BPIFB1 (LPLUNC1) is upregulated in cystic fibrosis lung disease

Abstract

Although the biology the PLUNC (recently renamed BPI fold, BPIF) family of secreted proteins is poorly understood, multiple array based studies have suggested that some are differentially expressed in lung diseases. We have examined the expression of BPIFB1 (LPLUNC1), the prototypic two-domain containing family member, in lungs from CF patients and in mouse models of CF lung disease. BPIFB1 was localized in CF lung samples along with BPIFA1, MUC5AC, CD68 and NE and directly compared to histologically normal lung tissues and that of bacterial pneumonia. We generated novel antibodies to mouse BPIF proteins to conduct similar studies on ENaC transgenic (ENaC-Tg) mice, a model for CF-like lung disease. Small airways in CF demonstrated marked epithelial staining of BPIFB1 in goblet cells but staining was absent from alveolar regions. BPIFA1 and BPIFB1 were not co-localised in the diseased lungs. In ENaC-Tg mice there was strong staining of both proteins in the airways and luminal contents. This was most marked for BPIFB1 and was noted within 2 weeks of birth. The two proteins were present in distinct cells within epithelium. BPIFB1 was readily detected in BAL from ENaC-Tg mice but was absent from wild-type mice. Alterations in the expression of BPIF proteins is associated with CF lung disease in humans and mice. It is unclear if this elevation of protein production, which results from phenotypic alteration of the cells within the diseased epithelium, plays a role in the pathogenesis of the disease.

Fig. 1

Increased BPIFB1 and BPIFA1 in CF lung. Immunohistochemistry for BPIFB1 (a, d, e, g) and BPIFA1 (b, f, h) was performed as described in "Materials and methods". Sections show staining in samples of normal large airway (a–c) and in sections from patients with cystic fibrosis (d–i). Sections c and i are controls (no primary antibody). The black arrows in panel a indicated BPIFB1 positive cells in the normal airway. Scale bars are present on each individual panel

Fig. 2

BPIFB1 is predominantly expressed in a goblet cell population in CF airways and is absent from neutrophils and macrophages. Immunohistochemistry for BPIFB1 (a, c, g), BPIFA1 (b), MUC5AC (d) NE (e, i) and CD68 (f, h) was performed as described in "Materials and methods" using sections from patients with cystic fibrosis (a–f) or from bacterial pneumonia (g–i). Scale bars are present on each individual panel

Fig. 3

BPIFA1 and BPIFB1 do not co-localise in the adult mouse respiratory tract. Immunohistochemistry was performed as described in "Materials and methods" using antibodies specific for murine BPIFA1 (a, c, e) and BPIFB1 (b, d, f). Sections show serial samples of the trachea (a, b), large airways (c, d) and smaller airways and peripheral lung (e, f). The inset figure in f represents staining of a further section with the Clara cell marker, SCGB1A1 (CCSP). The black arrows identify the rare BPIFB1 positive cells in the normal mouse trachea. Scale bars are present on each individual panel

Fig. 4

BPIFB1 and BPIFA1 are increased in the occluded airways of scgb1a1-ENaC transgenic mice. Immunohistochemistry was performed as described in "Materials and methods" using antibodies specific for murine BPIFB1 (a, c, e, g, i, j) and BPIFA1 (d, f, h, k, l) and SCGB1A1 (b). Sections show serial samples of lungs from 2 week old wild type mice (a, b), 2 week old transgenic mice (c, d) 6 week old wild type mice (e–f) and 6 week old transgenic mice (g–l). Scale bars are present on each individual panel

Fig. 5

BPIFB1 is significantly increased in the BAL fluid of Scgb1a1-ENaC transgenic mice. Replicate SDS-PAGE gels of BAL fluid samples from three wild type and three transgenic mice (both at 6 weeks of age) were generated as outlined in "Materials and methods". The resultant blots were subjected to western blotting and detected with specific antibodies against mouse BPIFB1 and BPIFA1 as described. The position of the molecular mass markers are indicated by the black arrows