGN proteins modulate the expression and chromatin accessibility of ameloblast-specific genes and affect the binding of transcription factors epiprofin and PITX2 to ameloblast-specific genes. Our results suggest that HMGN proteins regulate ameloblast differentiation and enamel mineralization by modulating lineage-specific chromatin accessibility and transcription factor binding to ameloblast regulatory sites.
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Proteínas del Esmalte Dental , Proteína HMGN1 , Proteína HMGN2 , Animales , Ratones , Ameloblastos/metabolismo , Proteína HMGN2/genética , Proteína HMGN2/metabolismo , Proteína HMGN1/genética , Proteína HMGN1/metabolismo , Epigénesis Genética , Diferenciación Celular/genética , Proteínas HMGN/genética , Proteínas HMGN/metabolismo , Factores de Transcripción/metabolismo , Proteínas del Esmalte Dental/genética , Proteínas del Esmalte Dental/metabolismo , Cromatina/metabolismo , Amelogenina/metabolismoRESUMEN
Bone sialoprotein (BSP) is an extracellular matrix (ECM) protein associated with mineralized tissues, particularly bone and cementum. BSP includes functional domains implicated in collagen binding, hydroxyapatite nucleation, and cell signaling, although its function(s) in osteoblast and osteoclast differentiation and function remain incompletely understood. Genetic ablation of BSP in Ibsp knockout (Ibsp-/-) mice results in developmental bone mineralization and remodeling defects, with alveolar bone more severely affected than the femurs and tibias of the postcranial skeleton. The role of BSP in alveolar bone healing has not been studied. We hypothesized that BSP ablation would cause defective alveolar bone healing. We employed a maxillary first molar extraction socket healing model in 42-d postnatalIbsp-/- and wild-type (WT) control mice. Tissues were collected at 0, 7, 14, 21, and 56 d postprocedure (dpp) for analysis by micro-computed tomography (microCT), histology, in situ hybridization (ISH), immunohistochemistry (IHC), and quantitative polymerase chain reaction (qPCR) array. As expected, alveolar bone healing progressed in WT mice with increasing bone volume fraction (BV/TV), bone mineral density (BMD), and tissue mineral density (TMD), transitioning from woven to mature bone from 7 to 56 dpp. Ibsp messenger RNA (mRNA) and BSP protein were strongly expressed during alveolar bone healing in parallel with other osteogenic markers. Compared to WT, Ibsp-/- mice exhibited 50% to 70% reduced BV/TV and BMD at all time points, 7% reduced TMD at 21 dpp, abnormally increased Col1a1 and Alpl mRNA expression, and persistent presence of woven bone and increased bone marrow in healing sockets. qPCR revealed substantially dysregulated gene expression in alveolar bone of Ibsp-/- versus WT mice, with significantly disrupted expression of 45% of tested genes in functional groups, including markers for osteoblasts, osteoclasts, mineralization, ECM, cell signaling, and inflammation. We conclude that BSP is a critical and nonredundant factor for alveolar bone healing, and its absence disrupts multiple major pathways involved in appropriate healing.
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Cemento Dental , Osteopontina , Animales , Ratones , Sialoproteína de Unión a Integrina/genética , Osteopontina/metabolismo , Microtomografía por Rayos X , Cemento Dental/metabolismo , ARN Mensajero , Sialoglicoproteínas/metabolismoAsunto(s)
Protocolos de Quimioterapia Combinada Antineoplásica/efectos adversos , Inhibidores de Puntos de Control Inmunológico/efectos adversos , Ipilimumab/efectos adversos , Melanoma/tratamiento farmacológico , Nivolumab/efectos adversos , Síndrome Uveomeningoencefálico/inducido químicamente , Femenino , Humanos , Persona de Mediana Edad , Síndrome Uveomeningoencefálico/patologíaRESUMEN
BACKGROUND: Newer flow diverters are enhanced with antithrombogenic surface modifications like the Pipeline Embolization Device with Shield Technology and the Derivo Embolization Device and are purported to facilitate deployment and reduce ischemic events. PURPOSE: Our aim was to review the safety and efficacy of surface-modified flow diverters in treating patients with cerebral aneurysms. DATA SOURCES: We used Preferred Reporting Items for Systematic Reviews and Meta-Analyses-compliant systematic review and meta-analysis covering 3 major data bases and gray literature between 2014 and 2019. STUDY SELECTION: Two reviewers independently reviewed human studies of surface-modified flow diverters for eligibility based on predetermined criteria. DATA ANALYSIS: The random effects model and Freeman-Tukey arcsine transformation were used to pool efficacy outcomes (technical success, aneurysm occlusion at 6 and 12 months) and safety outcomes (mortality, morbidity, all ischemia, and serious ischemia). Subgroup analysis was performed to compare outcomes between 2 different flow diverters. DATA SYNTHESIS: Eight single-arm case series involving 911 patients and 1060 aneurysms were included. The median follow-up was 8.24 months. Pooled estimate for technical success was 99.6%, while the aneurysm occlusion at 6 and 12 months were 80.5%, and 85.6%, respectively. Pooled estimates for mortality, morbidity, total ischemia, and serious ischemia rates were 0.7%, 6.0%, 6.7%, and 1.8%, respectively. Most studies were of good quality, and no significant heterogeneity was observed. LIMITATIONS: Limitations include a retrospective, observational design in some studies; heterogeneous and underreported antiplatelet therapy; and potential performance and ecologic bias. CONCLUSIONS: Early-to-midterm safety and efficacy for surface-modified flow diverters appear comparable with older devices, especially for small, unruptured anterior circulation aneurysms. Long-term clinical data are required to further corroborate these results.
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Prótesis Vascular , Embolización Terapéutica/instrumentación , Procedimientos Endovasculares/instrumentación , Aneurisma Intracraneal/terapia , Adulto , Anciano , Prótesis Vascular/efectos adversos , Embolización Terapéutica/métodos , Procedimientos Endovasculares/métodos , Femenino , Humanos , Masculino , Persona de Mediana Edad , Estudios Retrospectivos , Resultado del TratamientoRESUMEN
Biomineralization is regulated by inorganic pyrophosphate (PPi), a potent physiological inhibitor of hydroxyapatite crystal growth. Progressive ankylosis protein (ANK) and ectonucleotide pyrophosphatase/phosphodiesterase 1 (ENPP1) act to increase local extracellular levels of PPi, inhibiting mineralization. The periodontal complex includes 2 mineralized tissues, cementum and alveolar bone (AB), both essential for tooth attachment. Previous studies demonstrated that loss of function of ANK or ENPP1 (reducing PPi) resulted in increased cementum formation, suggesting PPi metabolism may be a target for periodontal regenerative therapies. To compare the effects of genetic ablation of Ank, Enpp1, and both factors concurrently on cementum and AB regeneration, mandibular fenestration defects were created in Ank knockout (Ank KO), Enpp1 mutant (Enpp1asj/asj), and double KO (dKO) mice. Genetic ablation of Ank, Enpp1, or both factors increased cementum regeneration compared to controls at postoperative days (PODs) 15 and 30 (Ank KO: 8-fold, 3-fold; Enpp1asj/asj: 7-fold, 3-fold; dKO: 11-fold, 4-fold, respectively) associated with increased fluorochrome labeling and expression of mineralized tissue markers, dentin matrix protein 1 (Dmp1/DMP1), osteopontin (Spp1/OPN), and bone sialoprotein (Ibsp/BSP). Furthermore, dKO mice featured increased cementum thickness compared to single KOs at POD15 and Ank KO at POD30. No differences were noted in AB volume between genotypes, but osteoblast/osteocyte markers were increased in all KOs, partially mineralized osteoid volume was increased in dKO versus controls at POD15 (3-fold), and mineral density was decreased in Enpp1asj/asj and dKOs at POD30 (6% and 9%, respectively). Increased numbers of osteoclasts were present in regenerated AB of all KOs versus controls. These preclinical studies suggest PPi modulation as a potential and novel approach for cementum regeneration, particularly targeting ENPP1 and/or ANK. Differences in cementum and AB regeneration in response to reduced PPi conditions highlight the need to consider tissue-specific responses in strategies targeting regeneration of the entire periodontal complex.
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Difosfatos , Anquilosis del Diente , Diente , Animales , Huesos , Cemento Dental , Ratones , Ratones NoqueadosRESUMEN
Nasopharyngeal carcinoma (NPC) is of high prevalence in Hong Kong and southern China. The pathogenesis of NPC is closely associated with Epstein-Barr virus (EBV) infection via regulation of viral oncoprotein latent membrane protein 1 (LMP1). The conventional treatment for NPC is chemo-radiotherapy, but the prognosis remains poor for advanced stage, recurrent and metastatic NPC. Photodynamic therapy (PDT) is a therapeutic approach to combat tumors. PDT effectiveness depends on the interaction of photosensitizers, light and molecular oxygen. 5- aminolevulinic acid hexyl derivative (H-ALA) is one of the photosensitizers derived from 5-ALA. H-ALA with improved lipophilic properties by adding a long lipophilic chain (hexyl group) to 5-ALA, resulted in better penetration into cell cytoplasm. In this study, the effect of H-ALA-PDT on NPC cells (EBV positive C666-1 and EBV negative CNE2) was investigated. The H-ALA mediated cellular uptake and cytotoxicity was revealed via flow cytometry analysis and MTT assay respectively. H-ALA PDT mediated protein modulation was analysed by western blot analysis. Our finding reported that the cellular uptake of H-ALA in C666-1 and CNE2 cells was in a time dependent manner. H-ALA PDT was effective to C666-1 and CNE2 cells. EBV LMP1 proteins was expressed in C666-1 cells only and its expression was responsive to H-ALA PDT in a dose dependent manner. This work revealed the potential of H-ALA PDT as a treatment regiment for EBV positive NPC cells. Understanding the mechanism of H-ALA mediated PDT could develop improved strategies for the treatment of NPC.
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Ácido Aminolevulínico/análogos & derivados , Terapia por Luz de Baja Intensidad/métodos , Neoplasias Nasofaríngeas/tratamiento farmacológico , Fotoquimioterapia/métodos , Fármacos Fotosensibilizantes/farmacología , Proteínas de la Matriz Viral/efectos de los fármacos , Ácido Aminolevulínico/farmacocinética , Ácido Aminolevulínico/farmacología , Línea Celular Tumoral , Supervivencia Celular , Herpesvirus Humano 4/efectos de los fármacos , Humanos , Láseres de Semiconductores/uso terapéutico , Fármacos Fotosensibilizantes/farmacocinética , Protoporfirinas/farmacocinéticaRESUMEN
Pyrophosphate (PPi) serves as a potent and physiologically important regulator of mineralization, with systemic and local concentrations determined by several key regulators, including: tissue-nonspecific alkaline phosphatase (ALPL gene; TNAP protein), the progressive ankylosis protein (ANKH; ANK), and ectonucleotide pyrophosphatase/phosphodiesterase 1 (ENPP1; ENPP1). Results to date have indicated important roles for PPi in cementum formation, and we addressed several gaps in knowledge by employing genetically edited mouse models where PPi metabolism was disrupted and pharmacologically modulating PPi in a PPi-deficient mouse model. We demonstrate that acellular cementum growth is inversely proportional to PPi levels, with reduced cementum in Alpl KO (increased PPi levels) mice and excess cementum in Ank KO mice (decreased PPi levels). Moreover, simultaneous ablation of Alpl and Ank results in reestablishment of functional cementum in dKO mice. Additional reduction of PPi by dual deletion of Ank and Enpp1 does not further increase cementogenesis, and PDL space is maintained in part through bone modeling/remodeling by osteoclasts. Our results provide insights into cementum formation and expand our knowledge of how PPi regulates cementum. We also demonstrate for the first time that pharmacologic manipulation of PPi through an ENPP1-Fc fusion protein can regulate cementum growth, supporting therapeutic interventions targeting PPi metabolism.
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Cementogénesis , Difosfatos , Animales , Cemento Dental , Ratones , OsteoclastosRESUMEN
Mutations in PHEX cause X-linked hypophosphatemia (XLH), a form of hypophosphatemic rickets. Hyp (Phex mutant) mice recapitulate the XLH phenotype. Dental disorders are prevalent in individuals with XLH; however, underlying dentoalveolar defects remain incompletely understood. We analyzed Hyp mouse dentoalveolar defects at 42 and 90 d postnatal to comparatively define effects of XLH on dental formation and function. Phex mRNA was expressed by odontoblasts (dentin), osteocytes (bone), and cementocytes (cellular cementum) in wild-type (WT) mice. Enamel density was unaffected, though enamel volume was significantly reduced in Hyp mice. Dentin defects in Hyp molars were indicated histologically by wide predentin, thin dentin, and extensive interglobular dentin, confirming micro-computed tomography (micro-CT) findings of reduced dentin volume and density. Acellular cementum was thin and showed periodontal ligament detachment. Mechanical testing indicated dramatically altered periodontal mechanical properties in Hyp versus WT mice. Hyp mandibles demonstrated expanded alveolar bone with accumulation of osteoid, and micro-CT confirmed decreased bone volume fraction and alveolar bone density. Cellular cementum area was significantly increased in Hyp versus WT molars owing to accumulation of hypomineralized cementoid. Histology, scanning electron microscopy, and nanoindentation revealed hypomineralized "halos" surrounding Hyp cementocyte and osteocyte lacunae. Three-dimensional micro-CT analyses confirmed larger cementocyte/osteocyte lacunae and significantly reduced perilacunar mineral density. While long bone and alveolar bone osteocytes in Hyp mice overexpressed fibroblast growth factor 23 (Fgf23), its expression in molars was much lower, with cementocyte Fgf23 expression particularly low. Expression and distribution of other selected markers were disturbed in Hyp versus WT long bone, alveolar bone, and cementum, including osteocyte/cementocyte marker dentin matrix protein 1 (Dmp1). This study reports for the first time a quantitative analysis of the Hyp mouse dentoalveolar phenotype, including all mineralized tissues. Novel insights into cellular cementum provide evidence for a role for cementocytes in perilacunar mineralization and cementum biology.
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Raquitismo Hipofosfatémico Familiar , Hipofosfatemia , Animales , Calcificación Fisiológica , Raquitismo Hipofosfatémico Familiar/diagnóstico por imagen , Raquitismo Hipofosfatémico Familiar/genética , Femenino , Factor-23 de Crecimiento de Fibroblastos , Masculino , Ratones , Ratones Endogámicos BALB C , Endopeptidasa Neutra Reguladora de Fosfato PHEX/genética , Microtomografía por Rayos XRESUMEN
Previous studies revealed that cementum formation is tightly regulated by inorganic pyrophosphate (PPi), a mineralization inhibitor. Local PPi concentrations are determined by regulators, including ectonucleotide pyrophosphatase/phosphodiesterase 1 (ENPP1), which increases PPi concentrations by adenosine triphosphate hydrolysis. Orthodontic forces stimulate alveolar bone remodelling, leading to orthodontic tooth movement (OTM). To better understand how disturbed mineral metabolism and the resulting altered periodontal structures affect OTM, we employed Enpp1 mutant mice that feature reduced PPi and increased cervical cementum in a model of OTM induced by a stretched closed-coil spring ligated between the maxillary left first molar and maxillary incisors. We analyzed tooth movement, osteoclast/odontoclast response, and tooth root resorption by micro-computed tomography, histology, histomorphometry, and immunohistochemistry. Preoperatively, we noted an altered periodontium in Enpp1 mutant mice, with significantly increased periodontal ligament (PDL) volume and thickness, as well as increased PDL-bone/tooth root surface area, compared to wild-type (WT) controls. After 11 d of orthodontic treatment, Enpp1 mutant mice displayed 38% reduced tooth movement versus WT mice. Molar roots in Enpp1 mutant mice exhibited less change in PDL width in compression and tension zones compared to WT mice. Root resorption was noted in both groups with no difference in average depths, but resorption lacunae in Enpp1 mutant mice were almost entirely limited to cementum, with 150% increased cementum resorption and 92% decreased dentin resorption. Osteoclast/odontoclast cells were reduced by 64% in Enpp1 mutant mice, with a predominance of tartrate-resistant acid phosphatase (TRAP)-positive cells on root surfaces, compared to WT mice. Increased numbers of TRAP-positive cells on root surfaces were associated with robust immunolocalization of osteopontin (OPN) and receptor-activator of NF-κB ligand (RANKL). Collectively, reduced response to orthodontic forces, decreased tooth movement, and altered osteoclast/odontoclast distribution suggests Enpp1 loss of function has direct effects on clastic function/recruitment and/or indirect effects on periodontal remodeling via altered periodontal structure or tissue mineralization.
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Hipercementosis/fisiopatología , Técnicas de Movimiento Dental/métodos , Animales , Cemento Dental/metabolismo , Inmunohistoquímica , Ratones , Osteoclastos/metabolismo , Ligamento Periodontal/fisiopatología , Hidrolasas Diéster Fosfóricas , Pirofosfatasas , Resorción Radicular/diagnóstico por imagen , Resorción Radicular/fisiopatología , Fosfatasa Ácida Tartratorresistente/metabolismo , Microtomografía por Rayos XRESUMEN
The periodontal complex is essential for tooth attachment and function and includes the mineralized tissues, cementum and alveolar bone, separated by the unmineralized periodontal ligament (PDL). To gain insights into factors regulating cementum-PDL and bone-PDL borders and protecting against ectopic calcification within the PDL, we employed a proteomic approach to analyze PDL tissue from progressive ankylosis knock-out (Ank-/-) mice, featuring reduced PPi, rapid cementogenesis, and excessive acellular cementum. Using this approach, we identified the matrix protein osteopontin (Spp1/OPN) as an elevated factor of interest in Ank-/- mouse molar PDL. We studied the role of OPN in dental and periodontal development and function. During tooth development in wild-type (WT) mice, Spp1 mRNA was transiently expressed by cementoblasts and strongly by alveolar bone osteoblasts. Developmental analysis from 14 to 240days postnatal (dpn) indicated normal histological structures in Spp1-/- comparable to WT control mice. Microcomputed tomography (micro-CT) analysis at 30 and 90dpn revealed significantly increased volumes and tissue mineral densities of Spp1-/- mouse dentin and alveolar bone, while pulp and PDL volumes were decreased and tissue densities were increased. However, acellular cementum growth was unaltered in Spp1-/- mice. Quantitative PCR of periodontal-derived mRNA failed to identify potential local compensators influencing cementum in Spp1-/- vs. WT mice at 26dpn. We genetically deleted Spp1 on the Ank-/- mouse background to determine whether increased Spp1/OPN was regulating periodontal tissues when the PDL space is challenged by hypercementosis in Ank-/- mice. Ank-/-; Spp1-/- double deficient mice did not exhibit greater hypercementosis than that in Ank-/- mice. Based on these data, we conclude that OPN has a non-redundant role regulating formation and mineralization of dentin and bone, influences tissue properties of PDL and pulp, but does not control acellular cementum apposition. These findings may inform therapies targeted at controlling soft tissue calcification.
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Proceso Alveolar/fisiología , Calcificación Fisiológica/fisiología , Dentina/metabolismo , Osteogénesis/fisiología , Osteopontina/metabolismo , Animales , Cementogénesis/fisiología , Femenino , Masculino , Ratones , Ratones Noqueados , Ligamento Periodontal/fisiologíaRESUMEN
Mineralization of bones and teeth is tightly regulated by levels of extracellular inorganic phosphate (Pi) and pyrophosphate (PPi). Three regulators that control pericellular concentrations of Pi and PPi include tissue-nonspecific alkaline phosphatase (TNAP), progressive ankylosis protein (ANK), and ectonucleotide pyrophosphatase/phosphodiesterase 1 (ENPP1). Inactivation of these factors results in mineralization disorders affecting teeth and their supporting structures. This study for the first time analyzed the effect of decreased PPi on dental development in individuals with generalized arterial calcification of infancy (GACI) due to loss-of-function mutations in the ENPP1 gene. Four of the 5 subjects reported a history of infraocclusion, overretained primary teeth, ankylosis, and/or slow orthodontic tooth movement, suggesting altered mineral metabolism contributing to disrupted tooth movement and exfoliation. All subjects had radiographic evidence of unusually protruding cervical root morphology in primary and/or secondary dentitions. High-resolution micro-computed tomography (micro-CT) analyses of extracted primary teeth from 3 GACI subjects revealed 4-fold increased cervical cementum thickness ( P = 0.00007) and a 23% increase in cementum density ( P = 0.009) compared to age-matched healthy control teeth. There were no differences in enamel and dentin densities between GACI and control teeth. Histology revealed dramatically expanded cervical cementum in GACI teeth, including cementocyte-like cells and unusual patterns of cementum resorption and repair. Micro-CT analysis of Enpp1 mutant mouse molars revealed 4-fold increased acellular cementum thickness ( P = 0.002) and 5-fold increased cementum volume ( P = 0.002), with no changes in enamel or dentin. Immunohistochemistry identified elevated ENPP1 expression in cementoblasts of human and mouse control teeth. Collectively, these findings reveal a novel dental phenotype in GACI and identify ENPP1 genetic mutations associated with hypercementosis. The sensitivity of cementum to reduced PPi levels in both human and mouse teeth establishes this as a well-conserved and fundamental biological process directing cementogenesis across species (ClinicalTrials.gov NCT00369421).
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Hipercementosis/diagnóstico por imagen , Hipercementosis/genética , Mutación con Pérdida de Función , Hidrolasas Diéster Fosfóricas/genética , Pirofosfatasas/genética , Calcificación Vascular/genética , Adulto , Animales , Niño , Femenino , Genotipo , Humanos , Masculino , Ratones , Linaje , Radiografía Panorámica , Diente Primario , Microtomografía por Rayos XRESUMEN
Although acellular cementum is essential for tooth attachment, factors directing its development and regeneration remain poorly understood. Inorganic pyrophosphate (PPi), a mineralization inhibitor, is a key regulator of cementum formation: tissue-nonspecific alkaline phosphatase (Alpl/TNAP) null mice (increased PPi) feature deficient cementum, while progressive ankylosis protein (Ank/ANK) null mice (decreased PPi) feature increased cementum. Bone sialoprotein (Bsp/BSP) and osteopontin (Spp1/OPN) are multifunctional extracellular matrix components of cementum proposed to have direct and indirect effects on cell activities and mineralization. Studies on dentoalveolar development of Bsp knockout (Bsp-/-) mice revealed severely reduced acellular cementum, however underlying mechanisms remain unclear. The similarity in defective cementum phenotypes between Bsp-/- mice and Alpl-/- mice (the latter featuring elevated PPi and OPN), prompted us to examine whether BSP is operating by modulating PPi-associated genes. Genetic ablation of Bsp caused a 2-fold increase in circulating PPi, altered mRNA expression of Alpl, Spp1, and Ank, and increased OPN protein in the periodontia. Generation of a Bsp knock-out (KO) cementoblast cell line revealed significantly decreased mineralization capacity, 50% increased PPi in culture media, and increased Spp1 and Ank mRNA expression. While addition of 2µg/ml recombinant BSP altered Spp1, Ank, and Enpp1 expression in cementoblasts, changes resulting from this dose were not dependent on the integrin-binding RGD motif or MAPK/ERK signaling pathway. Decreasing PPi by genetic ablation of Ank on the Bsp-/- mouse background reestablished cementum formation, allowing >3-fold increased acellular cementum volume compared to wild-type (WT). However, deleting Ank did not fully compensate for the absence of BSP. Bsp-/-; Ank-/- double-deficient mice exhibited mean 20-27% reduced cementum thickness and volume compared to Ank-/- mice. From these data, we conclude that the perturbations in PPi metabolism are not solely driving the cementum pathology in Bsp-/- mice, and that PPi is more potent than BSP as a cementum regulator, as shown by the ability to override loss of BSP by lowering PPi. We propose that BSP and PPi work in concert to direct mineralization in cementum and likely other mineralized tissues.
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Calcificación Fisiológica , Cementogénesis/efectos de los fármacos , Difosfatos/farmacología , Sialoproteína de Unión a Integrina/metabolismo , Animales , Calcificación Fisiológica/efectos de los fármacos , Cemento Dental/efectos de los fármacos , Cemento Dental/metabolismo , Eliminación de Gen , Regulación de la Expresión Génica/efectos de los fármacos , Sialoproteína de Unión a Integrina/deficiencia , Ratones Noqueados , Periodoncio/metabolismo , Fenotipo , Proteínas de Transporte de Fosfato/metabolismo , Fosforilación/efectos de los fármacosRESUMEN
Mutations and common polymorphisms in interferon regulatory factor 6 ( IRF6) are associated with both syndromic and nonsyndromic forms of cleft lip/palate (CLP). To date, much of the focus on this transcription factor has been on identifying its direct targets and the gene regulatory network in which it operates. Notably, however, IRF6 is found predominantly in the cytoplasm, with its import into the nucleus tightly regulated like other members of the IRF family. To provide further insight into the role of IRF6 in the pathogenesis of CLP, we sought to identify direct IRF6 protein interactors using a combination of yeast 2-hybrid screens and co-immunoprecipitation assays. Using this approach, we identified NME1 and NME2, well-known regulators of Rho-type GTPases, E-cadherin endocytosis, and epithelial junctional remodeling, as bona fide IRF6 partner proteins. The NME proteins co-localize with IRF6 in the cytoplasm of primary palatal epithelial cells in vivo, and their interaction with IRF6 is significantly enhanced by phosphorylation of key serine residues in the IRF6 C-terminus. Furthermore, CLP associated IRF6 missense mutations disrupt the ability of IRF6 to bind the NME proteins and result in elevated activation of Rac1 and RhoA, compared to wild-type IRF6, when ectopically expressed in 293T epithelial cells. Significantly, we also report the identification of 2 unique missense mutations in the NME proteins in patients with CLP (NME1 R18Q in an IRF6 and GRHL3 mutation-negative patient with van der Woude syndrome and NME2 G71V in a patient with nonsyndromic CLP). Both variants disrupted the ability of the respective proteins to interact with IRF6. The data presented suggest an important role for cytoplasmic IRF6 in regulating the availability or localization of the NME1/2 complex and thus the dynamic behavior of epithelia during lip/palate development.
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Labio Leporino/genética , Fisura del Paladar/genética , Factores Reguladores del Interferón/genética , Nucleósido Difosfato Quinasas NM23/genética , Animales , Embrión de Pollo , Variación Genética , Humanos , Inmunoprecipitación , Mutación , Fosforilación , Reacción en Cadena de la Polimerasa , Adherencias Tisulares/genética , Factores de Transcripción/genéticaRESUMEN
Cyclooxygenase (COX)-2 is upregulated in hepatocellular carcinoma (HCC). However, the direct causative effect of COX-2 in spontaneous HCC formation remains unknown. We thus investigate the role and molecular pathogenesis of COX-2 in HCC by using liver-specific COX-2 transgenic (TG) mice. We found spontaneous HCC formation with elevated inflammatory infiltrates and neovessels in male TG mice (3/21, 14.3%), but not in any of male WT mice (0/19). Reduced representation bisulfite sequencing (RRBS) and gene expression microarrays were performed in the HCC tumor and non-HCC liver tissues to investigate the molecular mechanisms of COX-2-driven HCC. By RRBS, DNA promoter hypermethylation was identified in HCC from TG mice. Induction of promoter hypermethylation was associated with reduced tet methylcytosine dioxygenase 1 (TET1) expression by COX-2. TET1 could catalyze the conversion of 5-methylcytosine into 5-hydroxymethylcytosine (5hmC) and prevents DNA hypermethylation. In keeping with this, loss of 5hmC was demonstrated in COX-2-induced HCC. Consistently, COX-2 overexpression in human HCC cell lines could reduce both TET1 expression and 5hmc levels. Integrative analyses of DNA methylation and gene expression profiles further identified significantly downregulated genes including LTBP1, ADCY5 and PRKCZ by promoter methylation in COX-2-induced HCC. Reduced expression of LTBP1, ADCY5 and PRKCZ by promoter hypermethylation was further validated in human HCCs. Bio-functional investigation revealed that LTBP1 inhibited cell proliferation in HCC cell lines, suggesting its potential role as a tumor suppressor in HCC. Gene expression microarrays revealed that signaling cascades (AKT (protein kinase B), STK33 (Serine/Threonine kinase 33) and MTOR (mechanistic target of rapamycin) pathways) were enriched in COX-2-induced HCC. In conclusion, this study demonstrated for the first time that enhanced COX-2 expression in hepatocytes is sufficient to induce HCC through inducing promoter hypermethylation by reducing TET1, silencing tumor-suppressive genes and activating key oncogenic pathways. Inhibition of COX-2 represents a mechanism-based target for HCC prevention.
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Carcinoma Hepatocelular/etiología , Ciclooxigenasa 2/fisiología , Neoplasias Hepáticas/etiología , Hígado/enzimología , Animales , Línea Celular Tumoral , Metilación de ADN , Proteínas de Unión al ADN/fisiología , Humanos , Proteínas de Unión a TGF-beta Latente/fisiología , Masculino , Ratones , Regiones Promotoras Genéticas , Proteínas Proto-Oncogénicas/fisiologíaRESUMEN
BACKGROUND: To evaluate the safety, tolerability, pharmacokinetics, and maximum tolerated dose (MTD) of copanlisib, a phosphatidylinositol 3-kinase inhibitor, in patients with advanced solid tumors or non-Hodgkin's lymphoma (NHL). PATIENTS AND METHODS: Phase I dose-escalation study including patients with advanced solid tumors or NHL, and a cohort of patients with type 2 diabetes mellitus. Patients received three weekly intravenous infusions of copanlisib per 28-day cycle over the dose range 0.1-1.2 mg/kg. Plasma copanlisib levels were analyzed for pharmacokinetics. Biomarker analysis included PIK3CA, KRAS, BRAF, and PTEN mutational status and PTEN immunohistochemistry. Whole-body [(18)F]-fluorodeoxyglucose positron emission tomography ((18)FDG-PET) was carried out at baseline and following the first dose to assess early pharmacodynamic effects. Plasma glucose and insulin levels were evaluated serially. RESULTS: Fifty-seven patients received treatment. The MTD was 0.8 mg/kg copanlisib. The most frequent treatment-related adverse events were nausea and transient hyperglycemia. Copanlisib exposure was dose-proportional with no accumulation; peak exposure positively correlated with transient hyperglycemia post-infusion. Sixteen of 20 patients treated at the MTD had reduced (18)FDG-PET uptake; 7 (33%) had a reduction >25%. One patient achieved a complete response (CR; endometrial carcinoma exhibiting both PIK3CA and PTEN mutations and complete PTEN loss) and two had a partial response (PR; both metastatic breast cancer). Among the nine NHL patients, all six with follicular lymphoma (FL) responded (one CR and five PRs) and one patient with diffuse large B-cell lymphoma had a PR by investigator assessment; two patients with FL who achieved CR (per post hoc independent radiologic review) were on treatment >3 years. CONCLUSION: Copanlisib, dosed intermittently on days 1, 8, and 15 of a 28-day cycle, was well tolerated and the MTD was determined to be 0.8 mg/kg. Copanlisib exhibited dose-proportional pharmacokinetics and promising anti-tumor activity, particularly in patients with NHL. CLINICALTRIALSGOV: NCT00962611; https://clinicaltrials.gov/ct2/show/NCT00962611.
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Fosfatidilinositol 3-Quinasa Clase I/antagonistas & inhibidores , Inhibidores Enzimáticos/administración & dosificación , Linfoma no Hodgkin/tratamiento farmacológico , Neoplasias/tratamiento farmacológico , Pirimidinas/administración & dosificación , Quinazolinas/administración & dosificación , Administración Intravenosa , Adulto , Anciano , Fosfatidilinositol 3-Quinasa Clase I/genética , Relación Dosis-Respuesta a Droga , Inhibidores Enzimáticos/efectos adversos , Inhibidores Enzimáticos/farmacocinética , Femenino , Humanos , Linfoma no Hodgkin/enzimología , Linfoma no Hodgkin/patología , Masculino , Persona de Mediana Edad , Estadificación de Neoplasias , Neoplasias/enzimología , Neoplasias/patología , Pirimidinas/efectos adversos , Pirimidinas/farmacocinética , Quinazolinas/efectos adversos , Quinazolinas/farmacocinéticaRESUMEN
Clefting of the lip, with or without palatal involvement (CLP), is associated with a higher incidence of developmental tooth abnormalities, including hypodontia and supernumerary teeth, aberrant crown and root morphologies, and enamel defects, although the underlying mechanistic link is poorly understood. As most CLP genes are expressed throughout the oral epithelium, the authors hypothesized that the expression of CLP genes may persist in the dental epithelium and thus, in addition to their earlier role in labiopalatine development, may play an important functional role in subsequent tooth patterning and amelogenesis. To address this, the authors generated a unique conditional knockout model involving the major CLP gene, Irf6, that overcomes the previously reported perinatal lethality to enable assessment of any posteruption dental phenotypes. A dental epithelium-specific Irf6 conditional knockout (Irf6-cKO) mouse was generated via a Pitx2-Cre driver line. Dental development was analyzed by microcomputed tomography, scanning electron microscopy, histology, immunohistochemistry, and quantitative polymerase chain reaction. Irf6-cKO mice displayed variable hypodontia, occasional supernumerary incisors and molars, as well as crown and root patterning anomalies, including peg-shaped first molars and taurodontic and C-shaped mandibular second molars. Enamel density was reduced in preeruption Irf6-cKO mice, and some shearing of enamel rods was noted in posteruption incisors. There was also rapid attrition of Irf6-cKO molars following eruption. Histologically, Irf6-cKO ameloblasts exhibited disturbances in adhesion and polarity, and delayed enamel formation was confirmed immunohistochemically. Altered structure of Hertwig's epithelial root sheath was also observed. These data support a role for IRF6 in tooth number, crown and root morphology and amelogenesis that is likely due to a functional role of Irf6 in organization and polarity of epithelial cell types. This data reinforce the notion that various isolated tooth defects could be considered part of the CLP spectrum in relatives of an affected individual.
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Labio Leporino/complicaciones , Labio Leporino/diagnóstico por imagen , Factores Reguladores del Interferón/genética , Anomalías Dentarias/complicaciones , Amelogénesis/genética , Animales , Labio Leporino/genética , Esmalte Dental/crecimiento & desarrollo , Modelos Animales de Enfermedad , Factores Reguladores del Interferón/fisiología , Ratones , Ratones Noqueados , Microscopía Electrónica de Rastreo , Fenotipo , Anomalías Dentarias/diagnóstico por imagen , Anomalías Dentarias/genética , Microtomografía por Rayos XRESUMEN
Epidemiological studies showed that obesity and its related non-alcoholic fatty liver disease (NAFLD) promote hepatocellular carcinoma (HCC) development. We aimed to uncover the genetic alterations of NAFLD-HCC using whole-exome sequencing. We compared HCC development in genetically obese mice and dietary obese mice with wild-type lean mice fed a normal chow after treatment with diethylnitrosamine. HCC tumor and adjacent normal samples from obese and lean mice were then subjected to whole-exome sequencing. Functional and mechanistic importance of the identified mutations in Carboxyl ester lipase (Cel) gene and Harvey rat sarcoma virus oncogene 1 (Hras) was further elucidated. We demonstrated significantly higher incidences of HCC in both genetic and dietary obese mice with NAFLD development as compared with lean mice without NAFLD. The mutational signatures of NAFLD-HCC and lean HCC were distinct, with <3% overlapped. Eight metabolic or oncogenic pathways were found to be significantly enriched by mutated genes in NAFLD-HCC, but only two of these pathways were dysregulated by mutations in lean HCC. In particular, Cel was mutated significantly more frequently in NAFLD-HCC than in lean HCC. The multiple-site mutations in Cel are loss-of-function mutations, with effects similar to Cel knock-down. Mutant Cel caused accumulation of cholesteryl ester in liver cells, which led to induction of endoplasmic reticulum stress and consequently activated the IRE1α/c-Jun N-terminal kinase (JNK)/c-Jun/activating protein-1 (AP-1) signaling cascade to promote liver cell growth. In addition, single-site mutations in Hras at codon 61 were found in NAFLD-HCC but none in lean HCC. The gain-of-function mutations in Hras (Q61R and Q61K) significantly promoted liver cell growth through activating the mitogen-activated protein kinase (MAPK) and phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3K)/3-phosphoinositide-dependent protein kinase-1 (PDK1)/Akt pathways. In conclusion, we have identified mutation signature and pathways in NAFLD-associated HCC. Mutations in Cel and Hras have important roles in NAFLD-associated hepatocellular carcinogenesis.
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Carcinoma Hepatocelular/genética , Neoplasias Hepáticas/genética , Obesidad/genética , Animales , Carcinoma Hepatocelular/complicaciones , Carcinoma Hepatocelular/patología , Línea Celular Tumoral , Modelos Animales de Enfermedad , Humanos , Neoplasias Hepáticas/complicaciones , Neoplasias Hepáticas/patología , Neoplasias Hepáticas Experimentales/complicaciones , Neoplasias Hepáticas Experimentales/genética , Neoplasias Hepáticas Experimentales/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Mutación , Obesidad/complicaciones , Obesidad/patología , Transducción de SeñalRESUMEN
An inherited germline mutation in CDKN2A is the most common cause of familial atypical multiple mole melanoma (FAMMM) syndrome. Although it is well known that CDKN2A mutations confer an increased risk for melanoma and pancreatic carcinoma, the association with an increased risk for nerve sheath tumours and other tumour types is under-recognized. We report a family with a missense mutation (c.151-1G>C) at the acceptor splice site of intron 1 of CDKN2A, resulting in loss of function of both tumour suppressor proteins p16(INK) (4) and p14(ARF) . This mutation is associated with a clinical phenotype of FAMMM syndrome in which patients develop numerous benign and malignant mutations, brain tumours, sarcomas and other solid tumours, in addition to melanoma and dysplastic naevi. Our proband initially presented with multiple nerve sheath tumours, leading to diagnostic confusion with Neurofibromatosis type 1. Loss of p14 expression results in increased MDM2-mediated degradation of the tumour suppressor protein p53, and predisposes mutation carriers to multiple benign and malignant neoplasms. This article highlights the importance of considering CDKN2A mutations in patients with dysplastic naevi, melanoma and multiple nerve sheath tumours, specifically those with histological features of both neurofibromas and schwannomas. We also present a discussion of medical management for patients with this high-risk cancer susceptibility syndrome.