microRNA-based identification of pulmonary signaling pathways in experimental asthma

Asthma is characterized by chronic airway inflammation; mucus
hypersecretion and episodes of airway obstruction. It is the most
common chronic disease in children and its incidence and prevalence
have been markedly increasing over the last few decades. While the
pathophysiology has been intensively studied, the underlying causes
for asthma development remain largely unknown. Hence there is an
unmet, clinical need to identify novel, so far non-characterized
pathways, relevant for this disease, to establish new therapies or
preventive approaches. We used miRNAs to do so as they are critical
regulators of key molecules within signalling pathways. Our group
previously identified a pulmonary increase of miR-17 and -144 in
ovalbumin-induced allergic airway inflammation in mice and they
were shown to target the transcription factor cAMP-responsive
element binding protein 1 (Creb1). In this work, we found and
validated additional binding sites for the two miRNAs and the newly
included miR-21 in the 3’UTR of Creb1 co-activators CRTC1, -2 and
-3. Creb1 and the three Crtcs were further shown to be
down-regulated in two models of murine experimental asthma
(ovalbumin and house dust mite). We postulated that a joint
down-regulation of Creb plus its co-activators by the miRNAs might
have detrimental, additive effects on the expression of target
genes. In this respect, we identified the putative Creb1 target
gene Sec14-like 3 (Sec14l3) which was down-regulated in both models
for murine experimental asthma and its loss seemed to be associated
with goblet cell metaplasia. Furthermore, we found that IL-13
treatment of primary normal human epithelial (NHBE) cells led to a
decrease of CREB1/CRTCs and SEC14L3, while the latter correlated
with forkheadbox protein J1 (FOXJ1), a marker for ciliated cells.
miR-17, -144 and -21 were actively secreted into exosomes by the
primary NHBE cells upon IL-13 stimuli, which could be uptaken by a
bronchial epithelial cell line and a T cell line . We could also
detect miRNA-containing exosomes in nasal washes from children with
allergic asthma and broncho-alveolar lavage fluid of mice with
OVA-induced allergic airway inflammation. In nasal epithelial cells
of children with allergic asthma, the three miRNAs were highly
increased compared to healthy controls, while SEC14l3 expression
was reduced. In conclusion, this work identified a novel molecular
axis (miRNA-Creb/Crtc/Sec14l3) relevant for murine and paediatric
asthma and gives first hints that miR-17, -144 and -21 in exosomes
might be able to perpetuate the asthmatic response between
different cell types.