| Rabies, which is caused by rabies virus, has been known as a deadly neurological zoonosis of bothhumans and other warm-blooded animals and remains a major threat to public health. According to thereport from WHO, rabies causes about26,000to55,000deaths worldwide each year, most of whichoccur in Asia and Africa. Canine rabies is responsible for more than90%of the human cases in Asia,Africa and Latin America. In the United States, dog rabies has been largely brought under controlthrough pet vaccination programs. Most of the human cases in the USA have been associated withrabies virus (RABV) found in wildlife, especially bats. The postexposure prophylaxis (PEP) of rabiesmainly includes vaccination and administration of anti-rabies immunoglobulin. Although it is widelyaccepted that there is no effective treatment and rabies is almost always fatal once neurologicalsymptoms develop, PEP is very effective if it is initiated promptly after exposure, because it permitsimmune effectors and antibodies to enter the central nervous system (CNS) tissues and clear virus fromCNS and prevent the development of rabies even if virus exist in the brain. Although human survivorshave been reported recently after treatment with the Milwaukee Protocol or a modification thereof, theeffectiveness of the method has been questioned.Our previous studies revealed that intracerebral administration of rRABV-GMCSF or a combinationof inactivated rRABV and monocyte chemotactic protein-1(MCP-1) can prevent mice from developingrabies by activating and recruiting dentritic cells, stimulating the production of virus neutralizingantibody (VNA), enhancing the BBB permeability, and clearing RABV from the CNS. To investigate ifintravenous administration of VNA together with (MCP-1) can protect mice from RABV infection, ICRmice were infected intramuscularly with10IMLD50(50%mouse intramuscular lethal dose) of a streetRABV (DRV-Mexico) then treated with VNA(i.v.) and (or) MCP-1(i.c.) at different time points afterinfection. It was found that significantly more mice treated with VNA and MCP-1at days3and5afterinfection with DRV were protected from developing rabies than sham-treated mice. No virus could bedetected in the brains of surviving mice, in contrast, high titers of virus were measured in the brains ofdead mice, by virus titration, pathology and immunopathochemestry. On the other hand, no antibodiescould be detected in the brains of dead mice, whereas high titers of VNA and Ig G were measured in thebrains of surviving animals, using FAVN and ELISA testes. The data showed that administration ofVNA and MCP-1can effectively clear virus from the CNS and prevent mice from developing rabies.Since high level of κ light chain mRNA was measured in the brains of surviving ICR mice, in orderto exclude the possibility that B cells entering into the CNS are absolutely required to clear RABV fromthe CNS, the experiment as summarized in ICR mice was repeated in B-cell deficient mice. The resultsshowed that only25%B cells k.o. mice survived when treated with MCP-1once at5d.p.i, however, thesurvivorship was significantly enhanced (up to75%) as long as MCP-1was repeatedly treated at day5and7post infection. Similarly, no virus and high level of neutralizing antibodies could be detected inthe brains of surviving mice, whereas no antibodies and high titers of virus was measured in the brains of dead mice. The data showed that administration of VNA and MCP-1can effectively clear virus fromthe CNS and prevent developing rabies in B cells k.o. mice once the blood-brain barrier (BBB)permeability is enhanced, even absence of VNA-producing plasma cells.To investigate the protection mechanism of administration of VNA and MCP-1for PEP of rabies,answers can be searched in possible two situations: with infection and without infection of rabies virus.In the case without DRV infection, the significant high neutralizing antibody was detected in the brainsof mice treated with VNA and MCP-1, while no VNA was measured in the VNA only-treated group.The result indicated that MCP-1enhanced the BBB permeability, which allows VNA entering the CNSand increases the level of VNA in the brains. In the case of DRV infection, results demonstrated thatBBB permeability started to increase at12h after MCP-1treatment and peeked at36h, then declined tonormal at48h. The expression of tight junction proteins (Occludin, Claudin-5and ZO-1) was reducedas the level of NaF in the brains increased. On the other hand，increasing BBB permeability enhancedthe level of VNA, and which reduced the titers of virus in the brains. In summary, MCP-1enhanced theBBB permeability, permitted VNA enter the CNS and cleared RABV from the CNS, then prevented thedevelopment of rabies. It is confirmed that it is needed for both VNA and enhanced BBB permeabilityin clearing virus from the CNS.Previous studies showed that pathogenicity of rabies virus variants inversely correlates withglycoprotein expression levels. There is an obvious difference in G protein expression levels betweenB2C (low virulent) and DRV (highly virulent) virus strains. To investigate the correlation oftransmembrane anchor (TM) and cytoplasmic domain (CT) of G protein invovled in the glycoproteinexpression levels and pathogenicity of rabies virus, we construct B2C recombinant virus with chimericDRV G protein (TM and CT) using the B2C reverse genetics system. The chimeric viruses wererescued and designated as rB2C-(DRV-TM+CT), rB2C-(DRV-CT) and rB2C-(DRV-TM). It was foundthat no significant difference of virus growth kinetics between the chimeric viruses and parental virus.Further studies need to be conducted to address the virus glycoprotein expression and its correlationwith the viral pathogenicity.In conclusion, intravenous administration of virus neutralizing antibodies can clear rabies virus fromthe CNS in both immunocompetent and immunocompromised mice as long as the blood-brain barrierpermeability is enhanced. |
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