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1.
Int J Nanomedicine ; 15: 3771-3790, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32547027

RESUMO

Introduction: Rapamycin has been considered as a potential treatment for osteoarthritis (OA). Drug carriers fabricated from liposomes can prolong the effects of drugs and reduce side effects of drugs. Low-intensity pulsed ultrasound (LIPUS) has been found to possess anti-OA effects. Materials and Methods: The anti-osteoarthritic effects of liposome-encapsulated rapamycin (L-rapa) combined with LIPUS were examined by culture of normal and OA chondrocytes in alginate beads and further validated in OA prone Dunkin-Hartley guinea pigs. Results: L-rapa with LIPUS largely up-regulated aggrecan and type II collagen mRNA in human OA chondrocytes (HOACs). L-rapa with LIPUS caused significant enhancement in proteoglycan and type II collagen production in HOACs. Large decreases in both MMP-13 and IL-6 proteins were found in the HOACs exposed to L-rapa with LIPUS. Intra-articular injection of 40 µL L-rapa at both 5 µM and 50 µM twice a week combined with LIPUS thrice a week for 8 weeks significantly increased GAGs and type II collagen in the cartilage of knee. Results on OARSI score showed that intra-articular injection of 5 µM L-rapa with LIPUS displayed the greatest anti-OA effects. Immunohistochemistry revealed that L-rapa with or without LIPUS predominantly reduced MMP-13 in vivo. The values of complete blood count and serum biochemical examinations remained in the normal ranges after the injections with or without LIPUS. These data indicated that intra-articular injection of L-rapa collaborated with LIPUS is not only effective against OA but a safe OA therapy. Conclusion: Taken together, L-rapa combined with LIPUS possessed the most consistently and effectively anabolic and anti-catabolic effects in HOACs and the spontaneous OA guinea pigs. This study evidently revealed that liposome-encapsulation collaborated with LIPUS is able to reduce the effective dose and administration frequency of rapamycin and further stably reinforce its therapeutic actions against OA.


Assuntos
Osteoartrite/terapia , Sirolimo/uso terapêutico , Ondas Ultrassônicas , Animais , Peso Corporal/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/efeitos da radiação , Células Cultivadas , Condrócitos/efeitos dos fármacos , Condrócitos/patologia , Condrócitos/efeitos da radiação , Colágeno Tipo II/metabolismo , Liberação Controlada de Fármacos , Cobaias , Humanos , Injeções Intra-Articulares , Interleucina-6/metabolismo , Lipossomos/ultraestrutura , Masculino , Metaloproteinase 13 da Matriz/metabolismo , Pessoa de Meia-Idade , Osteoartrite/sangue , Osteoartrite/patologia , Proteoglicanas/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Sirolimo/administração & dosagem , Sirolimo/farmacologia
2.
Life Sci ; 255: 117763, 2020 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-32389831

RESUMO

AIMS: To explored the potential of human umbilical cord mesenchymal stem cells (hUCMSCs) as seed cells for dental pulp regeneration and the possibility of cotransplantation hUCMSCs and endothelial cells (ECs) for angiogenesis and pulp regeneration in vivo. MATERIALS AND METHODS: hUCMSCs and human umbilical vein endothelial cells (HUVECs) were cocultured for matrigel angiogenesis assay in vitro and Matrigel plug assay in vivo. Next, we used the transwell coculture system to coculture hUCMSCs and HUVECs in vitro for RNA- sequencing (RNA-seq). Last, encapsulated hUCMSCs and HUVECs in scaffolds were injected into the root segments, and transplanted into immunodeficient mice for dental pulp regeneration. KEY FINDINGS: In vitro Matrigel angiogenesis assay and in vivo Matrigel plug assay indicated that cocultured hUCMSCs and HUVECs promote vascular formation of HUVECs, especially in 1:5 (hUCMSCs:HUVECs) coculture group. The RNA-seq result indicated that cocultured HUVECs exhibited high Hif-1 signaling pathway activity. We performed the cell transfection assay to knock down HIF1A-AS2 in HUVECs and then coculture with hUCMSCs, and the expression of VEGFA, HIF1A and PECAM1 were reduced. In pulp regeneration assay, Cotransplantation of hUCMSCs and HUVECs (1,5) group showed pulp-like tissue regeneration. SIGNIFICANCE: Cocultured hUCMSCs and HUVECs can promote vascular formation of HUVECs, and the optimal coculture ration is 1:5 (hUCMSCs:HUVECs). hUCMSCs promote angiogenesis of HUVECs through the long noncoding RNA HIF1A-AS2-activation of the Hif-1 signaling pathway. Cotransplantation of hUCMSCs and HUVECs can regenerate dental pulp-like tissue in vivo.


Assuntos
Polpa Dentária/metabolismo , Células Endoteliais da Veia Umbilical Humana/transplante , Transplante de Células-Tronco Mesenquimais/métodos , Neovascularização Fisiológica/fisiologia , Animais , Técnicas de Cocultura , Colágeno/metabolismo , Polpa Dentária/citologia , Combinação de Medicamentos , Feminino , Técnicas de Silenciamento de Genes , Humanos , Fator 1 Induzível por Hipóxia/metabolismo , Laminina/metabolismo , Camundongos , Proteoglicanas/metabolismo , RNA Longo não Codificante/genética , Regeneração/fisiologia , Cordão Umbilical/citologia
3.
PLoS One ; 15(4): e0230943, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32240230

RESUMO

Pericellular and extracellular proteoglycans play an important role in modulating morphogen gradients and signal transductions. Chondroitin sulfate proteoglycan 4 (Cspg4) is a membrane spanning proteoglycan expressed in immature progenitor cells and cancer cells. Cspg4 participates in cellular events such as proliferation, migration and signal transduction, and these events are generally important for embryo development. In this study, we characterized Cspg4 for its roles in zebrafish embryonic development. Our results demonstrated that cspg4 was maternally expressed from 0 to 3 hours post fertilization (hpf) and expressed in the anterior and posterior embryo end after 9 hpf. Knocking-down cspg4 resulted in a shorter anterior-posterior axis than control embryo, which could be rescued by co-injecting wnt11 mRNA suggesting that Cspg4 regulates body axis organization through modulating the Wnt/planar cell polarity signaling pathway. In addition, overexpressing cspg4 caused cyclopia. The Cspg4 transmembrane domain mutant embryo phenocopied the global over-expression of cspg4 mRNA and led to cyclopia with a very low penetrance. Our results demonstrated that the quantitatively and spatially accurate distribution of Cspg4 is critical for body axis and midline development during gastrulation.


Assuntos
Antígenos/metabolismo , Polaridade Celular/fisiologia , Proteoglicanas/metabolismo , Via de Sinalização Wnt/fisiologia , Peixe-Zebra/metabolismo , Animais , Embrião não Mamífero/metabolismo , Desenvolvimento Embrionário/fisiologia , RNA Mensageiro/metabolismo
4.
Biochim Biophys Acta Gene Regul Mech ; 1863(6): 194491, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32006715

RESUMO

The molecular characteristics of aging that lead to increased disease susceptibility remain poorly understood. Here we present a transcriptomic profile of the human brain associated with age and aging, derived from a systematic integrative analysis of four independent cohorts of genome-wide expression data from 2202 brain samples (cortex, hippocampus and cerebellum) of individuals of different ages (from young infants, 5-10 years old, to elderly people, up to 100 years old) categorized in age stages by decades. The study provides a signature of 1148 genes detected in cortex, 874 genes in hippocampus and 657 genes in cerebellum, that present significant differential expression changes with age according to a robust gamma rank correlation profiling. The signatures show a significant large overlap of 258 genes between cortex and hippocampus, and 63 common genes between the three brain regions. Focusing on cortex, functional enrichment analysis and cell-type analysis provided biological insight about the aging signature. Response to stress and immune response were up-regulated functions. Synapse, neurotransmission and calcium signaling were down-regulated functions. Cell analysis, derived from single-cell data, disclosed an increase of neuronal activity in the young stages of life and a decline of such activity in the old stages. A regulatory analysis identified the transcription factors (TF) associated with the signature of 258 genes, common to cortex and hippocampus; revealing the role of MEF2(A,D), PDX1, FOSL(1,2) and RFX(5,1) as candidate regulators of the signature. Finally, a deep-learning neural network algorithm was used to build a biological age predictor based on the aging signature. This article is part of a Special Issue entitled: Transcriptional Profiles and Regulatory Gene Networks edited by Dr. Federico Manuel Giorgi and Dr. Shaun Mahony.


Assuntos
Envelhecimento/genética , Encéfalo/metabolismo , Regulação da Expressão Gênica , Redes Reguladoras de Genes , Adolescente , Idoso , Idoso de 80 Anos ou mais , Envelhecimento/imunologia , Envelhecimento/metabolismo , Astrócitos/metabolismo , Sítios de Ligação , Córtex Cerebral/metabolismo , Criança , Pré-Escolar , Aprendizado Profundo , Hipocampo/metabolismo , Humanos , Lactente , Microglia/metabolismo , Pessoa de Meia-Idade , Neurônios/metabolismo , Proteoglicanas/metabolismo , Fatores de Transcrição/metabolismo , Transcriptoma , Adulto Jovem
5.
J Biomed Sci ; 27(1): 2, 2020 Jan 02.
Artigo em Inglês | MEDLINE | ID: mdl-31898491

RESUMO

BACKGROUND: Serglycin (SRGN), previously recognized as an intracellular proteoglycan involved in the storage processes of secretory granules, has recently been shown to be upregulated in several solid tumors. We have previously shown that SRGN in non-small cell lung cancer (NSCLC) promotes malignant phenotypes in a CD44-dependent manner and increased expression of SRGN predicts poor prognosis of primary lung adenocarcinomas. However, the underlying mechanism remains to be defined. METHODS: Overexpression, knockdown and knockout approaches were performed to assess the role of SRGN in cell motility using wound healing and Boyden chamber migration assays. SRGN devoid of glycosaminoglycan (GAG) modification was produced by site-directed mutagenesis or chondroitinase treatment. Liquid chromatography/tandem mass spectrometry was applied for quantitative analysis of the disaccharide compositions and sulfation extent of SRGN GAGs. Western blot and co-immunoprecipitation analyses were performed to determine the expression and interaction of proteins of interest. Actin cytoskeleton organization was monitored by immunofluorescence staining. RESULTS: SRGN expressed by NSCLC cells is readily secreted to the extracellular matrix in a heavily glycosylated form attached with mainly chondroitin sulfate (CS)-GAG chains, and to a lesser extent with heparin sulfate (HS). The CS-GAG moiety serves as the structural motif for SRGN binding to tumor cell surface CD44 and promotes cell migration. SRGN devoid of CS-GAG modification fails to interact with CD44 and has lost the ability to promote cell migration. SRGN/CD44 interaction promotes focal adhesion turnover via Src-mediated paxillin phosphorylation and disassembly of paxillin/FAK adhesion complex, facilitating cell migration. In support, depletion of Src activity or removal of CS-GAGs efficiently blocks SRGN-mediated Src activation and cell migration. SRGN also promotes cell migration via inducing cytoskeleton reorganization mediated through RAC1 and CDC42 activation accompanied with increased lamellipodia and filopodia formation. CONCLUSIONS: Proteoglycan SRGN promotes NSCLC cell migration via the binding of its GAG motif to CD44. SRGN/CD44 interaction induces Rho-family GTPase-mediated cytoskeleton reorganization and facilitates Src-mediated focal adhesion turnover, leading to increased cell migration. These findings suggest that targeting specific glycans in tumor microenvironment that serve as ligands for oncogenic pathways may be a potential strategy for cancer therapy.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/genética , Glicosaminoglicanos/genética , Receptores de Hialuronatos/genética , Proteoglicanas/genética , Proteínas de Transporte Vesicular/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Adesão Celular/genética , Linhagem Celular Tumoral , Movimento Celular/genética , Quinase 1 de Adesão Focal/genética , Regulação Neoplásica da Expressão Gênica/genética , Técnicas de Silenciamento de Genes , Glicosaminoglicanos/metabolismo , Humanos , Receptores de Hialuronatos/metabolismo , Mutagênese Sítio-Dirigida , Ligação Proteica/genética , Proteoglicanas/metabolismo , Proteínas de Transporte Vesicular/metabolismo , Proteínas rho de Ligação ao GTP/genética , Quinases da Família src/genética
6.
Carbohydr Polym ; 230: 115612, 2020 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-31887952

RESUMO

This work reports on the preparation and systematic testing of a novel multi-layered coating, comprised of the non-steroid anti-inflammatory drug diclofenac and biopolymer carboxymethyl cellulose. Drug release testing was performed on an Automated Transdermal Diffusion Cells Sampling System in combination with UV-VIS spectroscopy as the released drug concentration determination method. The results showed that most of the drug is released in the first six hours, whereas the overall released amount could be tailored through changes in the multi-layered coating composition. Biocompatibility tests performed on human osteoblast cells, showed cell viability improvement between 7% and 17% compared to the control sample. The expression of proteins playing important roles in extracellular matrix production and functioning was performed in order to obtain additional proof of the prepared materials' osteointegration boosting capacity. Finally, electrochemical measurements confirmed that the coatings do not influence the corrosion susceptibility of AISI 316LVM stainless steel.


Assuntos
Anti-Inflamatórios não Esteroides/administração & dosagem , Carboximetilcelulose Sódica/química , Diclofenaco/administração & dosagem , Liberação Controlada de Fármacos , Osteoblastos/efeitos dos fármacos , Osteogênese , Aço/química , Anti-Inflamatórios não Esteroides/química , Anti-Inflamatórios não Esteroides/farmacologia , Materiais Biocompatíveis/química , Proteína Morfogenética Óssea 2/genética , Proteína Morfogenética Óssea 2/metabolismo , Proliferação de Células , Células Cultivadas , Diclofenaco/química , Diclofenaco/farmacologia , Matriz Extracelular/metabolismo , Proteínas da Matriz Extracelular/genética , Proteínas da Matriz Extracelular/metabolismo , Humanos , Osteoblastos/metabolismo , Osteoblastos/fisiologia , Proteoglicanas/genética , Proteoglicanas/metabolismo
7.
J Orthop Res ; 38(2): 368-377, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31429976

RESUMO

The dog is the most commonly used large animal model for the study of osteoarthritis. Optimizing methods for assessing cartilage health would prove useful in reducing the number of dogs needed for a valid study of osteoarthritis and cartilage repair. Twelve beagles had critical-sized osteochondral defects created in the medial femoral condyle of both knees. Eight dogs had T1ρ and T2 magnetic resonance imaging (MRI) performed approximately 6 months after defect creation. Following MRI evaluations, all 12 dogs were humanely euthanatized and cartilage samples were obtained from the medial and lateral femoral condyles, medial and lateral tibial plateaus, trochlear groove, and patella for proteoglycan and collagen quantification. Equilibrium partitioning of an ionic contrast (EPIC)-µCT was then performed followed by the histologic assessment of the knees. Correlations between T1ρ, T2, EPIC-µCT and proteoglycan, collagen, and histology scores were assessed using a multivariate analysis accounting for correlations from samples within the same knee and in the same dog. Pearson's correlation coefficients were calculated to assess the strength of significant relationships. Correlations between µCT values and biochemical or histologic assessment were weak to moderately strong (0.09-0.41; p < 0.0001-0.66). There was a weak correlation between the T2 values and cartilage proteoglycan (-0.32; p = 0.04). The correlation between T1ρ values and cartilage proteoglycan were moderately strong (-0.38; p < 0.05) while the strongest correlation was between the T1ρ values and histological assessment of cartilage with a correlation coefficient of 0.58 (p < 0.0001). These data suggest that T1ρ shows promise for possible utility in the translational study of cartilage health and warrants further development in this species. © 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 38:368-377, 2020.


Assuntos
Cartilagem Articular/diagnóstico por imagem , Traumatismos do Joelho/diagnóstico por imagem , Animais , Cartilagem Articular/metabolismo , Colágeno/metabolismo , Modelos Animais de Doenças , Cães , Feminino , Traumatismos do Joelho/metabolismo , Imagem por Ressonância Magnética , Masculino , Proteoglicanas/metabolismo , Microtomografia por Raio-X
8.
Clin Sports Med ; 39(1): 1-12, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31767101

RESUMO

The menisci are 2 fibrocartilaginous crescents anchored via bony and ligamentous attachments to surrounding structures. Their biochemical composition and multilayered structure make them ideal for converting compressive forces to tensile forces in addition to improving joint congruity and providing shock absorption to weight bearing. The medial meniscus maintains more attachments at both the horns and the midbody than the lateral meniscus, making it more susceptible to injury. Understanding of the gross anatomy, vascular anatomy, biochemical composition, and microstructure is key to understanding causes of meniscal pathology as well as treatment options for restoring its primary functions.


Assuntos
Meniscos Tibiais/anatomia & histologia , Meniscos Tibiais/fisiologia , Água Corporal/metabolismo , Colágeno Tipo I/metabolismo , Colágeno Tipo II/metabolismo , Glicosaminoglicanos/metabolismo , Humanos , Articulação do Joelho/anatomia & histologia , Articulação do Joelho/fisiologia , Propriocepção/fisiologia , Proteoglicanas/metabolismo , Líquido Sinovial/fisiologia , Lesões do Menisco Tibial/fisiopatologia , Suporte de Carga/fisiologia
9.
In Vivo ; 34(1): 461-467, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31882514

RESUMO

BACKGROUND/AIM: Endothelial cell-specific molecule-1 (ESM-1) is a soluble proteoglycan which has important role in various biological events. We investigated the impact of the ESM-1 expression in cancer tissues on outcomes in stage II/III gastric cancer patients who received adjuvant S-1 chemotherapy. PATIENTS AND METHODS: The ESM-1 mRNA expression in cancerous tissues and adjacent normal mucosa from 253 patients was measured. The associations between the ESM-1 gene expression and the survival and clinicopathological features were investigated. RESULTS: A significant association was observed between high ESM-1 expression and undifferentiated adenocarcinoma. The overall survival curve was significantly lower in patients with high ESM-1 expression than in those with low expression (p=0.005). High ESM-1 expression was a significant independent prognosticator (HR=2.291, p=0.007). CONCLUSION: ESM-1 gene expression in cancerous tissues is an important prognosticator in stage II/III gastric cancer patients who received adjuvant S-1 chemotherapy.


Assuntos
Adenocarcinoma/mortalidade , Biomarcadores Tumorais/metabolismo , Proteínas de Neoplasias/metabolismo , Ácido Oxônico/uso terapêutico , Proteoglicanas/metabolismo , Neoplasias Gástricas/mortalidade , Tegafur/uso terapêutico , Adenocarcinoma/tratamento farmacológico , Adenocarcinoma/metabolismo , Adenocarcinoma/patologia , Idoso , Antimetabólitos Antineoplásicos/uso terapêutico , Quimioterapia Adjuvante , Combinação de Medicamentos , Feminino , Seguimentos , Humanos , Masculino , Estadiamento de Neoplasias , Estudos Retrospectivos , Neoplasias Gástricas/tratamento farmacológico , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/patologia , Taxa de Sobrevida
10.
Methods Mol Biol ; 2043: 125-136, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31463908

RESUMO

Aggrecan is a major matrix component of articular cartilage, and its dysregulated proteolysis is a crucial event in the pathogenesis of arthritis. Aggrecanases, members of ADAMTS family, play a pivotal role in aggrecan degradation with ADAMTS-4 and ADAMTS-5 being key enzymes. Cleavage events mediated by ADAMTSs are highly specific and very well characterized; therefore, it is possible to investigate aggrecanolysis by using antibodies reacting with the new N- and C-termini of the cleavage products (neoepitope antibodies). Here, we present a method for analyzing dynamic aggrecanolysis by Western blotting using neoepitope antibodies in combination with antibodies against total aggrecan fragments. The protocol is robust and has a broad application for investigation of aggrecanase activity in vitro and ex vivo.


Assuntos
Proteínas ADAMTS/metabolismo , Anticorpos/metabolismo , Epitopos/imunologia , Agrecanas/química , Agrecanas/imunologia , Animais , Western Blotting , Epitopos/química , Humanos , Proteoglicanas/metabolismo
11.
Int J Mol Sci ; 20(24)2019 Dec 09.
Artigo em Inglês | MEDLINE | ID: mdl-31835434

RESUMO

Transforming growth factor-ßs (TGF-ßs) signal after binding to the TGF-ß receptors TßRI and TßRII. Recently, however, betaglycan (BG) was identified as an important co-receptor, especially for TGF-ß2. Both proteins are involved in several testicular functions. Thus, we analyzed the importance of BG for TGF-ß1/2 signaling in Sertoli cells with ELISAs, qRT-PCR, siRNA silencing and BrdU assays. TGF-ß1 as well as TGF-ß2 reduced shedding of membrane-bound BG (mBG), thus reducing the amount of soluble BG (sBG), which is often an antagonist to TGF-ß signaling. Treatment of Sertoli cells with GM6001, a matrix metalloproteinases (MMP) inhibitor, also counteracted BG shedding, thus suggesting MMPs to be mainly involved in shedding. Interestingly, TGF-ß2 but not TGF-ß1 enhanced secretion of tissue inhibitor of metalloproteinases 3 (TIMP3), a potent inhibitor of MMPs. Furthermore, recombinant TIMP3 attenuated BG shedding. Co-stimulation with TIMP3 and TGF-ß1 reduced phosphorylation of Smad3, while a combination of TIMP3/TGF-ß2 increased it. Silencing of BG as well as TIMP3 reduced TGF-ß2-induced phosphorylation of Smad2 and Smad3 significantly, once more highlighting the importance of BG for TGF-ß2 signaling. In contrast, this effect was not observed with TIMP3/TGF-ß1. Silencing of BG and TIMP3 decreased significantly Sertoli cell proliferation. Taken together, BG shedding serves a major role in TGF-ß2 signaling in Sertoli cells.


Assuntos
Proliferação de Células , Proteoglicanas/metabolismo , Receptores de Fatores de Crescimento Transformadores beta/metabolismo , Células de Sertoli/metabolismo , Transdução de Sinais , Fator de Crescimento Transformador beta2/metabolismo , Animais , Linhagem Celular , Dipeptídeos/farmacologia , Masculino , Inibidores de Metaloproteinases de Matriz/farmacologia , Ratos , Células de Sertoli/citologia , Proteínas Smad/metabolismo , Inibidor Tecidual de Metaloproteinase-3/metabolismo
12.
Acta Neurobiol Exp (Wars) ; 79(4): 338-351, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31885391

RESUMO

Over the last twenty years chondroitin sulfate (CS) has become a focus of interest of neuroscience due to its indubitable role in shaping axonal growth, synaptic plasticity and glial scar forming. Various patterns of sulfation give rise to various CS molecules with different properties that are capable of interactions with a plethora of molecules, including growth factors, receptors and guidance molecules. The involvement of CS chains has been implicated in visual critical period regulation, memory formation, spinal cord regeneration. As part of proteoglycan molecules, they are widely expressed in the central nervous system, however, little is known about the enzymatic machinery responsible for CS synthesis and degradation. In this review we attempt to extract and collect the available information concerning the expression and function of enzymes of CS metabolism in the brain.


Assuntos
Encéfalo/metabolismo , Sulfatos de Condroitina/metabolismo , Animais , Encéfalo/enzimologia , Configuração de Carboidratos , Retículo Endoplasmático/metabolismo , Matriz Extracelular/metabolismo , Glicosiltransferases/metabolismo , Humanos , Redes e Vias Metabólicas , Proteínas do Tecido Nervoso/metabolismo , Plasticidade Neuronal , Proteoglicanas/metabolismo , Xilose/metabolismo
13.
Dermatol Surg ; 45(12): 1580-1584, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31765337

RESUMO

BACKGROUND: Little literature exits on the mechanism of action of implanted polymethylmethacrylate (PMMA) filler. OBJECTIVE: To characterize PMMA-induced dermal extracellular matrix production in the skin. MATERIALS AND METHODS: Single-center, open-label prospective study in healthy volunteers undergoing removal of redundant skin was injected intradermally and subdermally with PMMA dermal filler (Bellafill). Punch biopsies were harvested over a time course and evaluated for the deposition of collagen-3 and procollagen-1, proteoglycans and elastin using immunohistochemistry. Blinded histopathologic readings were performed by a dermatopathologist to characterize the nature of the dermal extracellular matrix findings. RESULTS: Normal inflammatory infiltrate was exhibited at all timepoints after PMMA injection with an influx of fibroblasts and new vasculature. Tissue proteoglycans were noted within the injectate beginning at Week 1 and persisted through the study end point. Increased collagen Type 3 was evident following the first week after injection, peaked at Month 2 and diminished through Months 3 through 6. Procollagen-1 was noted at Month 1 and continued to increase in intensity and organization through the study end point (6 months). Elastin staining was inconclusive. Polymethylmethacrylate microspheres remained within the initial injection area and became encapsulated within new collagen fibers. The growth and pattern of new connective tissue mimicked a normal wound healing response. CONCLUSION: Polymethylmethacrylate-collagen gel filler stimulates collagen-3 and procollagen-1 when injected into human skin. This combination of neocollagenesis followed by microencapsulation of PMMA microspheres in the new tissue provides for long-lasting results.


Assuntos
Colágeno Tipo III/biossíntese , Colágeno Tipo I/biossíntese , Colágeno/administração & dosagem , Preenchedores Dérmicos/administração & dosagem , Derme/efeitos dos fármacos , Polimetil Metacrilato/administração & dosagem , Adulto , Biópsia , Derme/citologia , Derme/metabolismo , Elastina/metabolismo , Feminino , Fibroblastos/efeitos dos fármacos , Voluntários Saudáveis , Humanos , Injeções Intradérmicas , Microesferas , Pessoa de Meia-Idade , Estudos Prospectivos , Proteoglicanas/metabolismo
14.
Proc Natl Acad Sci U S A ; 116(43): 21354-21360, 2019 10 22.
Artigo em Inglês | MEDLINE | ID: mdl-31601738

RESUMO

Trichomonas vaginalis, a human-infective parasite, causes the most prevalent nonviral sexually transmitted infection worldwide. This pathogen secretes extracellular vesicles (EVs) that mediate its interaction with host cells. Here, we have developed assays to study the interface between parasite EVs and mammalian host cells and to quantify EV internalization by mammalian cells. We show that T. vaginalis EVs interact with glycosaminoglycans on the surface of host cells and specifically bind to heparan sulfate (HS) present on host cell surface proteoglycans. Moreover, competition assays using HS or removal of HS from the host cell surface strongly inhibit EV uptake, directly demonstrating that HS proteoglycans facilitate EV internalization. We identified an abundant protein on the surface of T. vaginalis EVs, 4-α-glucanotransferase (Tv4AGT), and show using isothermal titration calorimetry that this protein binds HS. Tv4AGT also competitively inhibits EV uptake, defining it as an EV ligand critical for EV internalization. Finally, we demonstrate that T. vaginalis EV uptake is dependent on host cell cholesterol and caveolin-1 and that internalization proceeds via clathrin-independent, lipid raft-mediated endocytosis. These studies reveal mechanisms used to drive host:pathogen interactions and further our understanding of how EVs are internalized by target cells to allow cross-talk between different cell types.


Assuntos
Endocitose , Vesículas Extracelulares/metabolismo , Proteoglicanas/metabolismo , Vaginite por Trichomonas/parasitologia , Trichomonas vaginalis/metabolismo , Transporte Biológico , Caveolinas/metabolismo , Colesterol/metabolismo , Feminino , Interações Hospedeiro-Parasita , Humanos , Proteínas de Protozoários/genética , Proteínas de Protozoários/metabolismo , Vaginite por Trichomonas/metabolismo , Vaginite por Trichomonas/fisiopatologia , Trichomonas vaginalis/genética
15.
Biomed Res ; 40(5): 197-205, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31597905

RESUMO

We investigated the effects of ibandronate, a bisphosphonate; eldecalcitol, an active vitamin D3 analogue; and combination treatment with both agents on secondary osteoporosis and arthritis using rats with adjuvant-induced arthritis. Arthritis was induced in 8-week-old male Lewis rats. Rats were randomized into four treatment groups and an untreated normal control group: ibandronate, eldecalcitol, ibandronate + eldecalcitol, vehicle, and control. Paw thickness was measured to evaluate arthritis. Joint destruction was evaluated histomorphometrically by the ankle joint stained with Fast Green and safranin O. The femur and lumbar spine were scanned using dual-energy X-ray absorptiometry, and the distal femur was scanned using micro-computed tomography for bone mineral density (BMD) and trabecular microstructural evaluations. Ibandronate and/or eldecalcitol increased BMD in both the lumbar vertebrae and femur and improved several microstructural parameters (bone volume/total volume, structure model index, trabecular number, and trabecular separation of the distal femur). In addition, there was an additive effect of combination treatment compared with single treatments for most trabecular parameters, including BMD and bone volume. However, ibandronate and/or eldecalcitol did not inhibit arthritis and joint destruction. Combination treatment with ibandronate and eldecalcitol may be effective for secondary osteoporosis associated with arthritis.


Assuntos
Artrite Experimental/diagnóstico , Artrite Experimental/etiologia , Ácido Ibandrônico/farmacologia , Osteoporose/diagnóstico , Osteoporose/etiologia , Vitamina D/análogos & derivados , Microtomografia por Raio-X , Animais , Artrite Experimental/tratamento farmacológico , Artrite Experimental/metabolismo , Biópsia , Modelos Animais de Doenças , Imageamento Tridimensional , Articulações/diagnóstico por imagem , Articulações/patologia , Osteoporose/tratamento farmacológico , Osteoporose/metabolismo , Fenótipo , Proteoglicanas/metabolismo , Ratos , Vitamina D/farmacologia
16.
Arch Med Res ; 50(5): 304-314, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31600602

RESUMO

BACKGROUND AND AIM: Endocan is a novel endothelium-derived proteoglycan and may play a role in endothelial cells activity under diabetic conditions. Here, we evaluated the effect of high glucose concentration (30 mmol glucose) on endocan level in presence or absence of metformin in human umbilical vein endothelial cells (HUVECs). METHODS: Cells were incubated with 30 mmol glucose for 72 h. High glucose content, metformin (2.5 to 500 mmol) and compound C (10 mmol) effects were assessed on cell viability. HUVECs migration was studied by scratch test. The changes in endocan expression and protein level were evaluated by RT-PCR, ELISA and flow cytometry assays. Griess reaction was used to measure NO levels. Functional activity of endothelial cells was monitored related to lipoprotein lipase activity using Dil-Ac-LDL uptake. p-AMPK/AMPK ratio was assessed by western blotting. RESULTS: Cells viability significantly was reduced under high glucose condition (p <0.05). 30 mmol glucose inhibited HUVECs migration, whereas these features were improved by 50 mmol metformin (p <0.05). Endocan transcription and protein levels were increased in diabetic HUVECs exposed to metformin (p <0.05). Metformin increased NO production in HUVECs under high glucose condition (p <0.001). Metformin increased LDL uptake capacity under high glucose condition (p <0.05). The addition of compound C blunted these effects. Western blot analysis confirmed the increase of p-AMPK/AMPK ratio in metformin-treated cells. CONCLUSION: Data demonstrated that metformin could promote angiogenic potential of endothelial cells which its reduction is a main cause in the development of diabetic foot ulcer, probably by the regulation of endocan dynamics under high glucose condition.


Assuntos
Diabetes Mellitus/tratamento farmacológico , Células Endoteliais/metabolismo , Hipoglicemiantes/uso terapêutico , Metformina/uso terapêutico , Proteínas de Neoplasias/metabolismo , Proteoglicanas/metabolismo , Humanos , Hipoglicemiantes/farmacologia , Metformina/farmacologia
17.
Biomed Pharmacother ; 119: 109365, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31525642

RESUMO

Circular RNAs (circRNAs) have been reported to play critical roles in tumorigenesis. However, the roles of circRNAs in laryngeal squamous cell carcinoma (LSCC) are still largely unknown. In our present study, we identified a novel circRNA hsa_circ_0042666, which was poorly expressed in LSCC. Low hsa_circ_0042666 expression was closely associated with advanced tumor stage, lymph-node metastasis, and poor overall survival. In vitro function assays, we showed that hsa_circ_0042666 dramatically reduced LSCC cells proliferation and invasion in vitro. In mechanism, our data indicated that hsa_circ_0042666 could competitively bind to miR-223 as a miRNA sponge to regulate TGFBR3 expression in LSCC progression. Altogether, these findings elucidated that hsa_circ_0042666 regulated LSCC cells proliferation and invasion by miR-223/TGFBR3 axis, which might provide a therapeutic strategy for the treatment of LSCC.


Assuntos
Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/patologia , Neoplasias Laríngeas/genética , Neoplasias Laríngeas/patologia , MicroRNAs/metabolismo , Proteoglicanas/metabolismo , RNA Circular/metabolismo , Receptores de Fatores de Crescimento Transformadores beta/metabolismo , Idoso , Sequência de Bases , Linhagem Celular Tumoral , Proliferação de Células/genética , Progressão da Doença , Regulação para Baixo/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , MicroRNAs/genética , Pessoa de Meia-Idade , Invasividade Neoplásica , Proteoglicanas/genética , RNA Circular/genética , Receptores de Fatores de Crescimento Transformadores beta/genética
18.
Int J Biochem Cell Biol ; 116: 105614, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31550547

RESUMO

Osteopontin (OPN) is an osteoblast-derived secretory protein that plays a role in bone remodeling, osteoblast responsiveness, and inflammation. We recently found that osteoblast differentiation is type-specific, with conditions of JNK inactivation inducing osteoblasts that preferentially express OPN (OPN-type). Since OPN-type osteoblasts highly express osteogenesis-inhibiting proteins and Rankl, an important inducer of osteoclastogenesis, an increased appearance of OPN-type osteoblasts may be associated with inefficient and poor-quality bone regeneration. However, whether specific osteogenic inducers can modulate OPN-type osteoblast differentiation is completely unknown. Here, we demonstrate that bone morphogenic protein 9 (BMP9) prevents induction of OPN-type osteoblast differentiation under conditions of JNK inhibition. Although JNK inactivation suppressed both BMP2- and BMP9-induced matrix mineralization and osteocalcin expression, the expression of Rankl and specific cytokines such as Gpha2, Esm1, and Sfrp1 under similar conditions was increased in all cells except those treated with BMP9. Increased expression of Id4, a critical transcriptional regulator of OPN-type osteoblast differentiation, was similarly prevented only in BMP9-treated cells. We also found that BMP9 specifically induces the expression of Hey1, a bHLH transcriptional repressor, and that Id4 inhibits the suppressive effects of Hey1 on Opn promoter activity by forming Id4-Hey1 complexes in osteoblasts. Using site-direct mutagenesis, ChIP, and immunoprecipitation, we elucidated that BMP9-induced overexpression of Hey1 can overcome the effects of Id4 and suppress OPN expression. We further found that p38 activation and JNK inactivation are involved in BMP9-induced Hey1 expression. Collectively, these data suggest that BMP9 is a unique osteogenic inducer that regulates OPN-type osteoblast differentiation.


Assuntos
Proteínas de Ciclo Celular/genética , Fator 2 de Diferenciação de Crescimento/farmacologia , Proteínas Inibidoras de Diferenciação/genética , Osteoblastos/efeitos dos fármacos , Osteogênese/efeitos dos fármacos , Osteopontina/genética , Animais , Animais Recém-Nascidos , Proteína Morfogenética Óssea 2/farmacologia , Proteínas de Ciclo Celular/metabolismo , Diferenciação Celular/efeitos dos fármacos , Regulação da Expressão Gênica , Glicerofosfatos/farmacologia , Glicoproteínas/genética , Glicoproteínas/metabolismo , Proteínas Inibidoras de Diferenciação/metabolismo , MAP Quinase Quinase 4/antagonistas & inibidores , MAP Quinase Quinase 4/genética , MAP Quinase Quinase 4/metabolismo , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Mutagênese Sítio-Dirigida , Osteoblastos/citologia , Osteoblastos/metabolismo , Osteocalcina/genética , Osteocalcina/metabolismo , Osteogênese/genética , Osteopontina/metabolismo , Cultura Primária de Células , Proteoglicanas/genética , Proteoglicanas/metabolismo , Ligante RANK/genética , Ligante RANK/metabolismo , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Transdução de Sinais , Proteínas Quinases p38 Ativadas por Mitógeno/genética , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo
19.
Arterioscler Thromb Vasc Biol ; 39(10): 2067-2081, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31366218

RESUMO

OBJECTIVE: Investigate the requirement of Aggrecan (Acan) cleavage during aortic wall development in a murine model with ADAMTS (a disintegrin-like and metalloprotease domain with thrombospondin-type motifs) 5 deficiency and bicuspid aortic valves. APPROACH: Mice with altered extracellular matrix remodeling of proteoglycans will be examined for anomalies in ascending aortic wall development. Neo-epitope antibodies that recognize ADAMTS cleaved Acan fragments will be used to investigate the mechanistic requirement of Acan turnover, in aortic wall development. RESULTS: Adamts5-/-;Smad2+/- mice exhibited a high penetrance of aortic anomalies (n=17/17); Adamts5-/-;Smad2+/- mice with bicuspid aortic valves (7/17) showed a higher number of anomalies than Adamts5-/-;Smad2+/- mice with tricuspid aortic valves. Single mutant Adamts5-/- mice also displayed a high penetrance of aortic anomalies (n=19/19) compared with wild type (n=1/11). Aortic anomalies correlated with Acan accumulation that was apparent at the onset of elastogenesis in Adamts5-/- mice. Neo-epitope antibodies that recognize the initial amino acids in the Acan cleaved fragments neo-FREEE, neo-GLGS, and neo-SSELE were increased in the Adamts5-/- aortas compared with WT. Conversely, neo-TEGE, which recognizes highly digested Acan core fragments, was reduced in Adamts5-/- mice. However, mice containing a mutation in the TEGE373↓374ALGSV site, rendering it noncleavable, had low penetrance of aortic anomalies (n=2/4). Acan neo-DIPEN and neo-FFGVG fragments were observed in the aortic adventitia; Acan neo-FFGVG was increased abnormally in the medial layer and overlapped with smooth muscle cell loss in Adamts5-/- aortas. CONCLUSIONS: Disruption of ADAMTS5 Acan cleavage during development correlates with ascending aortic anomalies. These data indicate that the mechanism of ADAMTS5 Acan cleavage may be critical for normal aortic wall development.


Assuntos
Proteína ADAMTS5/genética , Agrecanas/genética , Aorta/anormalidades , Valva Aórtica/anormalidades , Regulação da Expressão Gênica no Desenvolvimento , Doenças das Valvas Cardíacas/patologia , Malformações Vasculares/genética , Proteínas ADAM/genética , Animais , Valva Aórtica/patologia , Biópsia por Agulha , Doenças das Valvas Cardíacas/genética , Imuno-Histoquímica , Camundongos , Camundongos Transgênicos , Modelos Animais , Proteoglicanas/metabolismo , Sensibilidade e Especificidade , Proteína Smad2/metabolismo
20.
BMC Endocr Disord ; 19(1): 90, 2019 Aug 27.
Artigo em Inglês | MEDLINE | ID: mdl-31455321

RESUMO

BACKGROUND: Endothelial cell-specific molecule-1 (ESM-1) is a biomarker associated with tumor progression in pituitary adenoma. We specifically focused on one type of pituitary adenoma, namely null cell adenoma (NCA) and evaluated the relationship between invasion and ESM-1 expression in both vascular endothelial and adenoma tissues. METHODS: Tissue samples from 94 patients with pituitary NCA were obtained through microscopic transsphenoidal resection. Tumor size and invasion were determined through preoperative magnetic resonance imaging. Immunohistochemical staining was performed to detect ESM-1 expression. ESM-1 index of ≥3 was defined as high expression. RESULTS: Signs of invasion were observed in 46 (47.9%) of the 94 patients. Significant differences were observed in the invasion state and maximum tumor diameter between high and low expression of ESM-1 in vascular endothelial tissues (both P < 0.05). Significant positive associations were noted between ESM-1 expression in vascular endothelial tissues and tumor invasion (P = 0.002) and tumor size (P = 0.020). However, only tumor size was associated with ESM-1 expression in adenoma tissues (P = 0.016). CONCLUSION: In NCA, a significant positive association between tumor invasion and ESM-1 expression was observed only in vascular endothelial tissues, suggesting that tumor progression occurs mainly through ESM-1-associated mechanism.


Assuntos
Adenoma/patologia , Biomarcadores/metabolismo , Linfócitos Nulos/patologia , Proteínas de Neoplasias/metabolismo , Neoplasias Hipofisárias/patologia , Proteoglicanas/metabolismo , Adenoma/metabolismo , Adenoma/cirurgia , Feminino , Seguimentos , Humanos , Linfócitos Nulos/metabolismo , Masculino , Pessoa de Meia-Idade , Invasividade Neoplásica , Neoplasias Hipofisárias/metabolismo , Neoplasias Hipofisárias/cirurgia , Prognóstico
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