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1.
Braz Oral Res ; 33: e042, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31508725

RESUMO

This study evaluated the cytotoxicity and biocompatibility of a new bioceramic endodontic sealer (i.e., Sealer Plus BC) in comparison with those of MTA Fillapex and AH Plus. L929 fibroblasts were cultured and Alamar Blue was used to evaluate cell viability of diluted extracts (1:50, 1:100, and 1:200) from each sealer at 24 h. Polyethylene tubes that were filled with material or empty (as a control) were implanted in the subcutaneous tissue of rats. The rats were killed after 7 and 30 d (n = 8), and the tubes were removed for histological analysis. Parametric data was analyzed using a one-way ANOVA test, and nonparametric data was analyzed via the Kruskal-Wallis test followed by the Dunn test (p < 0.05). A reduction in cell viability was observed in the extracts that were more diluted for Sealer Plus BC when compared to that of Control and AH Plus (p < 0.05). However, the 1:50 dilution of the Sealer Plus BC was similar to that of the Control (p > 0.05). Conversely, more diluted extracts of MTA Fillapex (1:200) and AH Plus (1:100 and 1:200) were similar to the Control (p > 0.05). Histological analysis performed at 7 d did not indicate any significant difference between tissue response for all materials, and the fibrous capsule was thick (p > 0.05). At 30 d, Sealer Plus BC was similar to the Control (p > 0.05) and MTA Fillapex and AH Plus exhibited greater inflammation than the Control (p < 0.05). The fibrous capsule was thin for the Control and for most specimens of Sealer Plus BC and AH Plus. Thus, Sealer Plus BC is biocompatible when compared to MTA Fillapex and AH Plus, and it is less cytotoxic when less-diluted extracts are used.


Assuntos
Cimentos para Ossos/química , Hidróxido de Cálcio/química , Cerâmica/química , Materiais Restauradores do Canal Radicular/química , Compostos de Alumínio/química , Animais , Materiais Biocompatíveis , Cimentos para Ossos/farmacologia , Cimentos para Ossos/toxicidade , Compostos de Cálcio/química , Hidróxido de Cálcio/farmacologia , Hidróxido de Cálcio/toxicidade , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas/efeitos dos fármacos , Combinação de Medicamentos , Resinas Epóxi/química , Fibroblastos/efeitos dos fármacos , Técnicas In Vitro , Inflamação , Masculino , Teste de Materiais , Óxidos/química , Ratos Wistar , Materiais Restauradores do Canal Radicular/toxicidade , Silicatos/química , Tela Subcutânea/patologia
2.
Chem Pharm Bull (Tokyo) ; 67(8): 849-854, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31366834

RESUMO

Regenerative therapy with keratinocyte growth factor (KGF) is a novel therapeutic approach for treatment of chronic wounds. However, KGF cannot be used directly to the wound site due to its physicochemical instability. In previous study, sacran, a natural megamolecular polysaccharide, showed potential properties as a biomaterial for hydrogel film in wound healing. In this study, we fabricated sacran hydrogel film containing KGF (Sac/KGF-HF) and evaluated the effects of Sac/KGF-HF on fibroblasts migration and re-epithelialization process. We successfully prepared a homogenous and -amorphous Sac/KGF-HF by a casting method. In addition, Sac/KGF-HF had a high swelling ratio and flexibility. Sac/KGF-HF promoted a migration process of NIH3T3 cells and improved wound healing ability in mice with a percentage of wound closure reaching 90.4% at 9 d. Interestingly, the addition of KGF in Sac-HF considerably increased the number of epithelial cells compared to control, which is important in the re-epithelialization process. It could be concluded that KGF in Sac-HF has the potential for promoting Sac-HF abilities in wound healing process.


Assuntos
Fator 7 de Crescimento de Fibroblastos/metabolismo , Fibroblastos/efeitos dos fármacos , Metilgalactosídeos/farmacologia , Polissacarídeos/farmacologia , Cicatrização/efeitos dos fármacos , Animais , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Fator 7 de Crescimento de Fibroblastos/química , Metilgalactosídeos/química , Camundongos , Camundongos Endogâmicos BALB C , Células NIH 3T3 , Polissacarídeos/química
3.
Toxicol Lett ; 315: 87-95, 2019 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-31425726

RESUMO

Prenatal alcohol exposure (PAE) is often associated with congenital heart defects, most commonly septal, valvular, and great vessel defects. However, there have been no known studies on whether PAE affects the resulting fibroblast population after development, and whether this has any consequences in the postnatal period. Our previous study focused on the effects of PAE on the postnatal fibroblast population, which translated into changes in cardiac extracellular matrix (ECM) composition and cardiac function in the neonatal heart. Moreover, our lab has previously demonstrated that alcohol-induced fibrosis is mediated by oxidative stress mechanisms in adult rat hearts following chronic alcohol exposure. Thus, we hypothesize that PAE alters cardiac ECM composition that persists into the postnatal period, leading to cardiac dysfunction, and these effects are prevented by antioxidant treatment. To investigate these effects, pregnant mice were intraperitoneally injected with 2.9 g EtOH/kg body weight on gestation days 6.75 and 7.25. Controls were injected with vehicle saline. Randomly selected dams in both groups were then treated with 100 mg/kg body weight of the antioxidant N-acetylcysteine (NAC) immediately after EtOH or vehicle administration. Left ventricular (LV) chamber dimension and function were assessed in sedated animals on neonatal day 5 using echocardiography. Ejection fraction decreased in the PAE group. NAC treatment prevented this depression of systolic function in PAE neonates. Hearts were analyzed for expression of fibroblast activation markers. Alpha smooth muscle actin (α-SMA) increased in PAE neonatal hearts, and this increase was prevented by NAC treatment. In PAE pups, collagen I decreased, but collagen III expression increased compared to saline animals; the overall collagen I/III ratio significantly decreased. When PAE mice were treated with NAC, collagen I/III ratio did not change. Overall, our data demonstrate that prenatal alcohol exposure produces changes in collagen subtype in neonatal cardiac ECM and a decline in systolic function, and these adverse effects were prevented by NAC treatment.


Assuntos
Acetilcisteína/farmacologia , Alcoolismo/fisiopatologia , Proliferação de Células/efeitos dos fármacos , Colágeno Tipo I/metabolismo , Vasos Coronários/química , Etanol/toxicidade , Fibroblastos/efeitos dos fármacos , Efeitos Tardios da Exposição Pré-Natal/induzido quimicamente , Animais , Animais Recém-Nascidos , Modelos Animais de Doenças , Feminino , Camundongos , Gravidez
4.
Int J Nanomedicine ; 14: 5033-5050, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31371945

RESUMO

Background: Repairs to deep skin wounds continue to be a difficult issue in clinical practice. A promising approach is to fabricate full-thickness skin substitutes with functions closely similar to those of the natural tissue. For many years, a three-dimensional (3D) collagen hydrogel has been considered to provide a physiological 3D environment for co-cultivation of skin fibroblasts and keratinocytes. This collagen hydrogel is frequently used for fabricating tissue-engineered skin analogues with fibroblasts embedded inside the hydrogel and keratinocytes cultivated on its surface. Despite its unique biological properties, the collagen hydrogel has insufficient stiffness, with a tendency to collapse under the traction forces generated by the embedded cells. Methods: The aim of our study was to develop a two-layer skin construct consisting of a collagen hydrogel reinforced by a nanofibrous poly-L-lactide (PLLA) membrane pre-seeded with fibroblasts. The attractiveness of the membrane for dermal fibroblasts was enhanced by coating it with a thin nanofibrous fibrin mesh. Results: The fibrin mesh promoted the adhesion, proliferation and migration of the fibroblasts upwards into the collagen hydrogel. Moreover, the fibroblasts spontaneously migrating into the collagen hydrogel showed a lower tendency to contract and shrink the hydrogel by their traction forces. The surface of the collagen was seeded with human dermal keratinocytes. The keratinocytes were able to form a basal layer of highly mitotically-active cells, and a suprabasal layer. Conclusion: The two-layer skin construct based on collagen hydrogel with spontaneously immigrated fibroblasts and reinforced by a fibrin-coated nanofibrous membrane seems to be promising for the construction of full-thickness skin substitute.


Assuntos
Colágeno/farmacologia , Fibrina/farmacologia , Hidrogéis/farmacologia , Membranas Artificiais , Nanofibras/química , Poliésteres/farmacologia , Pele Artificial , Animais , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Derme/citologia , Fibroblastos/citologia , Fibroblastos/efeitos dos fármacos , Humanos , Recém-Nascido , Queratinócitos/citologia , Queratinócitos/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Ratos
5.
Biol Res ; 52(1): 45, 2019 Aug 19.
Artigo em Inglês | MEDLINE | ID: mdl-31426853

RESUMO

BACKGROUND: Resveratrol was reported to trigger the apoptosis of fibroblast-like synoviocytes in adjuvant arthritis rats but the subcellular mechanism remains unclear. Since ER stress, mitochondrial dysfunction and oxidative stress were involved in the effects of resveratrol with imbalance of calcium bio-transmission, store operated calcium entry (SOCE), a novel intracellular calcium regulatory pathway, may also participate in this process. RESULTS: In the present study, Resveratrol was found to suppress ORAI1 expression of a dose dependent manner while have no evident effects on STIM1 expressive level. Besides, resveratrol had no effects on ATP or TG induced calcium depletion but present partly dose-dependent suppression of SOCE. On the one hand, microinjection of ORAI1 overexpressed vector in sick toe partly counteracted the therapeutic effects of resveratrol on adjuvant arthritis and serum inflammatory cytokine including IL-1, IL-6, IL-8, IL-10 and TNF-α. On the other hand, ORAI1 SiRNA injection provided slight relief to adjuvant arthritis in rats. In addition, ORAI1 overexpression partly diminished the alleviation of hemogram abnormality induced by adjuvant arthritis after resveratrol treatment while ORAI1 knockdown presented mild resveratrol-like effect on hemogram in rats model. CONCLUSION: These results indicated that resveratrol reduced store-operated Ca2+ entry and enhanced the apoptosis of fibroblast-like synoviocytes in adjuvant arthritis rats model via targeting ORAI1-STIM1 complex, providing a theoretical basis for ORAI1 targeted therapy in future treatment with resveratrol on rheumatoid arthritis.


Assuntos
Apoptose/efeitos dos fármacos , Artrite Experimental/fisiopatologia , Canais de Cálcio/efeitos dos fármacos , Fibroblastos/efeitos dos fármacos , Proteína ORAI1/efeitos dos fármacos , Resveratrol/farmacologia , Molécula 1 de Interação Estromal/efeitos dos fármacos , Sinoviócitos/efeitos dos fármacos , Animais , Canais de Cálcio/fisiologia , Mitocôndrias/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Ratos , Resveratrol/administração & dosagem
6.
Medicine (Baltimore) ; 98(34): e16952, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31441893

RESUMO

BACKGROUND: Botulinum toxin type A (BoNTA) is known to prevent fibroblast proliferation and expression of transforming growth factor beta 1 (TGF-ß1). It also induces temporary muscle paralysis and decreases tension vectors. Fibroblasts induce scar contracture and hypertrophy by producing collagen fibers in wound healing processes. The aim of this study is to identify the effect of BoNTA on the scar formation. METHODS: Forty-five patients with forehead laceration were enrolled in this study and randomized into 2 groups with or without injection of BoNTA. When the patients presented to the clinic to remove the stitches, BoNTA was injected to the BoNTA group with 24 patients and saline was injected to the control group with 21 patients. The BoNTA was injected on dermal layer with 5 IU/cm. After that, follow-up was done in 1, 3, and 6 months. The scars were analyzed with the patient and observer scar assessment scale, Stony Brook scar evaluation scales (SBSESs), and visual analog scale (VAS) and analyzed with independent T-test, along with clinical photographs, cutometer, and biopsies. RESULTS: In all scar scales, the scores changed into favorable direction in both groups and the changes were larger in BoNTA group compared with the control group. On SBSES and VAS, better improvements on BoNTA group showed statistical significance. Skin biopsy showed less collagen deposition on dermal layer in BoNTA group. CONCLUSION: Improvement of aesthetic, functional, and emotional aspect of the scar formation in the groups treated with BoNTA was illustrated. The application of BoNTA may be expanded to prevent hypertrophic scar after trauma, burns, or operations.


Assuntos
Toxinas Botulínicas Tipo A/administração & dosagem , Cicatriz Hipertrófica/prevenção & controle , Testa/lesões , Lacerações/terapia , Fármacos Neuromusculares/administração & dosagem , Adulto , Toxinas Botulínicas Tipo A/farmacologia , Método Duplo-Cego , Feminino , Fibroblastos/efeitos dos fármacos , Humanos , Injeções , Masculino , Pessoa de Meia-Idade , Fármacos Neuromusculares/farmacologia , Estudos Prospectivos , Técnicas de Sutura , Resultado do Tratamento , Escala Visual Analógica , Adulto Jovem
7.
Chem Biol Interact ; 310: 108747, 2019 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-31301289

RESUMO

Pathological scarring is an intractable problem for both patients and clinicians. A major obstacle for the development of scar remediation therapies is the paucity of suitable in vivo and in vitro models. The "Scar-in-a-jar" model was previously established by our colleagues based on the principle of "Macromolecular crowding". This has been demonstrated to be an extracellular matrix-rich in vitro model offering a novel tool for studies related to the extracellular matrix. In the study reported herein, we have optimised this approach to model human dermal fibroblasts derived from hypertrophic tissues. This optimised in vitro model has been found to hold similar properties, such as increased collagen I, interleukins and transforming growth factor beta-1 expression, compared to that observed in hypertrophic scar tissue in vivo. In addition, Shikonin has been previously demonstrated to hold potential as a novel hypertrophic scar treatment due to its apoptosis-inducing property on hypertrophic scar fibroblasts. Other Shikonin analogues have also been reported to hold apoptosis-inducing properties in various cancer cell lines, however, the effects of these analogues on hypertrophic scar-related cells are unknown. We therefore evaluated the effects of Shikonin and its analogues on hypertrophic scar-derived human fibroblasts using the optimised "Macromolecular crowding" model. Our data indicates that Shikonin and Naphthazarin are the most effective molecules compared to related naphthoquinones. The data generated from the study offers a novel in vitro collagen-rich model of hypertrophic scar tissue. It also provides further evidences supporting the use of Shikonin and Naphthazarin as potential treatments for hypertrophic scars.


Assuntos
Cicatriz/tratamento farmacológico , Naftoquinonas/uso terapêutico , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular , Cicatriz/patologia , Cicatriz Hipertrófica/tratamento farmacológico , Colágeno/metabolismo , Matriz Extracelular/metabolismo , Fibroblastos/efeitos dos fármacos , Humanos , Modelos Biológicos , Pele/patologia
8.
J Photochem Photobiol B ; 197: 111539, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31301638

RESUMO

Treatment of burn injury is clinically challenging one, therefore several steps and noteworthy approaches have been taken to improve wound mechanisms. Citrus pectin plays a stabilizing agent to synthesis of ZnO nanoparticles (ZnO NPs). The present study is focused on ZnO loaded collagen/chitosan nanofibrous were synthesized by electrospinning method using ZnO NPs. The chemical structure, phase purity and morphological observation were investigated under spectroscopic and mircoscopic techniques and demonstrated their suitable properties as a wound healing material. In addition, that prepared nanoparticles loaded biopolymeric fibrous nanomaterial showed suitable antibacterial activity against S. aureus and E. coli bacterial pathogens and also in vitro studies was confirmed the enhanced proliferation, cell viability and biocompatibility. In vitro evaluations have been exhibited acceptable cell proliferation is observed throughout the ZnO loaded Coll/CS nanofibrous within 3 days, which was comparable to the control material. In vivo wound healing ability was monitored on the rat wound experimental model. From the in vivo observations, revealed that the loaded of ZnO NPs with Coll/CS nanofibrous can effectively quicken wound healing mechanism, expressed in the initial stage healing process. These results suggest that ZnO loaded collagen/chitosan nanofibrous is a potential candidate for wound healing applications with enhanced biological properties.


Assuntos
Queimaduras/patologia , Quitosana/química , Colágeno/química , Nanopartículas Metálicas/química , Nanofibras/química , Animais , Antibacterianos/química , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Bacteriemia/prevenção & controle , Queimaduras/veterinária , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Escherichia coli/efeitos dos fármacos , Fibroblastos/citologia , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Humanos , Nanofibras/uso terapêutico , Nanofibras/toxicidade , Ratos , Pele/efeitos dos fármacos , Pele/patologia , Staphylococcus aureus/efeitos dos fármacos , Cicatrização/efeitos dos fármacos , Óxido de Zinco/química
9.
Anticancer Res ; 39(7): 3543-3551, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31262878

RESUMO

BACKGROUND/AIM: Both bevacizumab (BEV) and soluble fms-like tyrosine kinase-1 (sFlt-1) have demonstrated anti-angiogenic effects, thereby causing hypertension and proteinuria. We hypothesized that anti-preeclamptic drugs that combat the action of sFlt-1 may reduce BEV's anti-tumor efficacy. MATERIALS AND METHODS: 3D co-cultured human mini-tumors consisting of endothelial cells, fibroblasts, and cancer cells were developed. The influence of anti-preeclamptic drugs and BEV on the invasion of mini-tumors embedded in collagen gel was evaluated. RESULTS: Mini-tumor spheroids that contained MDA-MB-231 cells showed higher invasion ability than spheroids with A549. Among the six anti-preeclamptic drugs investigated, only nicorandil enhanced the invasion of mini-tumors and inhibited the action of BEV. Glibenclamide, an ATP-sensitive potassium channel inhibitor, completely quenched the action of nicorandil on mini-tumors. CONCLUSION: In the human mini-tumor model, nicorandil aggravated the invasion of mini-tumors. These data raise the possibility that concomitant use of nicorandil counteracts the efficacy of BEV therapy.


Assuntos
Inibidores da Angiogênese/farmacologia , Anti-Hipertensivos/farmacologia , Bevacizumab/farmacologia , Esferoides Celulares/efeitos dos fármacos , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Técnicas de Cocultura , Feminino , Fibroblastos/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Humanos , Nicorandil/farmacologia , Pré-Eclâmpsia , Gravidez , Esferoides Celulares/fisiologia , Fator A de Crescimento do Endotélio Vascular/farmacologia
10.
Aquat Toxicol ; 213: 105229, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31255889

RESUMO

Although the global use of the 1,1,1-trichloro-2,2-bis (4-chlorophenyl) ethane (p,p'-DDT) has been prohibited, its persistence in the environment has caused long-lasting exposure on marine mammals. Our previous studies revealed exceedingly high residue levels of DDTs in Indo-Pacific humpback dolphins (Sousa chinensis) from the Pearl River Estuary region, China. However, the molecular mechanisms of p,p'-DDT toxicity on the dolphin are largely unknown. This study conducted the first cytotoxicity effect exploration of p,p'-DDT on the dolphin skin fibroblasts (ScSFs) to enhance the understanding of the cellular and molecular regulation impacts. ScSF cells were exposed to p,p'-DDT (28∼168 µM) for 24, 48 and 72 h. The exposure remarkably decreased viability of ScSF cells, possibly due to the synergetic effects of cell cycle arrest and apoptosis via DNA damage and mitochondria dysfunction. The DNA damage and mitochondria dysfunction were likely triggered by an increase of cellular reactive oxygen species (ROS), alteration in mitochondrial membrane potential, reduction in the cellular ATP levels, decreased expression of the genes CDK1, CDK4, cyclin B1, cyclin D1 and apoptosis regulator Bcl-2, release of cytochrome c, and activation of caspase-3, caspase-8 and caspase-9. Moreover, caspase inhibitor displayed protective activity against p,p'-DDT-induced apoptosis, indicating that caspases played a central role in p,p'-DDT-triggered apoptosis in the ScSF cells. We hypothesize apoptosis likely plays a minor role in cytocidal effects induced by p,p'-DDT exposure, but the mechanisms remain unclear. Overall, this research provides new evidence of the cytotoxic mechanisms underlying p,p'-DDT exposure on humpback dolphin skin cells, and suggests that p,p'-DDT contamination is one of key health concern issues for the protection of this marine mammal.


Assuntos
Apoptose/efeitos dos fármacos , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , DDT/toxicidade , Golfinhos/metabolismo , Exposição Ambiental , Fibroblastos/citologia , Mitocôndrias/metabolismo , Pele/citologia , Animais , Caspases/metabolismo , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Poluentes Químicos da Água/toxicidade
11.
Life Sci ; 231: 116674, 2019 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-31344427

RESUMO

Hypertrophic scar formation is a fibroproliferative disorder caused by abnormal wound healing. At present, there are limited treatment strategies for hypertrophic scars. In this study, we identified an endogenous peptide, LYENRL, through peptidomics screening that is downregulated in scar skin tissues. The peptide exhibited concentration dependent inhibitory effects on the proliferation, migration and extracellular matrix (ECM) production of scar fibroblasts. By eukaryotic transcriptome sequencing analysis, we noted that LYENRL downregulated gene sets in scar fibroblasts were associated with the transforming growth factor-ß (TGF-ß) signaling pathway. Further experiments revealed that LYENRL was able to inhibit the activation of TGF-ß1/Smad signaling and TGF-ß1-induced activation of scar fibroblasts at the source by blocking the binding of AP-1 to the corresponding region of the Tgfb1 promoter, which in turn inhibited gene expression of Tgfb1. Taken together, we concluded that the effects of LYENRL on scar fibroblasts make it a potential peptide drug for hypertrophic scar treatment.


Assuntos
Cicatriz Hipertrófica/metabolismo , Cicatriz Hipertrófica/patologia , Peptídeos/farmacologia , Actinas/metabolismo , Linhagem Celular , Células Cultivadas , Regulação para Baixo/efeitos dos fármacos , Matriz Extracelular/metabolismo , Feminino , Fibroblastos/efeitos dos fármacos , Fibroblastos/fisiologia , Humanos , Peptídeos/metabolismo , Cultura Primária de Células , Transdução de Sinais/efeitos dos fármacos , Pele/metabolismo , Proteínas Smad/metabolismo , Proteínas Smad/fisiologia , Proteína Smad3/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Fator de Crescimento Transformador beta1/metabolismo , Fator de Crescimento Transformador beta1/fisiologia
12.
DNA Cell Biol ; 38(9): 969-981, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31355672

RESUMO

Analysis of gene expression can be challenging, especially if it involves genetically diverse populations that exhibit high variation in their individual expression profile. Despite this variation, it is conceivable that in the same individuals a high degree of coordination is maintained between transcripts that belong to the same signaling modules and are associated with related biological functions. To explore this further, we calculated the correlation in the expression levels between each of ATF4, CHOP (DDIT3), GRP94, DNAJB9 (ERdj4), DNAJ3C (P58IPK), and HSPA5 (BiP/GRP78) with the whole transcriptome in primary fibroblasts from deer mice following induction of endoplasmic reticulum (ER) stress. Since these genes are associated with different transducers of the unfolded protein response (UPR), we postulated that their profile, in terms of correlation of transcripts, reflects distinct UPR branches engaged, and therefore different biological processes. Standard gene ontology analysis was able to predict major functions associated with the corresponding transcript, and of the UPR arm related to that, namely regulation of the apoptotic response by ATF4 (PERK arm) and the ER stress-associated degradation for GRP94 (IRE1). BiP, being a global regulator of the UPR, was associated with activation of ER stress in a rather global manner. Pairwise comparison in the correlation coefficients for these genes' associated transcriptome showed the relevance of selected genes in terms of expression profiles. Conventional assessment of differential gene expression was incapable of providing meaningful information and pointed only to a generic association with stress. Collectively, this approach suggests that by evaluating the degree of coordination in gene expression, in genetically diverse biological specimens, may be useful in assigning genes in transcriptome networks, and more importantly in linking signaling nodules to specific biological functions and processes.


Assuntos
Estresse do Retículo Endoplasmático/genética , Animais , Células Cultivadas , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Peromyscus , Transcriptoma , Tunicamicina/farmacologia
13.
J Toxicol Sci ; 44(6): 425-433, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31168029

RESUMO

Cardiac fibroblasts (CFs) could be activated after myocardial infarction (MI). Thus, it is necessary to explore effective drugs to suppress the activation of CFs following MI. This study was designed to investigate the impacts of ellagic acid on CFs and the underlying mechanisms. The expression of histone deacetylases (HDACs) and fibrosis-related genes was detected by qRT-PCR and western blot. The Masson's Trichrome Staining assay was used to evaluate the area of cardiac fibrosis. The proliferation and migration of CFs were measured by CCK8 Kit and Transwell assay, respectively. Our results showed that ellagic acid significantly reduced protein expression of HDAC1, mRNA expression of collagen I, collagen III, MMP-2 and MMP-9 and the area of cardiac fibrosis in MI rats. In Ang II-stimulated CFs, ellagic acid (60 µmol/L) decreased the protein expression of HDAC1, collagen I, collagen III, MMP-2 and MMP-9, and inhibited cell proliferation and migration. Further, HDAC1 over-expression reversed the inhibitor effects of ellagic acid on proteins expression (collagen I, collagen III, MMP-2 and MMP-9) and proliferation and migration of CFs. The present results suggested that ellagic acid suppressed proliferation and migration of CFs by down-regulating expression of HDAC1.


Assuntos
Cardiotônicos/farmacologia , Ácido Elágico/farmacologia , Fibroblastos/efeitos dos fármacos , Angiotensina II , Animais , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Colágeno Tipo I/genética , Colágeno Tipo III/genética , Regulação para Baixo , Fibroblastos/fisiologia , Regulação da Expressão Gênica/efeitos dos fármacos , Histona Desacetilase 1/genética , Masculino , Metaloproteinase 2 da Matriz/genética , Metaloproteinase 9 da Matriz/genética , Miocárdio/citologia , Ratos Sprague-Dawley
14.
J Toxicol Sci ; 44(6): 435-440, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31168030

RESUMO

Fas/CD95 plays a pivotal role in T cell-mediated cytotoxicity. Accumulating evidence has suggested that resistance to Fas-mediated apoptosis contributes to the escape of cancer cells from immune destruction, and allows to undergo proliferation and outgrowth of cancer cells. In this study, we found that the anti-cancer drug gefitinib, a tyrosine kinase inhibitor of epidermal growth factor receptor (EGFR), has an ability to enhance Fas-mediated cytotoxicity. In the presence of nontoxic concentrations of gefitinib, Fas-induced activation of caspase-8 and subsequent apoptosis was dramatically promoted, suggesting that gefitinib increases the sensitivity to Fas-mediated apoptosis. Interestingly, the effects of gefitinib were observed in EGFR or p53 knockout (KO) cells. These observations indicate that both EGFR and p53 are dispensable for the enhancement. On the other hand, gefitinib clearly downregulated heat shock protein 70 (HSP70) as previously reported. Considering that HSP70 contributes to protection of cells against Fas-mediated apoptosis, gefitinib may increase the sensitivity to Fas-mediated apoptosis by downregulating HSP70. Thus, our findings reveal novel properties of gefitinib, which may provide insight into the alternative therapeutic approaches of gefitinib for Fas-resistant tumors.


Assuntos
Antineoplásicos/farmacologia , Caspase 8/metabolismo , Proteína Ligante Fas/fisiologia , Gefitinibe/farmacologia , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Fibroblastos/efeitos dos fármacos , Fibroblastos/fisiologia , Proteínas de Choque Térmico HSP72/metabolismo , Humanos , Camundongos
15.
Phytother Res ; 33(6): 1726-1735, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31155798

RESUMO

Rheumatoid arthritis (RA) is a complex chronic inflammatory disease that is associated with the aberrant activation of fibroblast-like synoviocytes (FLS). Kaempferitrin is a natural flavonoid glycoside that possesses anti-inflammatory bioactivity. However, the effect of kaempferitrin on RA has not yet been revealed. The aim of the present study was to investigate the effect of kaempferitrin on human RA-FLS MH7A cell line. We found that kaempferitrin inhibited proliferation and induced apoptosis of MH7A cells. Kaempferitrin decreased the levels of interleukin (IL)-1ß, IL-6, tumor necrosis factor (TNF)-α, matrix metalloproteinase (MMP)-1, and MMP-3 in MH7A cells. Moreover, kaempferitrin blocked the activation of nuclear factor-κB (NF-κB) and protein kinase B (Akt)/mammalian target of rapamycin (mTOR) pathways. Furthermore, treatment with kaempferitrin decreased paw thickness and arthritis scores, and reduced the serum levels of IL-1ß, IL-6, and TNF-α in a collagen-induced arthritis mouse model. In conclusion, kaempferitrin inhibited cell proliferation, induced cell apoptosis, and ameliorated inflammation of RA-FLS by suppressing the NF-κB and Akt/mTOR pathways.


Assuntos
Apoptose/efeitos dos fármacos , Artrite Reumatoide/patologia , Proliferação de Células/efeitos dos fármacos , Inflamação/prevenção & controle , Quempferóis/farmacologia , Sinoviócitos/efeitos dos fármacos , Animais , Artrite Experimental/metabolismo , Artrite Experimental/patologia , Artrite Experimental/prevenção & controle , Artrite Reumatoide/metabolismo , Artrite Reumatoide/prevenção & controle , Células Cultivadas , Feminino , Fibroblastos/efeitos dos fármacos , Fibroblastos/fisiologia , Humanos , Inflamação/metabolismo , Inflamação/patologia , Interleucina-1beta/metabolismo , Metaloproteinase 3 da Matriz , Camundongos , Camundongos Endogâmicos DBA , NF-kappa B/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Sinoviócitos/patologia , Sinoviócitos/fisiologia , Fator de Necrose Tumoral alfa/metabolismo
16.
Eur J Med Chem ; 178: 141-153, 2019 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-31177074

RESUMO

Anaplastic lymphoma kinase (ALK) is a receptor tyrosine kinase responsible for development of various tumor types. In this study, we synthesized a series of novel 2,4-diarylaminopyrimidine derivatives possessing a unique N-(3-pyridinylmethyl)urea moiety as ALK inhibitors. The most promising analog 5m bearing a 3-methoxy-4-morpholinophenyl substituent significantly inhibited proliferation of ALK positive H3122 and Karpas-299 cells with IC50 values about 10 nM, which were comparable with positive control LDK378. Compound 5m suppressed phosphorylation of ALK and its downstream proteins, and showed low cytotoxicity on normal human primary fibroblast cells (BJ cells). The binding mode of 5m was proposed by docking simulation, which explains the important role of N-(3-pyridinylmethyl)urea moiety. Furthermore, compound 5m exhibited favorable liver microsomal stability and significant efficacy in H3122 xenograft mice model. Interestingly, compound 5m also showed broader anti-proliferative activity on other human tumor cell lines, which was different from other ALK inhibitors.


Assuntos
Quinase do Linfoma Anaplásico/antagonistas & inibidores , Antineoplásicos/uso terapêutico , Compostos de Fenilureia/uso terapêutico , Inibidores de Proteínas Quinases/uso terapêutico , Piridinas/uso terapêutico , Pirimidinas/uso terapêutico , Adenocarcinoma de Pulmão/tratamento farmacológico , Quinase do Linfoma Anaplásico/química , Animais , Antineoplásicos/síntese química , Antineoplásicos/farmacologia , Antineoplásicos/toxicidade , Sítios de Ligação , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Estabilidade de Medicamentos , Feminino , Fibroblastos/efeitos dos fármacos , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Camundongos Endogâmicos BALB C , Microssomos Hepáticos/metabolismo , Simulação de Acoplamento Molecular , Estrutura Molecular , Compostos de Fenilureia/síntese química , Compostos de Fenilureia/farmacologia , Compostos de Fenilureia/toxicidade , Inibidores de Proteínas Quinases/síntese química , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/toxicidade , Piridinas/síntese química , Piridinas/farmacologia , Piridinas/toxicidade , Pirimidinas/síntese química , Pirimidinas/farmacologia , Pirimidinas/toxicidade , Relação Estrutura-Atividade , Ensaios Antitumorais Modelo de Xenoenxerto
17.
BMC Vet Res ; 15(1): 214, 2019 Jun 25.
Artigo em Inglês | MEDLINE | ID: mdl-31238913

RESUMO

BACKGROUND: Per- and polyfluoroalkyl substances (PFASs) are environmentally persistent and bioaccumulative chemicals. Immunomodulation is among the most concerning of toxic effects linked with PFAS exposure in mammalian models. However, no studies had yet shown this to be true in birds. Thus, we designed and conducted the first study to determine if PFASs could cause immunomodulation in birds. Secondly, we wanted to determine the effects on an avian host when exposed not only to immunomodulating chemicals, but also to a viral challenge. The aim, to determine if PFAS mediated immunmodulation functionally affects a pathogen challenge for a host. As innate immune system signalling pathways initiate crucial responses against a pathogen challenge, and are lesser studied than their adaptive counterparts, we focused on these pathways. To provide the first information on this, an in vitro experiment was designed and performed using chicken embryo fibroblasts exposed to perfluorooctane sulfonate (PFOS) (22 ppm) and immune markers characterised before and after being infected with gallid herpesvirus-2 (GaHV-2). RESULTS: The expression of two pro-inflammatory cytokines, namely interleukin 8 (IL-8) and tumor necrosis factor alpha (TNF-α), the nuclear factor 'kappa-light-chain-enhancer' of activated B-cells (NF-κB), and the anti-inflammatory cytokine interleukin 4 (IL-4) were investigated in various scenarios. These results showed that exposure to PFOS decreased immune gene expression in chicken fibroblasts from 36 h post-exposure. Next, it was shown that this decrease could be mitigated by infection with gallid herpesvirus-2, which increased gene expression back to the baseline/control levels. CONCLUSIONS: Not only is this the first study to provide the expected evidence that PFOS has immunomodulatory potential in birds, it also provides unexpected data that virus infections can mitigate this negative effect. Thereby, further research, including in vivo and in situ studies, on the impact of PFOS on host-virus interactions is now warranted, as it has been overlooked and might contribute to our understanding of recent disease outbreaks in wildlife. The mechanisms by which gallid herpesvirus mitigates immunomodulation were beyond the scope of this study, but are now of interest for future study.


Assuntos
Ácidos Alcanossulfônicos/toxicidade , Fluorcarbonetos/toxicidade , Herpesvirus Galináceo 2/imunologia , Imunomodulação/efeitos dos fármacos , Doença de Marek/imunologia , Viroses/veterinária , Ácidos Alcanossulfônicos/imunologia , Animais , Linhagem Celular , Embrião de Galinha , DNA Complementar , Fibroblastos/efeitos dos fármacos , Fluorcarbonetos/imunologia , Expressão Gênica/efeitos dos fármacos , Imunomodulação/genética , Mediadores da Inflamação/metabolismo , RNA Viral/isolamento & purificação , Reação em Cadeia da Polimerase em Tempo Real/veterinária , Transdução de Sinais/efeitos dos fármacos , Viroses/imunologia
18.
Braz Oral Res ; 33: e035, 2019 Apr 25.
Artigo em Inglês | MEDLINE | ID: mdl-31038569

RESUMO

Eugenia dysenterica is a Brazilian tree investigated for its properties and bioactive compounds, which are believed to have both pharmacological and phytochemical therapeutic effects. The leaves of this tree contain tannins, flavonoids, terpenes, and saponins, with reportedly beneficial effects to the human body. Despite these therapeutic applications, its effects have never been tested on oral tissues. Therefore, the aim of the present study was to evaluate the cytotoxic and antioxidant effects and the anti-inflammatory and repair properties of the acetone fraction of E. dysenterica on primary culture of human gingival fibroblasts and on the immortalized murine macrophage cell line (RAW 264.7). For this purpose, a metabolic activity assay, a wound healing assay, a nitric oxide assay, and RT-qPCR were performed. The assays revealed a cytoprotective effect of this plant, suggested by the increase in the expression of SOD1 and NRF2. An antioxidant potential effect was observed in the DPPH• assay. However, the fraction of E. dysenterica did not show anti-inflammatory activity. In conclusion, Eugenia dysenterica may promote cytoprotection when associated with chlorhexidine digluconate because of its antioxidant effect. However, additional studies are necessary on other human dental tissues using other parts of the plant in order to develop a possible mouthwash to assist patients with oral disorders.


Assuntos
Anti-Inflamatórios/farmacologia , Antioxidantes/farmacologia , Eugenia/química , Fibroblastos/efeitos dos fármacos , Gengiva/citologia , Extratos Vegetais/farmacologia , Animais , Brasil , Células Cultivadas , Clorexidina/análogos & derivados , Clorexidina/farmacologia , Humanos , Camundongos , Óxido Nítrico/análise , Folhas de Planta/química , Células RAW 264.7 , Valores de Referência , Reprodutibilidade dos Testes , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fatores de Tempo , Cicatrização/efeitos dos fármacos
19.
MBio ; 10(3)2019 05 14.
Artigo em Inglês | MEDLINE | ID: mdl-31088925

RESUMO

Viruses commandeer host cell 26S proteasome activity to promote viral entry, gene expression, replication, assembly, and egress. Proteasomal degradation activity is critical for herpes simplex virus (HSV) infection. The proteasome inhibitor bortezomib (also known as Velcade and PS-341) is a clinically effective antineoplastic drug that is FDA approved for treatment of hematologic malignancies such as multiple myeloma and mantle cell lymphoma. Low nanomolar concentrations of bortezomib inhibited infection by HSV-1, HSV-2, and acyclovir-resistant strains. Inhibition coincided with minimal cytotoxicity. Bortezomib did not affect attachment of HSV to cells or inactivate the virus directly. Bortezomib acted early in HSV infection by perturbing two distinct proteasome-dependent steps that occur within the initial hours of infection: the transport of incoming viral nucleocapsids to the nucleus and the virus-induced disruption of host nuclear domain 10 (ND10) structures. The combination of bortezomib with acyclovir demonstrated synergistic inhibitory effects on HSV infection. Thus, bortezomib is a novel potential therapeutic for HSV with a defined mechanism of action.IMPORTANCE Viruses usurp host cell functions to advance their replicative agenda. HSV relies on cellular proteasome activity for successful infection. Proteasome inhibitors, such as MG132, block HSV infection at multiple stages of the infectious cycle. Targeting host cell processes for antiviral intervention is an unconventional approach that might limit antiviral resistance. Here we demonstrated that the proteasome inhibitor bortezomib, which is a clinically effective cancer drug, has the in vitro features of a promising anti-HSV therapeutic. Bortezomib inhibited HSV infection during the first hours of infection at nanomolar concentrations that were minimally cytotoxic. The mechanism of bortezomib's inhibition of early HSV infection was to halt nucleocapsid transport to the nucleus and to stabilize the ND10 cellular defense complex. Bortezomib and acyclovir acted synergistically to inhibit HSV infection. Overall, we present evidence for the repurposing of bortezomib as a novel antiherpesviral agent and describe specific mechanisms of action.


Assuntos
Antivirais/farmacologia , Bortezomib/farmacologia , Herpesvirus Humano 1/efeitos dos fármacos , Herpesvirus Humano 2/efeitos dos fármacos , Inibidores de Proteassoma/farmacologia , Internalização do Vírus/efeitos dos fármacos , Aciclovir/farmacologia , Animais , Núcleo Celular/metabolismo , Cercopithecus aethiops , Sinergismo Farmacológico , Fibroblastos/efeitos dos fármacos , Fibroblastos/virologia , Prepúcio do Pênis/citologia , Herpes Simples/tratamento farmacológico , Herpesvirus Humano 1/fisiologia , Herpesvirus Humano 2/fisiologia , Humanos , Masculino , Nucleocapsídeo/metabolismo , Células Vero
20.
Yonsei Med J ; 60(6): 585-591, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31124343

RESUMO

PURPOSE: The aim of this study was to explore the function of microRNA-27b (miR-27b) in fibroblast-like synoviocytes (FLSs) stimulated by tumor necrosis factor α (TNF-α). MATERIALS AND METHODS: mRNA expression of miR-27b in FLS cells (MH7A) treated with or without TNF-α was determined by q-PCR. MiR-27b mimics was transfected into MH7A cells to upregulate miR-27b expression. MTT assay and flow cytometry analysis were performed to investigate the effect of miR-27b on MH7A cell viability and apoptosis. The targets of miR-27b were predicted by TargetScan. The direct regulation of miR-27b on IL-1ß expression was verified by luciferase assay. The protein expression levels of apoptosis-related proteins, IL-1ß, and NF-κB signaling-related proteins were detected by Western blot. RESULTS: We discovered that miR-27b expression was decreased in MH7A cells stimulated by TNF-α. Upregulation of miR-27b by miR-27b mimics significantly inhibited the proliferation and promoted the apoptosis of TNF-α-stimulated MH7A cells. Consistently, upregulation of miR-27 decreased the level of Bcl-2 and increased Bax and caspase-3 expression in MH7A cells stimulated by TNF-α. Luciferase assay revealed that IL-1ß was indeed a target of miR-27b. By quantitative real-time PCR and Western blot, we found that the expression of IL-1ß is negatively regulated by miR-27b. Moreover, the NF-κB signaling pathway was significantly inhibited by miR-27b. CONCLUSION: Taken together, our results illustrated that enhanced miR-27b expression results in the suppression of proliferation and the promotion of apoptosis in FLSs stimulated by TNF-α, partially by regulating IL-1ß expression and NF-κB signaling.


Assuntos
Apoptose , Fibroblastos/patologia , MicroRNAs/genética , Sinoviócitos/patologia , Fator de Necrose Tumoral alfa/farmacologia , Regulação para Cima/genética , Apoptose/efeitos dos fármacos , Apoptose/genética , Sequência de Bases , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Humanos , Interleucina-1beta/genética , Interleucina-1beta/metabolismo , MicroRNAs/metabolismo , NF-kappa B/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Transdução de Sinais/efeitos dos fármacos , Sinoviócitos/efeitos dos fármacos , Sinoviócitos/metabolismo , Regulação para Cima/efeitos dos fármacos
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