Detection of host and donor cells in sex-mismatched rat nerve allografts using RT-PCR for a Y chromosome (H-Y) marker.
The donor and host source of support cells, such as Schwann cells, in nerve allograft segments have been the subject of debate. The objective of the present study was to assess the utility of a molecular technique that probes for a Y chromosome expressed gene (H-Y) in distinguishing host from donor tissue in sex-mismatched nerve allograft segments. Forty-two Lewis rats received bilateral syngeneic Lewis or allogeneic ACI rat peroneal nerve grafts, with or without cyclosporin A (CsA) treatment. At different times thereafter animals were sacrificed and samples were harvested. We transplanted males and females reciprocally, to study both survival of donor cells (persisting H-Y mRNA in male grafts by transcription polymerase chain reaction (RT-PCR), and graft infiltration by host cells (detectable H-Y mRNA in female grafts). A kinetic analysis revealed a progressive loss of viable donor cells (loss of H-Y mRNA signal) from allografts, beginning 2-3 weeks, and culminating at 4 weeks, with little detectable H-Y in the absence of CsA treatment. CsA treatment led to prolonged survival of allograft cells, confirmed by detectable H-Y mRNA. By studying female grafts in male rats we could confirm that loss of viable donor tissue in allografts was accompanied by infiltration of host (H-Y mRNA positive) cells, whereas no H-Y mRNA signal was seen in males receiving autografts from females or in immunosuppressed allograft segments. These data suggest that reverse RT-PCR analysis for a Y chromosome gene product can be a valuable tool to assess the origin of viable cells in sex-mismatched nerve allotransplantation tissue.
Midha
R
R
Division of Neurosurgery, Sunnybrook and Women's College Health Sciences Centre, University of Toronto, Ontario, Canada. rajiv.midha@swchsc.on.ca
Ramakrishna
V
V
Munro
C A
CA
Matsuyama
T
T
Gorczynski
R M
RM
D000818:Animals; D016572:Cyclosporine / Q000494:pharmacology; D005260:Female; D006084:Graft Rejection / Q000188:drug therapy / Q000473:pathology; D006085:Graft Survival / Q000235:genetics*; D006182:H-Y Antigen / Q000235:genetics; D007166:Immunosuppressive Agents / Q000494:pharmacology; D008297:Male; D010543:Peroneal Nerve / Q000473:pathology* / Q000637:transplantation*; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D011912:Rats, Inbred ACI; D011917:Rats, Inbred Lew; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012737:Sex Factors; D014184:Transplantation, Homologous; D014998:Y Chromosome
D006182:H-Y Antigen; D007166:Immunosuppressive Agents; D012333:RNA, Messenger; D016572:Cyclosporine