| Background and objective RSV is worldwide in distribution and is a major cause of life-threatening viral pneumonia and bronchiolitis in infants and young children. The first three months of life is peak period of RSV infection with high mortality. All the population are susceptibility to RSV infection, and almost all children during the first 2 yr of life have infected once, and half of them have infected twice. The more younger, the state of illness is more severer. Protective immunity against RSV infection after infection only last several months, so that reinfection often occurred. Long-term and repeated RSV infection will result in chronic abnormal changes in pulmonary function. RSV-induced disease is associated with recurrent episodes of wheezing and asthma in children Effective therapy and vaccines are not available at present. RSV genome is a single strand of negative-sense RNA in which genomic RNA sequences are clear now. A M2 protein key factor in RSV gene expression encodes the essential viral polymerase for viral replication. Investigators found three targeted sites inM2 mRNA, and proved antisense oligodeoxynucleotides targete those sites have most efficient antiviral effects in infected cells.DNAzyme is a catalytic active DNA molecular isolated from artifical random-sequence population using evolutionary process in vitro selection protocols. The find of DNAzyme is a leap of knowledge to understand enzyme. A general RNA-cleaving DNA enzyme is 10-23 consisting of a internal loop as its catalytic domain and two flanking substrate-recognition domains each which is complementary to substrate. It could cleave almost any RNA sequence contains a purine-pyrimidine under simulated physiological conditions. Compared with synthetic RNA enzymes, DNA enzymes are smaller, more efficiently, easier to prepare and less sensitive to chemical and enzymatic degradation. Those features of DNAzymes endow it with tremendous potential for applications both in vitro and in vivo. DNAzyme has been applied in fields such as anti-tumor, cardiovascular diseases and antiviral infection.Based on the conservative sequence NS1-NS2 and M2 mRNA of RSV, our laboratory have designed six DNAzymes selected the efficient and specific catalytic active DNA molecular both in cell-free and in cell cultured system against a RSV mRNA. In this study, we used a selected substrate cleavage DNAzyme DZ8269, which was targeting M2 RNA of RSV, to observe it effect on the anti-RSV infection in mice. The overallobjective of this study is to found the groundwork for proventing and treating patients with RSV infection by using DNAzyme.Method We use 8-12 week-old specific-pathogen-free BALB/c mice. Mice were anesthetized and inoculated through intranasal dripping of 100μl of different titers of RSV, after then sacrificed at different times. Right lungs were removed in aseptic condition to detect the quantity of virus consecutively. Left lungs were fixed for immunohistochemical stain of ICAM-1, in situ hybridization, HE stain and histopathological electromicroscopic examination. Mice were anesthetized for surgically exposing their trachea, in which 100-μl volume Chinese ink was inoculated through a bent needle attached via polyethylene tubing. Other mice were inhaled Chinese ink by using high frequently nebulizer, or intranasal inoculated under light anesthetized 100μl Chinese ink. Sacrificed the mice, remove the lungs for comparing the quantity of Chinese ink in the lung of mice inoculated Chinese ink in different pathways. The mice infected with 104 PFU RSV were used to be tested. After 12h infection, same volume and different dose of DNAzyme were given to the mice through intranasal twice every 12h. Then the general condition of mice was observed everyday, and killed to have right lungs for plaque forming assay, left lungs fixed for immunohistochemical stain of ICAM-1, HE stain to pathological examination and smeares for RT-PCR of RSV G mRNA measurement. Results When mice were infected with 103 PFU RSV...
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