title: Role of protein kinase C alpha and cyclin D1 in the proliferation of airway smooth muscle in asthmatic rats.
abstract: BACKGROUND
  Airway smooth muscle (ASM) is suspected to be a determining factor in the structural change of asthma. However, the role of protein kinase C alpha (PKCalpha) and cyclin D1 involved in the dysfunction of ASM leading to asthmatic symptoms is not clear. In this study, the central role of PKCalpha and cyclin D1 in ASM proliferation in asthmatic rats was explored.
  
  
  METHODS
  Thirty-six pathogen-free male Brown Norway (BN) rats were randomly divided into 2 groups: control groups (group N1, N2 and N3) and asthmatic groups (group A1, A2, and A3). Groups A1, A2 and A3 were challenged with ovalbumin (OA) for 2 weeks, 4 weeks and 8 weeks respectively. Control animals were exposed to an aerosolized sterile phosphate buffered saline (PBS). The ASM mass and nucleus numbers were studied to estimate the degree of airway remodeling by the hematoxylin-eosin staining method. PKCalpha and cyclin D1 expression in the ASM cells was detected by reverse transcription polymerase chain reaction (RT-PCR) and immunohistochemistry. The relation between PKCalpha and cyclin D1 was assessed by linear regression analysis. PKC agonist phorbol 12-myristate 13-acetate (PMA), PKC inhibitor Ro31-8220 and an antisense oligonucleotide against cyclin D1 (ASOND) were used to treat ASM cells (ASMCs) obtained from the 2 weeks asthmatic rats. The cyclin D1 protein expression level was detected by Western blotting.
  
  
  RESULTS
  Compared with the control group, the PKCalpha and cyclin D1 mRNA levels were increased in the asthmatic group. Similar to RT-PCR results, immunohistochemistry analysis for PKCalpha and cyclin D1 expression revealed an increased production in ASMCs after allergen treatment for 2, 4 and 8 weeks compared with the respective control groups. No difference in expression of PKCalpha and cyclin D1 in ASM were found in the 2, 4 or 8 weeks asthmatic rats. There were significant positive correlations between PKCalpha and cyclin D1 expression, both transcriptionally (r = 0.944, P < 0.01) and translationally (r = 0.826, P < 0.01), in ASM. The content of cyclin D1 in asthmatic ASMCs increased after being stimulated by PMA, and decreased when induced by Ro31-8220. ASOND targeting for cyclin D1 lowered the expression of cyclin D1 induced by PMA.
  
  
  CONCLUSIONS
  Increased expression of PKCalpha and cyclin D1 in ASM along with smooth muscle structure changes might implicate PKCalpha and cyclin D1 participation in the proliferation of ASM and contribute to the pathogenesis of asthma after repeated allergen exposure in rats. The results suggested that cyclin D1 might be downstream of PKC signal transduction pathway.
authors.affiliation: Department of Respiratory Medicine, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei 430030, China.
authors.forename: Li-fen
authors.initials: LF
authors.lastname: Qiao
authors.forename: Yong-jian
authors.initials: YJ
authors.lastname: Xu
authors.forename: Xian-sheng
authors.initials: XS
authors.lastname: Liu
authors.forename: Jun-gang
authors.initials: JG
authors.lastname: Xie
authors.forename: Jin
authors.initials: J
authors.lastname: Wang
authors.forename: Chun-ling
authors.initials: CL
authors.lastname: Du
authors.forename: Jian
authors.initials: J
authors.lastname: Zhang
authors.forename: Wang
authors.initials: W
authors.lastname: Ni
authors.forename: Shi-xin
authors.initials: SX
authors.lastname: Chen
mesh_terms: D000818:Animals; D001249:Asthma / Q000473:pathology*; D049109:Cell Proliferation; D019938:Cyclin D1 / Q000235:genetics / Q000502:physiology*; D008168:Lung / Q000473:pathology*; D008297:Male; D032389:Myocytes, Smooth Muscle / Q000473:pathology*; D051571:Protein Kinase C-alpha / Q000235:genetics / Q000502:physiology*; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D011914:Rats, Inbred BN
chemical_list: C530106:Ccnd1 protein, rat; D012333:RNA, Messenger; D019938:Cyclin D1; D051571:Protein Kinase C-alpha