| Abstrakt: |
Background Previously, we demonstrated that murine mesenchymal stem cells (MSCs) injected intracranially into neonatal mice expand throughout the central nervous system (CNS). This paper describes real-time PCR (RT-PCR) assays that enable accurate quantification of transplanted cells in vivo . Methods RT-PCR assays that amplify sequences in the mouse Y chromosome or human Alu repeats were developed and used to quantify the number of male, murine or human MSCs in the CNS at various times after intracranial injection into neonatal mice, or in various organs of adult mice after i.p. injection of cells into 13 day-old embryos. Results In the CNS, levels of male mouse DNA in female transplant recipients increased on average 30-fold between 3 and 60 days post-injection, but then was unchanged at 140 days post-transplant (P=0.107). Male DNA accounted for up to 0.309% of the total DNA content of brain, representing maximally 600 000 donor cells. Human DNA was detected in the CNS up to 300 days post-transplant, but levels never exceeded 7.63×10 -4 % of total brain DNA content. After in utero transplantation, human DNA levels ranged from 0.36×10 -5 % to 2.14×10 -5 % of the total DNA content of liver, kidney and spleen. Significantly higher levels were found in heart (P=0.06), femur and brain (P=0.02), and lung (P=0.0025). Discussion RT-PCR assays were developed to quantify levels of male, murine and human cells in vivo following sex-mismatched or xeno-transplants. Due to their accuracy, precision, and sensitivity, these assays provide a versatile alternative to measuring stem-cell engraftment in vivo. [ABSTRACT FROM AUTHOR] |
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