RESUMO
To establish the relevance of targeting disease-associated T cells in anti-RNP-associated glomerulonephritis, mice developing nephritis following immunization with U1-70-kd small nuclear ribonucleoprotein (snRNP) were treated with a single dose of irradiated antigen-selected T cell vaccine. T cell receptor usage in nephritic kidneys revealed oligoclonal use of T Cell Receptor V Beta (TRBV) genes as previously found in spleens and lungs of immunized mice with pulmonary disease. The CDR3 regions from T cell isolates showed sequence homology to those in humans with anti-RNP autoimmunity. Following T cell vaccination, urinalysis returned to normal in 5/7 treated mice (71% response rate) whereas all mock-treated mice continued to have an active urinary sediment (Fisher's Exact p=0.02). An oligoclonal population of T cells homologous to those identified in humans with anti-RNP autoimmunity is implicated in disease pathogenesis, and T cell vaccination is associated with a high rate of clinical improvement in established nephritis.
Assuntos
Nefrite Lúpica/terapia , Ribonucleoproteína Nuclear Pequena U1/imunologia , Linfócitos T/imunologia , Vacinação/métodos , Sequência de Aminoácidos/genética , Animais , Autoanticorpos/sangue , Autoanticorpos/imunologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/patologia , Proliferação de Células , Regiões Determinantes de Complementaridade/genética , Antígenos HLA-DR/genética , Humanos , Glomérulos Renais/patologia , Nefrite Lúpica/imunologia , Nefrite Lúpica/patologia , Nefrite Lúpica/urina , Ativação Linfocitária/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/imunologia , Ribonucleoproteína Nuclear Pequena U1/genética , Baço/citologia , Baço/imunologia , Linfócitos T/patologia , UrináliseRESUMO
We previously showed that rat taste buds express several adenylyl cyclases (ACs) of which only AC8 is known to be stimulated by Ca2+. Here we demonstrate by direct measurements of cAMP levels that AC activity in taste buds is stimulated by treatments that elevate intracellular Ca2+. Specifically, 5 microM thapsigargin or 3 microM A-23187 (calcium ionophore), both of which increase intracellular Ca2+ concentration ([Ca2+]i), lead to a significant elevation of cAMP levels. This calcium stimulation of AC activity requires extracellular Ca2+, suggesting that it is dependent on Ca2+ entry rather than release from stores. With immunofluorescence microscopy, we show that the calcium-stimulated AC8 is principally expressed in taste cells that also express phospholipase Cbeta2 (i.e., cells that elevate [Ca2+]i in response to sweet, bitter, or umami stimuli). Taste transduction for sucrose is known to result in an elevation of both cAMP and calcium in taste buds. Thus we tested whether the cAMP increase in response to sucrose is a downstream consequence of calcium elevation. Even under conditions of depletion of stored and extracellular calcium, the cAMP response to sucrose stimulation persists in taste cells. The cAMP signal in response to monosodium glutamate stimulation is similarly unperturbed by calcium depletion. Our results suggest that tastant-evoked cAMP signals are not simply a secondary consequence of calcium modulation. Instead, cAMP and released Ca2+ may represent independent second messenger signals downstream of taste receptors.