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BACKGROUND: Porcine reproductive and respiratory syndrome virus (PRRSV) causes devastating disease characterized by reproductive failure and respiratory problems in the swine industry. To understand the recent prevalence and genetic diversity of field PRRSVs in the Republic of Korea, open reading frames (ORFs) 5 and 7 of PRRSV field isolates from 631 PRRS-affected swine farms nationwide in 2013-2016 were analyzed along with 200 Korean field viruses isolated in 2003-2010, and 113 foreign field and vaccine strains. RESULTS: Korean swine farms were widely infected with PRRSVs of a single type (38.4 and 37.4% for Type 1 and Type 2 PRRSV, respectively) or both types (24.2%) with up to approximately 83% nucleotide sequence similarity to prototype PRRSVs (Lelystad or VR2332). Phylogenetic analysis based on the ORF5 nucleotide sequence revealed that Korean Type 1 field isolates were classified as subgroups A, B, and C under subtype 1, while Korean Type 2 field isolates were classified as lineages 1 and 5 as well as three Korean lineages (kor A, B, and C) with the highest infection prevalence in subgroup A (50.5%) and lineage 5 (15.3%) for Type 1 and Type 2 PRRSV, respectively, among ORF5-positive farms. In particular, the lineages kor B and C were identified as novel lineages in this study, and lineage kor B comprised only the field viruses isolated from Gyeongnam Province in 2014-2015, establishing regionally unique genetic characteristics. It has also recently been confirmed that commercialized vaccine-like viruses (subgroup C) of Type 1 PRRSV and NADC30-like viruses of Type 2 PRRSV (lineage 1) are spreading rapidly in Korean swine farms. The Korean field viruses were also expected to be antigenically variable as shown in the high diversity of neutralizing epitopes and N-glycosylation sites. CONCLUSIONS: This up-to-date information regarding recent field PRRSVs should be taken into consideration when creating strategies for the application of PRRS control measures, including vaccination in the field.
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Síndrome Respiratória e Reprodutiva Suína/epidemiologia , Vírus da Síndrome Respiratória e Reprodutiva Suína/genética , Sequência de Aminoácidos , Animais , Epitopos , Fazendas , Variação Genética , Tipagem Molecular/veterinária , Fases de Leitura Aberta , Filogenia , Síndrome Respiratória e Reprodutiva Suína/virologia , Vírus da Síndrome Respiratória e Reprodutiva Suína/classificação , Prevalência , República da Coreia/epidemiologia , SuínosRESUMO
Recent studies have demonstrated the therapeutic potential of mesenchymal stem cells (MSCs) for the treatment of acute inflammatory injury and bacterial pneumonia, but their therapeutic applications in mycobacterial infections have not been investigated. In this study, we demonstrated the use of MSCs as a novel therapeutic strategy against Mycobacterium abscessus (M. abscessus), which is the most drug-resistant and difficult-to-treat mycobacterial pathogen. The systemic intravenous injection of MSCs not only improved mouse survival but also enhanced bacterial clearance in the lungs and spleen. Additionally, MSCs enhanced IFN-γ, TNF-α, IL-6, MCP-1, nitric oxide (NO) and PGE2 production and facilitated CD4(+) /CD8(+) T cell, CD11b(high) macrophage, and monocyte recruitment in the lungs of M. abscessus-infected mice. To precisely elucidate the functions of MSCs in M. abscessus infection, an in vitro macrophage infection system was used. MSCs caused markedly increased NO production via NF-κB activation in M. abscessus-infected macrophages cultured in the presence of IFN-γ. Inhibiting NO or NF-κB signaling using specific inhibitors reduced the antimycobacterial activity of MSCs. Furthermore, the cellular crosstalk between TNF-α released from IFN-γ-stimulated M. abscessus-infected macrophages and PGE2 produced by MSCs was necessary for the mycobacterial-killing activity of the macrophages. Finally, the importance of increased NO production in response to MSC administration was confirmed in the mouse M. abscessus infection model. Our results suggest that MSCs may offer a novel therapeutic strategy for treating this drug-resistant mycobacterial infection by enhancing the bacterial-killing power of macrophages. Stem Cells 2016;34:1957-1970.
Assuntos
Transplante de Células-Tronco Mesenquimais , Células-Tronco Mesenquimais/citologia , Infecções por Mycobacterium não Tuberculosas/microbiologia , Infecções por Mycobacterium não Tuberculosas/terapia , Mycobacterium abscessus/fisiologia , Animais , Comunicação Celular/efeitos dos fármacos , Citocinas/biossíntese , Dinoprostona/metabolismo , Guanidinas/farmacologia , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Células-Tronco Mesenquimais/efeitos dos fármacos , Células-Tronco Mesenquimais/metabolismo , Camundongos Endogâmicos C57BL , Infecções por Mycobacterium não Tuberculosas/patologia , Mycobacterium abscessus/efeitos dos fármacos , Mycobacterium abscessus/crescimento & desenvolvimento , NF-kappa B/metabolismo , Óxido Nítrico/biossíntese , Transdução de Sinais/efeitos dos fármacos , Análise de Sobrevida , Regulação para Cima/efeitos dos fármacosRESUMO
Mesenchymal stem cells (MSCs) have the potential to differentiate into multi-lineage cells, suggesting their future applicability in regenerative medicine and biotechnology. The immunomodulatory properties of MSCs make them a promising replacement therapy in various fields of animal research including in canine atopic dermatitis (AD), a skin disease with 10-15% prevalence. We investigated the immunomodulatory effects of MSCs in an experimental canine AD model induced by Dermatophagoides farinae extract ointment. Canine adipose tissue-derived MSCs (cAT-MSCs) were differentiated into mesodermal cell lineages at the third passage. Alterations in immunomodulatory factors in control, AD, and MSC-treated AD groups were evaluated using flow cytometric analysis, enzyme-linked immunosorbent assay, and quantitative reverse transcription PCR. In the MSC-treated AD group, the number of eosinophils decreased, and the number of regulatory T cells (Tregs) increased compared to those in the AD group. In addition, the immunoglobulin E (IgE) and prostaglandin E2 levels were reduced in the MSC-treated AD group compared to those in the AD group. Furthermore, the filaggrin, vascular endothelial growth factor, and interleukin-5 gene expression levels were relatively higher in the MSC-treated AD group than in the AD group, however, not significantly. cAT-MSCs exerted immunomodulatory effects in an AD canine model via a rebalancing of type-1 and -2 T helper cells that correlated with increased levels of Tregs, IgE, and various cytokines.
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Although edible bird's nest (EBN) has been shown to potentiate mitogenic responses, scientific evidence of its efficacy is still limited. In addition, human adipose-derived stem cells (hADSCs) are increasingly accepted as a source for stem cell therapy. Therefore, the aim of this study was to investigate the effects of the EBN extract (EBNE) on the proliferation of hADSCs and its action mechanisms. We found that EBNE strongly promoted the proliferation of hADSCs. In addition, EBNE-induced proliferation was found to be mediated through the production of IL-6 and VEGF, which was induced by activation of AP-1 and NF-κB. Specially, we found that production of IL-6 and VEGF was induced by EBNE. In addition, EBNE-induced production of IL-6 and VEGF was inhibited by PD98059 (a p44/42 MAPK inhibitor), SB203580 (a p38 MAPK inhibitor), and PDTC (a NF-κB inhibitor), but not SP600125 (a JNK inhibitor). Similarly, EBNE-induced proliferation of hADSCs was also attenuated by PD98059, SB203580, and PDTC but not SP600125. Taken together, these findings suggest that the EBNE-induced proliferation of hADSCs primarily occurs through increased expression of IL-6 and VEGF genes, which is mediated by the activation of NF-κB and AP-1 through p44/42 MAPK and p38 MAPK.
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Adipocyte dysfunction is strongly associated with the development of obesity, which is a major risk factor for many disorders, including diabetes, hypertension, and heart disease. This study shows that ultraviolet A (UVA) inhibits adipogenic differentiation of human adipose tissue-derived mesenchymal stem cells and its action mechanisms. The mRNA levels of peroxidase proliferator-activated receptor (PPAR) γ and CCAAT/enhancer-binding protein α (C/EBPα), but not CCAAT/enhancer-binding protein ((C/EBP) ß and δ, were reduced by UVA. Moreover, the mRNA levels of PPAR γ target genes (lipoprotein lipase (LPL), CD36, adipocyte protein (aP2), and liver X receptor α (LXR)) were down-regulated by UVA. Additionally, attempts to elucidate a possible mechanism underlying the UVA-mediated effects revealed that UVA induced migration inhibitory factor (MIF) gene expression, and this was mediated through activation of AP-1 (especially JNK and p42/44 MAPK) and nuclear factor-κB. In addition, reduced adipogenesis by UVA was recovered upon the treatment with anti-MIF antibodies. AMP-activated protein kinase phosphorylation and up-regulation of Kruppel-like factor 2 (KLF2) were induced by UVA. Taken together, these findings suggest that the inhibition of adipogenic differentiation of human adipose tissue-derived mesenchymal stem cells by UVA occurs primarily through the reduced expression of PPAR γ, which is mediated by up-regulation of KLF2 via the activation of MIF-AMP-activated protein kinase signaling.
Assuntos
Adipogenia/efeitos da radiação , Tecido Adiposo/citologia , Diferenciação Celular/efeitos da radiação , Fatores de Transcrição Kruppel-Like/metabolismo , Células-Tronco Mesenquimais/fisiologia , Células-Tronco Mesenquimais/efeitos da radiação , Raios Ultravioleta , Células 3T3-L1 , Proteínas Quinases Ativadas por AMP/metabolismo , Animais , Citocinas/genética , Citocinas/metabolismo , Humanos , Fatores de Transcrição Kruppel-Like/genética , Células-Tronco Mesenquimais/citologia , Camundongos , PPAR gama/metabolismo , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Regulação para CimaRESUMO
Adipocyte dysfunction is associated with the development of obesity. This study shows that 6-thioinosine inhibits adipocyte differentiation. The mRNA levels of PPAR gamma and C/EBPalpha, but not C/EBPbeta and delta, were reduced by 6-thioinosine. Moreover, the mRNA levels of PPAR gamma target genes (LPL, CD36, aP2, and LXRalpha) were down-regulated by 6-thioinosine. We also demonstrated that 6-thioinosine inhibits the transactivation activity and the mRNA level of PPAR gamma. Additionally, attempts to elucidate a possible mechanism underlying the 6-thioinosine-mediated effects revealed that 6-thioinosine induced iNOS gene expression without impacting eNOS expression, and that this was mediated through activation of AP-1, especially, JNK. In addition, 6-thioinosine was found to operate upstream of MEKK-1 in JNK activation signaling. Taken together, these findings suggest that the inhibition of adipocyte differentiation by 6-thioinosine occurs primarily through the reduced expression of PPAR gamma, which is mediated by upregulation of iNOS via the activation of JNK.
Assuntos
Adipogenia/efeitos dos fármacos , Antimetabólitos Antineoplásicos/farmacologia , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Óxido Nítrico Sintase Tipo II/metabolismo , PPAR gama/metabolismo , Tioinosina/farmacologia , Células 3T3-L1 , Animais , Antracenos/farmacologia , Proteína alfa Estimuladora de Ligação a CCAAT/genética , Proteína alfa Estimuladora de Ligação a CCAAT/metabolismo , Regulação para Baixo , Humanos , Proteínas Quinases JNK Ativadas por Mitógeno/antagonistas & inibidores , Proteínas Quinases JNK Ativadas por Mitógeno/genética , MAP Quinase Quinase Quinase 1/metabolismo , Camundongos , Óxido Nítrico Sintase Tipo II/genética , Óxido Nítrico Sintase Tipo III/metabolismo , PPAR gama/genética , Células-Tronco/citologia , Fator de Necrose Tumoral alfa/metabolismo , Regulação para CimaRESUMO
Natural killer (NK) cells are key immune cells engaged in fighting infection and malignant transformation. In this study, we found that canine NK cell-derived exosomes (NK-exosomes) separated from activated cytotoxic NK cell supernatants express specific markers including CD63, CD81, Alix, HSP70, TSG101, Perforin 1, and Granzyme B. We examined the antitumor effects of NK-exosomes in an experimental murine mammary tumor model using REM134 canine mammary carcinoma cell line. We observed changes in tumor size, tumor initiation, progression, and recurrence-related markers in the control, tumor group, and NK-exosome-treated tumor group. We found that the tumor size in the NK-exosome-treated tumor group decreased compared with that of the tumor group in the REM134-driven tumorigenic mouse model. We observed significant changes including the expression of tumorigenesis-related markers, such as B cell-specific Moloney murine leukemia virus insertion site 1 (Bmi-1), vascular endothelial growth factor (VEGF), matrix metallopeptidase-3 (MMP-3), interleukin-1ß (IL-1ß), IL-6, tumor necrosis factor-α (TNF-α), multidrug resistance protein (MDR), tumor suppressor protein p53 (p53), proliferating cell nuclear antigen (PCNA), and the apoptotic markers, B cell lymphoma-2 associated X (Bax) and B cell lymphoma-extra large (Bcl-xL) belonging to the Bcl-2 family, in the tumor group compared with those in the control group. The expression of CD133, a potent cancer stem cell marker, was significantly higher than that of the control. By contrast, the NK-exosome-treated tumor group exhibited a significant reduction in Bmi-1, MMP-3, IL-1ß, IL-6, TNF-α, Bax, Bcl-xL, and PCNA expression compared with that in the tumor group. Furthermore, the expression of CD133, which mediates tumorigenesis, was significantly decreased in the NK-exosome-treated tumor group compared with that in the tumor group. These findings indicate that canine NK-exosomes represent a promising therapeutic tool against canine solid tumors, including mammary carcinoma.
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Exossomos/imunologia , Células Matadoras Naturais/imunologia , Neoplasias Mamárias Animais/imunologia , Animais , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Biomarcadores Tumorais , Neoplasias da Mama/imunologia , Neoplasias da Mama/metabolismo , Neoplasias da Mama/terapia , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Modelos Animais de Doenças , Cães , Exossomos/metabolismo , Exossomos/fisiologia , Feminino , Células Matadoras Naturais/metabolismo , Células Matadoras Naturais/transplante , Neoplasias Mamárias Animais/metabolismo , Neoplasias Mamárias Animais/terapia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Cultura Primária de Células , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Age-related cartilage loss is worsened by the limited regenerative capacity of chondrocytes. The role of cell-based therapies using mesenchymal stem cells is gaining interest. Adipose tissue-derived mesenchymal stem cells (ADSCs) are an attractive source to generate the optimal number of chondrocytes required to repair a cartilage defect and regenerate hyaline articular cartilage. Here, we report an outstanding technique to prepare chondrocytes for cartilage repair using canine ADSCs. We hypothesized that external electrical fields promote prechondrogenic condensation without requiring genetic modifications or exogenous factors. We analyzed the effect of electrical stimulation (ES) on the differentiation of ADSC micromass into chondrocytes. Highly compact structures were formed within 3 days of ES of canine ADSC micromass. The expression of type I collagen gene was abolished in these cells compared with that in control micromass cultures and monolayer cultures. We further found that ES enhanced the production of proteoglycan, a highly produced extracellular matrix component in chondrocytes. Additionally, single-cell RNA sequencing analysis showed that canine ADSC micromass undergoing ES developed a prechondrogenic cell aggregation, suggesting their metabolic conversion, biogenesis, and calcium ion change. Collectively, our findings demonstrate the capacity of ES to drive the chondrogenesis of ADSCs in the absence of exogenous factors and confirm its commercial potential as a budget-friendly therapy for the repair of cartilage defects.
Assuntos
Cartilagem ArticularRESUMO
Mesenchymal stem cells (MSCs) possess the ability to differentiate into multiple cell lineages, and thus, confer great potential for use in regenerative medicine and biotechnology. In the present study, we attempted to isolate and characterize bovine tongue tissue epithelium-derived MSCs (boT-MSCs) and investigate the culture conditions required for long-term culturing of boT-MSCs. boT-MSCs were successfully isolated by the collagenase digestion method and their proliferative capacity was maintained for up to 20 or more passages. We observed a significant increase in the proliferation of boT-MSCs during the 20 consecutive passages under low-glucose Dulbecco's modified Eagle's medium culture condition among the three culture conditions. These boT-MSCs presented pluripotency markers (octamer-binding transcription factor 3/4 (Oct3/4) and sex determining region Y-box2 (Sox2)) and cell surface markers, which included CD13, CD29, CD44, CD73, CD90, CD105, CD166, and major histocompatibility complex (MHC) class I (MHC-I) but not CD11b, CD14, CD31, CD34, CD45, CD80, CD86, CD106, CD117, and MHC-II at third passage. Moreover, these boT-MSCs could differentiate into mesodermal (adipocyte, osteocyte, and chondrocyte) cell lineages. Thus, the present study suggests that the tongue of bovines could be used as a source of bovine MSCs.
Assuntos
Células Epiteliais/fisiologia , Células-Tronco Mesenquimais/fisiologia , Mucosa Bucal/citologia , Língua/citologia , Animais , Bovinos , Diferenciação Celular , Proliferação de Células , Células Cultivadas , Cultura Primária de CélulasRESUMO
The present study was undertaken to investigate the effect of dietary, synthetic lycopene or tomato paste on laying performance and egg qualities in laying hens, and on lipid oxidation of stored eggs. One hundred and sixty 38-week-old Hy-line Brown laying hens were randomly housed in cages (two birds per cage, five cages per replicate) equipped with nipples and a trough-type feeder and subjected to one of four experimental diets. Each treatment had four replicates. A corn and soybean meal base diet was added with or without either synthetic lycopene to contain 10 or 20 mg per kg of diet, or with 17 g of tomato paste per kg of diet. The feeding trial lasted four weeks. Feed intake did not differ between dietary treatments. Laying hens fed diets containing lycopene or tomato paste laid lighter eggs (P<0.05) compared with those fed on the control diet. Egg production was higher (P<0.05) in tomato paste-fed layers, but lower (P<0.05) in those fed on a diet containing 20 mg/kg of lycopene compared with the control diet-fed counterparts. Dietary lycopene did not affect egg quality, except for yolk color, nor did serum lipid profiles. Malondialdehyde (MDA) content in serum samples and eggs that had been stored at 24°C for four weeks was reduced (P<0.05) by lycopene or tomato paste. Adding lycopene or tomato paste into a diet of laying hens increased the incorporation of lycopene into the liver and egg yolk. Collectively, the present study shows that addition of low levels of lycopene or tomato paste into the layers' diet is an effective nutritional strategy to enhance oxidative stability of fresh eggs.
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Melanogenesis is the biological process which the skin pigment melanin is synthesized to protect the skin against ultraviolet irradiation and other external stresses. Abnormal biology of melanocytes is closely associated with depigmented skin disorders such as vitiligo. In this study, we examined the effects of maclurin on melanogenesis and cytoprotection. Maclurin enhanced cellular tyrosinase activity as well as cellular melanin levels. We found that maclurin treatment increased the expression of microphthalmia-associated transcription factor (MITF), tyrosinase-related protein- (TRP-) 1, TRP-2, and tyrosinase. Mechanistically, maclurin promoted melanogenesis through cyclic adenosine monophosphate (cAMP) response element binding (CREB) protein-dependent upregulation of MITF. CREB activation was found to be mediated by p38 mitogen-activated protein kinase (MAPK) or cAMP-protein kinase A (PKA) signaling. In addition, maclurin-induced CREB phosphorylation was mediated through the activation of both the cAMP/PKA and the p38 MAPK signaling pathways. Maclurin-induced suppression of p44/42 MAPK activation also contributed to its melanogenic activity. Furthermore, maclurin showed protective effects against H2O2 treatment and UVB irradiation in human melanocytes. These findings indicate that the melanogenic effects of maclurin depend on increased MITF gene expression, which is mediated by the activation of both p38 MAPK/CREB and cAMP/PKA/CREB signaling. Our results thus suggest that maclurin could be useful as a protective agent against hypopigmented skin disorders.
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Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , AMP Cíclico/metabolismo , Epiderme/metabolismo , Melaninas/biossíntese , Melanócitos/metabolismo , Lectinas de Plantas/farmacologia , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Células Cultivadas , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/genética , Proteínas Quinases Dependentes de AMP Cíclico/genética , Epiderme/efeitos dos fármacos , Regulação da Expressão Gênica , Humanos , Melanócitos/efeitos dos fármacos , Oxirredutases/genética , Oxirredutases/metabolismo , Fosforilação , Transdução de Sinais , Proteínas Quinases p38 Ativadas por Mitógeno/genéticaRESUMO
In this study, we used a murine intestinal inflammation model that mimics immunologic characteristics of human Crohn's disease (CD) to investigate the anti-inflammatory effects of mycophenolate mofetil (MMF) on intestinal injury and tissue inflammation. When these colitic mice were pretreated with MMF, we observed a significant decrease in mortality rates and body weight loss as well as an improvement in both wasting and histopathologic signs of colonic inflammation, relative to untreated colitic mice. To determine the mechanisms of action of MMF, we compared various immunological characteristics of the untreated and MMF-pretreated colitic mice. MMF-pretreated colitic mice showed an 18% decrease in the proportion of CD19+ B cells compared with untreated colitic mice 3 days. As a result, MMF pretreatment increases proportion of apoptotic T and B cells, especially CD19+ B cells. Also, down-regulation of Th1 cytokines (TNF-alpha, IFN-gamma) and augmentation of CD4+CD45RB(low) regulatory T (Treg) cells were observed in MMF-pretreated colitic mice compared with untreated colitic mice. Furthermore, mycophenolic acid (MPA) reduced TNF-alpha-stimulated NF-kappaB activation in HT-29 colon epithelial cells. Also, MMF-pretreated colitic mice significantly reduced expression of MD-1 compared with untreated colitic mice on B cells and dendritic cells (DCs). These studies show that MMF pretreatment can improve experimental colitis by down-regulation of expanded B cells population through apoptosis and augmentation of Treg cells. Through these mechanisms, MMF might also be an effective agent for the treatment of other diseases characterized by mucosal inflammation.
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Linfócitos B/efeitos dos fármacos , Colite/fisiopatologia , Ácido Micofenólico/análogos & derivados , Fator de Necrose Tumoral alfa/biossíntese , Animais , Antígenos CD19/metabolismo , Antígenos de Superfície/biossíntese , Apoptose/efeitos dos fármacos , Colite/induzido quimicamente , Colite/tratamento farmacológico , Modelos Animais de Doenças , Regulação para Baixo , Feminino , Glicoproteínas de Membrana/biossíntese , Camundongos , Camundongos Endogâmicos BALB C , Ácido Micofenólico/farmacologia , NF-kappa B/efeitos dos fármacos , NF-kappa B/fisiologia , Linfócitos T Reguladores/efeitos dos fármacos , Ácido TrinitrobenzenossulfônicoRESUMO
Matrix metalloproteinase-1 (MMP-1) plays an important role in the maintenance and turnover of extracellular matrix (ECM) macromolecules. Remodelling of extracellular matrix by MMPs is a hallmark feature of physiological and pathological processes. In this study, in order to establish the therapeutic potential of matrine, we investigated its effect on MMP-1 expression in human dermal fibroblast cells. We found that matrine inhibited both MMP-1 mRNA and protein expression induced by PMA (phorbol myristate acetate). Therefore, we characterized the inhibitory mechanism of matrine on PMA-induced MMP-1 expression. Matrine inhibited PMA-induced activation of the AP-1 promoter, an important nuclear transcription factor in MMP-1 expression. Additionally, we detected that matrine suppressed the PMA-induced phosphorylation of two mitogen-activated protein kinases, extracellular signal-regulated protein kinase and c-Jun N-terminal kinase, but did not suppress the PMA-induced phosphorylation of p38 kinase. These results suggest that matrine suppresses PMA-induced MMP-1 expression through inhibition of the AP-1 signaling pathway and also may be beneficial for treatment of some inflammatory skin disorders.
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Alcaloides/farmacologia , Derme/enzimologia , Fibroblastos/enzimologia , Metaloproteinase 1 da Matriz/metabolismo , Quinolizinas/farmacologia , Acetato de Tetradecanoilforbol/farmacologia , Carcinógenos/farmacologia , Derme/citologia , Relação Dose-Resposta a Droga , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Humanos , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Metaloproteinase 1 da Matriz/genética , Inibidores de Metaloproteinases de Matriz , Fosforilação , RNA Mensageiro/metabolismo , Transdução de Sinais , Fator de Transcrição AP-1/genética , Fator de Transcrição AP-1/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , MatrinasRESUMO
The epidermis, the outermost layer of the skin, is a stratified epithelium that protects the body from the external environment. Keratinocyte stem cells (KSCs) are involved in epidermis homeostasis by maintaining epidermal integrity through a process of constant regeneration. Ultraviolet B (UVB) radiation is a major inducer of cellular damage in the epidermis. In this study, we investigated the effects of zingerone (a phenolic compound derived from spices) on UVB-induced cellular damage in KSCs. We found that zingerone significantly inhibited cellular senescence of KSCs in response to UVB irradiation. These effects were confirmed by the senescence-associated ß-galactosidase and comet assays. Zingerone decreased the production of proinflammatory cytokines such as tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß), and interleukin-6 (IL-6) in UVB-irradiated KSCs. Moreover, UVB-induced expression of p21, a cell cycle arrest-related gene, was reduced by zingerone treatment, whereas zingerone upregulated the expression of proliferation-related genes such as proliferating cell nuclear antigen (PCNA) and vascular endothelial growth factor (VEGF), in addition to anti-senescence-related genes including telomerase reverse transcriptase (TERT), histone deacetylase 1 (HDAC1), and DNA (cytosine-5)-methyltransferase 1 (DNMT1). The UVB-protective effects of zingerone were mediated by inhibition of p42/44 MAPK and p38 MAPK. Therefore, zingerone could potentially be used to protect the epidermis from UVB-induced damage.
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Guaiacol/análogos & derivados , Queratinócitos/efeitos dos fármacos , Queratinócitos/efeitos da radiação , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Pontos de Checagem do Ciclo Celular/efeitos da radiação , Morte Celular/efeitos dos fármacos , Morte Celular/efeitos da radiação , Células Cultivadas , Citocinas/genética , Citocinas/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos da radiação , Guaiacol/farmacologia , Humanos , Inflamação/metabolismo , Raios UltravioletaRESUMO
Mesenchymal stem cells (MSCs) are useful candidates for tissue engineering and cell therapy fields. We optimize culture conditions of equine adipose tissue-derived MSCs (eAD-MSCs) for treatment of horse fractures. To investigate enhancing properties of three-dimensional (3D) culture system in eAD-MSCs, we performed various sized spheroid formation and determined changes in gene expression levels to obtain different sized spheroid for cell therapy. eAD-MSCs were successfully isolated from horse tailhead. Using hanging drop method, spheroid formation was generated for three days. Quantitative real-time PCR was performed to analyze gene expression. As results, expression levels of pluripotent markers were increased depending on spheroid size and the production of PGE2 was increased in spheroid formation compared to that in monolayer. Ki-67 showed a remarkable increase in the spheroid formed with 2.0 × 105 cells/drop as compared to that in the monolayer. Expression levels of angiogenesis-inducing factors such as VEGF, IL-6, IL-8, and IL-18 were significantly increased in spheroid formation compared to those in the monolayer. Expression levels of bone morphogenesis-inducing factors such as Cox-2 and TGF-ß1 were also significantly increased in spheroid formation compared to those in the monolayer. Expression levels of osteocyte-specific markers such as RUNX2, osteocalcin, and differentiation potential were also significantly increased in spheroid formation compared to those in the monolayer. Therefore, spheroid formation of eAD-MSCs through the hanging drop method can increases the expression of angiogenesis-inducing and bone morphogenesis-inducing factors under optimal culture conditions.
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Técnicas de Cultura de Células/veterinária , Diferenciação Celular/fisiologia , Cavalos/fisiologia , Células-Tronco Mesenquimais/citologia , Esferoides Celulares/fisiologia , Tecido Adiposo/citologia , Animais , Células Cultivadas , Feminino , MasculinoRESUMO
The stemness of stem cells is negatively affected by ultraviolet A (UVA) irradiation. This study was performed to examine the effects of arctigenin on UVA-irradiation-induced damage to the stemness of human mesenchymal stem cells (hMSCs) derived from adipose tissue. The mechanisms of action of arctigenin were also investigated. A BrdU-incorporation assay demonstrated that arctigenin attenuated the UVA-induced reduction of the cellular proliferative potential. Arctigenin also increased the UVA-induced reduction in stemness of hMSCs by upregulating stemness-related genes such as SOX2, OCT4, and NANOG. In addition, the UVA-induced reduction in the mRNA expression level of hypoxia-inducible factor (HIF)-1α was significantly recovered by arctigenin. The antagonizing effect of arctigenin on UVA irradiation was mediated by reduced PGE2 production through the inhibition of MAPKs (p42/44 MAPK, p38 MAPK, and JNK) and NF-κB. Overall, these findings suggest that arctigenin can ameliorate the reduced stemness of hMSCs induced by UVA irradiation. The effects of arctigenin are mediated by PGE2-cAMP signaling-dependent upregulation of HIF-1α. Therefore, arctigenin could be used as an antagonist to attenuate the effects of UVA irradiation.
Assuntos
Furanos/farmacologia , Lignanas/farmacologia , Células-Tronco Mesenquimais/efeitos dos fármacos , Proteínas Quinases Ativadas por Mitógeno/metabolismo , NF-kappa B/metabolismo , Substâncias Protetoras/farmacologia , Transdução de Sinais/efeitos dos fármacos , Raios Ultravioleta/efeitos adversos , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , AMP Cíclico/metabolismo , Dinoprostona/metabolismo , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Células-Tronco Mesenquimais/metabolismo , Proteína Homeobox Nanog/metabolismo , Fator 3 de Transcrição de Octâmero/metabolismo , RNA Mensageiro/metabolismo , Fatores de Transcrição SOXB1/metabolismo , Regulação para Cima/efeitos dos fármacosRESUMO
BACKGROUND: The novel cytokine, interleukin (IL)-18, is a strong interferon-gamma inducer and costimulatory factor in Th1 cell activation. IL-18 triggers IFN-gamma production and enhances cytolytic activity in both T and NK cells. However, the exact mechanism of antitumor action of IL-18 remains to be clarified. To determine the effects of IL-18 plasmid DNA on hepatic cancer in mice, CT26 murine colon adenocarcinoma cells were established in mouse liver. METHODS: Plasmid vectors encoding IL-18 were transferred directly into the liver 7 days after tumor injection to restrict IL-18 expression within the tumor site. The IL-18 protein level was increased in the liver 4 days after plasmid injection, and a marked antitumoral effect was observed at day 7. Antitumor effects were evaluated by measuring tumor regression, immune cell population, and IFN-gamma production. RESULTS: The IL-18 plasmid controlled the growth of hepatic tumors and proliferation of splenic immune cells. Moreover, treatment of CT26 tumors with the IL-18 plasmid significantly enhanced the population of the effector T and NK cells in the spleen and peripheral blood. In spleen, the population of CD4+CD62Low cells was augmented in response to IL-18 on day 7. These results are consistent with the increase in CD4+ T cells secreting IFN-gamma, but not CD8+ T cells. The marked reduction of tumor growth in tumor-bearing mice was associated with the maintenance of IFN-gamma production in spleen in response to IL-18. These antitumoral effects were maintained until 14 days after plasmid injection. CONCLUSION: Our results suggest that direct plasmid DNA transfer of IL-18 with no accompanying reagents to augment transfection efficiency may be useful in tumor immunotherapy.
Assuntos
Adenocarcinoma/terapia , DNA/administração & dosagem , Terapia Genética/métodos , Interleucina-18/genética , Interleucina-18/farmacologia , Neoplasias Hepáticas/terapia , Linfócitos T/imunologia , Adenocarcinoma/imunologia , Adenocarcinoma/patologia , Animais , Modelos Animais de Doenças , Citometria de Fluxo , Técnicas de Transferência de Genes , Vetores Genéticos , Injeções Intralesionais , Neoplasias Hepáticas/imunologia , Neoplasias Hepáticas/patologia , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos BALB C , Plasmídeos , Sensibilidade e Especificidade , Células Th1/imunologia , Resultado do TratamentoRESUMO
The immunosuppressive drug 15-deoxyspergualin (DSG) is currently being used in clinical trials to prolong graft survival and reverse graft rejection. Here we evaluated whether DSG has a potential for ameliorating diseases characterized by mucosal inflammation. Using a murine model of 2,4,6-trinitrobenzene sulfonic acid (TNBS)-induced colitis, we were able to demonstrate that DSG reduced the severity of colitis. Therefore, colitic mice pretreated with DSG showed a striking improvement of the wasting disease compared with colitic mice, as assessed by weight loss as well as clinical, macroscopic and microscopic analysis. Also, we observed the significant change occurred in the CD19(+) B cell subset, which was decreased 15% in DSG pretreated colitic mice compared with colitic mice. However, DSG pretreatment does not influence the apoptotic population of T and B cells. Compared with colitic mice, down-regulation of TNF-alpha production was observed in DSG pretreated colitic mice. In addition, DSG pretreated colitic mice significantly reduced expression of MD-1 compared with colitic mice on B cells and dendritic cells (DCs). Therefore, pretreatment with DSG resulted in a significant histologic improvement, protecting against mucosal ulcerations and reduced inflammatory response by modulating expression of MD-1, which plays a very important role in immune response on B cells and DCs. Also, this improvement was paralleled by a reduction in TNF-alpha levels. Collectively, current results demonstrate that DSG may be an effective agent for the treatment of diseases characterized by mucosal inflammation.
Assuntos
Antígenos de Superfície/metabolismo , Colite/prevenção & controle , Guanidinas/farmacologia , Mucosa Intestinal/efeitos dos fármacos , Glicoproteínas de Membrana/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Administração Retal , Animais , Antígenos CD19/metabolismo , Apoptose/efeitos dos fármacos , Linfócitos B/citologia , Linfócitos B/efeitos dos fármacos , Linfócitos B/metabolismo , Proliferação de Células/efeitos dos fármacos , Colite/induzido quimicamente , Colite/metabolismo , Colo/efeitos dos fármacos , Colo/lesões , Colo/metabolismo , Modelos Animais de Doenças , Regulação para Baixo/efeitos dos fármacos , Feminino , Guanidinas/administração & dosagem , Imunossupressores/administração & dosagem , Imunossupressores/farmacologia , Injeções Intraperitoneais , Interferon gama/metabolismo , Mucosa Intestinal/lesões , Mucosa Intestinal/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Receptores Toll-Like/metabolismo , Ácido Trinitrobenzenossulfônico/administração & dosagem , Ácido Trinitrobenzenossulfônico/toxicidadeRESUMO
Tuberculosis, a chronic progressive disease, has been reported in bovine, swine, and primate species. Here, we report the first case of Mycobacterium tuberculosis infection in a Korean wild boar ( Sus scrofa coreanus). The owners this domesticated boar brought it to the Gyeongbuk Veterinary Service Laboratory in Korea after it was found dead and severely emaciated. Demarcated yellowish white nodules were found around the larynx and retropharyngeal lymph node during necropsy. The lungs had diffuse fibrinous pleuritis, severe congestion, and scattered nodules. More nodules were found in the spleen. Tuberculosis is characterized by massive macrophage infiltration and central caseous necrosis; both characteristics were found in the lungs. Histopathologic examination revealed that the alveolar lumen had marked fibrosis and exudates. Examination of the fluid revealed extensive macrophage permeation. To confirm a Mycobacterium infection, PCR was performed using two primer sets specific to the rpoB gene of Mycobacterium; Mycobacterium was detected in the lungs and spleen. To identify the species of Mycobacterium, immunohistochemical evaluation was performed using antibodies against Mycobacterium tuberculosis and Mycobacterium bovis . The results revealed immunoreactivity against M. tuberculosis but not against M. bovis . The consumption of undercooked or raw meat from game animals may expose humans and other animals to sylvatic infection. Consequently, Koreans who ingest wild boar may be at risk of a tuberculosis infection. To reduce the risk of foodborne infection and maintain public health, continuous monitoring and control strategies are required.
Assuntos
Mycobacterium tuberculosis , Sus scrofa , Animais , Bovinos , República da Coreia , Suínos , Doenças dos Suínos/microbiologia , Tuberculose/microbiologiaRESUMO
Research into adipose tissue-derived mesenchymal stem cells (AD-MSCs) has demonstrated the feasibility of their use in clinical applications due to their ease of isolation and abundance in adipose tissue. We isolated AD-MSCs from young and old dogs, and the cells were subjected to sequential sub-passaging from passage 1 (P1) to P7. Canine AD-MSCs (cAD-MSCs) were examined for proliferation kinetics, expression of molecules associated with self-renewal, expression of cell surface markers, and differentiation potentials at P3. Cumulative population doubling level was significantly higher in cAD-MSCs of young donors than in those of old donors. In addition, expressions of CD73, CD80, Oct3/4, Nanog, cell survival genes and differentiation potentials were significantly higher in young donors than in old donors. The present study suggests that donor age should be considered when developing cell-based therapies for clinical application of cAD-MSCs.