title: Downregulation of mouse CCR3 by lentiviral shRNA inhibits proliferation and induces apoptosis of mouse eosinophils.
abstract: RNA interference has been considered as an effective gene silencing method in basic and preclinical investigations. The aims of the present study were to construct a lentiviral vector expressing a short hairpin RNA (shRNA) targeting the murine CC chemokine receptor 3 (mCCR3), and to investigate its effects on the proliferation and apoptosis of mouse eosinophils. A recombinant lentiviral vector expressing four fragments of mouse CCR3 shRNA (pLVX‑mCCR3‑1+2+3+4‑shRNA) was constructed using subcloning techniques. This novel lentivirus was then packaged into 293T cells by co‑transduction with plasmids, including Baculo p35, pCMV R8.2 and VSV. The interference effects of the vector were verified using polymerase chain reaction (PCR) and western blot analyses. The effects of the interference on the proliferation and apoptosis of mouse eosinophils were investigated using 3‑(4,5‑dimethylthiazol‑2‑yl)‑5‑(3‑carboxymethoxyphenyl)‑2‑(4‑sulfophenyl)‑2H‑tetrazolium and terminal deoxynucleotidyl transferase dUTP nick end labeling methods, respectively. The results of the PCR and western blot analyses confirmed that the novel recombinant vector, pLVX‑mCCR3‑1+2+3+4‑shRNA, had high efficiency in inhibiting the mRNA and protein expression levels of mCCR3 in mouse eosinophils. The downregulation of mCCR3 significantly inhibited proliferation of the eosinophils. Furthermore, the present study found that the downregulation of mCCR3 significantly promoted apoptosis of the eosinophils. Therefore, the downregulation of mCCR3 led to the inhibition of proliferation and induction of apoptosis in mouse eosinophils. The predominant characteristics of allergic rhinitis are eosinophil infiltration and release of inflammatory mediators, which appear in a variety of clinical manifestations. The results of the present study indicate that mCCR3 silencing may serve as a putative approach for the treatment of allergic rhinitis.
authors.affiliation: Department of Otolaryngology Head and Neck Surgery, The Second Affiliated Hospital of Nanchang University, Nanchang, Jiangxi 330006, P.R. China.
authors.forename: Xin-Hua
authors.initials: XH
authors.lastname: Zhu
authors.affiliation: Department of Head and Neck Surgery, The Tumor Hospital of Jiangxi Province, Nanchang, Jiangxi 330006, P.R. China.
authors.forename: Bing
authors.initials: B
authors.lastname: Liao
authors.affiliation: Department of Otolaryngology Head and Neck Surgery, The Second Affiliated Hospital of Nanchang University, Nanchang, Jiangxi 330006, P.R. China.
authors.forename: Yi
authors.initials: Y
authors.lastname: Xu
authors.affiliation: Department of Otolaryngology Head and Neck Surgery, The Second Affiliated Hospital of Nanchang University, Nanchang, Jiangxi 330006, P.R. China.
authors.forename: Ke
authors.initials: K
authors.lastname: Liu
authors.affiliation: Department of Otolaryngology Head and Neck Surgery, The Second Affiliated Hospital of Nanchang University, Nanchang, Jiangxi 330006, P.R. China.
authors.forename: Yun
authors.initials: Y
authors.lastname: Huang
authors.affiliation: Department of Otolaryngology Head and Neck Surgery, The Second Affiliated Hospital of Nanchang University, Nanchang, Jiangxi 330006, P.R. China.
authors.forename: Quan-Long
authors.initials: QL
authors.lastname: Huang
authors.affiliation: Department of Otolaryngology Head and Neck Surgery, The Second Affiliated Hospital of Nanchang University, Nanchang, Jiangxi 330006, P.R. China.
authors.forename: Yue-Hui
authors.initials: YH
authors.lastname: Liu
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000235:genetics*; D001854:Bone Marrow Cells / Q000166:cytology; D049109:Cell Proliferation / Q000235:genetics; D015536:Down-Regulation; D004804:Eosinophils / Q000166:cytology* / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D016086:Lentivirus / Q000235:genetics; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D010957:Plasmids / Q000235:genetics / Q000378:metabolism; D034622:RNA Interference; D012333:RNA, Messenger / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism; D054397:Receptors, CCR3 / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism*
chemical_list: C515351:Ccr3 protein, mouse; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D054397:Receptors, CCR3