TY - JOUR
T1 - Correction of anemia in uremic mice by genetically modified peritoneal mesothelial cells
AU - Einbinder, Tom
AU - Sufaro, Yuval
AU - Yusim, Igor
AU - Byk, Gerardo
AU - Passlick-Deetjen, Jutta
AU - Chaimovitz, Cidio
AU - Douvdevani, Amos
PY - 2003/6/1
Y1 - 2003/6/1
N2 - Background. During peritoneal dialysis, mesothelial cells become detached from the peritoneum and accumulate in the dialysate. Our aim was to evaluate the potential of peritoneal effluent (PF)-derived human peritoneal mesothelial cells (HPMC) as target for gene therapy. We used erythropoietin (EPO) as our target gene. Methods. Various extracellular matrixes (ECM) were tested for optimal adhesion and growth of HPMC. The EPO gene was introduced to mouse peritoneal mesothelial cells (MPMC) and HPMC by transfection or retroviral transduction. EPO secretion from PMC was measured by enzyme-linked immunosorbent assay (ELISA) and by the TF-1 cell proliferation assay. We performed intraperitoneal or intramuscular transplantations of the genetically modified cells into regular or 5/6 nephrectomized Balb/c mice and nude mice. Finally, we measured serum EPO and hematocrit levels. Results. ECM-coated plates provided up to sixfold increase in the efficiency of PMC isolation from PF. Gelatin coated dishes (20 μg/cm2) were found optimal for isolation of PF-HPMC. RPR-120535 liposome was found to be best for PMC transduction. In vitro studies showed EPO secretion from modified HPMC over 6 months. Intraperitoneal transplantation aided with collagen matrix was the most effective. EPO, in MPMC transplanted mice, was detected up to 3 weeks (peak at 13 ± 1 mIU/mL), and anemia of uremic mice was corrected (35.3 ± 0.9 mIU/mL to 41.9 ± 1.1 mIU/mL). Conclusion. PF-HPMC can be considered as an appropriate target for gene therapy since these cells can be efficiently isolated, modified, and transplanted. Nevertheless, implantation techniques in the peritoneum should be directed at obtaining longer duration of transgene expression in vivo, and means should be developed for enabling regulated expression of the gene.
AB - Background. During peritoneal dialysis, mesothelial cells become detached from the peritoneum and accumulate in the dialysate. Our aim was to evaluate the potential of peritoneal effluent (PF)-derived human peritoneal mesothelial cells (HPMC) as target for gene therapy. We used erythropoietin (EPO) as our target gene. Methods. Various extracellular matrixes (ECM) were tested for optimal adhesion and growth of HPMC. The EPO gene was introduced to mouse peritoneal mesothelial cells (MPMC) and HPMC by transfection or retroviral transduction. EPO secretion from PMC was measured by enzyme-linked immunosorbent assay (ELISA) and by the TF-1 cell proliferation assay. We performed intraperitoneal or intramuscular transplantations of the genetically modified cells into regular or 5/6 nephrectomized Balb/c mice and nude mice. Finally, we measured serum EPO and hematocrit levels. Results. ECM-coated plates provided up to sixfold increase in the efficiency of PMC isolation from PF. Gelatin coated dishes (20 μg/cm2) were found optimal for isolation of PF-HPMC. RPR-120535 liposome was found to be best for PMC transduction. In vitro studies showed EPO secretion from modified HPMC over 6 months. Intraperitoneal transplantation aided with collagen matrix was the most effective. EPO, in MPMC transplanted mice, was detected up to 3 weeks (peak at 13 ± 1 mIU/mL), and anemia of uremic mice was corrected (35.3 ± 0.9 mIU/mL to 41.9 ± 1.1 mIU/mL). Conclusion. PF-HPMC can be considered as an appropriate target for gene therapy since these cells can be efficiently isolated, modified, and transplanted. Nevertheless, implantation techniques in the peritoneum should be directed at obtaining longer duration of transgene expression in vivo, and means should be developed for enabling regulated expression of the gene.
KW - Erythropoietin
KW - Mesothelial cells
KW - Peritoneal dialysis
UR - http://www.scopus.com/inward/record.url?scp=0038527357&partnerID=8YFLogxK
U2 - 10.1046/j.1523-1755.2003.00014.x
DO - 10.1046/j.1523-1755.2003.00014.x
M3 - Article
C2 - 12753297
AN - SCOPUS:0038527357
SN - 0085-2538
VL - 63
SP - 2103
EP - 2112
JO - Kidney International
JF - Kidney International
IS - 6
ER -