RESUMEN
In cortical synaptosomes of Experimental Autoimmune Encephalomyelitis (EAE) mice at the early stage of disease (13 days post immunization, d.p.i.), the Regulated upon Activation Normal T cell Expressed and Secreted (RANTES, CCL5)-mediated control of [3H]D-aspartate ([3H]D-ASP) exocytosis elicited by a mild depolarizing stimulus (12 mM KCl) shifted from inhibition to facilitation. By using selective antagonists for the chemokine receptor (CCR) 1, 3, and 5 subtypes, we found that the pharmacological profile of the receptor(s) accounting for CCL5-mediated effect was unaltered when compared to control. Inasmuch, CCR protein expression was unaltered. This studies was not extended at 21 d.p.i. since, at this stage, CCL5 failed to affect [3H]D-ASP exocytosis. At 13 d.p.i., the expression of CCR proteins was largely conserved when compared to control. In spinal cord synaptosomes of EAE mice at 21 d.p.i., when presynaptic defects became evident, the [3H]D-ASP exocytosis elicited by 15 mM KCl was significantly increased when compared to control and it was significantly potentiated by 1 nM CCL5. The antagonist pharmacological profile and the western blot analysis of the CCR proteins unveiled that the receptor repertoire involved was unmodified. Differently from controls, however, the CCR1 antagonist BX513 efficiently inhibited on its own [3H]D-ASP exocytosis suggesting that this receptor could have adopted an active conformation. Altogether, our observations favor the use of CCR antagonists to the cure of neurological symptoms in patients suffering from demyelinating syndrome.
Asunto(s)
Adaptación Fisiológica , Corteza Cerebral/metabolismo , Quimiocina CCL5/metabolismo , Encefalomielitis Autoinmune Experimental/metabolismo , Terminales Presinápticos/metabolismo , Receptores CCR1/metabolismo , Médula Espinal/metabolismo , Animales , Ácido Aspártico/metabolismo , Quimiocina CCL5/agonistas , Quimiocina CCL5/genética , Exocitosis , Femenino , Ratones , Ratones Endogámicos C57BL , Receptores CCR1/antagonistas & inhibidores , Sinaptosomas/metabolismoRESUMEN
CCR5 is the major HIV-1 entry coreceptor. RANTES/CCL5 analogs are more potent inhibitors of infection than native chemokines; one class activates and internalizes CCR5, one neither activates nor internalizes, and a third partially internalizes without activation. Here we show that mutations in CCR5 transmembrane domains differentially impact the activity of these three inhibitor classes, suggesting that the transmembrane region of CCR5, a key interaction site for inhibitors, is a sensitive molecular switch, modulating receptor activity.
Asunto(s)
Quimiocina CCL5/fisiología , Mutación , Receptores CCR5/genética , Receptores CCR5/fisiología , Secuencia de Aminoácidos , Sustitución de Aminoácidos , Fármacos Anti-VIH/farmacología , Línea Celular , Quimiocina CCL5/agonistas , Quimiocina CCL5/antagonistas & inhibidores , Quimiocina CCL5/genética , VIH-1/patogenicidad , VIH-1/fisiología , Interacciones Huésped-Patógeno , Humanos , Datos de Secuencia Molecular , Mutagénesis Sitio-Dirigida , Estructura Terciaria de Proteína , Receptores CCR5/química , Transducción de Señal , Internalización del Virus/efectos de los fármacosRESUMEN
We previously identified a novel transcript, mouse (m)IL-20R1a, generated by alternative splicing of the mIL-20R1 gene and studied its possible in vitro functions. However, the function of mIL-20R1a in vivo is unknown. Overexpression of mIL-20R1a in transgenic FvB/N mice resulted in the pathological change of excess calcium deposited in the kidneys. The interplay between renal epithelial cells and calcium oxalate (CaOx) was important in the crystallization involved in the formation of renal stones (nephrolithiasis). Thus, we investigated and compared the responses of mouse renal proximal (TKPTS) and collecting (M-1) tubule cell lines to CaOx with or without mIL-20R1a. The renal epithelial cell lines exposed to CaOx in the presence of mIL-20R1a showed significantly increased lactate dehydrogenase release; loss of cell viability through apoptosis; increased CaOx internalization; higher tumor necrosis factor (TNF)-alpha, MCP-1 and RANTES expression; and higher reactive oxygen species production. Interleukin-6, TNF-alpha and MCP-1 were also upregulated in the kidneys of mIL-20R1a transgenic mice. These effects of mIL-20R1a on CaOx-exposed renal epithelial cells showed that mIL-20R1a functioned as an aggravating factor in promoting calcium deposition in kidney of mice.