Morpholino oligomer-mediated protection of porcine pulmonary alveolar macrophages from arterivirus-induced cell death.
BACKGROUND
Porcine reproductive and respiratory syndrome (PRRS) causes extensive economic losses in the swine industry. Current strategies and vaccines to control the disease are inadequate. We previously demonstrated that peptide-conjugated phosphorodiamidate morpholino oligomers (PPMOs) could potently inhibit PRRS virus (PRRSV) replication in cell cultures. PPMOs are single-stranded DNA analogues containing a modified backbone and cell-penetrating peptide. PPMOs are nuclease-resistant, water-soluble, can enter cells readily and exhibit highly specific binding to complementary RNA. In this study, we examined PPMO-mediated inhibition of PRRSV replication in a primary culture of porcine pulmonary alveolar macrophages (PAMs).


METHODS
PAMs were collected from piglets, pre-incubated in culture and infected with PRRSV. Viability, cytopathic effects, virus yield and apoptosis of PAMs in the presence or absence of a PPMO (5UP2) were examined. The 5UP2 PPMO is complementary to a conserved sequence in the 5'-terminal region of the PRRSV genome. The level of several interferon-associated gene products and activity of caspases were monitored.


RESULTS
PRRSV infection induced the activity of caspases-3/7, -8 and -9 significantly. Treatment of PAMs with 5UP2 resulted in protection of the cells from PRRSV-induced cell death for at least 7 days and avoided the activation of the caspases evaluated. 5UP2 treatment of PRRSV-infected PAMs also prevented the vigorous induction of interferon-beta and chemokines observed in infected and mock-treated PAMs.


CONCLUSIONS
PPMO-mediated suppression of PRRSV replication in PAMs was associated with a reduction of apoptotic and inflammatory responses. These results provide further rationale for the development of PPMO 5UP2 as an antiviral to control PRRSV infection.
Patel
Deendayal
D
University of Maryland, College Park, MD, USA.
Stein
David A
DA
Zhang
Yan-Jin
YJ
D000818:Animals; D020169:Caspases / Q000096:biosynthesis; D018929:Cell Culture Techniques; D016923:Cell Death / Q000187:drug effects; D018925:Chemokines / Q000096:biosynthesis / Q000235:genetics; D004790:Enzyme Induction / Q000187:drug effects; D015967:Gene Expression Regulation, Viral / Q000187:drug effects; D054884:Host-Pathogen Interactions / Q000187:drug effects; D016899:Interferon-beta / Q000096:biosynthesis / Q000235:genetics; D016676:Macrophages, Alveolar / Q000187:drug effects* / Q000502:physiology / Q000821:virology; D009025:Morpholines / Q000494:pharmacology*; D060172:Morpholinos; D019318:Porcine Reproductive and Respiratory Syndrome / Q000188:drug therapy* / Q000378:metabolism; D019316:Porcine respiratory and reproductive syndrome virus / Q000187:drug effects* / Q000502:physiology; D013552:Swine; D014779:Virus Replication / Q000187:drug effects
D018925:Chemokines; D009025:Morpholines; D060172:Morpholinos; D016899:Interferon-beta; D020169:Caspases