Compartmentalized transgene expression of granulocyte-macrophage colony-stimulating factor (GM-CSF) in mouse lung enhances allergic airways inflammation

Abstract

To investigate the role of GM-CSF in asthmatic airways inflammation, we have targeted GM-CSF transgene to the airway cells in a mouse model of ovalbumin (OVA)-induced allergic airways inflammation, a model in which there is marked induction of endogenous IL-5 and IL-4 but not GM-CSF. Following intranasal delivery of a replication-deficient adenoviral gene transfer vector (Ad), transgene expression was found localized primarily to the respiratory epithelial cells. Intranasal delivery of 0.03 x 10(9) plaque-forming units (PFU) of AdGM-CSF into naive BALB/c mice resulted in prolonged and compartmentalized release of GM-CSF transgene protein with a peak concentration of approximately 80 pg/ml detected in bronchoalveolar lavage fluid (BALF) at day 7, but little in serum. These levels of local GM-CSF expression per se resulted in no eosinophilia and only a minimum of tissue inflammatory responses in the lung of naive mice, similar to those induced by the control vector. However, such GM-CSF expression in the airways of OVA-sensitized mice resulted in a much greater and sustained accumulation of various inflammatory cell types, most noticeably eosinophils, both in BALF and airway tissues for 15-21 days post-OVA aerosol challenge, at which times airways inflammation had largely resolved in control mice. While the levels of IL-5 and IL-4 in BALF and the rate of eosinophil apoptosis were found similar between different treatments, there was an increased number of proliferative leucocytes in the lung receiving GM-CSF gene transfer. Our results thus provide direct experimental evidence that GM-CSF can significantly contribute to the development of allergic airways inflammation through potentiating and prolonging inflammatory infiltration induced by cytokines such as IL-5 and IL-4.

Fig. 1

(a) Bronchoalveolar lavage fluid (BALF) and serum levels of GM-CSF in normal mice at various time points after intranasal delivery of AdGM-CSF or Add170-3. ○, BALF from AdGM-CSF-treated mice; •, serum from AdGM-CSF-treated mice; □, BALF from Add170-3-treated mice. (b) BALF levels of GM-CSF at various time points in ovalbumin (OVA)-sensitized mice after OVA aerosol challenge. Results are expressed as mean ± s.e.m. from four to six mice per point.

Fig. 2

Total cell number (TCN) in bronchoalveolar lavage (BAL) from AdGM-CSF- or Add170-3-treated normal mice. Results are expressed mean ± s.e.m. from four to six mice per point.

Fig. 3

Schematic illustration of experimental protocol. BALB/c mice were intraperitoneally sensitized twice with ovalbumin (OVA) 17 and 12 days before OVA aerosol challenge (S1and S2), and OVA aerosol-challenged 12 days after S2 (day 0; Ch). AdGM-CSF, Add170-3 or PBS was intranasally administered into these mice 1 day before OVA aerosol challenge. Bronchoalveolar lavage fluid (BALF) and lung tissue samples were collected at days 1, 5, 14 and 21 post-OVA aerosol challenge.

Fig. 4

Effects of AdGM-CSF over-expression in ovalbumin (OVA)-sensitized/challenged mice. Data demonstrate changes in inflammatory cells in bronchoalveolar lavage fluid (BALF) at various time points after OVA aerosol challenge. (a) Total cell number (TCN). (b) Macrophages/monocytes (macro/mono.). (c) Lymphocytes. (d) Neutrophils. (e) Eosinophils. Data are expressed as mean ± s.e.m. from four to six mice/time point. Statistical analysis was performed using Minitab software; *P ≤ 0.05.

Fig. 5

Light photomicrographs of paraffin-embedded sections of lung tissues stained with haematoxylin–eosin obtained from ovalbumin (OVA)-sensitized/challenged mice at 6 days (a,b,c) and 15 days (d,e,f) after intranasal delivery of PBS (a,d), or Addl70-3 (b,e), or AdGM-CSF (c,f), i.e. 5 days and 14 days after OVA aerosol challenge. Some of eosinophils are marked by arrows. (Mag. × 200.)