ABSTRACT
<p><b>OBJECTIVE</b>To assess the accuracy and reliability of the nest-PCR-sequence specific primer(SSP) method in HLA-A site genotyping of single blastomeres retrieved from human pre-implantation embryos.</p><p><b>METHODS</b>By nest PCR on HLA-A exon 2, the success rate of first-round amplification was estimated for single blastomeres. Based on the first-round amplification, the HLA-A genotype of every single blastomeres was analyzed by commercially available PCR-SSP kits.</p><p><b>RESULTS</b>The amplification of HLA-A exon 2 were performed to 120 blasotmeres retrieved from in vitro fertilization(IVF) surplus embryos donated by 10 couples. The average success rate of family 1-5 and 6-10 was 78.2%(43/55) and 93.8%(61/65), respectively. And 86.7%(104/120) in total. Eighty blastomeres were further tested by nest-PCR-SSP, among which 11 blastomeres failed to HLA-A exon 2 amplification and then failed to genotyping while the other 69 blastomeres succeed in HLA-A exon 2 amplification and succeed in genotyping. Except for 6 blastomeres that were uncertain for allele lost because of parents' homozygosity, the left 63 blastomeres had accurate HLA genotyping. Among these 63 blastomeres, 59 blastomeres had genotypes confirmed from their parents(93.6%), 3 blastomeres lost one of parents' alleles(4.8%), and only one blastomere had two more than parents' alleles(1.6%).</p><p><b>CONCLUSION</b>The above research results indicated that based on the successful first round amplification of single blastomeres, nest-PCR-SSP strategy offers a convenient and reliable option for HLA genotyping on single blastomeres, which is a key process in pre-selecting HLA-identical sibling for allogeneic cord blood cell transplantation.</p>
Subject(s)
Female , Humans , Male , Base Sequence , Blastomeres , Metabolism , DNA , DNA Fingerprinting , Methods , DNA Mutational Analysis , HLA Antigens , HLA-A Antigens , Genetics , Histocompatibility Antigens Class I , Genetics , Polymerase Chain Reaction , Methods , Single PersonABSTRACT
Objective To investigate the protective effects of alcoholic extracts of Ganoderma lucidum (GL) and its combination with Rad ix Salviae Miltiorrhizae (RSM), Radix Bupleuri (RB), and Fructus Schisandrae Chi nensis (FSC) respectively on experimental hepatic injuries. Methods Hepatic injury models were established with the injections of carbon tetrac hloride (CCl4) or D-galactosamine(D-GlaN) into mouse, and then the therapeut ic agents at same dose were given respectively. The activity of cholinesterase(C hE) and alaline aminotransferase(ALT) in the serum and hepatic homogenate, the c ontents of malonyl dialdehyde (MDA), alkaline phosphatase (ALP) and reduced glut athione (GSH) in hepatic homogenate from all mouse were determined. The morpholo gic changes of the livers were also observed with VG staining. Results In CCl4 treated mice, serum ALT activity was increased markedly wh ile ChE acti vity decreased significantly. GL and GL+RB, GL +FSC could relieve these changes ; GL combined with all other agents could inhibit the GSH accumulation, but only GL+FSC had a suppressive effect on MDA increase and made it return to normal. For D-GlaN treated mice, all agent groups had little effect on the activity of ChE; Only GL dramatically reduced the increased activity of ALT; GL +RSM could e liminate the GSH, but not significant; GL and all GL combinations could decrease the increased MDA (P<0.01). Pathological observation showed that hepatocyte damage in GL group and GL+FSC group incline d to recover. Conclusion The results indicate that Ganoderma lucidum is an effective agent against hepatic injuries, especially combined wi th F ructus Schisandrae Chinensis, which may be associated with its anti-hepatocyte oxidation.
ABSTRACT
Objective To investigate the protective effects of alcoholic extracts of Ganoderma lucidum (GL) and its combination with Rad ix Salviae Miltiorrhizae (RSM), Radix Bupleuri (RB), and Fructus Schisandrae Chi nensis (FSC) respectively on experimental hepatic injuries. Methods Hepatic injury models were established with the injections of carbon tetrac hloride (CCl4) or D-galactosamine(D-GlaN) into mouse, and then the therapeut ic agents at same dose were given respectively. The activity of cholinesterase(C hE) and alaline aminotransferase(ALT) in the serum and hepatic homogenate, the c ontents of malonyl dialdehyde (MDA), alkaline phosphatase (ALP) and reduced glut athione (GSH) in hepatic homogenate from all mouse were determined. The morpholo gic changes of the livers were also observed with VG staining. Results In CCl4 treated mice, serum ALT activity was increased markedly wh ile ChE acti vity decreased significantly. GL and GL+RB, GL +FSC could relieve these changes ; GL combined with all other agents could inhibit the GSH accumulation, but only GL+FSC had a suppressive effect on MDA increase and made it return to normal. For D-GlaN treated mice, all agent groups had little effect on the activity of ChE; Only GL dramatically reduced the increased activity of ALT; GL +RSM could e liminate the GSH, but not significant; GL and all GL combinations could decrease the increased MDA (P<0.01). Pathological observation showed that hepatocyte damage in GL group and GL+FSC group incline d to recover. Conclusion The results indicate that Ganoderma lucidum is an effective agent against hepatic injuries, especially combined wi th F ructus Schisandrae Chinensis, which may be associated with its anti-hepatocyte oxidation.