Effects Of Genetic Polymorphism Of Cytochrome P450Oxidoreductase On Its MRNA And Activity

Research backgroundCytochrome P450oxidoreductase (POR), a kind of flavoprotein, provides electrons to all microsomal cytochrome P450(CYP) enzymes and is essential for CYP activities. To our knowledge, there is no detail report of POR biological parameters in large samples.AimsThis study was planed to determine the mRNA level and activity of POR to rich the physiological data in HLM and analyzed the affected factors. A correlation analysis of POR mRNA level and POR activity were performed.MethodsPOR mRNA levels were identified by qPCR. POR activity was determined by measuring cytochrome C reduction in liver microsomes and POR polymorphisms were identified by SnapShort. ResultsThere was a substantial variation in POR mRNA level (0.047～1.25with a median of0.28). The POR activity in HLM protein ranged from26.91to125.77nmol·min-l*mg-1with median of56.05nmol·min-1·mg-1, the difference between the maximum and minimum values is4.67times. There was a substantial variation in the POR activity in liver tissue proteins ranged from215to12323nmol·min-1·g-1with median of2152nmol·min-1·g-1.We define the same bases of the eight SNPs of POR gene as a kind of genotype.123cases of donor can be divided into14halotypes, and analysed the difference of POR activity between halotypes. The results showed:comparing to halotype1(n=24) and halotype7(n=7), the difference of four bases (rs1057868, rs10954732, rs1135612, rs41301394) leaded to POR activity of each mg HLM was statistically significant difference between two kind of halotype samples (P=0.011). Comparing to halotype8(n=8) and halotype10(n=36), the difference of three bases (rs1135612, rs4732515, rs4732516) leaded to POR activity of each mg HLM was statistically significant difference between two kind of halotype samples (P=0.028). Furthermore, comparing to the POR activity of halotype6and halotype11, halotype8and halotype11, halotype1and halotype11, halotype6and halotype8, there were statistically significant difference (P<0.05). With each g liver tissue for POR activity, comparing to halotype7(n=7) and halotype1(n=24) or halotype6(n=6), the POR activity of each g liver tissue was statistically significant difference (P<0.05).We analysed the difference of POR mRNA expression level between14halotypes in the same way. The results showed:comparing to halotype3and halotype1or halotype7, halotype8, halotype10, the POR mRNA expression level were statistically significant difference (P<0.05). Analyzing demography factors, the results showed:there was no influence of age, gender, smoking to POR mRNA expression level.Furthermore, we analyzed the statistically correlation of POR mRNA expression level and POR activity only if POR gene SNP locus mutation results were the same. The results showed:when the donors (n=11) of POR genotype of rs1057868and rs41301394were TT homozygous mutations, there was statistically significant correlation of POR mRNA expression level and POR activity (r=0.675, P=0.023). POR mRNA expression level and POR activity of halotype8(rs1057868-rs4732516: TC-G-A-G-C-CT-TC-CG)(n=6) were statistically significant correlation (r=0.885, P=0.019). The results of demography factors analysis showed:POR mRNA expression level and POR activity of male were statistically significant correlation (r=0.384, P=0.036). POR mRNA expression level and POR activity of contemporary were not statistically significant correlation. POR mRNA expression level and POR activity of smoker were statistically significant correlation (r=0.658, P=0.028). POR mRNA expression level and POR activity of drinker were not statistically significant correlation.Conclusions1. POR gene mutation influence POR mRNA and activity.2. Smoking and drinking significant decreased POR activity.3. There was no correlation of POR mRNA and POR activity in general.4. The POR mRNA level was significantly associated with the POR activity of men (r=0.384, P=0.036) and smokers (r=0.658, P=0.028)5. Biological data of POR in normal large liver samples are established. These data provide fundamental and useful information for the pharmacokinetics studies of drugs metabolized by CYPs in the Chinese population.