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1.
PLoS Comput Biol ; 16(6): e1007693, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32520928

RESUMO

Understanding cellular remodeling in response to mechanical stimuli is a critical step in elucidating mechanical activation of biochemical signaling pathways. Experimental evidence indicates that external stress-induced subcellular adaptation is accomplished through dynamic cytoskeletal reorganization. To study the interactions between subcellular structures involved in transducing mechanical signals, we combined experimental data and computational simulations to evaluate real-time mechanical adaptation of the actin cytoskeletal network. Actin cytoskeleton was imaged at the same time as an external tensile force was applied to live vascular smooth muscle cells using a fibronectin-functionalized atomic force microscope probe. Moreover, we performed computational simulations of active cytoskeletal networks under an external tensile force. The experimental data and simulation results suggest that mechanical structural adaptation occurs before chemical adaptation during filament bundle formation: actin filaments first align in the direction of the external force by initializing anisotropic filament orientations, then the chemical evolution of the network follows the anisotropic structures to further develop the bundle-like geometry. Our findings present an alternative two-step explanation for the formation of actin bundles due to mechanical stimulation and provide new insights into the mechanism of mechanotransduction.


Assuntos
Citoesqueleto de Actina/fisiologia , Resistência à Tração , Actinas/fisiologia , Animais , Anisotropia , Fenômenos Biomecânicos , Células Cultivadas , Simulação por Computador , Fibronectinas/fisiologia , Processamento de Imagem Assistida por Computador , Imageamento Tridimensional , Mecanotransdução Celular , Microscopia de Força Atômica , Miócitos de Músculo Liso/metabolismo , Miosinas/fisiologia , Ratos , Estresse Mecânico
2.
PLoS One ; 15(6): e0234465, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32544194

RESUMO

Obesity leads a crucial importance in metabolic disorders, as well as type 2 diabetes mellitus. Our present study was designed to assess the potential role of irisin, adiponectin, leptin and gene polymorphism of PNPLA3, leptin and adiponectin as predictive markers of diabetes associated with obesity. One hundred eighty subjects were distributed to three groups including; healthy non-diabetic non obese volunteers as a control group, diabetic non obese group, and diabetic obese group (n = 60 for each group). Fasting blood samples of all groups were collected to determine fasting blood glucose, insulin levels, insulin resistance, total cholesterol, high density lipoprotein cholesterol (HDL-C), low density lipoprotein cholesterol (LDL-C), triacylglycerol, irisin, adiponectin, leptin; as well as, polymorphism of PNPLA3, adiponectin and leptin. The results showed that glucose, insulin resistance, total cholesterol, irisin, leptin, LDL-C, triacylglycerol concentrations were significantly increased, however, insulin, HDL-C, adiponectin were significantly decreased in diabetic obese patients in relation to diabetic non-obese patients as well as in healthy volunteers. The polymorphism of PNPLA3 rs738409 was linearly related to irisin and leptin but was not related with circulating concentrations of adiponectin. We concluded that increased irisin and leptin levels can predict the insulin resistance in obese patients. Moreover, patients who have mutant genotype of PNPLA3 I148 gene (rs738409) C>G, ADIPOQ gene (rs266729) G>C and LEP gene (rs2167270) G>A showed a significant higher susceptibility rate for DM in obese people than those with wild type. This could be considered as an adjustable retort to counter the impact of obesity on glucose homeostasis.


Assuntos
Adiponectina/genética , Diabetes Mellitus Tipo 2/etiologia , Predisposição Genética para Doença , Resistência à Insulina/genética , Leptina/genética , Lipase/genética , Proteínas de Membrana/genética , Obesidade/complicações , Obesidade/genética , Adiponectina/sangue , Adulto , Feminino , Fibronectinas/sangue , Fibronectinas/genética , Marcadores Genéticos , Humanos , Leptina/sangue , Lipase/sangue , Masculino , Proteínas de Membrana/sangue , Polimorfismo Genético , Adulto Jovem
3.
PLoS One ; 15(6): e0234617, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32555665

RESUMO

Semicarbazide-sensitive amine oxidase (SSAO) is an enzyme with a unique dual function in controlling inflammation as well as reactive oxygen species (ROS) generation. We have demonstrated benefit of SSAO inhibition in acute kidney fibrosis. However the function of SSAO in chronic kidney disease (CKD) and diabetic kidney disease (DKD) is yet to be determined. We aimed to assess the effectiveness of a SSAO inhibitor (SSAOi; PXS-4728A) as an antifibrotic agent using a diabetic model of CKD. Diabetic mice were treated with SSAOi for 24 weeks and outcomes compared with untreated diabetic mice and telmisartan treated animals as a standard of care comparator. Extracellular matrix markers, fibronectin and oxidative stress, were downregulated in diabetic mice treated with SSAOi compared with untreated diabetic mice. Expression of the pan-leukocyte marker CD45 was also supressed by SSAOi. SSAO inhibition in diabetic mice resulted in a significant reduction in glomerulosclerosis and associated albuminuria compared to untreated diabetic mice. However, the effect of SSAO inhibition was less obvious in the tubulointerstitial compartment than in the glomeruli. Therefore, SSAO may be a potential target for diabetic glomerulosclerosis.


Assuntos
Albuminúria/tratamento farmacológico , Alilamina/análogos & derivados , Amina Oxidase (contendo Cobre)/antagonistas & inibidores , Benzamidas/uso terapêutico , Nefropatias Diabéticas/tratamento farmacológico , Inibidores Enzimáticos/uso terapêutico , Glomérulos Renais/patologia , Túbulos Renais/patologia , Insuficiência Renal Crônica/tratamento farmacológico , Alilamina/farmacologia , Alilamina/uso terapêutico , Animais , Benzamidas/farmacologia , Diabetes Mellitus Experimental , Nefropatias Diabéticas/patologia , Inibidores Enzimáticos/farmacologia , Fibronectinas/metabolismo , Fibrose , Glomérulos Renais/efeitos dos fármacos , Túbulos Renais/efeitos dos fármacos , Antígenos Comuns de Leucócito/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Estresse Oxidativo/efeitos dos fármacos , Insuficiência Renal Crônica/patologia , Telmisartan/farmacologia , Telmisartan/uso terapêutico
4.
PLoS One ; 15(6): e0234638, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32569325

RESUMO

Hematopoietic stem cell transplantation is successfully applied since the late 1950s; however, its efficacy still needs to be increased. A promising strategy is to transplant high numbers of pluripotent hematopoietic stem cells (HSCs). Therefore, an improved ex vivo culture system that supports proliferation and maintains HSC pluripotency would override possible limitations in cell numbers gained from donors. To model the natural HSC niche in vitro, we optimized the HSC medium composition with a panel of cytokines and valproic acid and used an artificial 3D bone marrow-like scaffold made of polydimethylsiloxane (PDMS). This 3D scaffold offered a suitable platform to amplify human HSCs in vitro and, simultaneously, to support their viability, multipotency and ability for self-renewal. Silicon oxide-covering of PDMS structures further improved amplification of CD34+ cells, although the conservation of naïve HSCs was better on non-covered 3D PDMS. Finally, we found that HSC cultivated on non-covered 3D PDMS generated most pluripotent colonies within colony forming unit assays. In conclusion, by combining biological and biotechnological approaches, we optimized in vitro HSCs culture conditions, resulting in improved amplification, multipotency maintenance and vitality of HSCs.


Assuntos
Materiais Biomiméticos/farmacologia , Células-Tronco Hematopoéticas/citologia , Nicho de Células-Tronco , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Colágeno/farmacologia , Dimetilpolisiloxanos/farmacologia , Feminino , Fibronectinas/farmacologia , Células-Tronco Hematopoéticas/efeitos dos fármacos , Humanos , Masculino , Purinas/farmacologia , Nicho de Células-Tronco/efeitos dos fármacos , Ácido Valproico/farmacologia
5.
Arch Endocrinol Metab ; 64(3): 201-204, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32555986

RESUMO

OBJECTIVE: Autonomic nervous system, especially the sympathetic nervous system, may stimulate the expression of peroxisome proliferator-activated receptor γ coactivator-1α, which regulates irisin. This study aimed to explore whether there was any association between autonomic function as assessed by heart rate related indices and irisin release following acute exercise. SUBJECTS AND METHODS: Seventeen healthy adults were asked to perform an incremental exhaustive cycling as well as an incremental exhaustive running separately on different days. Heart rate was monitored, and blood samples were collected before, immediately, 10-, and 60-minutes post-exercise. Serum irisin was measured using ELISA kit. RESULTS: Markers for autonomic function, such as heart rate at rest, peak, or recovery, heart rate reserve, heart rate recovery, and chronotropic index, were comparable between cycling and running (all P > 0.10). Irisin was increased immediately following both exercise. No significant association was observed between heart rate at rest, peak, or recovery and irisin level at the corresponding time-point, as well as between heart rate reserve, heart rate recovery, or chronotropic index and exercise induced irisin release, with or without controlling for age, body mass index, and glucose (all P > 0.10). CONCLUSIONS: Autonomic function might not be associated with irisin release in healthy adults. Arch Endocrinol Metab. 2020;64(3):201-4.


Assuntos
Sistema Nervoso Autônomo/irrigação sanguínea , Sistema Nervoso Autônomo/fisiologia , Fibronectinas/sangue , Frequência Cardíaca/fisiologia , Corrida/fisiologia , Adulto , Estudos Cross-Over , Ensaio de Imunoadsorção Enzimática , Feminino , Humanos , Masculino , Distribuição Aleatória , Adulto Jovem
6.
PLoS Pathog ; 16(5): e1008516, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32413091

RESUMO

Lyme disease, caused by Borrelia burgdorferi, B. afzelii and B. garinii, is a chronic, multi-systemic infection and the spectrum of tissues affected can vary with the Lyme disease strain. For example, whereas B. garinii infection is associated with neurologic manifestations, B. burgdorferi infection is associated with arthritis. The basis for tissue tropism is poorly understood, but has been long hypothesized to involve strain-specific interactions with host components in the target tissue. OspC (outer surface protein C) is a highly variable outer surface protein required for infectivity, and sequence differences in OspC are associated with variation in tissue invasiveness, but whether OspC directly influences tropism is unknown. We found that OspC binds to the extracellular matrix (ECM) components fibronectin and/or dermatan sulfate in an OspC variant-dependent manner. Murine infection by isogenic B. burgdorferi strains differing only in their ospC coding region revealed that two OspC variants capable of binding dermatan sulfate promoted colonization of all tissues tested, including joints. However, an isogenic strain producing OspC from B. garinii strain PBr, which binds fibronectin but not dermatan sulfate, colonized the skin, heart and bladder, but not joints. Moreover, a strain producing an OspC altered to recognize neither fibronectin nor dermatan sulfate displayed dramatically reduced levels of tissue colonization that were indistinguishable from a strain entirely deficient in OspC. Finally, intravital microscopy revealed that this OspC mutant, in contrast to a strain producing wild type OspC, was defective in promoting joint invasion by B. burgdorferi in living mice. We conclude that OspC functions as an ECM-binding adhesin that is required for joint invasion, and that variation in OspC sequence contributes to strain-specific differences in tissue tropism displayed among Lyme disease spirochetes.


Assuntos
Borrelia burgdorferi/metabolismo , Dermatan Sulfato/metabolismo , Matriz Extracelular/metabolismo , Artropatias/metabolismo , Articulações/metabolismo , Doença de Lyme/metabolismo , Animais , Antígenos de Bactérias , Aderência Bacteriana , Proteínas da Membrana Bacteriana Externa , Borrelia burgdorferi/genética , Borrelia burgdorferi/patogenicidade , Dermatan Sulfato/genética , Matriz Extracelular/genética , Matriz Extracelular/microbiologia , Matriz Extracelular/patologia , Feminino , Fibronectinas/genética , Fibronectinas/metabolismo , Artropatias/genética , Artropatias/microbiologia , Artropatias/patologia , Articulações/microbiologia , Articulações/patologia , Doença de Lyme/genética , Doença de Lyme/microbiologia , Doença de Lyme/patologia , Camundongos , Camundongos SCID , Mutação , Especificidade de Órgãos
7.
Proc Natl Acad Sci U S A ; 117(21): 11387-11398, 2020 05 26.
Artigo em Inglês | MEDLINE | ID: mdl-32385149

RESUMO

Altered microarchitecture of collagen type I is a hallmark of wound healing and cancer that is commonly attributed to myofibroblasts. However, it remains unknown which effect collagen microarchitecture has on myofibroblast differentiation. Here, we combined experimental and computational approaches to investigate the hypothesis that the microarchitecture of fibrillar collagen networks mechanically regulates myofibroblast differentiation of adipose stromal cells (ASCs) independent of bulk stiffness. Collagen gels with controlled fiber thickness and pore size were microfabricated by adjusting the gelation temperature while keeping their concentration constant. Rheological characterization and simulation data indicated that networks with thicker fibers and larger pores exhibited increased strain-stiffening relative to networks with thinner fibers and smaller pores. Accordingly, ASCs cultured in scaffolds with thicker fibers were more contractile, expressed myofibroblast markers, and deposited more extended fibronectin fibers. Consistent with elevated myofibroblast differentiation, ASCs in scaffolds with thicker fibers exhibited a more proangiogenic phenotype that promoted endothelial sprouting in a contractility-dependent manner. Our findings suggest that changes of collagen microarchitecture regulate myofibroblast differentiation and fibrosis independent of collagen quantity and bulk stiffness by locally modulating cellular mechanosignaling. These findings have implications for regenerative medicine and anticancer treatments.


Assuntos
Colágeno/ultraestrutura , Miofibroblastos/citologia , Células Estromais/citologia , Tecido Adiposo/citologia , Fenômenos Biomecânicos , Diferenciação Celular , Células Cultivadas , Colágeno/metabolismo , Matriz Extracelular/ultraestrutura , Fibronectinas/metabolismo , Humanos , Mecanotransdução Celular , Miofibroblastos/metabolismo , Miofibroblastos/ultraestrutura , Células Estromais/metabolismo , Células Estromais/ultraestrutura
8.
Nat Commun ; 11(1): 2416, 2020 05 15.
Artigo em Inglês | MEDLINE | ID: mdl-32415208

RESUMO

Chemoresistance is a major obstacle in triple negative breast cancer (TNBC), the most aggressive breast cancer subtype. Here we identify hypoxia-induced ECM re-modeler, lysyl oxidase (LOX) as a key inducer of chemoresistance by developing chemoresistant TNBC tumors in vivo and characterizing their transcriptomes by RNA-sequencing. Inhibiting LOX reduces collagen cross-linking and fibronectin assembly, increases drug penetration, and downregulates ITGA5/FN1 expression, resulting in inhibition of FAK/Src signaling, induction of apoptosis and re-sensitization to chemotherapy. Similarly, inhibiting FAK/Src results in chemosensitization. These effects are observed in 3D-cultured cell lines, tumor organoids, chemoresistant xenografts, syngeneic tumors and PDX models. Re-expressing the hypoxia-repressed miR-142-3p, which targets HIF1A, LOX and ITGA5, causes further suppression of the HIF-1α/LOX/ITGA5/FN1 axis. Notably, higher LOX, ITGA5, or FN1, or lower miR-142-3p levels are associated with shorter survival in chemotherapy-treated TNBC patients. These results provide strong pre-clinical rationale for developing and testing LOX inhibitors to overcome chemoresistance in TNBC patients.


Assuntos
Antineoplásicos/farmacologia , Resistencia a Medicamentos Antineoplásicos , Proteína-Lisina 6-Oxidase/antagonistas & inibidores , Neoplasias de Mama Triplo Negativas/tratamento farmacológico , Neoplasias de Mama Triplo Negativas/enzimologia , Animais , Apoptose , Biomarcadores Tumorais/metabolismo , Linhagem Celular Tumoral , Colágeno/química , Regulação para Baixo , Matriz Extracelular/metabolismo , Feminino , Fibronectinas/metabolismo , Quinase 1 de Adesão Focal/metabolismo , Regulação Neoplásica da Expressão Gênica , Humanos , Hipóxia , Integrinas/metabolismo , Camundongos , Camundongos Nus , MicroRNAs/metabolismo , Transplante de Neoplasias , RNA-Seq , Transdução de Sinais
9.
Med Oral Patol Oral Cir Bucal ; 25(4): e532-e540, 2020 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-32388521

RESUMO

BACKGROUND:  The aim of this histomorphometric study was to assess the bone regeneration potential of beta-tricalcium phosphate with fibronectin (ß-TCP-Fn) in critical-sized defects (CSDs) in rats calvarial, to know whether Fn improves the new bone formation in a short time scope. MATERIAL AND METHODS:  CSDs were created in 30 Sprague Dawley rats, and divided into four groups (2 or 6 weeks of healing) and type of filling (ß-TCP-Fn, ß-TCP, empty control). Variables studied were augmented area (AA), gained tissue (GT), mineralized/non mineralized bone matrix (MBM/NMT) and bone substitute (BS). RESULTS:  60 samples at 2 and six weeks were evaluated. AA was higher for treatment groups comparing to controls (p < 0.001) and significant decrease in BS area in the ß-TCP-Fn group from 2 to 6 weeks (p = 0.031). GT was higher in the ß-TCP-Fn group than in the controls expressed in % (p = 0.028) and in mm2 (p = 0.011), specially at two weeks (p=0.056). CONCLUSIONS:  Both ß-TCP biomaterials are effective as compared with bone defects left empty in maintaining the volume. GT in defects regeneration filed with ß-TCP-Fn are significantly better in short healing time when comparing with controls but not for ß-TCP used alone in rats calvarial CSDs.


Assuntos
Fibronectinas , Crânio , Animais , Regeneração Óssea , Fosfatos de Cálcio , Ratos , Ratos Sprague-Dawley
10.
Proc Natl Acad Sci U S A ; 117(19): 10131-10141, 2020 05 12.
Artigo em Inglês | MEDLINE | ID: mdl-32350144

RESUMO

Over the course of the aging process, fibroblasts lose contractility, leading to reduced connective-tissue stiffness. A promising therapeutic avenue for functional rejuvenation of connective tissue is reprogrammed fibroblast replacement, although major hurdles still remain. Toward this, we recently demonstrated that the laterally confined growth of fibroblasts on micropatterned substrates induces stem-cell-like spheroids. In this study, we embedded these partially reprogrammed spheroids in collagen-I matrices of varying densities, mimicking different three-dimensional (3D) tissue constraints. In response to such matrix constraints, these spheroids regained their fibroblastic properties and sprouted to form 3D connective-tissue networks. Interestingly, we found that these differentiated fibroblasts exhibit reduced DNA damage, enhanced cytoskeletal gene expression, and actomyosin contractility. In addition, the rejuvenated fibroblasts show increased matrix protein (fibronectin and laminin) deposition and collagen remodeling compared to the parental fibroblast tissue network. Furthermore, we show that the partially reprogrammed cells have comparatively open chromatin compaction states and may be more poised to redifferentiate into contractile fibroblasts in 3D-collagen matrix. Collectively, our results highlight efficient fibroblast rejuvenation through laterally confined reprogramming, which has important implications in regenerative medicine.


Assuntos
Diferenciação Celular , Reprogramação Celular , Fibroblastos/citologia , Medicina Regenerativa , Rejuvenescimento/fisiologia , Idoso , Animais , Proliferação de Células , Células Cultivadas , Colágeno Tipo I/metabolismo , Matriz Extracelular/metabolismo , Fibroblastos/metabolismo , Fibronectinas/metabolismo , Humanos , Laminina/metabolismo , Camundongos , Células NIH 3T3 , Esferoides Celulares/citologia , Esferoides Celulares/metabolismo
11.
Cancer Sci ; 111(6): 1991-2003, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32232887

RESUMO

Alternative polyadenylation (APA), which induces shortening of the 3'-UTR, is emerging as an important feature in cancer development and progression. Nevertheless, the effects and mechanisms of APA-induced 3'-UTR shortening in nasopharyngeal carcinoma (NPC) remain largely unclear. Fibronectin type III domain containing 3B (FNDC3B) tended to use proximal polyadenylation site and produce shorter 3'-UTR according to our previous sequencing study. Herein, we found that FNDC3B with shorter 3'-UTR could escape from miRNA-mediated gene repression, and caused its increased expression in NPC. Knocking down of FNDC3B inhibited NPC cell proliferation, migration, invasion, and metastasis in vitro and in vivo. Overexpression of FNDC3B, especially those with shorter 3'-UTR, promoted NPC progression. Furthermore, the mechanism study revealed that FNDC3B could bind to and stabilize myosin heavy chain 9 (MYH9) to activate the Wnt/ß-catenin signaling pathway. In addition, MYH9 could reverse the inhibitory effects of FNDC3B knockdown in NPC. Altogether, our results suggested that the 3'-UTR shortening of FNDC3B mRNA mediated its overexpression in NPC and promoted NPC progression by targeting MYH9. This newly identified FNDC3B-MYH9-Wnt/ß-catenin axis could represent potential targets for individualized treatment in NPC.


Assuntos
Fibronectinas/metabolismo , Regulação Neoplásica da Expressão Gênica/fisiologia , Cadeias Pesadas de Miosina/metabolismo , Carcinoma Nasofaríngeo/patologia , Neoplasias Nasofaríngeas/patologia , Regiões 3' não Traduzidas , Animais , Movimento Celular/genética , Proliferação de Células/genética , Progressão da Doença , Fibronectinas/genética , Xenoenxertos , Humanos , Camundongos , MicroRNAs , Cadeias Pesadas de Miosina/genética , Carcinoma Nasofaríngeo/genética , Carcinoma Nasofaríngeo/metabolismo , Neoplasias Nasofaríngeas/genética , Neoplasias Nasofaríngeas/metabolismo , Via de Sinalização Wnt/fisiologia
12.
Invest Ophthalmol Vis Sci ; 61(3): 44, 2020 03 09.
Artigo em Inglês | MEDLINE | ID: mdl-32232343

RESUMO

Purpose: To determine the composition of extracellular matrix (ECM) proteins secreted by a conjunctival epithelial cell line and to identify components that aid conjunctival epithelial cell culture. Methods: Human conjunctival epithelial cell line (HCjE-Gi) cells were cultured in serum-free media and their ECM isolated using ammonium hydroxide. Growth characteristics were evaluated for fresh HCjE-Gi cells plated onto ECMs obtained from 3- to 28-day cell cultures. Mass spectrometry was used to characterize the ECM composition over 42 culture days. Cell adhesion and growth on pre-adsorbed fibronectin and α-2-HS-glycoprotein (α-2-HS-GP) were investigated. Results: Day 3 ECM provided the best substrate for cell growth compared to ECM obtained from 5- to 28-day cell cultures. Mass spectrometry identified a predominantly laminin 332 matrix throughout the time course, with progressive changes to matrix composition over time: proportional decreases in matrix-bound growth factors and increases in proteases. Fibronectin and α-2-HS-GP were 5- and 200-fold enriched as a proportion of the early ECM relative to the late ECM, respectively. Experiments on these proteins in isolation demonstrated that fibronectin supported rapid cell adhesion, whereas fibronectin and α-2-HS-GP both supported enhanced cell growth compared to tissue culture polystyrene. Conclusions: These data reveal α-2-HS-GP as a candidate protein to enhance the growth of conjunctival epithelial cells and raise the possibility of exploiting these findings for targeted improvement to synthetic tissue engineered conjunctival substrates.


Assuntos
Túnica Conjuntiva/metabolismo , Proteínas da Matriz Extracelular/metabolismo , alfa-2-Glicoproteína-HS/metabolismo , Biomarcadores/metabolismo , Adesão Celular/fisiologia , Contagem de Células , Linhagem Celular , Proliferação de Células/fisiologia , Túnica Conjuntiva/citologia , Meios de Cultura Livres de Soro , Células Epiteliais/metabolismo , Matriz Extracelular/metabolismo , Fibronectinas/metabolismo , Humanos , Espectrometria de Massas
13.
PLoS One ; 15(4): e0231791, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32302361

RESUMO

Periprosthetic joint infections (PJIs) are a devastating complication that occurs in 2% of patients following joint replacement. These infections are costly and difficult to treat, often requiring multiple corrective surgeries and prolonged antimicrobial treatments. The Gram-positive bacterium Staphylococcus aureus is one of the most common causes of PJIs, and it is often resistant to a number of commonly used antimicrobials. This tolerance can be partially attributed to the ability of S. aureus to form biofilms. Biofilms associated with the surface of indwelling medical devices have been observed on components removed during chronic infection, however, the development and localization of biofilms during PJIs remains unclear. Prior studies have demonstrated that synovial fluid, in the joint cavity, promotes the development of bacterial aggregates with many biofilm-like properties, including antibiotic resistance. We anticipate these aggregates have an important role in biofilm formation and antibiotic tolerance during PJIs. Therefore, we sought to determine specifically how synovial fluid promotes aggregate formation and the impact of this process on surface attachment. Using flow cytometry and microscopy, we quantified the aggregation of various clinical S. aureus strains following exposure to purified synovial fluid components. We determined that fibrinogen and fibronectin promoted bacterial aggregation, while cell free DNA, serum albumin, and hyaluronic acid had minimal effect. To determine how synovial fluid mediated aggregation affects surface attachment, we utilized microscopy to measure bacterial attachment. Surprisingly, we found that synovial fluid significantly impeded bacterial surface attachment to a variety of materials. We conclude from this study that fibrinogen and fibronectin in synovial fluid have a crucial role in promoting bacterial aggregation and inhibiting surface adhesion during PJI. Collectively, we propose that synovial fluid may have conflicting protective roles for the host by preventing adhesion to surfaces, but by promoting bacterial aggregation is also contributing to the development of antibiotic tolerance.


Assuntos
Aderência Bacteriana/fisiologia , Biofilmes/crescimento & desenvolvimento , Staphylococcus aureus/fisiologia , Líquido Sinovial/microbiologia , Aderência Bacteriana/efeitos dos fármacos , Biofilmes/efeitos dos fármacos , Fibrinogênio/farmacologia , Fibronectinas/farmacologia , Humanos , Staphylococcus aureus/efeitos dos fármacos , Líquido Sinovial/efeitos dos fármacos , Fatores de Tempo
15.
Invest Ophthalmol Vis Sci ; 61(4): 40, 2020 04 09.
Artigo em Inglês | MEDLINE | ID: mdl-32340032

RESUMO

Purpose: To determine whether high glucose (HG) compromises internalization of lysyl oxidase (LOX) through excess binding of LOX with extracellular matrix (ECM) proteins. Methods: To determine whether HG promotes binding of LOX with ECM proteins, fibronectin (FN) and collagen IV (Coll IV), total or ECM-only proteins from rat retinal endothelial cells grown in normal (N; 5 mM) or HG (30 mM) medium were analyzed by coimmunoprecipitation and Western blot (WB). In parallel, coimmunostaining was performed to determine changes in LOX binding to FN or Coll IV. To determine the effect of HG on extracellular LOX levels, medium in which cells were grown for 1, 3, 5, and 7 days were assessed for LOX levels. Results: WB analysis using total protein showed LOX overexpression and elevated levels of LOX bound to Coll IV or FN in HG condition. Similarly, a significant increase in LOX bound to FN or Coll IV was observed in ECM-only protein. These data were supported by Z-stack confocal microscopy images from coimmunostaining. Furthermore, immunostaining performed on ECM layer revealed increased presence of LOX bound to Coll IV or FN. Additionally, when media from cells grown in HG was monitored, a maximal increase in LOX level was observed by day 3, which declined by day 7. Conclusions: Findings indicate that HG promotes binding of LOX to FN and Coll IV extracellularly that results in reduced LOX internalization, attenuation of negative feedback, and upregulation of LOX expression associated with diabetic retinopathy.


Assuntos
Retinopatia Diabética/metabolismo , Matriz Extracelular/genética , Hiperglicemia/metabolismo , Proteína-Lisina 6-Oxidase/genética , Regulação para Cima/genética , Animais , Western Blotting/métodos , Células Cultivadas , Colágeno Tipo IV/metabolismo , Diabetes Mellitus Experimental/metabolismo , Retinopatia Diabética/patologia , Modelos Animais de Doenças , Células Endoteliais , Fibronectinas/metabolismo , Regulação da Expressão Gênica/genética , Humanos , Hiperglicemia/fisiopatologia , Ligação Proteica/genética , Ratos
16.
Int J Sports Med ; 41(7): 427-442, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32252102

RESUMO

Exercise is commonly utilized for weight loss, yet research has focused less on specific modifications to adipose tissue metabolism. White adipose tissue (WAT) is the storage form of fat, whereas brown adipose tissue (BAT) is a thermogenic tissue whose uncoupling increases energy expenditure. The most established BAT activator is cold exposure, which also transforms WAT into "beige cells" that express uncoupling protein 1 (UCP1). Preliminary evidence in rodents suggests exercise elicits similar effects. The purpose of this review is to parallel and examine differences between exercise and cold exposure on BAT activation and beige induction. Like cold exposure, exercise stimulates the sympathetic nervous system and activates molecular pathways responsible for BAT/beige activation, including upregulation of BAT activation markers (UCP1, proliferator-activated receptor-gamma coactivator-1α) and stimulation of endocrine activators (fibroblast growth factor-21, irisin, and natriuretic peptides). Further, certain BAT activators are altered exclusively by exercise (interleukin-6, lactate). Markers of BAT activation increase from both cold exposure and exercise, whereas effects in WAT are compartment-specific. Stimulation of endocrine activators depends on numerous factors, including stimulus intensity and duration. Evidence of these analogous, albeit not mirrored, mechanisms is demonstrated by increases in adipose activity in rodents, while effects remain challenging to quantify in humans.


Assuntos
Tecido Adiposo Marrom/metabolismo , Temperatura Baixa , Exercício Físico/fisiologia , Termogênese , Tecido Adiposo Bege/metabolismo , Tecido Adiposo Branco/metabolismo , Animais , Fator Natriurético Atrial/metabolismo , Biomarcadores/metabolismo , Metabolismo Energético , Fatores de Crescimento de Fibroblastos/metabolismo , Fibronectinas/metabolismo , Humanos , Peptídeo Natriurético Encefálico/metabolismo , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/metabolismo , Proteína Desacopladora 1/metabolismo
17.
Adv Exp Med Biol ; 1245: 85-96, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32266654

RESUMO

Fibronectin (FN) is a large glycoprotein that plays a diverse set of biological roles. This chapter discusses the structural biology, the normal biological functions, and the molecular role of FN and its splice variants in cancer cell proliferation, metastasis, and chemoresistance. The potential role of FN in cancer imaging is discussed in detail. The chapter also discusses the future directions of basic and translational research of fibronectin in the context of the tumor microenvironment and its role in tumor biology.


Assuntos
Fibronectinas , Neoplasias , Microambiente Tumoral , Proliferação de Células/genética , Fibronectinas/genética , Humanos , Neoplasias/genética , Neoplasias/patologia
18.
Am J Physiol Renal Physiol ; 318(4): F1017-F1029, 2020 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-32116017

RESUMO

Diabetic kidney disease is a worldwide epidemic, and therapies are incomplete. Clinical data suggest that improved renal outcomes by Na+-glucose cotransporter 2 inhibitor (SGLT2i) are partly beyond their antihyperglycemic effects; however, the mechanisms are still elusive. Here, we investigated the effect of the SGLT2i dapagliflozin (DAPA) in the prevention of elevated O-GlcNAcylation and tubular hypoxia as contributors of renal fibrosis. Type 1 diabetes was induced by streptozotocin in adult male Wistar rats. After the onset of diabetes, rats were treated for 6 wk with DAPA or DAPA combined with losartan (LOS). The effect of hyperglycemia was tested in HK-2 cells kept under normal or high glucose conditions. To test the effect of hypoxia, cells were kept in 1% O2 for 2 h. Cells were treated with DAPA or DAPA combined with LOS. DAPA slowed the loss of renal function, mitigated renal tubular injury markers (kidney injury molecule-1 and neutrophil gelatinase-associated lipocalin), and reduced tubulointerstitial fibrosis. DAPA diminished high glucose-induced protein O-GlcNAcylation and moderated the tubular response to hypoxia through the hypoxia-inducible factor pathway. DAPA alone was as effective as combined treatment with LOS in all outcome parameters. These data highlight the role of ameliorated O-GlcNAcylation and diminished tubular hypoxia as important benefits of SGLT2i treatment. Our results support the link between glucose toxicity, tubular hypoxia, and fibrosis, a vicious trio that could be targeted by SGLT2i in kidney diseases of other origins as well.


Assuntos
Compostos Benzidrílicos/farmacologia , Diabetes Mellitus Experimental/tratamento farmacológico , Diabetes Mellitus Tipo 1/tratamento farmacológico , Nefropatias Diabéticas/prevenção & controle , Glucosídeos/farmacologia , Glicosilação/efeitos dos fármacos , Túbulos Renais Proximais/efeitos dos fármacos , Inibidores do Transportador 2 de Sódio-Glicose/farmacologia , Animais , Glicemia/efeitos dos fármacos , Glicemia/metabolismo , Hipóxia Celular , Linhagem Celular , Colágeno/metabolismo , Diabetes Mellitus Experimental/induzido quimicamente , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Tipo 1/induzido quimicamente , Diabetes Mellitus Tipo 1/metabolismo , Nefropatias Diabéticas/etiologia , Nefropatias Diabéticas/metabolismo , Nefropatias Diabéticas/patologia , Fibronectinas/metabolismo , Fibrose , Humanos , Túbulos Renais Proximais/metabolismo , Túbulos Renais Proximais/patologia , Masculino , Ratos Wistar , Estreptozocina
19.
Toxicol Appl Pharmacol ; 394: 114953, 2020 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-32165127

RESUMO

Exercise training is one of the major non-pharmacological treatments for hypertension. However, the central mechanism by which exercise training attenuates the hypertensive responses remains unclear. Irisin is a muscle-secreted cytokine derived from fibronectin type III domain containing 5 (FNDC5) that will be released into the circulation during exercise. We hypothesized that irisin may play a role in the blood pressure regulation by exercise. To examine the hypothesis, our study investigated the effect of irisin on hypertension and its central mechanism. The study was performed in spontaneously hypertensive rats (SHRs) and normotensive Wistar-Kyoto rats. We found that intravenous injection of irisin effectively reduced blood pressure, plasma norepinephrine, paraventricular nucleus (PVN) levels of neuronal activation, oxidative stress and inflammation in SHRs. Moreover, irisin activated nuclear factor E2-related factor-2 (Nrf2) and restored the imbalance of neurotransmitters in the PVN. Our study also found PVN knockdown of Nrf2 abolished the protective effects of irisin on hypertension. These findings demonstrate irisin can improve hypertension via Nrf2-mediated antioxidant in the PVN.


Assuntos
Anti-Hipertensivos/farmacologia , Fibronectinas/farmacologia , Fator 2 Relacionado a NF-E2/efeitos dos fármacos , Núcleo Hipotalâmico Paraventricular/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Animais , Antioxidantes/metabolismo , Citocinas/metabolismo , Neurotransmissores/metabolismo , Norepinefrina/sangue , Estresse Oxidativo/efeitos dos fármacos , Esforço Físico , Ratos , Ratos Endogâmicos SHR , Ratos Endogâmicos WKY
20.
Nat Commun ; 11(1): 1571, 2020 03 26.
Artigo em Inglês | MEDLINE | ID: mdl-32218432

RESUMO

Estrogens and progesterone control breast development and carcinogenesis via their cognate receptors expressed in a subset of luminal cells in the mammary epithelium. How they control the extracellular matrix, important to breast physiology and tumorigenesis, remains unclear. Here we report that both hormones induce the secreted protease Adamts18 in myoepithelial cells by controlling Wnt4 expression with consequent paracrine canonical Wnt signaling activation. Adamts18 is required for stem cell activation, has multiple binding partners in the basement membrane and interacts genetically with the basal membrane-specific proteoglycan, Col18a1, pointing to the basement membrane as part of the stem cell niche. In vitro, ADAMTS18 cleaves fibronectin; in vivo, Adamts18 deletion causes increased collagen deposition during puberty, which results in impaired Hippo signaling and reduced Fgfr2 expression both of which control stem cell function. Thus, Adamts18 links luminal hormone receptor signaling to basement membrane remodeling and stem cell activation.


Assuntos
Proteínas ADAMTS/metabolismo , Hormônios/farmacologia , Glândulas Mamárias Animais/citologia , Nicho de Células-Tronco , Células-Tronco/metabolismo , Proteínas ADAMTS/deficiência , Proteínas ADAMTS/genética , Animais , Antígenos CD/metabolismo , Linhagem Celular , Autorrenovação Celular/efeitos dos fármacos , Epitélio/metabolismo , Matriz Extracelular/efeitos dos fármacos , Matriz Extracelular/metabolismo , Feminino , Fibronectinas/metabolismo , Glicoproteínas/metabolismo , Humanos , Camundongos Endogâmicos C57BL , Modelos Biológicos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores de Progesterona/metabolismo , Regeneração/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Nicho de Células-Tronco/efeitos dos fármacos , Células-Tronco/citologia , Células-Tronco/efeitos dos fármacos , Transcrição Genética/efeitos dos fármacos
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