ABSTRACT
Purpose To investigate the expression of CYP4 A11 and CYP4 A22 in triple-negative breast carcinoma (TNBC) and its relationship with clinicopathological features and M2 tumor-associated macrophages (TAMs). Methods 72 cases of TNBC with clinical and pathological data were collected. The expression of CYP4 A11 and CYP4 A22 in the carcinoma cells and the expression of CD68 and CD163 of the TAMs were detected by immunohistochemically and analyzed with image processing software. The relationship between the expressions of CYP4 A11 and CYP4 A22 with clinicopathologic features and its correlation of the M2 state of TAMs was studied. Results Both the immunohistochemically staining scores of CYP4 A11 and CYP4 A22 were higher in cancer tissues than that in breast tissues (P<0.001, P<0.001). The higher expression of CYP4 A11 was associated with tumor diameter increase (P<0.001), lymph node metastasis (P<0.001), higher clinical stage (P<0.001) and higher Ki-67 index (P=0.011). Both the positive rates of CD68 and CD163 in the high expression group of CYP4 A11 were higher than those in the low expression group of CYP4 A11 (P=0.021, P<0.001). The higher expression of CYP4 A22 was associated with lymph node metastasis (P<0.001), higher clinical stage (P=0.006), higher recurrence rate (P<0.001), and higher Ki-67 index (P=0.040).The positive rates of CD163 in the high expression group of CYP4 A22 was higher than that in the low expression group of CYP4 A22 (P<0.001). Conclusion Both the expression of CYP4 A11 and CYP4 A22 may be associated with M2 polarization state of TAMs, high proliferative activity and lymph node metastasis in the TNBC.
ABSTRACT
Objective To investigate the association between single nucleotide polymorphisms (SNPs) in CYP4A11 and ApoB、 ApoE genesandthe risk of essential hypertension. Methods By means of correlation analysis, this case-controlled study included a total of 350 essential hypertension patients admitted in the first Hospital of Xi'an from June 2012 to December 2016, and another 350 cases with matched ages and sexes enrolled for routine check-ups as the healthy control group. The genotypes of CYP4A11 and ApoB、 ApoE genes were determined by MassARRAY method. SPSS 21.0 software was used to determine the correlation between SNPs and the risk of primary hypertension.Results Comparing the allele frequencies of SNPs, we found rs1126742 (OR=1. 45, 95 % CI, 1.09-1.92,P=0.008)、 rs3890011 (OR=1.98, 95 % CI, 1.06-1.85, P=0.001) and rs9332978 (OR=1.54, 95%CI, 1.27-1.91,P=0.004) on CYP4A11 were significantly associated with an increased risk of essential hypertension; XbaⅠpolymorphism in ApoB gene was associated with essential hypertension risks (OR=1.55,95%CI, 1.15-2.55,P=0.001);ε4 in ApoE gene was also found associated with essential hypertension risks (OR=1.49, 95 % CI, 1.09-2.35, P=0.012), while ε2 andε3 not associated.Comparing the genotype frequencies of SNPs,we found the GG genotype of rs9332978, TC and CC genotype of rs1126742, CG and GG genotypes of rs3890011 were associated with increased risks of essential hypertension (P＜0.05) .The X -X- genotype of XbaⅠin ApoB gene were associated with increased risks of essential hypertension (OR=2.45, 95%CI, 1.25-3.25, P=0.035) . The E2/E4、E3/E4、E4/E4 genotype of ApoE were associated with increased risks of essential hypertension (P＜0.05).In the genetic model analysis, we found that the minor allele "A" of rs9332978 was associated with an increased risk of essential hypertension under dominant model (P＜0.05) . The minor allele "T" of rs1126742 was associated with increased risks essential hypertension under dominant and recessive models (P＜0.05) . Conclusion Gene polymorphisms of CYP4A11、ApoB and ApoE play an important role in the occurrence and development of essential hypertension.
ABSTRACT
Cytochrome P450 4A11 (CYP4A11) is a fatty acid hydroxylase enzyme expressed in human liver. It catalyzes not only the hydroxylation of saturated and unsaturated fatty acids, but the conversion of arachidonic acid to 20-hydroxyeicosatetraenoic acid (20-HETE), a regulator of blood pressure. In this study, we performed a directed evolution analysis of CYP4A11 using the luminogenic assay system. A random mutant library of CYP4A11, in which mutations were made throughout the entire coding region, was screened with luciferase activity to detect the demethylation of luciferin-4A (2-[6-methoxyquinolin-2-yl]-4,5-dihydrothiazole-4-carboxylic acid) of CYP4A11 mutants in Escherichia coli. Consecutive rounds of random mutagenesis and screening yielded three improved CYP4A11 mutants, CP2600 (A24T/T263A), CP2601 (T263A), and CP2616 (A24T/T263A/V430E) with ~3-fold increase in whole cells and >10-fold increase in purified proteins on the luminescence assay. However, the steady state kinetic analysis for lauric acid hydroxylation showed the significant reductions in enzymatic activities in all three mutants. A mutant, CP2600, showed a 51% decrease in catalytic efficiency (k cat/K m) for lauric acid hydroxylation mainly due to an increase in K m. CP2601 and CP2616 showed much greater reductions (>75%) in the catalytic efficiency due to both a decrease in k cat and an increase in K m. These decreased catalytic activities of CP2601 and CP2616 can be partially attributed to the changes in substrate affinities. These results suggest that the enzymatic activities of CYP4A11 mutants selected from directed evolution using a luminogenic P450 substrate may not demonstrate a direct correlation with the hydroxylation activities of lauric acid.