RESUMO
BACKGROUND: Postinflammatory hyperpigmentation (PIH) is a common, acquired pigmentary disorder of the skin associated with significant quality-of-life impairment, especially in individuals with skin of colour. Current treatment for PIH is limited, largely due to a poor understanding of disease pathogenesis and the lack of a representative disease model. OBJECTIVES: This study is intended to further develop, update and validate our previously designed in vivo model of acne-induced PIH/postinflammatory erythema (PIE) using different concentrations of trichloroacetic acid (TCA), a medium-depth chemical peel. METHODS: Twenty-nine patients with skin types II-VI and clinician-confirmed presence of two or more truncal acne pustules and PIH/PIE were included. On the basis of Investigator's Global Assessment (IGA), clinical polarized photography (CPP), colorimetry and Skindex, we experimentally determined an optimum TCA concentration and assessed our model's ability to exhibit a dose-response relationship between degree of inciting insult and severity of resulting pigmentation. We also performed differential microRNA profiling and pathway analysis to explore the potential of microRNAs as molecular adjuncts to our model. RESULTS: Application of TCA 30% produced lesions indistinguishable from acne-induced PIH and PIE lesions on the basis of colorimetry data without causing epidermal necrosis. Application of progressively increasing TCA doses from 20% to 30% resulted in concentration-dependent increases in CPP, IGA and colorimetry scores at all timepoints during the study. miRNA-31 and miRNA-23b may play a role in PIH pathogenesis, although further validation is required. CONCLUSIONS: Our TCA-based in vivo model, using TCA concentrations between 20% and 30% with an optimum of 30%, enables the quantitative assessment of the pigmentary response to varying degrees of cutaneous inflammation in a fashion that mirrors natural acne-induced PIH and PIE.
Assuntos
Acne Vulgar , Hiperpigmentação , MicroRNAs , Acne Vulgar/complicações , Acne Vulgar/patologia , Colorimetria , Eritema/etiologia , Humanos , Hiperpigmentação/patologia , Imunoglobulina A , Ácido TricloroacéticoRESUMO
Langerhans cells (LC) represent a specialized subset of evolutionarily conserved dendritic cells (DC) that populate stratified epithelial tissues, which are essential for the induction of skin and mucosal immunity and tolerance, including allergy. Transforming growth factor-ß1 (TGF-ß1) has been confirmed to be a predominant factor involved in LC development. Despite great advances in the understanding of LC ontogeny and diverse replenishment patterns, the underlying molecular mechanisms remain elusive. This review focuses on the recent discoveries in TGF-ß1-mediated LC development and maintenance, with special attention to the involved transcription factors and related regulators.
Assuntos
Células de Langerhans/imunologia , Células de Langerhans/metabolismo , Fatores de Transcrição/metabolismo , Fator de Crescimento Transformador beta1/metabolismo , Animais , Diferenciação Celular , Regulação da Expressão Gênica , Humanos , Células de Langerhans/citologia , Ligação Proteica , Transcrição GênicaRESUMO
Cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) is a costimulatory molecule that negatively regulates T-cell activation. Originally identified in murine CD8(+) T cells, it has been found to be rapidly induced on human T cells. Furthermore, CTLA-4 is expressed on regulatory T cells. Clinically, targeting CTLA-4 has clinical utility in the treatment of melanoma. Whether the expression of CTLA-4 is differentially regulated in CD8(+) vs CD4(+) human T cells is unclear. Here, we analyzed CTLA-4 in normal human CD4(+) and CD8(+) T-cell subsets and show for the first time that CTLA-4 is expressed significantly higher in the CD4(+) T cells than in CD8(+) T cells. CTLA-4 is higher at the protein and the transcriptional levels in CD4(+) T cells. This increase is due to the activation of the CTLA-4 promoter, which undergoes acetylation at the proximal promoter. Furthermore, we show that blocking CTLA-4 on CD4(+) T cells permits greater proliferation in CD4(+) vs CD8(+) cells. These findings demonstrate a differential regulation of CTLA-4 on CD4(+) and CD8(+) T-cell subsets, which is likely important to the clinical efficacy for anti-CTLA-4 therapies. The findings hint to strategies to modulate CTLA-4 expression by targeting epigenetic transcription to alter the immune response.
Assuntos
Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD8-Positivos/metabolismo , Antígeno CTLA-4/metabolismo , Fatores de Transcrição NFATC/metabolismo , Acetilação , Antígeno CTLA-4/genética , Proliferação de Células , Células Cultivadas , Regulação da Expressão Gênica , Histonas/metabolismo , Humanos , Ativação Linfocitária , Regiões Promotoras Genéticas , Linfócitos T Reguladores/metabolismo , Regulação para CimaRESUMO
Langerhans cells (LCs) are bone marrow-derived immature skin-residential dendritic cells (DCs) with a life cycle distinct from that of other types of DCs. The mechanisms involved in LC homeostasis and immunological functions are still not clear. MicroRNAs (miRNAs) are a class of short noncoding RNAs that regulate gene expression through either translational repression or mRNA degradation. A recent study showed that specific deletion of total miRNAs in DCs affects the homeostasis and function of only LCs, but not of other types of DCs. The roles of specific individual miRNA in LC development are still lacking. The miRNA miR-17-92 class, encoding miR-17, miR-18, miR-19a, miR-19b, miR-20 and miR-92, plays a very important role in B- and T-cell development and function. Here, we first report that epidermal LCs highly express the miR-17-92 class compared with spleen naive T cells. To further characterize the role of miR-17-92 in LC development, we generated LC-specific miR-17-92 knockout and knock-in mice. Interestingly, LC-specific gain- and loss-of-function of miR-17-92 cluster did not significantly change LC homeostasis, maturation ability, antigen capture and migration to draining lymph nodes. Thus, the miR-17-92 cluster may be functionally redundant and not critically required for LC development and function.
Assuntos
Células de Langerhans/fisiologia , MicroRNAs/genética , Animais , Movimento Celular , Dermatite Alérgica de Contato/genética , Dermatite Alérgica de Contato/imunologia , Dinitrofluorbenzeno/imunologia , Dinitrofluorbenzeno/toxicidade , Células Epidérmicas , Regulação da Expressão Gênica , Homeostase , Camundongos , Camundongos Knockout , Família Multigênica , Baço/citologia , Linfócitos T/fisiologiaRESUMO
OBJECTIVE: The aim of this study was to examine serum non-coding RNAs as potential biomarkers for cartilage damage associated with anterior cruciate ligament (ACL) injury. METHODS: Serum was obtained from 80 patients 1 year after surgery for ACL injury and 60 normal donors without overt skeletal injury. Total serum RNA was isolated, small non-coding RNAs profiled by TaqMan array MicroRNA (miRNA) analysis and individual small RNA assays performed by quantitative TaqMan RT-PCR (qPCR). Semi-quantitative magnetic resonance imaging (MRI) analysis was performed using Whole Organ Magnetic Resonance Knee Score (WORMS) scoring for analysis of cartilage damage. RESULTS: Initial TaqMan array miRNA profiling showed an increased serum concentration of a small nucleolar RNA (snoRNA), U48, in five patients with cartilage damage compared with that in five patients without cartilage damage and six normal donors. Independent qPCR analysis of snoRNAs in serum from all patients and normal donors showed a strong association between the serum level of another snoRNA, U38, and cartilage damage in ACL injury patients and together with snoRNA, U48, clear distinction between ACL injury patients and normal donors. CONCLUSION: SnoRNAs U38 and U48 are significantly elevated in the serum of patients developing cartilage damage at 1 year after ACL injury. Serum levels of U38 have the potential to facilitate early diagnosis of patients with cartilage damage after ACL injury. This study suggests serum non-coding RNAs may serve as novel noninvasive biomarkers for the detection and assessment of cartilage damage after ACL injury.
Assuntos
Lesões do Ligamento Cruzado Anterior , Cartilagem Articular/lesões , Traumatismos do Joelho/complicações , Osteoartrite do Joelho/sangue , RNA não Traduzido/sangue , Adulto , Idoso , Biomarcadores/sangue , Cartilagem Articular/patologia , Progressão da Doença , Feminino , Humanos , Traumatismos do Joelho/sangue , Traumatismos do Joelho/diagnóstico , Imageamento por Ressonância Magnética , Masculino , Pessoa de Meia-Idade , Osteoartrite do Joelho/etiologia , Osteoartrite do Joelho/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Adulto JovemRESUMO
Type 1 diabetes (T1D) in non-obese diabetic (NOD) mice may be favored by immune dysregulation leading to the hyporesponsiveness of regulatory T cells and activation of effector T-helper type 1 (Th1) cells. The immunoregulatory activity of natural killer T (NKT) cells is well documented, and both interleukin (IL)-4 and IL-10 secreted by NKT cells have important roles in mediating this activity. NKT cells are less frequent and display deficient IL-4 responses in both NOD mice and individuals at risk for T1D (ref. 8), and this deficiency may lead to T1D (refs. 1,6-9). Thus, given that NKT cells respond to the alpha-galactosylceramide (alpha-GalCer) glycolipid in a CD1d-restricted manner by secretion of Th2 cytokines, we reasoned that activation of NKT cells by alpha-GalCer might prevent the onset and/or recurrence of T1D. Here we show that alpha-GalCer treatment, even when initiated after the onset of insulitis, protects female NOD mice from T1D and prolongs the survival of pancreatic islets transplanted into newly diabetic NOD mice. In addition, when administered after the onset of insulitis, alpha-GalCer and IL-7 displayed synergistic effects, possibly via the ability of IL-7 to render NKT cells fully responsive to alpha-GalCer. Protection from T1D by alpha-GalCer was associated with the suppression of both T- and B-cell autoimmunity to islet beta cells and with a polarized Th2-like response in spleen and pancreas of these mice. These findings raise the possibility that alpha-GalCer treatment might be used therapeutically to prevent the onset and recurrence of human T1D.
Assuntos
Diabetes Mellitus Tipo 1/prevenção & controle , Galactosilceramidas/farmacologia , Células Matadoras Naturais/imunologia , Animais , Anticorpos Monoclonais/farmacologia , Antígenos CD1/genética , Ciclofosfamida/toxicidade , Diabetes Mellitus Tipo 1/induzido quimicamente , Diabetes Mellitus Tipo 1/imunologia , Interferon gama/metabolismo , Interleucina-4/metabolismo , Interleucina-7/farmacologia , Células Matadoras Naturais/efeitos dos fármacos , Selectina L/metabolismo , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos NOD , Camundongos Mutantes , Receptores de Interleucina/efeitos dos fármacos , Receptores de Interleucina/imunologia , Receptores de Interleucina-10 , Baço/efeitos dos fármacos , Baço/metabolismoRESUMO
Skin and corneal wounds in diabetics are a major healthcare burden. MicroRNAs are small, non-coding RNAs that post-transcriptionally regulate the expression of protein-coding genes. Studies have identified microRNAs involved in all phases of wound healing. The dysregulation of microRNAs can contribute to impaired or delayed skin and corneal wound healing in diabetics. Here, we present a comprehensive review of the literature involving microRNAs in diabetic skin and corneal wound healing as well as those serving as potential biomarkers for diabetic wound healing.
RESUMO
At high concentrations, the tubule poison paclitaxel is able to kill cancer cells that express Bcl-2; it inhibits the antiapoptotic activity of Bcl-2 by inducing its phosphorylation. To localize the site on Bcl-2 regulated by phosphorylation, mutant forms of Bcl-2 were constructed. Mutant forms of Bcl-2 with an alteration in serine at amino acid 70 (S70A) or with deletion of a 60-aa loop region between the alpha1 and alpha2 helices (Deltaloop Bcl-2, which also deletes amino acid 70) were unable to be phosphorylated by paclitaxel treatment of MDA-MB-231 cells into which the genes for the mutant proteins were transfected. The Deltaloop mutant completely inhibited paclitaxel-induced apoptosis. In cells expressing the S70A mutant, paclitaxel induced about one-third the level of apoptosis seen with wild-type Bcl-2. To evaluate the role of mitogen-activated protein kinases (MAPKs) in Bcl-2 phosphorylation, the activation of c-Jun N-terminal kinase (JNK), extracellular signal-regulated kinase (ERK), and p38 was examined. Paclitaxel-induced apoptosis was associated with phosphorylation of Bcl-2 and activation of ERK and JNK MAPKs. If JNK activation was blocked by transfections with either a stress-activated protein kinase kinase dominant-negative (K-->R) gene (which prevents the activation of a kinase upstream of JNK) or MAPK phosphatase-1 gene (which dephosphorylates and inactivates JNK), Bcl-2 phosphorylation did not occur, and the cells were not killed by paclitaxel. By contrast, neither an ERK inhibitor (PD098059) nor p38 inhibitors (SB203580 and SB202190) had an effect on Bcl-2 phosphorylation. Thus, our data show that the antiapoptotic effects of Bcl-2 can be overcome by phosphorylation of Ser-70; forms of Bcl-2 lacking the loop region are much more effective at preventing apoptosis than wild-type Bcl-2 because they cannot be phosphorylated. JNK, but not ERK or p38 MAPK, appear to be involved in the phosphorylation of Bcl-2 induced by paclitaxel.
Assuntos
Apoptose/efeitos dos fármacos , Proteínas Quinases Ativadas por Mitógeno , Paclitaxel/toxicidade , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Alanina , Substituição de Aminoácidos , Apoptose/fisiologia , Neoplasias da Mama , Proteínas Quinases Dependentes de Cálcio-Calmodulina/metabolismo , Citosol/metabolismo , Feminino , Humanos , Proteínas Quinases JNK Ativadas por Mitógeno , Mitocôndrias/metabolismo , Mutagênese Sítio-Dirigida , Fosforilação , Mutação Puntual , Proteínas Proto-Oncogênicas c-bcl-2/química , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Deleção de Sequência , Serina , Transdução de Sinais , Transfecção , Células Tumorais Cultivadas , Vincristina/toxicidadeRESUMO
Chromosomal regions near the mu opioid receptor gene are implicated in morphine preference by quantitative trait loci studies. Differences in expression of the mu opioid receptor are expected to contribute to differences in inter-individual (humans) or strain-specific (mice) responses to painful stimuli, opiate drugs, and addictive behaviors. The search for relevant genetic elements is hindered by a lack of inter-strain (or inter-individual) genomic sequence information. This work describes 9.3 kb of DNA sequence surrounding exons 2 and 3 of the murine mu opioid receptor gene from both 129/Sv and C57BL/6 strains. While the exons are perfectly conserved, intronic sequences demonstrate approximately a 2.5% divergence between the strains. Polymorphism within these intronic regions may effect either primary transcript stability or C-terminal splicing. Homologous recombination frequencies of targeting vectors harboring mu opioid receptor gene sequences have also been compared in embryonic stem cells derived from these strains. Non-isogenic targeting reduces homologous recombination in both 129/Sv and C57BL/6 embryonic stem cells by greater than 15-fold. These findings are the first to examine C57BL/6 embryonic stem cells for non-isogenic targeting frequencies and to define polymorphisms that exist between these mouse strains which might contribute to opioid behaviors.
Assuntos
Marcação de Genes , Camundongos/classificação , Camundongos/genética , Polimorfismo Genético/genética , Receptores Opioides mu/genética , Recombinação Genética/genética , Células-Tronco/metabolismo , Alelos , Animais , Sequência de Bases , Embrião de Mamíferos/citologia , Embrião de Mamíferos/metabolismo , Frequência do Gene/genética , Vetores Genéticos/genética , Genoma , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos , Dados de Sequência Molecular , Homologia de Sequência do Ácido Nucleico , Especificidade da EspécieRESUMO
Phosphocholine (PC) is the immunodominant epitope found on the surface of Streptococcus pneumoniae (SPn). T15-idiotype Abs, whose heavy (H) chain variable region is encoded by the V1 gene, are dominant in the anti-PC response in adult mice and protect mice from lethal pneumococcal infection. The ability of anti-PC Abs using H chains other than the V1 H chain to protect against pneumococcal infection remains controversial. We generated V1(-/-) knockout mice to determine whether protective anti-PC Abs could be produced in the absence of the V1 gene. No anti-PC Abs were produced in V1(-/-) mice immunized with avirulent SPn; however, PC-BSA binding Abs were induced after immunization with PC-keyhole limpet hemocyanin but at significantly lower levels than those in wild-type mice. These Abs provided poor protection against virulent SPn; thus, <25% of V1(-/-) mice survived challenge with 10(4) bacteria as compared with 100% survival of V1(+/+) mice. The anti-PC Abs in V1(-/-) mice were heteroclitic, binding to nitrophenyl-PC better than to PC. None of nine hybridomas produced from V1(-/-) mice provided passive protection. However, the V1(-/-) mice produced normal amounts of Ab to SPn proteins that can partially protect mice against SPn. These data indicate that the V1 gene is critical for the production of anti-PC Abs providing optimum protection against infection with SPn, and the V1(-/-) mice could be useful in unmasking epitopes other than the immunodominant PC epitope on SPn capable of providing cross protection.