RESUMEN
Renal ischemia-reperfusion injury (IRI) rapidly induces production of inflammatory mediators including, and in particular, tumor necrosis factor (TNF). Possible sources include resident parenchymal and bone marrow-derived cells as well as recruited leukocytes. Cell suspensions from kidneys subjected to IRI were examined by cell separation followed by in vitro culture and enzyme-linked immunosorbent assay (ELISA), immunoperoxidase and immunofluorescence microscopy, and multicolor flow cytometry to determine the contribution of dendritic cells (DCs) to early production of TNF and other inflammatory mediators. Secretion of TNF, interleukin (IL-6), monocyte chemoattractant protein-1 (MCP-1), and regulated on activation normal T cell expressed and secreted (RANTES) was increased in cell suspensions from IRI compared with control kidneys and was higher in DC-enriched preparations. Immunostaining identified TNF(+ve) cells that coexpressed the DC marker CD11c. Flow cytometry of bone marrow-derived (CD45(+ve)) cell populations at 24 h post-IRI demonstrated that F4/80(+ve)/CD11c(+ve) DCs remained proportionately stable and exhibit higher levels of DC maturation markers, whereas the proportion of F4/80(-ve) DCs, monocytes, neutrophils, and T cells increased. Intracellular staining for TNF confirmed that F4/80(+ve) DCs were the predominant TNF(+ve) cell and expressed higher levels than other TNF(+ve) cells. In vivo depletion of DCs from the kidney substantially attenuated TNF secretion by total and CD45(+ve) cells following IRI. The results uncover a role for resident F4/80(+ve) DCs as the predominant secretors of TNF within 24 h of IRI.
Asunto(s)
Células Dendríticas/metabolismo , Riñón/irrigación sanguínea , Daño por Reperfusión/inmunología , Factor de Necrosis Tumoral alfa/metabolismo , Animales , Antígenos CD11/biosíntesis , Ratones , Ratones Endogámicos C57BLRESUMEN
Dendritic cells (DCs) play a central role in regulating immune activation and responses to self. DC maturation is central to the outcome of antigen presentation to T cells. Maturation of DCs is inhibited by physiological levels of 1alpha,25 dihydroxyvitamin D(3) [1alpha,25(OH)(2)D(3)] and a related analog, 1alpha,25(OH)(2)-16-ene-23-yne-26,27-hexafluoro-19-nor-vitamin D(3) (D(3) analog). Conditioning of bone marrow cultures with 10(-10) M D(3) analog resulted in accumulation of immature DCs with reduced IL-12 secretion and without induction of transforming growth factor beta1. These DCs retained an immature phenotype after withdrawal of D(3) analog and exhibited blunted responses to maturing stimuli (CD40 ligation, macrophage products, or lipopolysaccharide). Resistance to maturation depended on the presence of the 1alpha,25(OH)(2)D(3) receptor (VDR). In an in vivo model of DC-mediated antigen-specific sensitization, D(3) analog-conditioned DCs failed to sensitize and, instead, promoted prolonged survival of subsequent skin grafts expressing the same antigen. To investigate the physiologic significance of 1alpha,25(OH)(2)D(3)/VDR-mediated modulation of DC maturity we analyzed DC populations from mice lacking VDR. Compared with wild-type animals, VDR-deficient mice had hypertrophy of subcutaneous lymph nodes and an increase in mature DCs in lymph nodes but not spleen. We conclude that 1alpha,25(OH)(2)D(3)/VDR mediates physiologically relevant inhibition of DC maturity that is resistant to maturational stimuli and modulates antigen-specific immune responses in vivo.
Asunto(s)
Calcitriol/farmacología , Células Dendríticas/efectos de los fármacos , Receptores de Calcitriol/fisiología , Animales , Antígenos CD/análisis , Antígeno B7-1/análisis , Antígeno B7-2 , Calcitriol/análogos & derivados , Células Dendríticas/fisiología , Femenino , Masculino , Glicoproteínas de Membrana/análisis , Ratones , Ratones Endogámicos C57BL , Trasplante de Piel , Factor de Crecimiento Transformador beta/biosíntesisRESUMEN
PURPOSE: To measure the concentration of ascorbic acid in the human corneal epithelium. METHODS: Corneal epithelium was removed from postmortem eyes 4 to 16 hours after death and ascorbate measured by high-performance liquid chromatography. RESULTS: The concentration of ascorbate was 1.33 +/- 0.48 mg/gm wet weight (mean +/- SD), estimated to be 14 times its concentration in the aqueous humor. CONCLUSIONS: Ascorbate can protect the basal layer of the epithelium by absorption of incident ultraviolet radiation.
Asunto(s)
Ácido Ascórbico/análisis , Epitelio Corneal/química , Adulto , Anciano , Anciano de 80 o más Años , Humor Acuoso/química , Cromatografía Líquida de Alta Presión , Femenino , Humanos , Masculino , Persona de Mediana EdadRESUMEN
We show that the immunosuppressive effects of 1alpha, 25-dihydroxyvitamin D(3) (1alpha,25(OH)(2)D(3)) are due, in part, to inhibition of the T cell stimulatory functions of dendritic cells (DCs). Addition of 10(-12) and 10(-8) M 1alpha,25(OH)(2)D(3) to murine DC cultures resulted in a concentration-dependent reduction in levels of class II MHC and the co-stimulatory ligands B7-1, B7-2, and CD40 without affecting the number of DCs generated. Higher concentrations of 1alpha,25(OH)(2)D(3) reduced DC yield. The capacity of DCs to induce proliferation of purified allogeneic T cells was reduced by 1alpha,25(OH)(2)D(3). The vitamin D(3) analog, 1alpha,25(OH)(2)-16-ene-23-yne-26,27-hexafluoro-19-nor -D(3), exerted identical effects at 100-fold lower concentrations. Inhibition of DC maturation and stimulatory function was absent in cultures from mice genetically lacking vitamin D receptors (VDR). Vitamin D analogs effectively reduce DC function via VDR-dependent pathways.
Asunto(s)
Calcitriol/farmacología , Diferenciación Celular/efectos de los fármacos , Colecalciferol/análogos & derivados , Células Dendríticas/citología , Células Madre Hematopoyéticas/citología , Receptores de Calcitriol/fisiología , Linfocitos T/inmunología , Animales , Calcitriol/análogos & derivados , Células Cultivadas , Colecalciferol/farmacología , Células Dendríticas/efectos de los fármacos , Células Dendríticas/inmunología , Femenino , Células Madre Hematopoyéticas/efectos de los fármacos , Células Madre Hematopoyéticas/inmunología , Antígenos de Histocompatibilidad Clase II/análisis , Integrina alfaXbeta2/análisis , Activación de Linfocitos/efectos de los fármacos , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Receptores de Calcitriol/deficiencia , Receptores de Calcitriol/genética , Linfocitos T/citología , Linfocitos T/efectos de los fármacosRESUMEN
The purpose of this study was to measure the effect of topical ibopamine and epinephrine on intraocular pressure in undisturbed rabbits. Six pigmented rabbits were outfitted with a telemetric transducer system connected via catheter to the vitreous cavity of one eye for continuous monitoring of intraocular pressure. After entrainment to a 12 hour light, 12 hour dark cycle and after stabilization of the circadian rhythm of intraocular pressure, the animals were tested with 2% ibopamine and 2% epinephrine. Each drug was instilled during the light phase and during the dark phase of the circadian cycle. When administered during the light phase, both drugs caused a transient pressure rise followed by prolonged hypotension. When administered during the dark phase, neither drug caused a pressure rise but both drugs caused prolonged hypotension. It was concluded that ibopamine and epinephrine cause identical intraocular pressure changes in the normal rabbit eye. The effect was dependent on the timing of administration during the circadian cycle. Since ibopamine is a pro-drug of deoxyepinephrine (N-methyl dopamine, epinine), its effects are assumed to be due to this metabolite, a metabolite that is structurally similar to epinephrine.