PURPOSE: The purpose of this study was to determine, when assuming the worst case scenario of bone contamination, the sterility assurance level in bone transplantation treated by irradiation and after donor screening. MATERIAL AND METHODS: The virus: We employed the HIV-1, LAV-1 as our reference strain, which has always been cultured on normal human cells (and has therefore never been in contact with cell lines). To test viral virulence, we used a very sensitive cell line: MT2 cell line, a T lymphoblastoid cell line which is HTLV-I positive and is sensitive to HIV-1/LAV-1 infection. It was maintained in RPMI 1640 culture medium containing 10 per cent heat-inactivated fetal calf serum (Boehringer Mannheim), 2 mM L-glutamine (Boehringer Mannheim) and 1 per cent antibiotics (PSN 100 x, Gibco). Cell concentration was 100,000 cells per milliliter. Irradiation was performed using an accelerator delivering electrons of 6.2 MeV providing several tens of kilograys in a few seconds. The dose delivered was checked by placing alanine dosimeters at the site of the HIV aliquots. Alanine is a solid amino acid which, when irradiated, gives rise to free radicals; these were counted using electron paramagnetic resonance (relative standard deviation of less than 1 per cent). RESULTS: 25 kilograys irradiation of 316,227 TCID50/ml at -80 degrees C led to a considerable titer reduction (3 logs) with 316 TCID50/ml remaining. According to the technique's sensitivity (100 TCID50/ml) and to the sensitivity with which the dose of irradiation can be measured (less than 1 per cent), the D10 value with "99 per cent confidence limits" was between 8.3 kilograys and 7.2 kilograys. Assuming the worst characteristics of irradiation, we have chosen 8.3 kilograys as the D10 value for -80 degrees C. DISCUSSION: Considering a mean level of contamination of 30 TCID50 per milliliter of plasma during the asympomatic phase of infection, the reduction of the transmission risk of HIV by a femoral head allograft was calculated in the best case to be only 5 per cent when performed at 25 kilograys. Irradiation cannot be considered as a process able to achieve sterility of the graft and has to be associated to an extensive screening. However irradiation associated with appropriate screening of donors decreases the risk of transmission of the disease under one out of one million in femoral allograft implantation.
PURPOSE: The purpose of this study was to determine, when assuming the worst case scenario of bone contamination, the sterility assurance level in bone transplantation treated by irradiation and after donor screening. MATERIAL AND METHODS: The virus: We employed the HIV-1, LAV-1 as our reference strain, which has always been cultured on normal human cells (and has therefore never been in contact with cell lines). To test viral virulence, we used a very sensitive cell line: MT2 cell line, a T lymphoblastoid cell line which is HTLV-I positive and is sensitive to HIV-1/LAV-1 infection. It was maintained in RPMI 1640 culture medium containing 10 per cent heat-inactivated fetal calf serum (Boehringer Mannheim), 2 mM L-glutamine (Boehringer Mannheim) and 1 per cent antibiotics (PSN 100 x, Gibco). Cell concentration was 100,000 cells per milliliter. Irradiation was performed using an accelerator delivering electrons of 6.2 MeV providing several tens of kilograys in a few seconds. The dose delivered was checked by placing alanine dosimeters at the site of the HIV aliquots. Alanine is a solid amino acid which, when irradiated, gives rise to free radicals; these were counted using electron paramagnetic resonance (relative standard deviation of less than 1 per cent). RESULTS: 25 kilograys irradiation of 316,227 TCID50/ml at -80 degrees C led to a considerable titer reduction (3 logs) with 316 TCID50/ml remaining. According to the technique's sensitivity (100 TCID50/ml) and to the sensitivity with which the dose of irradiation can be measured (less than 1 per cent), the D10 value with "99 per cent confidence limits" was between 8.3 kilograys and 7.2 kilograys. Assuming the worst characteristics of irradiation, we have chosen 8.3 kilograys as the D10 value for -80 degrees C. DISCUSSION: Considering a mean level of contamination of 30 TCID50 per milliliter of plasma during the asympomatic phase of infection, the reduction of the transmission risk of HIV by a femoral head allograft was calculated in the best case to be only 5 per cent when performed at 25 kilograys. Irradiation cannot be considered as a process able to achieve sterility of the graft and has to be associated to an extensive screening. However irradiation associated with appropriate screening of donors decreases the risk of transmission of the disease under one out of one million in femoral allograft implantation.
Authors: P Díaz-de-Rada; A Barriga; J L Barroso; E García-Barrecheguren; M Alfonso; J R Valentí Journal: Knee Surg Sports Traumatol Arthrosc Date: 2003-06-19 Impact factor: 4.342