Recombinant bovine respiratory syncytial virus with deletion of the SH gene induces increased apoptosis and pro-inflammatory cytokines in vitro, and is attenuated and induces protective immunity in calves

Abstract

Bovine respiratory syncytial virus (BRSV) causes inflammation and obstruction of the small airways, leading to severe respiratory disease in young calves. The virus is closely related to human (H)RSV, a major cause of bronchiolitis and pneumonia in young children. The ability to manipulate the genome of RSV has provided opportunities for the development of stable, live attenuated RSV vaccines. The role of the SH protein in the pathogenesis of BRSV was evaluated in vitro and in vivo using a recombinant (r)BRSV in which the SH gene had been deleted. Infection of bovine epithelial cells and monocytes with rBRSVΔSH, in vitro, resulted in an increase in apoptosis, and higher levels of TNF-α and IL-1β compared with cells infected with parental, wild-type (WT) rBRSV. Although replication of rBRSVΔSH and WT rBRSV, in vitro, were similar, the replication of rBRSVΔSH was moderately reduced in the lower, but not the upper, respiratory tract of experimentally infected calves. Despite the greater ability of rBRSVΔSH to induce pro-inflammatory cytokines, in vitro, the pulmonary inflammatory response in rBRSVΔSH-infected calves was significantly reduced compared with that in calves inoculated with WT rBRSV, 6 days previously. Virus lacking SH appeared to be as immunogenic and effective in inducing resistance to virulent virus challenge, 6 months later, as the parental rBRSV. These findings suggest that rBRSVΔSH may be an ideal live attenuated virus vaccine candidate, combining safety with a high level of immunogenicity.

Mumps Virus SH Protein Inhibits NF-κB Activation by Interacting with Tumor Necrosis Factor Receptor 1, Interleukin-1 Receptor 1, and Toll-Like Receptor 3 Complexes

“…Interference with NF-B activation often results in the reduction of apoptosis and a reduced expression of proinflammatory cytokines such as TNF-␣ or IL-1␤, providing more time for virus replication (21,36,53). We did not observe MuV SH to have a severe impact on virus replication in cell culture, which is in line with previous findings by Malik et al (33).…”

“…Earlier studies usually investigated the function of paramyxoviral SH proteins using viruses lacking the complete SH gene. Cytopathic effect (CPE) and apoptosis induction was enhanced in cells infected with those viruses, which was mainly attributed to an increase of NF-B-dependent secretion of TNF-␣ (19,21,22,(34)(35)(36). Similarly, we found NF-B activation to be enhanced in cells infected with rMuV deficient in SH, confirming that the observed effect was due to the interference of MuV SH with NF-B activation and not a consequence of the disruption of the genomic structure.…”

Partial Attenuation of Respiratory Syncytial Virus with a Deletion of a Small Hydrophobic Gene Is Associated with Elevated Interleukin-1β Responses

“…The observation that deletion of RSV SH leads to an increase in IL-1␤ is supported by studies in bRSV, where a ⌬SH vaccine strain induced higher levels of IL-1␤ in the lungs of infected cattle (17). In the current study, blockade of IL-1␤ prior to infection increased the viral load, supporting the idea that SH might enable immunomodulation.…”

“…This suggests it may play a role in modulating the immune response, with earlier studies showing that RSV SH inhibits tumor necrosis factor (TNF) signaling (16). In a recent study, recombinant bovine RSV (bRSV) with an SH gene deletion was attenuated (17), inducing increased levels of the cytokines interleukin-1␤ (IL-1␤) and TNF. Viral pore proteins have been proposed to modulate the inflammasome, a multiprotein pattern recognition complex that catalyzes the cleavage of the proforms of IL-1␤ and IL-18 into their active forms via caspase 1 (18).…”

“…SH genes are found in all members of the subfamily Pneumovirinae and some of the Paramyxovirinae, including mumps virus (48) and human parainfluenza virus 5 (hPIV5) (49). The SH proteins from human metapneumovirus (hMPV) (50), bRSV (51), hPIV5 (49), and mumps virus (52) are nonessential for in vitro growth, and an immunomodulatory function has been described for the SH proteins from PIV5 (49), bRSV (17), and hMPV (53), which also encodes a viroporin (54). We hypothesize that the pore structure of the SH protein (10) facilitates this immunomodulatory function, counteracting the activation of NLRP3 by ionic changes induced by RSV infection (27).…”