PPARβ/δ-Interfering Peptide Enhanced Mesenchymal Stromal Cell Immunoregulatory Properties

Figure 4

PP11 peptide significantly enhanced hAD-MSC immunosuppressive properties. (a–d) Effect of PPARβ/δ activity modulation on hAD-MSC immunosuppressive properties. PBMC stained with CTV were collected and analyzed by FACS to determine their proliferation rate (b–d) and calculate the percentage of inhibition (a) of PHA-activated PBMC cocultured in the presence of untreated hAD-MSC, hAD-MSC pretreated with GSK3787 (GSK hAD-MSC), or hAD-MSC pretreated with PP11 (PP11 hAD-MSC). (e, f) Impact of PPARβ/δ inactivation on immunomodulatory mediators. (e) IDO activity determined by a spectrophotometric assay for kynurenine in supernatant of cocultured hAD-MSC:PBMC. Nontreated (Untreated), GSK3787 (GSK), or PP11- (PP11-) pretreated hAD-MSC were cocultured with PHA-activated PBMC for 96 h. The kynurenine concentration in the presence of pretreated cells is expressed relative to that measured in the presence of untreated cells (%). (f) COX2 mRNA expression level in hAD-MSC was pretreated either with GSK3787 (GSK) or PP11 peptide (PP11), compared to untreated hAD-MSC. COX2 expression level was normalized to housekeeping gene (RPS9) expression. mRNA expression in treated cells was standardized to expression in untreated hAD-MSC, and results are expressed using the method. The results for proliferation inhibition were obtained with hAD-MSC from 4 donors (). The statistical analysis was performed using the Mann–Whitney test. Statistical significance was noted as ns for and for . The results for IDO activity were obtained with hAD-MSC from 3 donors () on 2 independent experiments. The statistical analysis was performed using the Wilcoxon test. Statistical significance was noted as ns for and for . The results for COX2 expression were obtained with hAD-MSC from 3 donors () on 2 independent experiments. The statistical analysis was performed using the Mann–Whitney test. Statistical significance was noted as ns for and for .

(a)

(b)

(c)

(d)

(e)

(f)