Abstract
This study aims to explore the effect of gene polymorphisms of 5a-reduction enzyme (SRD5A2), steroidogenic cytochrome P-450 17alpha-hydroxylase (CYP17), aromatase cytochrome P450 family 19 (CYP19) and vita-min D receptor (VDR) on benign prostatic hyperplasia (BPH) susceptibility and clinical progress. A total of 452 BHP patients and 501 healthy individuals were selected in Harbin Medical University Daqing School from October 2014 and December 2015 as the case and control groups. All BPH patients received drug treatment and were subsequently divided into the progression and non-progression groups based on their therapeutic efficacy. PCR-RFLP was applied to detect the genotype distributions of SRD5A2/CYP17/CYP19/VDR, which were further tested with Hardy-Weinberg (H-W) equilibrium. Logistic regression analysis was applied to determine the risk factors for BPH progression. Compared with subjects carrying VV genotype and V allele at SRD5A2 V89L, those with LL genotype and L allele at SRD5A2 V89L may have reduced risk of BPH susceptibility or progression (all P < 0.05). Compared with subjects carrying TT genotype and T allele at CYP17 -34T>C, those with CC genotype and C allele at CYP17 -34T>C may have increased risk of BPH susceptibility or progression (all P < 0.05). Compared with individuals carrying FF genotype and F allele at VDRVDR Fok I, those with ff genotype and f allele at VDRVDR Fok I may have increased susceptibility to BPH (all P < 0.05). Logistic regression analysis showed that SRD5A2 V89L and CYP17 -34T>C polymorphisms and CYP17 -34T>C (TC + CC)/SRD5A2 V89L (VV) combined genotypes were significantly related with the clinical progression of BHP. These results revealed that SRD5A2 V89L and CYP17 -34T>C polymorphisms were associated with the risk of BPH and its clinical progression.
Keywords: SRD5A2, CYP17, CYP19, VDR, benign prostatic hyperplasia, gene polymorphisms
Introduction
Benign prostatic hyperplasia (BPH) usually represents the urologic diagnosis that occurs most frequently in elderly males [1]. It is a histologic diagnosis which refers to epithelial cell and smooth muscle proliferation in the prostatic transition zone [2]. BPH is also a kind of proliferative abnormality of the human prostate, which is often related with age [3]. BPH occurs in about 50% of men by the age of 50, the incidence of which usually increases with age [4]. Report suggested that the incidence rate of this disease is about 2.96 per 1000 man among patients in their forties, while 34.46 per 1000 man among patients in their seventies [5]. There are mainly two established risk factors in the development of this disease: aging and androgens [3]. Laser technology has been adopted to treat BPH for years [6]. Pharmacological therapy is most commonly applied for patients with moderate to severe BPH, which aims to improve symptoms and identify patients who are at risk of disease progression so as to optimize their management [7,8]. And the number of BPH patients seeking treatment is expected to increase in the next few years because of the ageing male population [9]. Certain alleles of some genes are associated with BPH [10,11].
The gene encoding steroid 5-alpha reductase type II (SRD5A2) is proved to be located on chromosome 2p23 and has ive exons and four introns [12,13]. Steroidogenic cytochrome P-450 17alpha-hydroxylase (CYP17) encodes an enzyme with activities of 17-hydroxylase and 17,20-lyase, which is a step with rate limiting in the biosynthesis of testosterone [14]. Aromatase Cytochrome P450 family 19 (CYP19) is a key enzyme in the estrogen metabolism and CYP19 polymorphisms usually cause changes in enzyme activity and can influence the estrogen synthesis [15]. Vita-min D receptor (VDR) belongs to the trans-acting transcriptional regulatory factors family and can modulate many biological activities of the endocrine, immune and neural systems, including apoptosis, cell differentiation, calcium and phosphorous homeostasis [16]. VDR polymorphisms have been proved to be associated with many diseases such as the leprosy phenotypes and ovarian cancer [17,18]. Similarly, the variants in SRD5A2 are reported to be connected with the quality of semen [19]. It is also proved that variants in CYP17 and CYP19 genes are related with the onset of Alzheimer’s disease [20].
However, little can be found on the genetics and mechanism of SRD5A2, CYP17, CYP19 and VDR polymorphisms influencing BPH, especially in northern Chinese population. In order to further understand the genetic characteristics of BPH, this study targets to explore the association of BPH with SRD5A2, CYP17, CYP19 and VDR polymorphisms among northern Chinese men, hoping to provide a new sight for the diagnosis and treatment of BPH.
Materials and methods
Ethical statement
The present study was performed in accordance with the guidelines established by Medicine Ethics Review Committee at Harbin Medical University Daqing School. All patients have signed written forms of consent.
Study subjects
A total of 452 BPH patients were selected into the case group in the urological department at Harbin Medical University Daqing School from October 2014 to December 2015. The inclusion criteria were as follows: patients who (1) met the BPH diagnosis criteria recommended by the 5th International Benign Prostatic Hyperplasia Advisory Committee in 2001 [21]; (2) had no abnormal echo in abdominal or rectal prostate ultrasound; (3) had a prostate specific antigen (PSA) concentration greater than 4 ng/mL; (4) had a prostate volume (PV) greater than 30 mL; (5) had a postvoiding residue (PVR) greater than 30 mL; (6) went through pathological examination of prostate and were confirmed by two experienced pathophysicians in Harbin Medical University Daqing School; (7) were permanent residents of northern China (lived in the local community for more than 2 years); (8) received no formal treatment before this study. Exclusion criteria were as follows: patients who (1) were confirmed as prostate cancer and prostate sarcoma in immunohistochemical examination; (2) had previous history of surgery in the prostate, urethra and bladder; (3) had neurological diseases that may affect the urinary tract functions; (4) had urinary tract infection; (5) used medications that may affect the urinary functions. During the same period, 501 healthy individuals who underwent physical examinations at Harbin Medical University Daqing School were enrolled into the control group. The subjects in the control group were all permanent residents of northern China (lived in the local community for more than 2 years) and had no blood relationship with the case group. Blood samples from all subjects were collected and detailed clinical data were recorded.
Treatment regimen and grouping
BPH patients were treated with combined therapy of Terazosin (National medicine permission number: H20023659, Abbott (Shanghai) Pharmaceutical Co., Ltd., Shanghai, China) and Finasteride tablets (National medicine permission number: J20090145, Merck (Hangzhou) Pharmaceutical Co., Ltd., Hangzhou, China). Treatment regimen: one tablet of Terazosin (2 mg) and 1 tablet of Finasteride (5 mg) were given orally per day before sleep for 3 months consecutively. For patients who showed significant improvement, Finasteride tablets were given alone. The case group was further divided into clinical progression group and non-progression group according to the following assessment indicators for clinical progress of BPH after drug treatment [22]: (1) decreased dynamic maximum urinary flow rate; (2) presence of complications such as acute urinary retention, hematuria, urinary tract infection, bladder stones and renal dysfunction.
Sample collection
Ten ml fasting venous blood were collected from all subjects in the morning. Ethylenediamine tetraacetic acid (EDTA) was added to 4 ml blood samples as anticoagulant and stored in refrigerator at -80°C. The genomic DNA was extracted using a conventional phenol extraction method and was diluted to a final concentration of 10 ng/μl. Two ml blood samples were used for routine blood examination, which covered: total cholesterol (TC), low density lipoprotein cholesterol (LDL-C), triglyceride (TG), high density lipoprotein cholesterol (HDL-C) and PSA concentration. Clinical data, including age, height, weight, PV, maximum flow rate (Qmax) and PVR, were obtained from all subjects.
Single nucleotide polymorphism (SNP) screening and sequencing
The candidate loci identified in this study were 5 PV-associated SNPs selected in a previous prostate cancer genome-wide association study (GWAS), including SRD5A2 A49T and V89L, CYP17 -34T>C, CYP19 +19C>T, VDR Apa I and Fok I. The relevant gene sequences were obtained from Genbank, and their primers were designed by Oligo 6.0 and Primer 5.0 software. The relevant primers were shown in Table 1.
Table 1.
Primer sequence for each SNP
| Locus | Gene | Forward primer | Reverse primer |
|---|---|---|---|
| A49T | SRD5A2 | 5’-ACCCTTGGGGCACTGGCCTTG-3’ | 5’-GTCAGCTCCTGCAGGAACCAG-3’ |
| V89L | SRD5A2 | 5’-CCACCTGGGACGGTACTTCT-3’ | 5’- CTCCACGCTGCGCTCCTGGA-3’ |
| -34T>C | CYP17 | 5’-CATTCGCaccTCTGGAGTC-3’ | 5’-GGCTCTTGGGGTACTTG-3’ |
| +19C>T | CYP19 | 5’-GGTACTTAGTTAGCTACAATC-3’ | 5’-GGGTGATAGAGTCAGAGCCT-3’ |
| Apa I | VDR | 5’-CAACCAAGACTACAAGTACCGCGTCAGTGA-3’ | 5’- CACTTCGAGCACAAGGGGCGTTAGC-3’ |
| Fok I | VDR | 5’-AGCTGGCCCTGGCACTGACTCTGCTCT-3’ | 5’-ATGGAAACACCTTGCTTCTTCTCCCTC-3’ |
Note: SNP, single nucleotide polymorphism.
The genomic DNA was extracted using a phenol/chloroform method and analyzed by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). The PCR reaction system included: 10 ng of template DNA (batch number: KN1014, C&M Biolabs, Richmond, CA, USA), 10 × PCR buffer, 15 pmol of each primer, 0.2 mmol/L dNTPs, 0.12 U Tag enzyme (batch number: K1321, Shanghai Runcheng Biotechnology Co., Ltd., Shanghai, China). The above components were diluted with double distilled water to a final volume of 15 μl. A touchdown PCR reaction was performed on a C100 PCR apparatus (Bio-Rad, Hercules, CA, USA). The amplification conditions were: 94°C for 2 min, 94°C for 30 s, 63°C for 1 min and 72°C for 1 min; and 94°C for 30 s, 57°C for 1 min, 72°C for 1 min and 72°C for 7 min. The amplification products were transferred to 1.5% agarose gel electrophoresis (2117 Multiphor II, Bio-Rad, Hercules, CA, USA). The purity of the amplified bands were observed under an ultraviolet lamp and the concentration was estimated. A total of 3 μl of each PCR amplification product was added with different endonucleases at the following reaction condition was as follows: 45 min at 37°C and 15 min at 85°C. The PCR product was then kept at 4°C. A reverse primer (1 μl) and BDT (1 μl) were added into purified PCR products using a Prism BigDyeTerminator (BDT) Cycle Sequencing kit (lot number: N73144, ABI, Thermo Fisher, Waltham, MA, USA).
Follow-up
BPH patients were followed up every 3 months for 3 to 12 months through phone calls, outpatient service and medical record evaluation. The follow-up ended in February, 2016. Altogether, 6 patients were lost to follow-up and the follow-up rate was 99.50%. Detailed information about the clinical progression of BPH was closely monitored, including: (1) Qmax; (2) the incidence of complications such as acute urinary retention, hematuria, urinary tract infection, bladder stones and renal function impairment.
Statistical method
SPSS 17.0 software was used for data analysis. The measurement data were expressed as mean ± SD. The t test was used for comparison between two groups in normal distribution. One-way ANOVA was applied for comparison among multiple groups with variance homogeneity and normality. Nonparametric Kruskal-Wallis rank-sum test was used for comparison among multiple groups without variance homogeneity and normality. Count data were expressed in percentage or rate and were compared using X2 test. The comparison of alleles and genotypes and the Hardy-Weinberg equilibrium of alleles were verified using X2 test. The relative risk was expressed with odds ratio (OR) and 95% confidence interval (CI). Logistic regression analysis was carried out to analyze the risk factors for clinical progression of BPH. P < 0.05 was considered statistically significant.
Results
Comparisons of baseline characteristics between the control group and the BPH group
The BMI, HDL-C and Qmax in the BPH group were significantly lower than those in the control group (all P < 0.05). The levels of TG, PV, PSA and PVR in the BPH group were significantly higher than those in the control group (all P < 0.05). There was no significant difference in age, TC and LDC-C between the two groups (all P > 0.05) (Table 2).
Table 2.
Comparison of baseline characteristics between the control group and the BPH group
| Item | BPH group (n = 452) | Control group (n = 501) | P |
|---|---|---|---|
| Age (years) | 58.02 ± 7.38 | 57.28 ± 7.61 | 0.129 |
| BMI (kg/m2) | 22.46 ± 3.55 | 24.14 ± 3.63 | < 0.001 |
| TC (mmol/L) | 4.75 ± 0.91 | 4.72 ± 0.99 | 0.628 |
| TG (mmol/L) | 2.18 ± 0.91 | 1.98 ± 1.22 | 0.005 |
| LDL-C (mmol/L) | 3.21 ± 0.77 | 3.14 ± 0.81 | 0.173 |
| HDL-C (mmol/L) | 1.58 ± 0.31 | 1.63 ± 0.35 | 0.02 |
| PV (mL) | 69.59 ± 20.49 | 66.68 ± 17.76 | 0.019 |
| PSA (ng/L) | 2.02 ± 1.11 | 1.71 ± 0.81 | < 0.001 |
| Qmax (mL/s) | 6.35 ± 1.91 | 6.59 ± 1.79 | 0.046 |
| PVR (mL) | 62.89 ± 20.71 | 58.51 ± 21.05 | 0.001 |
Notes: BMI, body mass index; TC, total cholesterol; TG, triglyceride; LDL-C, low density lipoproteincholesterol; HDL-C, high-density lipoproteincholesterol; PV, prostate volume; PSA, prostate specific antigen concentration; Qmax, maximum flow rate; PVR, postvoiding residue; the t test was performed.
Electrophoresis results of SRD5A2 A49T and V89L, CYP17 -34T>C, CYP19 +19C>T, VDR Apa I and Fok I
In the SRD5A2 A49T locus, AA, AT and TT genotypes were detected after Rsai digestion: a single restriction site was found in the wild type homozygous (AA), which was cut into bands of 169 bp, 105 bp and 64 bp; the wild-type homozygous (TT) was cut into bands of 169 bp, 105 bp and 83 bp; the mutant heterozygote (AT) only showed a 169 bp band (Figure 1A).
Figure 1.
Electropherogram of PCR products at each gene locus. Notes: A: Electropherogram of SRD5A2 A49T locus after Rsai enzyme digestion, M: Marker; AA: Wild-type homozygote; AT: Mutant heterozygote; TT: Mutant homozygote; B: Electropherogram of SRD5A2 V89L locus after HYF10VI enzyme digestion; M: Marker; VV: Wild-type homozygote; VL: Mutant heterozygote; LL: Mutant homozygote; C: Electropherogram of CYP17 -34T>C locus after MspA1I enzyme digestion, CC: Mutant homozygote; TC: Mutant heterozygote; TT: Mutant homozygote; D: Electropherogram of CYP19 +19C>T locus after BSP1286 I enzyme digestion, CC: Mutant homozygote; TC: Mutant heterozygote; TT: Mutant homozygote; E: Electropherogram of VDR Apa I locus post Apa I enzyme digestion; M: Marker; AA: Wild-type homozygote; Aa: Mutant heterozygote; aa: Mutant homozygote; F: Electropherogram of VDR Fok I locus post Fok I enzyme digestion, FF: Wild-type homozygote; Ff: Mutant heterozygote; ff: Mutant heterozygote; M: Marker; PCR = polymerase chain reaction.
In the SRD5A2 V89L locus, VV, VL and LL genotypes were detected after HYF10VI digestion: wild-type homozygous (VV) had no digestion site and only showed a 89 bp band; the mutant homozygous (LL) only showed a 124 bp band; the mutant heterozygous (VL) was cut into bands of 124 bp and 89 bp (Figure 1B).
After MspA1I digestion, the CYP17 -34T>C site showed TT, CC and TC genotypes: the wild-type homozygote (TT) had no digestion site and only showed a 459 bp band; the mutant homozygote (CC) had one digestion site and was cut into bands of 336 bp and 123 bp; the mutant heterozygous (TC) was cut into bands of 459 bp, 336 bp and 123 bp (Figure 1C).
After Bsp1286 I digestion, the CYP19 +19C>T site showed TT, CC and TC genotypes: the mutant homozygote (TT) had no digestion site and only showed a 202 bp band; the wild-type homozygote (CC) had one digestion site and was cut into bands of 172 bp and 30 bp; the mutant heterozygous (TC) was cut into bands of 202 bp, 172 bp and 30 bp (Figure 1D).
In the VDR Apa I locus, AA, Aa and aa genotypes were detected after Apa I digestion: the mutant homozygous (aa) and wild-type homozygous (AA) both only showed a 495 bp band; the mutant heterozygote (Aa) had one digestion site and was cut into bands of 495 bp, 280 bp and 215 bp (Figure 1E).
In the VDR Fok I locus, FF, Ff and ff genotypes were detected after Fok I digestion: the mutant homozygous (ff) genotype had two digestion sites and was cut into two bands of 199 bp and 59 bp; the wild-type homozygous (FF) had no digestion site and only showed a 246 bp band; the mutant heterozygous (Ff) had one digestion site and was cut into bands of 59 bp, 199 bp and 246 bp (Figure 1F).
Hardy-Weinberg equilibrium test for SRD5A2 A49T and V89L, CYP17 -34T>C, CYP19 +19C>T, VDR Apa I and Fok I
Hardy-Weinberg equilibrium analysis was carried out for different loci in the control group. No significant difference was found between the actual and expected values of SRD5A2 A49T, SRD5A2 V89L, CYP17 -34T>C, CYP19 +19C>T, VDR Apa I and VDR Fok I (all P > 0.05) (Table 3), indicating that the distribution frequency of each gene locus is representative.
Table 3.
Hardy-Weinberg equilibrium for the representativeness of SRD5A2 A49T and V89L, CYP17 -34T>C, CYP19 +19C>T, VDR Apa I and Fok I
| Locus | Genotype | Predicted value n (%) | Actual value n (%) | X 2 | P |
|---|---|---|---|---|---|
| SRD5A2 A49T | AA | 421 (84.03) | 417 (83.23) | 4.322 | 0.115 |
| AT | 77 (14.78) | 84 (16.77) | |||
| TT | 4 (0.80) | 0 (0.00) | |||
| SRD5A2 V89L | VV | 320 (63.87) | 326 (65.07) | 1.303 | 0.521 |
| VL | 161 (32.14) | 149 (29.74) | |||
| LL | 20 (3.99) | 26 (5.19) | |||
| CYP17 -34T>C | TT | 382 (76.25) | 377 (75.25) | 2.737 | 0.255 |
| TC | 111 (22.16) | 121 (25.15) | |||
| CC | 8 (1.60) | 3 (0.60) | |||
| CYP19 +19C>T | CC | 337 (67.27) | 343 (68.46) | 1.507 | 0.471 |
| TC | 148 (29.54) | 136 (27.15) | |||
| TT | 16 (3.19) | 22 (4.39) | |||
| VDR Apa I | AA | 356 (71.06) | 363 (72.46) | 2.321 | 0.313 |
| Aa | 132 (26.35) | 119 (23.75) | |||
| aa | 12 (2.40) | 19 (3.79) | |||
| VDR Fok I | FF | 390 (77.78) | 390 (77.78) | 0 | 1 |
| Ff | 104 (20.76) | 104 (20.76) | |||
| ff | 7 (1.40) | 7 (1.40) |
Note: the chi-square test was performed.
Distribution frequency of SRD5A2/CYP17/CYP19/VDR SNPs between the BPH group and the control group
There was no significant difference in the distribution frequency of SRD5A2 A49T, VDR Apa I and CYP19 +19C>T between the BPH and control group (all P > 0.05). The frequency of SRD5A2 V89L was significantly different between the control group and the BPH group (all P < 0.05). Compared with the VV genotype, the LL genotype may reduce the risk of BPH (OR = 0.401, 95% CI = 0.190-0.845, P = 0.013); compared with the V allele, the L allele also tended to reduce the risk of BPH (OR = 0.787, 95% CI = 0.622-0.994, P = 0.044). The distribution frequency of CYP17 -34T>C was significantly different between the BPH and control groups (all P < 0.05). Compared with the TT genotype, the CC genotype may increase the risk of BPH (OR = 9.064, 95% CI = 2.687-30.580, P < 0.001); compared with the T allele, the C allele also tended to increased the risk of BPH (OR = 1.584, 95% CI = 1.233-2.035, P < 0.001). The distribution frequency of VDR Fok I was significantly different between the control group and the BPH group (all P < 0.05). Compared with the VDR FF genotype, the ff genotype may increase the risk of BPH (OR = 1.587, 95% CI = 1.266-2.053, P = 0.007); compared with the F allele, the f allele also tended to increase the risk of BPH (OR = 1.587, 95% CI = 1.266-2.053, P < 0.001) (Table 4).
Table 4.
Distribution frequency of SRD5A2/CYP17/CYP19/VDR SNPs between the BPH group and the control group
| Genotype | BPH group (N = 452) | Control group (N = 501) | OR (95% CI) | P | ||
|---|---|---|---|---|---|---|
|
| ||||||
| Cases | % | Cases | % | |||
| SRD5A2 A49T | ||||||
| AA | 385 | 85.18 | 417 | 83.21 | 1 (Reference) | |
| AT | 64 | 14.16 | 84 | 16.79 | 1.137 (0.851-1.726) | 0.286 |
| TT | 3 | 0.66 | 0 | 0 | 7.851 (0.390-147.30) | 0.072 |
| AT + TT | 67 | 14.82 | 84 | 16.77 | 1.158 (0.816-1.642) | 0.412 |
| A | 834 | 92.26 | 918 | 91.62 | 1 (Reference) | |
| T | 70 | 7.74 | 84 | 8.38 | 1.090 (0.783-1.518) | 0.609 |
| SRD5A2 V89L | ||||||
| VV | 313 | 69.25 | 326 | 65.07 | 1 (Reference) | |
| VL | 129 | 25.54 | 149 | 29.74 | 0.902 (0.680-1.196) | 0.472 |
| LL | 10 | 2.21 | 26 | 5.19 | 0.401 (0.190-0.845) | 0.013 |
| VL + LL | 121 | 30.75 | 175 | 34.93 | 0.720 (0.545-0.952) | 0.021 |
| V | 755 | 85.52 | 801 | 79.94 | 1 (Reference) | |
| L | 149 | 16.48 | 201 | 20.06 | 0.787 (0.622-0.994) | 0.044 |
| CYP17 -34T>C | ||||||
| TT | 305 | 67.48 | 377 | 75.25 | 1 (Reference) | |
| TC | 125 | 27.65 | 121 | 24.15 | 1.277 (0.954-1.710) | 0.101 |
| CC | 22 | 4.87 | 3 | 0.6 | 9.064 (2.687-30.580) | < 0.001 |
| TC + CC | 147 | 32.52 | 124 | 24.75 | 1.454 (1.096-1.928) | 0.009 |
| T | 735 | 81.31 | 875 | 87.33 | 1 (Reference) | |
| C | 169 | 18.69 | 127 | 12.67 | 1.584 (1.233-2.035) | < 0.001 |
| CYP19 +19C>T | ||||||
| CC | 310 | 68.58 | 343 | 68.37 | 1 (Reference) | |
| TC | 121 | 26.77 | 136 | 27.25 | 0.984 (0.737-1.315) | 0.915 |
| TT | 21 | 4.65 | 22 | 4.38 | 0.947 (0.511-1.756) | 0.862 |
| TC + TT | 142 | 31.42 | 159 | 31.54 | 0.988 (0.752-1.299) | 0.932 |
| C | 741 | 81.97 | 823 | 82.04 | 1 (Reference) | |
| T | 163 | 18.03 | 181 | 17.96 | 1.000 (0.791-1.263) | 0.998 |
| VDR Apa I | ||||||
| AA | 307 | 67.92 | 363 | 72.46 | 1 (Reference) | |
| Aa | 125 | 27.65 | 119 | 23.75 | 1.242 (0.926-1.666) | 0.147 |
| aa | 20 | 4.42 | 19 | 3.79 | 1.245 (0.652-2.375) | 0.506 |
| Aa + aa | 145 | 32.08 | 137 | 27.54 | 1.251 (0.947-1.654) | 0.114 |
| A | 740 | 81.75 | 845 | 84.33 | 1 (Reference) | |
| a | 164 | 18.25 | 155 | 15.67 | 1.208 (0.950-1.537) | 0.123 |
| VDR Fok I | ||||||
| FF | 312 | 69.03 | 389 | 77.64 | 1 (Reference) | |
| Ff | 122 | 26.99 | 105 | 20.96 | 1.466 (1.085-1.981) | 0.012 |
| ff | 18 | 3.98 | 7 | 1.4 | 3.214 (1.325-7.795) | 0.007 |
| Ff + ff | 140 | 30.97 | 112 | 22.36 | 1.577 (1.179-2.108) | 0.002 |
| F | 746 | 82.52 | 883 | 88.12 | 1 (Reference) | |
| f | 158 | 17.48 | 119 | 11.88 | 1.587 (1.226-2.053) | < 0.001 |
Notes: BPH, benign prostatic hyperplasia; OR, odds ratio; 95% CI, 95% confidence interval; the chi-square test was performed.
Relationship between SRD5A2 A49T and V89L, CYP17 -34T>C, CYP19 +19C>T, VDR Apa I and Fok I gene polymorphisms and clinical progression of BPH
No significant difference was found in the distribution frequencies of SRD5A2 A49T, CYP19 +19C>T, VDR Apa I and VDR Fok I between the progression group and the non-progression group (all P > 0.05). The distribution frequency of SRD5A2 V89L genotypes and alleles was not significantly different between the progression group and the non-progression group (all P > 0.05). Compared with the VV genotype, the LL genotype might decrease the risk of clinical progression in BPH patients (OR = 0.235, 95% CI = 0.049-1.123, P = 0.049); compared with the V allele, the L allele might also reduce the risk of clinical progression (OR = 0.534, 95% CI = 0.330-0.865, P = 0.010). The distribution of genotype and allele of CYP17 -34T>C was significantly different between the progression group and the non-progression group (all P < 0.05). Compared with the TT genotype, the CC genotype may increase the risk of clinical progression in BPH patients (OR = 4.821, 95% CI = 1.732-13.420, P = 0.001); compared with the T allele, the C allele might also increase the risk of clinical progression in BPH patients (OR = 2.261, 95% CI = 1.546-3.306, P < 0.001) (Table 5).
Table 5.
Relationship between SRD5A2 A49T and V89L, CYP17 -34T>C, CYP19 +19C>T, VDR Apa I and Fok I gene polymorphisms and clinical progression of BPH
| Genotype | Progression group (N = 215) | Non-progression group (N = 237) | OR (95% CI) | P | ||
|---|---|---|---|---|---|---|
|
| ||||||
| Cases | % | Cases | % | |||
| SRD5A2 A49T | ||||||
| AA | 190 | 88.37% | 195 | 82.28% | 1 (Reference) | |
| AT | 24 | 11.16% | 40 | 16.88% | 0.616 (0.357-1.061) | 0.079 |
| TT | 1 | 0.47% | 2 | 0.84% | 0.513 (0.046-5.710) | 0.58 |
| AT + TT | 25 | 11.63% | 42 | 17.72% | 0.611 (0.358-1.042) | 0.068 |
| A | 404 | 93.95% | 430 | 90.72% | 1 (Reference) | |
| T | 37 | 8.60% | 44 | 9.28% | 0.895 (0.566-1.415) | 0.635 |
| SRD5A2 V89L | ||||||
| VV | 162 | 75.35% | 152 | 64.14% | 1 (Reference) | |
| VL | 51 | 23.72% | 77 | 32.49% | 0.621 (0.409-0.943) | 0.025 |
| LL | 2 | 0.93% | 8 | 3.38% | 0.235 (0.049-1.123) | 0.049 |
| VL + LL | 53 | 24.65% | 85 | 35.86% | 0.585 (0.389-0.880) | 0.01 |
| V | 232 | 53.95% | 324 | 68.35% | 1 (Reference) | |
| L | 26 | 6.05% | 68 | 14.35% | 0.534 (0.330-0.865) | 0.01 |
| CYP17 -34T>C | ||||||
| TT | 122 | 56.74% | 173 | 73.00% | 1 (Reference) | |
| TC | 76 | 35.35% | 59 | 24.89% | 1.827 (1.210-2.757) | 0.004 |
| CC | 17 | 7.91% | 5 | 2.11% | 4.821 (1.732-13.42) | 0.001 |
| TC + CC | 93 | 43.26% | 64 | 27.00% | 2.061 (1.390-3.055) | < 0.001 |
| T | 320 | 74.42% | 405 | 85.44% | 1 (Reference) | |
| C | 110 | 25.58% | 69 | 14.56% | 2.261 (1.546-3.306) | < 0.001 |
| CYP19 +19C>T | ||||||
| CC | 148 | 68.84% | 172 | 72.57% | 1 (Reference) | |
| TC | 58 | 26.98% | 53 | 22.36% | 1.272 (0.825-1.960) | 0.275 |
| TT | 9 | 4.19% | 12 | 5.06% | 0.872 (0.357-2.127) | 0.763 |
| TC + TT | 67 | 31.16% | 65 | 27.43% | 1.198 (0.798-1.798) | 0.383 |
| C | 354 | 82.33% | 327 | 68.99% | 1 (Reference) | |
| T | 76 | 17.67% | 65 | 13.71% | 1.080 (0.751-1.554) | 0.678 |
| VDR Apa I | ||||||
| AA | 145 | 67.44% | 162 | 68.35% | 1 (Reference) | |
| Aa | 61 | 28.37% | 64 | 27.00% | 0.939 (0.619-1.424) | 0.767 |
| aa | 9 | 4.19% | 11 | 4.64% | 0.914 (0.368-2.269) | 0.846 |
| Aa + aa | 70 | 32.56% | 75 | 3418.00% | 0.959 (0.646-1.424) | 0.836 |
| A | 351 | 81.63% | 388 | 81.86% | 1 (Reference) | |
| a | 79 | 18.37% | 86 | 18.14% | 0.985 (0.702-1.381) | 0.929 |
| VDR Fok I | ||||||
| FF | 149 | 69.30% | 163 | 68.78% | 1 (Reference) | |
| Ff | 59 | 27.44% | 63 | 26.58% | 0.976 (0.642-1.484) | 0.91 |
| ff | 7 | 3.26% | 11 | 4.64% | 0.696 (0.263-1.843) | 0.464 |
| Ff + ff | 66 | 30.70% | 74 | 31.22% | 0.976 (0.654-1.455) | 0.904 |
| F | 357 | 83.02% | 389 | 82.07% | 1 (Reference) | |
| f | 413 | 96.05% | 85 | 17.93% | 0.936 (0.663-1.320) | 0.706 |
Notes: OR, odds ratio; 95% CI, 95% confidence interval; the chi-square test was performed.
Relationship between a combination of SRD5A2 V89L/CYP17 -34T>C and clinical progression of BPH
The relationship between various combinations of SRD5A2 V89L/CYP17 -34T>C SNPs and the risk of clinical progression in BPH patients was anlyzed (Table 6). The distribution frequency of SRD5A2 V89L (VL + LL)/CYP17 -34T>C (TT) genotype combination was significantly lower in the progression group (14.4%) than that in the non-progression group (27.4%), indicating that SRD5A2 V89L (VL + LL)/CYP17 -34T>C (TT) genotype combination may be a protective factor during the clinical progression of BPH (OR = 0.429, 95% CI = 0.266-0.693, P < 0.001). The distribution frequency of genotype combination SRD5A2 V89L (VV)/CYP17 -34T>C (TC + CC) was significantly higher in the progression group (32.6%) than that in the non-progression group (18.6%), indicating that SRD5A2 V89L (VV)/CYP17 -34T>C (TC + CC) genotype combination may be a risk factor during the clinical progression of BPH (OR = 2.118, 95% CI = 1.371-3.629, P = 0.001). No significant correlation was found between the other genotype combinations and the clinical progression in BPH patients (all P > 0.05).
Table 6.
Relationship between a combination of SRD5A2 V89L/CYP17 -34T>C and clinical progression of BPH
| Combination of genotypes | Progression group (n = 215) | Non-progression group (n = 237) | OR | 95% CI | P |
|---|---|---|---|---|---|
| SRD5A2 V89L (VL + LL)/CYP17 -34T>C (TT) | 30 (14.0%) | 65 (27.4%) | 0.429 | 0.266-0.693 | < 0.001 |
| SRD5A2 V89L (VL + LL)/CYP17 -34T>C (TC + CC) | 24 (11.2%) | 23 (9.7%) | 1.169 | 0.639-2.140 | 0.612 |
| SRD5A2 V89L (VV)/CYP17 -34T>C (TC + CC) | 70 (32.6%) | 44 (18.6%) | 2.118 | 1.371-3.629 | 0.001 |
| SRD5A2 V89L (VV)/CYP17 -34T>C (TT) | 91 (42.3%) | 105 (44.3%) | 0.923 | 0.636-1.339 | 0.672 |
Notes: OR, odds ratio; 95% CI, 95% confidence interval; the chi-square test was performed.
Logistic regression analysis for risk factors for clinical progression of BPH
A logistic regression analysis was performed using BPH progression as a dependent variable. The following factors were included in the analysis as independent variables: varied factors in clinical data of subjects (such as BMI, TG, HDL-C, PV, PSA, Qmax and PVR), polymorphism of SRD5A2 V89L and CYP17 -34T>C, and the genotype combinations of SRD5A2 V89L (VL + LL)/CYP17 -34T>C (TT) and CYP17 -34T>C (TC + CC)/SRD5A2 V89L(VV). It was found that PSA, the VV genotype of SRD5A2 V89L, and the genotype combination of CYP17 -34T>C (TC + CC)/SRD5A2 V89L (VV) were all the risk factors for the clinical progression in BPH patients (all P < 0.05). TG, HDL-C, PV, Qmax, PVR and CYP17 -34T>C were all protective factors for the clinical progression in BPH patients (all P < 0.05) (Table 7).
Table 7.
Logistic regression analysis to determine the related risk factors for clinical progression in BPH patients
| Variables | B | Wald | Sig | Exp (B) | 95% CI |
|---|---|---|---|---|---|
| BMI | 0.236 | 2.065 | 0.151 | 1.267 | 0.918-1.749 |
| TG | -2.64 | 20.397 | < 0.001 | 0.071 | 0.023-0.224 |
| HDL-C | -8.94 | 18.445 | < 0.001 | 0.001 | 0.000-0.008 |
| PSA | 3.01 | 14.065 | < 0.001 | 20.289 | 4.208-97.822 |
| PV | 0.104 | 5.927 | 0.015 | 1.109 | 1.020-1.206 |
| Qmax | -0.47 | 5.151 | 0.013 | 0.626 | 0.431-0.906 |
| PVR | -0.05 | 3.975 | 0.046 | 0.955 | 0.913-0.999 |
| SRD5A2 V89L | 2.914 | 23.097 | < 0.001 | 18.438 | 5.617-60.521 |
| CYP17 -34T>C | -2.45 | 25.86 | < 0.001 | 0.087 | 0.034-0.222 |
| SRD5A2 V89L (VL + LL)/CYP17 -34T>C (TT) | -4.27 | 10.02 | 0.002 | 0.014 | 0.001-0.196 |
| SRD5A2 V89L (VV)/CYP17 -34T>C (TT) | 3.573 | 11.209 | 0.001 | 35.619 | 4.398-288.439 |
Notes: 95% CI, 95% confidence interval; BPH, benign prostatic hyperplasia; BMI, body mass index; TG, triglyceride; HDL-C, high-density lipoproteincholesterol; PV, prostate volume; PSA, prostate specific antigen concentration; Qmax, maximum flow rate; PVR, postvoiding residue; the logistic regression analysis was performed.
Discussion
BPH is a common urological problem among elderly men and is mainly perceived as a proliferative stromal disease featured with lower urinary tract and prostate enlargement symptoms [23]. Symptoms of BPH will not usually threat life, however, they often drastically affect life quality [9]. Previous studies have demonstrated that the gene polymorphisms were largely applied into the researches of BPH [24,25]. For further supplementation, this paper explored the association of BPH with SRD5A2/CYP17/CYP19/VDR gene polymorphisms in northern Chinese population.
It was found in this study that the BMI, HDL-C and Qmax were significantly lower, while the levels of TG, PV, PSA and PVR were significantly higher in the BPH group when compared with those the control group. Consistent with the result of our study, Mehmet Murat Baykam et al. have researched the correlation between prostatic resistive index and cardiovascular risk factors in BPH patients, in which HDL-C and Qmax were demonstrated to be risk factors of BPH [26]. And it is also reported that levels of BMI, TG were greater while HDL-C was lower in BPH patients [27]. Qmax and PVR were significantly correlated with BPH [28].
Furthermore, it was also found that in SRD5A2 V89L, compared with the VV genotype and V allele, LL genotype and L allele may reduce the risk of BPH and its clinical progression. As revealed in a previous study, SRD5A2 is the target of 5-reductase inhibitor which may affect the enzyme activity and lead to individual variability in the efficacy [29]. SRD5A2 can encode testosterone, a primary androgen in males, into dihydrotestosterone which is necessary for the prostatic growth and male external genitalia development [13]. Besides, L allele in the SRD5A2 is connected with reduced androstanediol glucuronide concentrations, and L/L genotype of V89L polymorphism was significantly related to lower concentrations of free testosterone and testosterone [30]. It was reported that when compared with the V/V genotype, the L/L genotype makes a 42% reduction in the activity of 5 a-reductase enzyme [31].
Another important finding from this study is that compared with the TT genotype and T allele at CYP17 -34T>C, CC genotype and C allele at CYP17 -34T>C may increase the risk of disease and its clinical progression. CYP17 is a key enzyme in the steroid biosynthesis which contains a heme prosthetic group at the active site and it can inhibit the treatment of prostatic diseases such as BPH [32,33]. CYP17 enzyme mediates two steps in steroid hormone biosynthesis, which are 17 a-hydroxylase and 17, 20-lyase activities; CYP17 contains a single-base pair (bp) T to C polymorphism which can create a Sp-1 site (CCACC box) at 34 bp upstream from the beginning of translation and downstream from the transcription site, which can provide an additional promoter activity with increased CYP17 transcription [24]. One study conducted by Tigli H et al. have demonstrated that in the risk allele, the T to C transition would create a new recognition site for restriction enzyme MspA1 [34]. All of these studies were in accordance with the result of this study.
This study also found that the SRD5A2 V89L and CYP17 -34T>C polymorphisms, CYP17 -34T>C (TC + CC)/SRD5A2 V89L (VV) combined genotypes were significantly related with the clinical progression of BHP. A relevant research has revealed that SRD5A2-V89L and CYP17-A1/A2 are involved in the biosynthesis of testosterone and dihydrotestosterone, V89L polymorphism of the SRD5A2 gene has been proved to be an important determinant of hypospadias risk, and CYP17-A1/A2 and SRD5A2-V89L are both involved in the biosynthesis of dihydrotestosterone and testosterone, which are in related with hypospadias [14].
In conclusion, these results have confirmed that SRD5A2/CYP17/CYP19/VDR gene polymorphisms are associated with the susceptibility and clinical progress of BPH. However, there are still some deficiencies in this study for various constraints. For examples, certain deviation in the statistical results may arise due to limited research time and sample size. Therefore, further studies are needed to verify our findings. Nevertheless, this study might shed some light on the future treatment of BPH.
Acknowledgements
This study was supported by Guiding Science and Technology Program of Daqing (szdy-2015-03).
Disclosure of conflict of interest
None.
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