Hum. Reprod. Advance Access originally published online on November 2, 2006
Human Reproduction 2007 22(3):733-742; doi:10.1093/humrep/del418
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The efficiency of magnetic-activated cell sorting and fluorescence-activated cell sorting in the decontamination of testicular cell suspensions in cancer patients
1 Research Centre for Reproduction and Genetics 2 Centre for Reproductive Medicine, University Hospital and Medical School, Vrije Universiteit Brussel, Brussels, Belgium
3 To whom correspondence should be addressed at: Research Centre for Reproduction and Genetics, University Hospital and Medical School, Vrije Universiteit Brussel, Laarbeeklaan 101, B-1090 Brussels, Belgium. E-mail: mieke.geens{at}az.vub.ac.be
BACKGROUND: Before clinical application, the feasibility and safety of autologous testicular stem cell transplantation should be explored. Apart from limitations in their numbers, spermatogonial stem cells may also be contaminated by malignant cells. Therefore, both enrichment and decontamination before transplantation may be necessary. This study aimed at evaluating the decontaminating potential of magnetic-activated cell sorting (MACS) and/or fluorescence-activated cell sorting (FACS) for both murine and human testicular cell suspensions. In the mouse, the effectiveness of the transplantation technique after cell sorting was also assessed. METHODS: Murine testicular cells were contaminated with 5% EL4 cells. Fresh and frozen–thawed suspensions were sorted using MACS (CD49f +) and FACS (CD49f +, H-2Kb–) and evaluated by FACS, cell culture and transplantation into W/Wv mice. Human testicular cells were contaminated with 5 or 0.05% CCRF-SB (SB) cells. Frozen–thawed suspensions were sorted using FACS (HLA class I–) and evaluated by FACS, cell culture and PCR for the B-cell receptor. RESULTS: In the mouse, the sorted fractions contained 0.39% H-2Kb-positive and 76.55% CD49f-positive cells. After transplantation, 1 in 20 recipient mice developed a malignancy. In the human experiments, an average of 0.58% SB cells was detected after sorting. In only 1 of 11 samples, there were no SB cells observed. CONCLUSION: MACS and/or FACS are insufficient for completely depleting testicular tissue of malignant cells. Although more research on alternative decontamination techniques is necessary, developing a reliable method to screen a priori testicular tissue for malignant cells may be equally important.
Key words: spermatogonia/decontamination/transplantation/cancer
Submitted on June 7, 2006; resubmitted on August 30, 2006; accepted on September 21, 2006.
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