ABSTRACT
Alport syndrome (AS) is a hereditary glomerulopathy caused by a mutation in type IV collagen genes, which disrupts glomerular basement membrane, leading to progressive glomerulosclerosis and end-stage renal failure. There is at present no cure for AS, and cell-based therapies offer promise to improve renal function. In this study, we found that human first trimester fetal chorionic stem cells (CSC) are able to migrate to glomeruli and differentiate down the podocyte lineage in vitro and in vivo. When transplanted into 7-week-old Alport 129Sv-Col4α3(tm1Dec)/J (-/-) mice, a single intraperitoneal injection of CSC significantly lowered blood urea and urine proteinuria levels over the ensuing 2 weeks. In addition, nearly two-thirds of transplanted -/- mice maintained their weight above the 80% welfare threshold, with both males and females weighing more than age-matched nontransplanted -/- mice. This was associated with less renal cortical fibrosis and interstitial inflammation compared to nontransplanted mice as shown by reduction in murine CD4, CD68, and CD45.2 cells. Transplanted CSC homed to glomeruli, where they expressed CR1, VEGFA, SYNAPTOPODIN, CD2AP, and PODOCIN at the RNA level and produced PODOCIN, CD2AP, and COLIVα3 proteins in nontransplanted -/- mice, indicating that CSC have adopted a podocyte phenotype. Together, these data indicate that CSC may be used to delay progression of renal pathology by a combination of anti-inflammatory effects and replacement of the defective resident podocytes.
Subject(s)
Cell Differentiation , Chorion/cytology , Nephritis, Hereditary/therapy , Podocytes/cytology , Stem Cells/cytology , Animals , Biomarkers/metabolism , Cell Differentiation/drug effects , Cell Movement/drug effects , Cells, Cultured , Chorion/transplantation , Coculture Techniques , Collagen Type IV/pharmacology , Down-Regulation/drug effects , Female , Fibrosis , Humans , Inflammation/pathology , Intracellular Signaling Peptides and Proteins/metabolism , Kidney Cortex/pathology , Kidney Glomerulus/drug effects , Kidney Glomerulus/metabolism , Male , Membrane Proteins/metabolism , Mice , Mutation/genetics , Nephritis, Hereditary/pathology , Phenotype , Podocytes/drug effects , Podocytes/metabolism , RNA, Messenger/genetics , RNA, Messenger/metabolism , Stem Cells/drug effects , Stem Cells/metabolismABSTRACT
INTRODUCTION: Microarray studies have shown elevated transcript levels of endothelial and natural killer (NK) cell-associated genes during antibody-mediated rejection (AMR) of the renal allograft. This study aimed to assess the use of quantitative real-time polymerase chain reaction as an alternative to microarray analysis on a subset of these elevated genes. METHODS: Thirty-nine renal transplant biopsies from patients with de novo donor-specific antibodies and eighteen 1-year surveillance biopsies with no histological evidence of rejection were analyzed for expression of 11 genes previously identified as elevated in AMR. RESULTS: Expression levels of natural killer markers were correlated to microcirculation inflammation and graft outcomes to a greater extent than endothelial markers. Creating a predictive model reduced the number of gene transcripts to be assessed to 2, SH2D1b and MYBL1, resulting in 66.7% sensitivity and 89.7% specificity for graft loss. DISCUSSION: This work demonstrates that elevated gene expression levels, proposed to be associated with AMR, can be detected by established quantitative real-time polymerase chain reaction technology, making transition to the clinical setting feasible. Transcript analysis provides additional diagnostic information to the classification schema for AMR diagnosis but it remains to be determined whether significant numbers of centres will validate transcript analysis in their laboratories and put such analysis into clinical use.