Virus-against-virus dominant-negative interference strategy targeting a viral CC chemokine prevents cytomegalovirus-related neurodevelopmental pathogenesis

Abstract

BackgroundCongenital cytomegalovirus (CMV) infections are one leading cause of human neurodevelopmental disorders. Increasing evidence for the pathogenic involvement of brain immune alterations was obtained in the recent years. Host and virus-encoded chemokines might play important roles in CMV-related neuropathogenesis by regulating leukocyte trafficking and microglia recruitment in the CMV-infected brains, and by interfering with key neurodevelopmental steps. In a rat model of CMV infection of the fetal brain in utero that leads to detrimental neurologic and other severe phenotypes postnatally, we reported on the early alteration of microglia and on the infiltration of the infected brains by lymphoid and myeloid cells. Particularly, expression of the r129 gene encoding the viral chemokine RCK3 was detected as early as 24h post-infection; together with the previously reported chemotaxis properties exerted by RCK3 on lymphocytes and on macrophages in vitro, this suggested that RCK3 might be involved in the brain immune cell alterations seen in the CMV-infected developing brains, and in the related neuropathogenesis. MethodsInfection of the rat fetal brain was done by intracerebroventricular injections in utero of either rat CMV encoding wild-type (wt) RCK3 (RCMV-wt), or a mutant CMV counterpart encoding RCK3 with a deletion in its chemokine domain (RCMV-r129{Delta}NT). As RCMV-r129{Delta}NT had shown dominant-negative effects on the chemotaxis properties of RCK3 in vitro, simultaneous and successive co-infection rescue assays were also performed. The detrimental postnatal phenotypes in vivo and the epileptiform activity ex vivo usually detected after infection of the rat developing brain with RCMV-wt were monitored in the RCMV-r129{Delta}NT condition and in co-infection assays. ResultsIn sharp contrast with RCMV-wt whose infection of the fetal brain led to decreased postnatal survival, impaired sensorimotor development, hindlimb hyperextension and epileptic seizures in neonatal pups, RCMV-r129{Delta}NT infection was not associated with any severe postnatal phenotype in vivo. Consistently, the epileptiform activity recorded in most neocortical slices from RCMV-wt-infected pups was not detected in any slice from RCMV-r129{Delta}NT-infected pups. Simultaneous co-infection assays led to dramatic prevention against the postnatal phenotypes in vivo and the altered network activity ex vivo, revealing a dose-dependent rescuing effect exerted by RCMV-r129{Delta}NT on RCMV-wt. Importantly, successful rescue was also obtained when the mutant RCMV-r129{Delta}NT was inoculated in the fetal brains either before or after infection with RCMV-wt. SignificanceOur data demonstrate the crucial neuropathogenic role of RCK3 CMV chemokine in vivo. The dramatic success and apparent safety of the dominant-negative RCK3 rescue assays in vivo provide a proof-of-principle for the beneficial use of a virus-against-virus approach against CMV-related pathogenesis.