Salvage treatments for prostate-specific antigen relapse of cT3N0M0 prostatic adenocarcinoma after radical prostatectomy combined with neoadjuvant androgen deprivation

Objective The aim of the study was to evaluate the efficiency of salvage treatments for prostate specific antigen(PSA)relapse of cT3N0M0 prostatic adenocarcinoma(PCa)after radical prostatectomy(RP)combined with neoadjuvant androgen deprivation(ADT).Methods A total of 332 patients with cT3N0M0 PCa were enrolled in the prospective study and received RP and pelvic lymph node dissection with neoadjuvant ADT for 3 months.All patients with PSA relapse were treated with salvage external beam radiation therapy(RT)and ADT for 6 months.Results The 5-year postoperative PSA relapse rate was 40.96%(136/332).The patients have been divided into the PSA relapse and PSA relapse-free groups in order to compare patient characteristics.The ratio of patients with Gleason score≥8 and positive surgical margin in the PSA relapse group were significantly higher than those of in the PSA relapse-free group(P=0.01).The mean duration between the start of operative treatment and PSA relapse was 31 months.Salvage treatment to all 136 PSA relapse patients led to favorable outcomes.PSA relapse was not observed after salvage treatment by the end of follow-up.The 5-year overall survival rates of the PSA relapse and PSA relapse-free groups were 94.9%and 93.9%,respectively.Conclusion In pursuit of curative treatment,our study showed that RP combined with neoadjuvant ADT is an aggressive multimodality strategy associated with lower PSA relapse and better survival outcomes for stage cT3N0M0 PCa patients.Patients with PSA relapse after RP may benefit from early aggressive salvage RT combined with short-term ADT.

Mass screening of prostate cancer in a Chinese population:the relationship between pathological features of prostate cancer and serum prostate specific antigen

Aim:To investigate the pathological features of the prostate biopsy through mass screening for prostate cancer in a Chinese cohort and their association with serum prostate specific antigen (PSA).Methods:A total of 12 027 Chinese men in Changchun were screened for prostate cancer by means of the serum total prostate specific antigen (tPSA) test (by Elisa assay).Transrectal ultrasound-guided systematic six-sextant biopsies were performed on those whose serum tPSA value was >4.0 ng/mL and those who had obstructive symptoms (despite their tPSA value) and were subject to subse- quent pathological analysis with the aid of the statistic software SPSS 10.0 (SPSS.Inc.,Chicago.USA).Results:Of the 12 027 cases,158 (including 137 patients whose serum tPSA values were >4.0 ng/mL and 21 patients [serum tPSA <4. 0 ng/mL] who had obstructive symptoms) undertook prostate biopsy.Of the 158 biopsies,41 cases of prostatic carci- noma were found (25.9 %,41/158).The moderately differentiated carcinoma and poorly differentiated carcinoma ac- counted for 61% and 34 %,respectively.A significant linear positive correlation between the serum tPSA and the Gleason scores in the 41 cases of prostatic carcinoma (r=0.312,P<0.01) was established.A significant linear positive correlation between the serum tPSA value of the 41 prostatic carcinoma and the positive counts of carcinoma in sextant biopsies was established (r=0.406,P<0.01),indicating a significant linear relationship between serum tPSA and the size of tumor. Condusion:This study was the first to conduct mass screening for prostate cancer by testing for serum tPSA values and the first to investigate the pathological features of prostate cancer in a cohort of Chinese men.Our results reveal that the moderately differentiated carcinoma is the most common type of prostate cancer.This study also has shown that the serum tPSA value in prostate cancer is associated with the Gleason score and the size of tumor.

Construction of Smac gene-containing and human prostate specific antigen promoter-regulated vector and its expression

To construct an eukaryotic expression vector containing Smac gene and study the expression efficiency and specificity of prostate specific antigen（PSA） enhancer/promoter in a possible targeted gene therapy scheme for prostate cancer. Methods： PSA enhancer （PSAE） and promoter （PSAP） sequences were amplified using PCR method. CMV and T7 promoters were deleted from pcDNA3.1-Smac and replaced by the two specific fragments to generate pPSAE-PSAP-Smac. After transfection into different cell lines, the status of cells was observed. And then, we determined the relative concentration of Smac mRNA in RT-PCR. Results： The recombinant plasmid of pPSAE-PSAP-Smac was successfully constructed. And only the prostate cancer cell line PC-3 was suppressed after transfection with pPSAE-PSAP-Smac. However, other nonprostate lines were not. Moreover, the concentration of Smac mRNA regulated by PSA promoter and enhancer was higher in comparison to the CMV promoter-driven control vectors. Conclusion： An expression vector containing the Smac gene （based on elements of the PSA gene regulatory sequences） has been developed and shown to function in prostate cancer cell lines which provides a solid platform for launching clinical studies.

Contemporary outcomes in the detection of prostate cancer using transrectal ultrasound-guided 12-core biopsy in Singaporean men with elevated prostate specific antigen and/or abnormal digital rectal examination

Objective:Despite being the third commonest cancer in Singaporean men,there is a dearth of basic data on the detection rate of prostate cancer and post-procedure complication rates locally using systematic 12-core biopsy.Our objective is to evaluate prostate cancer detection rates using 12-core prostate biopsy based on serum prostate specific antigen(PSA)levels and digital rectal examination(DRE)findings in Singaporean men presenting to a single tertiary centre.The secondary objective is to evaluate the complication rates of transrectal prostate biopsies.Methods:We retrospectively examined 804 men who underwent first transrectal-ultrasound(TRUS)guided 12-core prostate biopsies from January 2012 to April 2014.Prostate biopsies were performed on men presenting to a tertiary institution when their PSA levels were
4.0 ng/mL and/or when they had suspicious DRE findings.Results:Overall prostate cancer detection rate was 35.1%.Regardless of DRE findings,patients were divided into four subgroups based on their serum PSA levels:0e3.99 ng/mL,4.00 e9.99 ng/mL,10.00e19.99 ng/mL and
20.00 ng/mL and their detection rates were 9.5%,20.9%,38.4% and 72.3%,respectively.The detection rate of cancer based on suspicious DRE findings alone was 59.2% compared to 36.5% based on serum PSA cut-off of 4.0 ng/mL alone.The post-biopsy admission rate for sepsis was 1.5%.Conclusion:In conclusion,using contemporary 12-core biopsy methods,the local prostate cancer detection rate based on serum PSA and DRE findings has increased over the past decade presumably due to multiple genetic and environmental factors.Post-biopsy sepsis remains an important complication worldwide.

Early diagnosis of prostate cancer using free/total prostate specific antigen ratio: a population based screening data

Aim: To evaluate the use of free/total prostate specific antigen ratio (fPSA/tPSA ratio) in improving the early diagnosis of prostate cancer. Methods: The fPSA/tPSA ratio in the serum was analyzed in 187 men with tPSA ranging between 4.0 and 20.0 μg/L. All of them underwent ultrasound guided sextant prostatic biopsy. The results were calculated by SPSS 10.0 software. Results: (1) When the tPSA was within the ranges of 4.0 - 10.0 and 10.0 -20.0 μg/L, the prostate cancer detection rate was 18.1 % and 22.5 %, respectively; (2) The area under the curve (AUC) was bigger in fPSA/tPSA than in tPSA (P<0.05) in all the men; (3) When the cut off value of fPSA/tPSA ratio was set at 0.25 and the tPSA at 4.0 - 10.0 μg/L and 10.0 - 20.0 μg/L, the diagnostic sensitivity of tPSA was 90.5 % and 87.5 %, respectively. Thus at the tPSA ranges of 4.0 - 10.0 and 10.0 - 20.0 μg/L, 26.7 % and 11.3 % of biopsies could be avoided, respectively. Conclusion: The use of fPSA/tPSA ratio can improve the prostate cancer detection rate and reduce unnecessary biopsies when tPSA is within the range of 4.0 - 20.0 μg/L.

Prostate specific antigen bounce after intensity-modulated radiation therapy in an Asian population

Objective:Serum prostate specific antigen(PSA)is commonly used to evaluate treatment response after definitive radiation therapy(RT).However,PSA levels can temporarily rise without a clear reason,termed“PSA bounce”,and often engender great anxiety for both patients and physicians.The present study aimed to determine the prevalence and factors that predict“PSA bounce”after intensity-modulated radiation therapy(IMRT),and the relevance to biochemical failure and cancer recurrence in an Asian population.Methods:We retrospectively reviewed 206 patients who received IMRT for prostate cancer from 2004 to 2012 in the National Cancer Centre Singapore.These patients were followed up with regular PSA monitoring.We defined“PSA bounce”as a rise of 0.1 ng/mL,followed by two consecutive falls.Patients with biochemical failure(PSA nadir t 2 ng/mL)were further evaluated for cancer recurrence.Results:Sixty-one patients(29.6%)experienced“PSA bounce”,at a median time of 16 months and lasted for 12 months.Age remained the most consistent predictor of the incidence,duration and extent of“PSA bounce”.Other contributory factors included baseline PSA,Gleason score and PSA nadir.Hormonal therapy and prostate volume did not affect this phenomenon.Sixteen patients(7.8%)developed biochemical recurrence,at median time of 32 months,of which 11 were confirmed to have metastatic disease.The median follow-up time was 71 months.

Analysis of results related to the percent free prostate specific antigen among men without prostate diseases in Xi'an area

To measure the percent of free prostate specific antigen （fPSA） among men without prostate diseases in Xi＇an area, and to study the relationship of percent fPSA with age and pathological grade, clinical stage of prostate cancer （PCa） with percent fPSA, and to analyze the difference between the data in China and the.overseas data to determine appropriate reference range for Chinese male. Methods： A total of 713 participants were enrolled into the study, with PSA, fPSA in serum measured and the percent fPSA calculated. Out of 713 cases, 679 without prostate diseases were divided into 5 groups by age, and then the relationships of PSA, fPSA and percent fPSA with age were studied, respectively. The relationship of pathological grade and clinical stage with percent fPSA of the 34 participants with PCa was also studied. With the help of the related data of men without prostate disease, the appropriate reference range for Chinese male was established. Results： The increases in PSA or fPSA were correlated with age, while there was no significant correlation between age and percent fPSA. The percent fPSA was also correlated with pathological grade and clinical stage of PCa. The percent fPSA of men without prostate disease in Xi＇an area was significandy lower than that in the related overseas data. The reference range of percent fPSA for Chinese male was≥ 15%. Conclusion： Percent fPSA might be more useful than PSA in the detection of prostate cancer. As the percent fPSA is decreased, the pathological grade is decreased, and the clinical stage is increased, the malignant degree is increased. The reference range of ≥15% is more appropriate for Chinese male.

Combined Detection of Serum Heat Shock Protein-90<i>α</i>and Prostate Specific Antigen for Prostate Cancer Diagnosis

Objective: To explore the relationship between heat shock protein-90α (HSP-90α) and occurrence of prostate cancer, and clinical value of combined detection of serum HSP-90α and prostate specific antigen (PSA) in the diagnosis of prostate cancer. Method: A total of 30 patients with prostate cancer, 30 patients with benign prostatic hyperplasia (BPH) and 30 healthy men (control group) were selected from September 2018 to September 2019, then to detect levels of serum HSP-90α, total PSA and free PSA (FPSA) by ELISA, serum testosterone level by radioimmunoassay, prostate cancer tissue was removed by operation, and relative expression of tissue HSP-90α protein by Western blot. Results: The levels of serum HSP-90α and total PSA in prostate cancer group were significantly higher than other two groups, and testosterone level was lower than other two groups (P < 0.05);there was no difference of serum FPSA level between the three groups (P > 0.05). It was found by Pearson test that serum HSP-90α was positively correlated with total PSA level (r = 0.659, P = 0.005), while negatively correlated with testosterone level (r = -0.549, P = 0.006). According to TNM stage of prostate cancer, there were 17 cases of stage I - II, 13 cases of stage III - IV, 6 cases of Gleason score 1 - 4, 13 cases of 5 - 7, 11 cases of 8 - 10, tumor diameter range from 0.8 to 6.2 cm, with average of (3.9 ± 1.5) cm. The relative expression of HSP-90α protein in tumor tissue was closely related to TNM stage, Gleason score and tumor diameter (P < 0.05). By ROC analysis, it was found that accuracy of combined detection of serum HSP-90α and PSA levels for prostate cancer diagnosis was 0.896, and that of single PSA detection was 0.852. Conclusion: Higher expressions of HSP-90α in prostate cancer tissue and serum may be closely related to occurrence and development of prostate cancer, and combined detections of serum HSP-90α and PSA levels are of great significance in improving early diagnosis of prostate cancer.

Anti-tumor effects induced by gene vaccines co-expressing truncated human prostate specific membrane antigen gene and mouse 4-1BBL

Objective To investigate the influence of m4-1BBL on anti-tumor effects induced by truncated human prostate specific membrane antigen ( tPSMA ) gene in mice. Methods A eukaryotic expression plasmid encoding tPSMA and m4-1BBL ( pDC316-tPSMA-IRES m4-1BBL) ,pDC316-tPSMA and pDC316 were constructed.

Expression of IL-6 and TNF-α in benign prostatic hyperplasia combined with histological inflammation

Objective:To detect the expression of TNF-αand IL-6 in simple benign prostatic hyperplasia(BPH)and BPH combined with prostatitis,and to evaluate the effect of inflammation of the development and progression of BPH.Methods:Specimens of 90 patients with BPH underwent transurethral resection of prostate(TURP)were obtained.All speci-mens were divided histologically by HE staining into group of 35 cases of simple BPH(A group)and group of 55 cases of BPH combined with prostatitis(B group).Proteins levels of TNF-αand IL-6 were examined by immunohistochemistry.All patients’clinical indicators including PSA,prostate volume,age,urinary flow rate were recorded,and all patients were diagnosed as BPH by pathology.Results:All patients of B group were found combined with chronic prostatitis.The constituent ratio was 61%.Patients’prostate volume and PSA of B group were significant higher than A group(p<.05),but there were no significant differences in urine flow rate and patients’age(p>.05).As compared with A group,the expression of TNF-αand IL-6 significantly increased in B group(p<.05).The results of analysis were as follows:there existed a moderate correlation between prostate volume and patients’age(r=0.430,p<.001);there existed a moderate correlation between prostate volume and the degree of inflammation(r=0.610 and r=0.609,p<.01);there existed a moderate correlation between PSA and the degree of inflammation(r=0.572 and r=0.487,p<.01);there was not significant correlation between PSA and patients’age(r=0.065,p>.1);controlling influence of the age factor by partial correlation analysis,there was still a significant correlation between the prostate volume and level of inflammation.Conclusions:Most BPH patients are combined with prostatitis especially chronic inflammation.TNF-αand IL-6 are two-pro-inflammation cytokines,which obtain high expression in the prostate tissue combined with histological inflammation.Patients’prostate volume and PSA of BPH combined with prostatitis are significantly higher than those of simple BPH.However,there are no significant differences in urine flow rate and patients’age.These two cytokines may play an important part in promoting BPH and secretion of PSA.

From pro-prostate specific antigen, [-2]pro-prostate specific antigen to Beckman Coulter phi： the evolution of new biomarkers for early detection of prostatic carcinoma

Prostate specific antigen （PSA） has a wide clinical use for the early detection of prostatic carcinoma （PCa）; however, it has never been a perfect marker due to its low specificity and low positive predictive value which ranges between 4 ng/ml and 10 ng/ml. The discovery of different PSA molecular forms in serum in the early 1990s brought insight into searching for more specific markers. Since then free PSA （fPSA） has been used routinely to increase the specificity for PCa and to reduce unnecessary biopsies. More recently, promising data is emerging regarding one proenzyme molecular form of free PSA, proPSA, and a few truncated proPSA isoforms. The purpose of this article is to review the recent studies on clinical utility of proPSA, especially [-2]pPSA, an isoform of proPSA, and parameters involving [-2]pPSA as well as other PSA derivatives in early detection of PCa.

Inhibitory Effect of Matrine on the Expression of PSA and AR in Prostate Cancer Cell line LNCaP

In order to investigate the inhibitory effect of matrine on the expression of prostate specific antigen （PSA） and androgen receptor （AR） in prostate cancer cell line LNCaP in vitro, LNCaP cells were treated with matrine at different concentrations （0.5, 1.0, 1.5, 2.0 g/L） for 12-36 h. The growth activities of cancer cells were determined by MTT colorimetric assay. The AR level was measured by Western blotting. The expression of PSA was detected by using AXSYM system-chemical luciferase methods. The results showed that matrine could effectively inhibit the growth of androgen-dependent prostate cancer cell line LNCaP in vitro in a time-and dose-dependent manner （P〈0.05）. It could obviously decrease the level of AR （P〈0.01） and inhibit the expression of PSA in a dose-dependent manner （P〈0.05） in LNCaP cells. It was concluded that matrine could significantly suppress the growth of LNCaP cells and inhibit the expression of PSA and AR of prostate cancer cells.

Mass screening of 12027 elderly men for prostate carcinoma by measuring serum prostate specific antigen

Background The incidence of prostate carcinoma (Pca) has been increasing in China. We detected Pca in elderly men in Changchun,north China and the significance of prostate specific antigen (PSA) in mass screening and clinical staging of Pca. Methods Serum PSA from 12 027 men over 50 years old from Changchun was analyzed. In case of serum PSA greater than 4.0 ng/ml,the patient was suspected of potentially suffering from Pca,and transrectal six-point puncture prostate biopsies were performed under ultrasound guidance. Pathological examinations were performed on the biopsy tissue,and ABCD and TNM clinical stagings were used in accordance with international standards. Correlations between serum PSA level and clinical stage were analyzed. Results PSA was greater than 4.0 ng/ml in 813 patients (6.8% of the 12 027 men). Transrectal six-point prostate puncture biopsies guided by ultrasound were performed in 273 patients (33.6% of the 813 patients who were tested positive in the initial mass screening). Of these 273 patients,69 cases of Pca (25.3% of 273) were confirmed by biopsy in the second screening,with an overall detection rate for Pca of 0.57% (69/12 027). The total number of patients in stages A,B,T1,or T2 was 57.9%,and over 20% of them suffered from late stage Pca with lymph node and bone metastasis. An obvious positive correlation was observed between ABCD staging,TNM staging,and serum PSA level. Conclusions Serum PSA level is not only the golden standard for mass screening of Pca,but also the predictor for clinical stage of Pca. PSA testing revealed asymptomatic Pca cases in early,middle,and later stages in the elderly,suggesting that mass screening is of paramount importance.

Differentiation of lethal and non lethal prostate cancer： PSA and PSA isoforms and kinetics

Prostate-specific antigen （PSA） testing for the early diagnosis of prostate cancer has led to a decrease in cancer mortality. However, the high prevalence of low-grade prostate cancer and its long natural history, competing causes of death in older men and treatment patterns of prostate cancer, have led to dramatic overtreatment of the disease. Improved markers of prostate cancer lethality are needed to reduce the overtreatment of prostate cancer that leads to a reduced quality of life without extending life for a high proportion of men. The PSA level prior to treatment is routinely used in multivariable models to predict prostate cancer aggressiveness. PSA isoforms and PSA kinetics have been associated with more aggressive phenotypes, but are not routinely employed as part of prediction tools prior to treatment. PSA kinetics is a valuable marker of lethality post treatment and routinely used in determininE the need for salva=e theraov.

A phase 3,double-blind,randomized placebo-controlled efficacy and safety study of abiraterone acetate in chemotherapynaı¨ve patients with mCRPC in China,Malaysia,Thailand and Russia

Objective:This double-blind,placebo-controlled phase 3 study was designed to compare efficacy and safety of abiraterone acetate+prednisone(abiraterone)to prednisone alone in chemotherapy-naı¨ve,asymptomatic or mildly symptomatic metastatic castrationresistant prostate cancer(mCRPC)patients from China,Malaysia,Thailand and Russia.Methods:Adult chemotherapy-naı¨ve patients with confirmed prostate adenocarcinoma,Eastern Cooperative Oncology Group(ECOG)performance status(PS)grade 0e1,ongoing androgen deprivation(serum testosterone<50 ng/dL)with prostate specific antigen(PSA)or radiographic progression were randomized to receive abiraterone acetate(1000 mg,QD)t prednisone(5 mg,BID)or placebo t prednisone(5 mg,BID),until disease progression,unacceptable toxicity or consent withdrawal.Primary endpoint was improvements in time to PSA progression(TTPP).Results:Totally,313 patients were randomized(abiraterone:n Z 157;prednisone:n Z 156);and baseline characteristics were balanced.At clinical cut-off(median follow-up time:3.9 months),80% patients received treatment(abiraterone:n Z 138,prednisone:n Z 112).Median time to PSA progression was not reached with abiraterone versus 3.8 months for prednisone,attaining 58%reduction in PSA progression risk(HR=0.418;p<0.0001).Abirateronetreated patients had higher confirmed PSA response rate(50%vs.21%;relative odds=2.4;p<0.0001)and were 5 times more likely to achieve radiographic response than prednisonetreated patients(22.9%vs.4.8%,p=0.0369).Median survival was not reached.Most common(≥10% abiraterone vs.prednisone-treated)adverse events:bone pain(7%vs.14%),pain in extremity(6%vs.12%),arthralgia(10%vs.8%),back pain(7%vs.11%),and hypertension(15%vs.14%).Conclusion:Interim analysis confirmed favorable benefit-to-risk ratio of abiraterone in chemotherapy-naı¨ve men with mCRPC,consistent with global study,thus supporting use of abiraterone in this patient population.

An ultrasensitive aptasensor for prostate specific antigen assay based on Exonuclease T-aided cyclic cleavage

An ultrasensitive electrochemical aptasensor is presented for prostate specific antigen(PSA) detection. DNA tetrahedronaptamer is designed, which not only facilitates the molecular self-assembly events,but also improves the recognition efficiency between PSA and aptamer sequence on the electrode interface. The DNA conformation on top of DNA tetrahedron changes accordingly, which can be further digested by Exonuclease T(Exo T), a type of single-strand specific nuclease. Electrochemical species are removed synchronously and the initial PSA level can thus be determined. A linear range from 0.5 pg mL^(-1) to50 ng mL^(-1) is achieved with the limit of detection(LOD) as low as 0.15 pg mL^(-1). Moreover, this proposed method is highly selective and is successfully applied to determine PSA in human serum samples.

Combined tests of prostate specific antigen and testosterone will improve diagnosis and monitoring the progression of prostate cancer

Prostate-specific antigen （PSA） testing has been widely used to screen men for prostate cancer （PCa） and to monitor PCa progression. However, more studies have shown that around 15% of men with low or normal PSA levels have PCa. In this study, we aimed to investigate the relationship of androgen and PSA levels and to better understand the reason that some PCa patients have low serum PSA values. The in vitro data demonstrated that cultured LNCaP cells ceased to produce PSA after androgen withdrawal and resumed PSA production after androgen was re-added. The in vivo experiment results showed that 48% of PCa xenografts carrying mice have serum PSA level lower than 4 ng ml^-1. The serum PSA levels increased significantly with rises in testosterone （T） levels 1 week after T pellet implantation. These data indicated that the androgen is a key factor controlling the production of PSA. Low serum PSA levels in mice with PCa xenografts are associated with low serum T levels. Raising serum T levels in tumor caring mice will also significantly increase serum PSA level. This may have clinical implications when screening PSA in men, who have occult PCa.

Prostate chronic inflammation type Ⅳ and prostate cancer risk in patients undergoing first biopsy set:Results of a large cohort study

Objective:In prostate specimens,chronic inflammatory infiltrate(CII)type Ⅳ has been detected,but its association with prostate cancer(PCa)is controversial.The aim of the present study is to investigate on associations of CII with PCa detection in patients undergoing prostate first biopsy set.Methods:Ultrasound transrectal-guided biopsies by the transperineal approach were retrospectively evaluated in 441 consecutive patients.The study excluded patients who were in active surveillance,prostate specific antigen(PSA)
30 ng/mL,re-biopsies,incidental PCa after transurethral resection of the prostate(TURP),less than 14 cores or metastatic.Analysis of population and subpopulations(with or without PCa)was performed by statistical methods which included ManneWhitney(U test),KruskaleWallis test,Chi-squared statistic,logistic regression.Multivariate logistic regression models predicting mean probability of PCa detection were established.Results:PCa detection rate was 46.03%.Age,PSA,prostate volume(PV),prostate intraepithelial neoplasia(PIN)and CII were the significant independent predictors of PCa detection.PV(OR Z 0.934)and CII(OR Z 0.192)were both negative independent predictors.CII was a significant negative independent predictor in multivariate logistic regression models predicting the mean probability of PCa detection by age,PSA and PV.The inverse association of CII with PCa does not necessary mean protection because of PSA confounding.Conclusion:In a population of patients undergoing prostate first biopsy set,CII was a strong negative independent predictor of PCa detection.CII type Ⅳ should be considered as an adjunctive parameter in re-biopsy or active surveillance protocols.

Molecular mechanism of modulating the liquefaction epididymal protease inhibitor of human semen

Aim： To study the molecular mechanism of epididymal protease inhibitor （Eppin） modulating the process of prostate specific antigen （PSA） digesting semenogelin （Sg）. Methods： Human Sg cDNA （nucleotides 82-849） and Eppin cDNA （nucleotides 70-423） were generated by polymerase chain reaction （PCR） and cloned into pET-100D/TOPO. Recombinant Eppin and Sg （rEppin and rSg） were produced by BL21 （DE3）. The association of Eppin with Sg was studied by far-western immunoblot and radioautography. In vitro the digestion of rSg by PSA in the presence or absence of rEppin was studied. The effect of anti-Q20E （N-terminal） and C-terminal of Eppin on Eppin-Sg binding was monitored. Results： Eppin binds Sg on the surface of human spermatozoa with the C-terminal of Eppin （amino acids 75-133）. rSg was digested with PSA and many low molecular weight fragments were produced. When rEppin is bound to rSg, then digested by PSA, incomplete digestion and a 15-kDa fragment results. Antibody binding to the N-terminal of rEppin did not affect rSg digestion. Addition of antibodies to the C-terminal of rEppin inhibited the modulating effect of rEppin. Conclusion： Eppin protects a 15-kDa fragment of rSg from hydrolysis by PSA.

Molecular mechanism of epididymal protease inhibitor modulating the liquafication of human semen

To study the molecular mechanism of epididymal protease inhibitor （Eppin） modulating the liquafication of semen. Methods： Human semenogelin cDNA （nucleotides 82-849） and Eppin cDNA （nucleotides 70-423） were generated by PCR and cloned into pET-100D/TOPO.Recombinant Eppin and Sg were produced by BL21 （DE3）. The association of Eppin with Sg was studied by far-western and radioautography.In vitro the digestion of Sg by PSA in the presence or absence of recombinant Eppin was studied. The effect of anti-Q20E （N-terminal） and C-terminal of Eppin on Eppin-Sg binding was monitored. Results： Eppin binds Sg on the surface of human spermatozoa with C-terminal Eppin （aa75-133）.Recombinant Sg was digested with PSA ,many low molecular weight fragments were produced, when Eppin is bound to Sg ,then digested by PSA ,producing incomplete digestion and a 14.5-14.8 kDa fragmen. Antibody binding to the N-terminal of Eppin did not affect Sg digestion. Addition of antibodies to the C-terminal of Eppin inhibited the modulating effects of Eppin. Conclusion： Eppin modulates the digestion activity of PSA through binding Sg.The active site locates at C-terminal.