Abstract
Chronic rejection, bronchiolitis obliterans syndrome (BOS), is a major hurdle following human lung transplantation. Antibodies (Ab) to HLA (DSA) and lung-associated antigens (Collagen V (Col-V) and K-alpha-1 Tubulin (Kα1T) have been associated with development of BOS. Our goal was to demonstrate early events (genes and their role in inflammation) associated with administration of anti-MHC class I (H-2Kb) into murine (C57BL/6) lungs that precede cellular and humoral autoimmunity and chronic rejection.
We analyzed molecular signatures arising from anti-MHC administration by genechip microarray. Zbtb7a was selected for functional analysis. siRNA-Lentivirus was used to knockdown Zbtb7a in lungs to study its role in Ab induced chronic rejection. Kinetics of Ab development and T cell responses was tested by ELISA and ELISPOT. Changes in leukocyte profile were analyzed by flow cytometry and gene expression levels by real-time PCR.
In genechip assay, 12 genes including Zbtb7a were significantly (p<0.005) upregulated (>1.5-fold) at 4h post Ab administration. Use of siRNA-Lentivirus knocked down Zbtb7a expression. Following anti-MHC I, Zbtb7a knockdown demonstrated significant (p<0.001) reduction in anti-Kα1T and anti-Col-V, and remained free from inflammation and fibrosis. Further, anti-MHC elicited lower Kα1T and Col-V specific Th17 and Th1 cells. Moreover, less infiltration of neutrophil and B cell were seen in lungs and decreased levels of B cell (CXCL13) and neutrophil (CXCL15) chemoattractants were observed.
We demonstrate that DSA activates unique molecular signature involved in lung autoimmunity. Zbtb7a, as a transcription factor induced by DSA, is a “master regulator” of B and T cell development and has novel inflammatory functions leading to chronic rejection. By targeted knockdown, we established that Zbtb7a has an obligatory role in the amplification of inflammatory circuits where its loss rendered protection from Ab induced obliterative airway disease.