Feasibility of diagnosing subclinical renal allograft rejection in children by whole blood gene expression analysis.
BACKGROUND
Protocol biopsies are used to monitor allograft histology after transplantation. However, biopsy is an invasive procedure with potential complications, requires special facilities, and is unpractical for repeated monitoring of the graft. A noninvasive, robust, and rapid diagnostic method would be welcomed. Monitoring gene expression from blood samples could provide such a means.


METHODS
Whole blood samples taken at the time of 3- or 6-month protocol biopsy in 31 pediatric renal transplant recipients, 13 of whom had biopsy-proven subclinical rejection (SCR), were studied. The samples were collected into tubes containing an RNA stabilization reagent enabling feasible collection during a normal ward schedule. In all patients, the gene expression of candidate genes CD154 and inducible T-cell co-stimulator (ICOS) was measured. A low-density array containing 90 immunologic-related genes were measured with real-time quantitative PCR (RT-QPCR) in 10 patients. In addition, a whole genome microarray analysis was performed in eight patients.


RESULTS
Neither CD154 nor ICOS gene expression was diagnostic for SCR (median expression level 1.25 vs. 1.16 and 1.95 vs. 1.61 for CD154 and ICOS, respectively). In addition, expression levels of none of the genes on the low-density array were associated with SCR. Finally, in the microarray analysis none of the found differences between SCR and normal patients' gene expression could be validated with RT-QPCR in 17 genes.


CONCLUSIONS
In our relatively small series no robust whole blood gene expression biomarker for SCR was found. Further studies are needed to determine whether small changes in expression may provide a supporting diagnostic method.
Alakulppi
Noora
N
Research and Development, Finnish Red Cross Blood Service, Helsinki, Finland. noora.alakulppi@veripalvelu.fi
Seikku
Paula
P
Jaatinen
Taina
T
Holmberg
Christer
C
Laine
Jarmo
J
D000293:Adolescent; D000945:Antigens, Differentiation, T-Lymphocyte / Q000235:genetics / Q000378:metabolism; D015415:Biomarkers / Q000097:blood; D001706:Biopsy; D023201:CD40 Ligand / Q000235:genetics / Q000378:metabolism; D002648:Child; D002675:Child, Preschool; D005240:Feasibility Studies; D005260:Female; D006084:Graft Rejection / Q000097:blood* / Q000175:diagnosis*; D006801:Humans; D060889:Inducible T-Cell Co-Stimulator Protein; D007223:Infant; D007668:Kidney / Q000378:metabolism / Q000473:pathology; D016030:Kidney Transplantation; D008297:Male; D020411:Oligonucleotide Array Sequence Analysis; D014184:Transplantation, Homologous
D000945:Antigens, Differentiation, T-Lymphocyte; D015415:Biomarkers; C556692:ICOS protein, human; D060889:Inducible T-Cell Co-Stimulator Protein; D023201:CD40 Ligand