RESUMEN
rIL-10 plays a major role in restricting exaggerated inflammatory and immune responses, thus preventing tissue damage. However, the restriction of inflammatory and immune responses by IL-10 can also favor the development and/or persistence of chronic infections or neoplasms. Dogs that succumb to canine leishmaniasis (CanL) caused by L. infantum develop exhaustion of T lymphocytes and are unable to mount appropriate cellular immune responses to control the infection. These animals fail to mount specific lymphoproliferative responses and produce interferon gamma and TNF-alpha that would activate macrophages and promote destruction of intracellular parasites. Blocking IL-10 signaling may contribute to the treatment of CanL. In order to obtain a tool for this blockage, the present work endeavored to identify the canine casIL-10R1 amino acid sequence, generate a recombinant baculovirus chromosome encoding this molecule, which was expressed in insect cells and subsequently purified to obtain rcasIL-10R1. In addition, rcasIL-10R1 was able to bind to homologous IL-10 and block IL-10 signaling pathway, as well as to promote lymphoproliferation in dogs with leishmaniasis caused by L. infantum.
Asunto(s)
Interleucina-10/metabolismo , Leishmaniasis/tratamiento farmacológico , Receptores de Interleucina-10/metabolismo , Animales , Línea Celular , Citocinas/metabolismo , Enfermedades de los Perros/genética , Perros , Femenino , Inmunidad Celular/inmunología , Inmunidad Celular/fisiología , Interferón gamma/genética , Interleucina-10/agonistas , Interleucina-12/genética , Leishmania infantum/inmunología , Leishmania infantum/patogenicidad , Leishmaniasis/inmunología , Macrófagos/metabolismo , Masculino , Ratones , Receptores de Interleucina-10/efectos de los fármacos , Transducción de Señal , Linfocitos T/inmunología , Factor de Necrosis Tumoral alfaRESUMEN
Visceral leishmaniasis is a fatal parasitic neglected disease affecting 1.5 million people worldwide. Based on a drug repositioning approach, the aim of this work was to investigate the in vitro immunomodulatory potential of buparvaquone (BPQ) and to establish a safe regimen to evaluate the in vivo efficacy of BPQ entrapped by negatively charged nanoliposomes (BPQ-LP) in Leishmania infantum-infected hamsters. Small-angle X-ray scattering, dynamic light scattering, and the ζ-potential were applied in order to study the influence of BPQ on the liposome structure. Our data revealed that BPQ was located in the polar-apolar interface, snorkeling the polar region, and protected against aggregation inside the lipophilic region. The presence of BPQ also decreased the Z-average hydrodynamic diameter and increased the surface charge. Compared to intravenous and intramuscular administration, a subcutaneous route was a more effective route for BPQ-LP; at 0.4 mg/kg, BPQ-LP reduced infection in the spleen and liver by 98 and 96%, respectively. Treatment for 5 days resulted in limited efficacy, but 10 days of treatment resulted in an efficacy similar to that of a 15-day regimen. The nanoliposomal drug was highly effective, with a mean 50% effective dose of 0.25 mg/kg, reducing the parasite load in bone marrow by 80%, as detected using quantitative PCR analysis. In addition, flow cytometry studies showed that BPQ upregulated cytokines as tumor necrosis factor, monocyte chemoattractant protein 1, interleukin-10 (IL-10), and IL-6 in Leishmania-infected macrophages, eliminating the parasites via a nitric oxide-independent mechanism. This new formulation proved to be a safe and effective treatment for murine leishmaniasis that could be a useful candidate against visceral leishmaniasis.
Asunto(s)
Antiprotozoarios/farmacología , Factores Inmunológicos/farmacología , Leishmania infantum/efectos de los fármacos , Leishmaniasis Visceral/tratamiento farmacológico , Liposomas/química , Macrófagos/efectos de los fármacos , Naftoquinonas/farmacología , Administración Cutánea , Animales , Antiprotozoarios/química , Médula Ósea/efectos de los fármacos , Médula Ósea/inmunología , Médula Ósea/parasitología , Quimiocina CCL2/agonistas , Quimiocina CCL2/biosíntesis , Cricetinae , Modelos Animales de Enfermedad , Composición de Medicamentos/métodos , Factores Inmunológicos/química , Interleucina-10/agonistas , Interleucina-10/biosíntesis , Interleucina-6/agonistas , Interleucina-6/biosíntesis , Leishmania infantum/crecimiento & desarrollo , Leishmaniasis Visceral/inmunología , Leishmaniasis Visceral/parasitología , Liposomas/farmacocinética , Hígado/efectos de los fármacos , Hígado/inmunología , Hígado/parasitología , Macrófagos/inmunología , Macrófagos/parasitología , Masculino , Ratones , Nanoestructuras/administración & dosificación , Nanoestructuras/química , Naftoquinonas/química , Carga de Parásitos , Bazo/efectos de los fármacos , Bazo/inmunología , Bazo/parasitología , Electricidad Estática , Factor de Necrosis Tumoral alfa/agonistas , Factor de Necrosis Tumoral alfa/biosíntesisRESUMEN
Proteinuria is critical in the tubulointerstitial changes that ultimately lead to renal insufficiency. Increased protein filtration has direct toxic effects on tubular epithelial cells, leading to epithelial mesenchymal transition (EMT) to a myofibroblast phenotype. Angiotensin II and transforming growth factor (TGF)-ß1 are the main mediators of EMT. Calcitriol may exert a potential renoprotective effect by reducing the activity of the renin angiotensin system by suppressing renin gene expression and also by inhibiting the proinflammatory nuclear factor-κB pathway. The present study investigated the benefits of calcitriol treatment in a puromycin-induced proteinuric nephropathy model. Uninephrectomized adult male Wistar rats received intraperitoneal administration of a single dose of puromycin (100 mg/kg) or vehicle. After eight weeks, the animals were divided into two groups and received vehicle or calcitriol (0.5 µg/kg) for four weeks. The vehicle-treated, proteinuric rats developed progressive proteinuria and tubulointerstitial fibrosis after 12 weeks. Increased collagen deposition and fibrosis were significantly ameliorated by calcitriol treatment. Calcitriol was effective in preventing an increase in the EMT markers, α-smooth muscle actin and fibroblast-specific protein 1, reducing macrophage infiltration as evidenced by levels of ED-1. In addition, calcitriol increased the anti-inflammatory cytokine interleukin-10 and reduced the pro-oxidant p47 phox enzyme. These effects were paralleled by a reduction in TGF-ß/Smad3 expression. Calcitriol may have therapeutic potential in the proteinuric nephropathy model used in the present study by inhibiting the TGF-ß1 axis.
Asunto(s)
Calcitriol/farmacología , Riñón/efectos de los fármacos , Nefritis Intersticial/tratamiento farmacológico , Proteinuria/tratamiento farmacológico , Factor de Crecimiento Transformador beta1/antagonistas & inhibidores , Vitaminas/farmacología , Actinas/genética , Actinas/metabolismo , Animales , Biomarcadores/metabolismo , Proteínas de Unión al Calcio/genética , Proteínas de Unión al Calcio/metabolismo , Modelos Animales de Enfermedad , Ectodisplasinas/genética , Ectodisplasinas/metabolismo , Transición Epitelial-Mesenquimal/efectos de los fármacos , Fibrosis , Expresión Génica , Interleucina-10/agonistas , Interleucina-10/genética , Interleucina-10/metabolismo , Riñón/metabolismo , Riñón/patología , Masculino , NADPH Oxidasas/antagonistas & inhibidores , NADPH Oxidasas/genética , NADPH Oxidasas/metabolismo , Nefrectomía , Nefritis Intersticial/inducido químicamente , Nefritis Intersticial/metabolismo , Nefritis Intersticial/patología , Proteinuria/inducido químicamente , Proteinuria/metabolismo , Proteinuria/patología , Puromicina , Ratas , Ratas Wistar , Sistema Renina-Angiotensina/efectos de los fármacos , Proteína smad3/antagonistas & inhibidores , Proteína smad3/genética , Proteína smad3/metabolismo , Factor de Crecimiento Transformador beta1/genética , Factor de Crecimiento Transformador beta1/metabolismoRESUMEN
OBJECTIVE: The beneficial effects of kappa opioid agonist U-50,488 in preventing periodontal disease (PD) progression in rats have already been described, but its mechanism of action is unknown. The present study evaluated the expression of TNF-α, IL-6, IL-8 and IL-10 in the gingival tissues of rats with ligature-induced PD, treated with U-50,488. It also correlated the effects of this agonist with myeloperoxidase (MPO) activity and the presence of osteoclasts. DESIGN: Male Holtzman rats weighing 250-300 g were divided into four groups: (1) control, (2) ligature, (3) ligature+saline and (4) ligature+kappa agonist. Experimental PD was induced by placing a sterile silk ligature around the 2nd left upper molar. Rats from groups 3 to 4 were locally administered with either saline or U-50,488, respectively, from day 3 to day 5 following ligation. After 5 or 11 days, the rats were euthanized and periodontal tissue samples were collected for histological and morphometric analysis and for determination of TNF-α, IL-6, IL-8, IL-10 and MPO. RESULTS: Ligature placement induced significant alveolar bone loss. The number of osteoclasts, degree of MPO activity, IL-6, IL-8 and TNF-α expression were also increased by PD. U-50,488 reduced both bone loss and the number of osteoclasts, but did not alter histological inflammatory infiltrate or MPO activity. U-50,488 significantly reduced IL-6 and increased IL-10 levels, but did not affect TNF-α and IL-8. CONCLUSION: Lowering the levels of IL-6 and increasing IL-10 are important mechanisms by which U-50,488 reduces alveolar bone loss in ligature-induced periodontal disease.
Asunto(s)
3,4-Dicloro-N-metil-N-(2-(1-pirrolidinil)-ciclohexil)-bencenacetamida, (trans)-Isómero/farmacología , Pérdida de Hueso Alveolar/prevención & control , Interleucina-10/agonistas , Interleucina-6/antagonistas & inhibidores , Receptores Opioides kappa/agonistas , Análisis de Varianza , Animales , Modelos Animales de Enfermedad , Interleucina-8/antagonistas & inhibidores , Masculino , Osteoclastos/efectos de los fármacos , Peroxidasa/antagonistas & inhibidores , Ratas , Ratas Sprague-Dawley , Factor de Necrosis Tumoral alfa/antagonistas & inhibidoresRESUMEN
Tumor cells treated with IL-10 were shown to have decreased, but peptide-inducible expression of MHC class I, decreased sensitivity to MHC class I-restricted CTL, and increased NK sensitivity. These findings could be explained, at least partially, by a down-regulation of TAP1/TAP2 expression. In this study, IT9302, a nanomeric peptide (AYMTMKIRN), homologous to the C-terminal of the human IL-10 sequence, was demonstrated to mimic these previously described IL-10 effects on MHC class I-related molecules and functions. We observed a dose-dependent down-regulation of MHC class I at the cell surface of melanoma cells after 24-h treatment with IT9302. The IL-10 homologue peptide also caused a dose-dependent inhibition of the IFN-gamma-mediated surface induction of MHC class I in a melanoma cell line. We demonstrated, using Western blot and flow cytometry, that IT9302 inhibits the expression of TAP1 and TAP2 proteins, but not MHC class I H chain or low molecular protein molecules. Finally, peptide-treated melanoma cells were shown to be more sensitive to lysis by NK cells in a dose-dependent way. Taken together, these results demonstrate that a small synthetic peptide derived from IL-10 can mimic the Ag presentation-related effects mediated by this cytokine in human melanomas and increase tumor sensitivity to NK cells, which can be relevant in the designing of future strategies for cancer immune therapy.
Asunto(s)
Transportadoras de Casetes de Unión a ATP/biosíntesis , Neoplasias del Ojo/metabolismo , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Antígenos de Histocompatibilidad Clase I/biosíntesis , Interleucina-10/agonistas , Melanoma/metabolismo , Proteínas de Neoplasias/biosíntesis , Oligopéptidos/farmacología , Transportador de Casetes de Unión a ATP, Subfamilia B, Miembro 2 , Miembro 3 de la Subfamilia B de Transportadores de Casetes de Unión a ATP , Transportadoras de Casetes de Unión a ATP/genética , Línea Celular Tumoral/efectos de los fármacos , Línea Celular Tumoral/metabolismo , Cisteína Endopeptidasas/biosíntesis , Cisteína Endopeptidasas/genética , Citotoxicidad Inmunológica , Relación Dosis-Respuesta a Droga , Neoplasias del Ojo/patología , Genes MHC Clase I , Humanos , Interferón gamma/antagonistas & inhibidores , Interferón gamma/farmacología , Interleucina-10/química , Células Asesinas Activadas por Linfocinas/inmunología , Melanoma/patología , Proteínas de Neoplasias/genética , Estructura Terciaria de Proteína , Proteínas RecombinantesRESUMEN
Leishmaniases have a high prevalence in tropical countries. In order to improve existing diagnostic systems based on total Leishmania proteins, and to identify antigen candidates for vaccine development, an intensive search for the identification of antigens was performed using molecular biology techniques. In this study, the immune response to three L. infantum recombinant antigens was evaluated. Upon stimulation with KMP11, mononuclear cells from leishmaniasis patients produced high levels of IL-10, while a predominant IFN-gamma production could be observed in cultures stimulated with H2A and soluble Leishmania antigen. All the recombinant antigens induced very little IL-5. KMP11 decreased IFN-gamma production by 48% in cultures of peripheral blood mononuclear cells from cutaneous leishmaniasis patients who had been stimulated with soluble Leishmania antigen. Furthermore, antibodies to KMP11 were detected in the sera from all patients with visceral leishmaniasis and in the majority of the sera from patients with cutaneous leishmaniasis or individuals with asymptomatic L. chagasi infection. Thus, KMP11 is recognized by cells and sera of patients with different clinical forms of leishmaniasis, and KMP11, through IL-10 production, proved to be a potent antigen in modulating type 1 immune response.