title: In-vitro effect of human cathelicidin antimicrobial peptide LL-37 on dengue virus type 2.
abstract: Human Cathelicidin antimicrobial peptide LL-37 is known to have antiviral activity against many viruses. In the present study, we investigated the in-vitro effect of LL-37 on dengue virus type 2 (DENV-2) infection and replication in Vero E6 cells. To study the effect of pretreatment of virus or cells with LL-37, the virus was pretreated with different concentrations of LL-37 (2.5μM-15μM) or scrambled (Scr) LL-37(5μM-15μM) and used for infection or the cells were first treated with LL-37 and infected. To study the effect of LL-37 post infection (PI), the cells were infected first followed by addition of LL-37 to the culture medium 24h after infection. In all conditions, after the incubation, the culture supernatant was assessed for viral RNA copy number by real time RT-PCR, infectious virus particles by focus forming unit assay (FFU) and non structural protein 1 (NS1) antigen levels by ELISA. Percentage of infection was assessed using immunoflourescence assay (IFA). The results revealed that pretreatment of virus with 10-15μM LL-37 significantly reduced its infectivity as compared to virus control (P<0.0001). Moreover, pretreatment of virus with 10-15μM LL-37 significantly reduced the levels of viral genomic RNA and NS1 antigen (P<0.0001). Treatment of virus with 10-15μM LL-37 resulted in two to three log reduction of mean log10FFU/ml as compared to virus control (P<0.0001). Treatment of the virus with scrambled LL-37 had no effect on percentage of infection and viral load as compared to virus control cultures (P>0.05). Pretreatment of cells before infection or addition of LL-37 to the culture 24h PI had no effect on viral load. Molecular docking studies revealed possible binding of LL-37 to both the units of DENV envelope (E) protein dimer. Together, the in-vitro experiments and in-silico analyses suggest that LL-37 inhibits DENV-2 at the stage of entry into the cells by binding to the E protein. The results might have implications for prophylaxis against DENV infections and need further in-vivo studies.
authors.affiliation: Dengue/Chikungunya Group, ICMR-National Institute of Virology, 20A, Dr. Ambedkar Road, Pune 411001, Maharashtra, India. Electronic address: alagarasu@gmail.com.
authors.forename: K
authors.initials: K
authors.lastname: Alagarasu
authors.affiliation: Dengue/Chikungunya Group, ICMR-National Institute of Virology, 20A, Dr. Ambedkar Road, Pune 411001, Maharashtra, India.
authors.forename: P S
authors.initials: PS
authors.lastname: Patil
authors.affiliation: Bioinformatics Group, Microbial Containment Complex, ICMR-National Institute of Virology, Sus Road, Pashan, Pune 411021, Maharashtra, India.
authors.forename: P
authors.initials: P
authors.lastname: Shil
authors.affiliation: Dengue/Chikungunya Group, ICMR-National Institute of Virology, 20A, Dr. Ambedkar Road, Pune 411001, Maharashtra, India.
authors.forename: M
authors.initials: M
authors.lastname: Seervi
authors.affiliation: Dengue/Chikungunya Group, ICMR-National Institute of Virology, 20A, Dr. Ambedkar Road, Pune 411001, Maharashtra, India.
authors.forename: M B
authors.initials: MB
authors.lastname: Kakade
authors.affiliation: Dengue/Chikungunya Group, ICMR-National Institute of Virology, 20A, Dr. Ambedkar Road, Pune 411001, Maharashtra, India.
authors.forename: H
authors.initials: H
authors.lastname: Tillu
authors.affiliation: Dengue/Chikungunya Group, ICMR-National Institute of Virology, 20A, Dr. Ambedkar Road, Pune 411001, Maharashtra, India.
authors.forename: A
authors.initials: A
authors.lastname: Salunke
mesh_terms: D000704:Analysis of Variance; D000818:Animals; D023181:Antimicrobial Cationic Peptides; D000998:Antiviral Agents / Q000494:pharmacology*; D054804:Cathelicidins / Q000494:pharmacology*; D002522:Chlorocebus aethiops; D003716:Dengue Virus / Q000187:drug effects* / Q000235:genetics; D004305:Dose-Response Relationship, Drug; D006801:Humans; D062105:Molecular Docking Simulation; D012367:RNA, Viral / Q000187:drug effects / Q000235:genetics; D014709:Vero Cells; D014759:Viral Envelope Proteins / Q000378:metabolism; D019562:Viral Load / Q000187:drug effects*; D017361:Viral Nonstructural Proteins / Q000235:genetics / Q000378:metabolism; D014771:Virion / Q000187:drug effects
chemical_list: D023181:Antimicrobial Cationic Peptides; D000998:Antiviral Agents; D054804:Cathelicidins; C079340:NS1 protein, Dengue virus type 2; D012367:RNA, Viral; D014759:Viral Envelope Proteins; D017361:Viral Nonstructural Proteins
keywords: Antimicrobial peptides; Cathelicidin; Dengue virus; LL-37