The effects of poly I:C stimulation of primary bronchial epithelial cells and TSLP secretion on CD34+ progenitor cell eosinophil and basophil differentiation
Nasal Polyposis. Allergy Asthma Immunol Res
The effects of poly I:C stimulation of primary bronchial epithelial cells and TSLP secretion on CD34+ progenitor cell eosinophil and basophil differentiation
Ashley Yu 0 1
Claudia CK Hui 0 1
Judah A Denburg 0 1
0 References 1. Shikotra A, Choy DF, Ohri CM, Doran E, Butler C, Hargadon B, et al: Increased expression of immunoreactive thymic stromal lymphopoietin in patients with severe asthma. J Allergy Clin Immunol 2012 , 129:104-11, e1-9.)rinters. 2. Wang WL, Li HY, Zhang MS, Gao PS, He SH, Zheng T, Zhu Z , Zhou LF: Thymic Stromal Lymphopoietin: A Promising Therapeutic Target for Allergic Diseases. Int Arch Allergy Immunol 2013, 160:18-26
1 Department of Medicine and Clinical Immunology, McMaster University , Hamilton, Ontario, L8S 4L8 , Canada
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Background
In asthmatic lungs, elevated levels of thymic stromal
lymphopoetin (TSLP) are linked to eosinophilic
inflammation and disease severity [1]. TSLP is involved in initiating
a TH2 inflammatory response, through activation of
T cells, and recently, CD34+ hemopoietic progenitor cells
[2,3]. However, the biological effects of epithelial-derived
TSLP on human peripheral blood (PB) CD34+ progenitor
eosinophil-basophil (Eo/B) lineage commitment have not
been described. The aim of the current study is to examine
the effects of primary bronchial epithelial cell-derived
TSLP on CD34+ hemopoietic progenitor differentiation.
Methods
Primary bronchial epithelial cells (PBEC) grown in
airliquid interface were apically stimulated with media or
varying doses of polyinosinic:polycytidylic acid (Poly I:C;
1, 10, 25, and 50g/mL) and cultured in the presence or
absence of PB CD34+ cells in the basolateral
compartment overnight. Supernatant was collected and analyzed
for cytokine/chemokine secretion using Luminex assays.
Overnight co-cultured PB CD34+ cells were (1) cultured
in methylcellulose colony assays to assess for the mean
numbers of Eo/B colony-forming units (CFU) (colonies
were defined as 40 cells) after 14 d; or (2) assessed for
TSLPR expression using flow cytometry.
Results
Preliminary data demonstrates that overnight stimulation
of PBEC with poly I:C in the absence of PB CD34+ cells
induced a dose-dependent release of IL-4, IL-5, IL-13,
TNF eotaxin-1, and MCP-1; however, failed to secrete
detectible levels of IL-1b and IFN-. Poly I:C at 10g/mL
enhanced TSLP and TARC secretion while at 50g/mL,
poly I:C enhanced IL-33 secretion from PBEC compared
to unstimulated control. Furthermore, basal levels of
IL-3, IL-6, IL-8, MDC, and RANTES were detected from
rested PBEC, with no observable trend in secretion
following poly I:C stimulation. Finally, PB CD34+ cells
co-cultured overnight with poly I:C-stimulated PBEC
have been cultured in methylcellulose colony assays and
waiting for Eo/B CFU to be counted.
Conclusions
In conclusion, our co-culture system will allow for the
establishment of epithelial-derived TSLP activity and its
influence on CD34+ progenitor Eo/B differentiation. In
the future, we would like to examine whether PBEC
obtained from atopic vs. non-atopic individuals results
in distinct progenitor response.
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