Steroidogenesis in peripheral and transition zones of human prostate cancer tissue

The peripheral zone (PZ) and transition zone (TZ) represent about 70% of the human prostate gland with each zone having differential ability to develop prostate cancer. Androgens and their receptor are the primary driving cause of prostate cancer growth and eventually castration-resistant prostate cancer (CRPC). De novo steroidogenesis has been identified as a key mechanism that develops during CRPC. Currently, there is very limited information available on human prostate tissue steroidogenesis. The purpose of the present study was to investigate steroid metabolism in human prostate cancer tissues with comparison between PZ and TZ. Human prostate cancer tumors were procured from the patients who underwent radical prostatectomy without any neoadjuvant therapy. Human prostate homogenates were used to quantify steroid levels intrinsically present in the tissues as well as formed after incubation with 2 µg/mL of 17-hydroxypregnenolone (17-OH-pregnenolone) or progesterone. A Waters Acquity ultraperformance liquid chromatography coupled to a Quattro Premier XE tandem quadrupole mass spectrometer using a C18 column was used to measure thirteen steroids from the classical and backdoor steroidogenesis pathways. The intrinsic prostate tissue steroid levels were similar between PZ and TZ with dehydroepiandrosterone (DHEA), dihydrotestosterone (DHT), pregnenolone and 17-OH-pregnenolone levels higher than the other steroids measured. Interestingly, 5-pregnan-3,20-dione, 5-pregnan-3-ol-20-one, and 5-pregnan-17-ol-3,20-dione formation was significantly higher in both the zones of prostate tissues, whereas, androstenedione, testosterone, DHT, and progesterone levels were significantly lower after 60 min incubation compared to the 0 min control incubations. The incubations with progesterone had a similar outcome with 5-pregnan-3,20-dione and 5-pregnan-3-ol-20-one levels were elevated and the levels of DHT were lower in both PZ and TZ tissues. The net changes in steroid formation after the incubation were more observable with 17-OH-pregnenolone than with progesterone. In our knowledge, this is the first report of comprehensive analyses of intrinsic prostate tissue steroids and precursor-driven steroid metabolism using a sensitive liquid chromatography-mass spectrometry assay. In summary, the PZ and TZ of human prostate exhibited similar steroidogenic ability with distinction in the manner each zone utilizes the steroid precursors to divert the activity towards backdoor pathway through a complex matrix of steroidogenic mechanisms

Application of electrostatic separation and differential scanning calorimetry for microplastic analysis in river sediments

A method with the potential for comprehensive microplastic monitoring in river sediments is presented in this study. We introduce a novel combination of electrostatic separation, density separation, and differential scanning calorimetry (DSC). Currently, microplastic analysis in sediments is limited in terms of sample masses, processing time, and analytical robustness. This work evaluated a method to process large sample masses efficiently and still obtain robust results. Four particulate matrices, including commercial sands and river sediments, were spiked with PCL, LD-PE, and PET microplastic particles (63–200 µm). Samples with a mass of 100 g and 1,000 g (sand only) contained 75 mg of each microplastic. After electrostatic separation, the mass of sand samples was reduced by 98%. Sediment samples showed a mass reduction of 70–78%. After density separation, the total mass reduction of sediment samples was above 99%. The increased concentration of total organic carbon seems to have the highest impact on mass reduction by electrostatic separation. Nevertheless, the recovery of microplastic was independent of the particulate matrix and was polymer-specific. In 100 g samples, the average recovery rates for PCL, LD-PE, and PET were 74 ± 9%, 93 ± 9%, and 120 ± 18%, respectively. The recoveries of microplastic from 1,000 g samples were 50 ± 8%, 114 ± 9%, and 82 ± 11%, respectively. In scale up experiments, high recoveries of all microplastics were observed with a decrease in standard deviation. Moreover, the biodegradable polymer PCL could be used as an internal standard to provide quality assurance of the process. This method can overcome the current limitations of routine microplastic analysis in particulate matrices. We conclude that this method can be applied for comprehensive microplastic monitoring in highly polluted sediments. More studies on electrostatic separation and polymer-specific recovery rates in complex matrices are proposed

Application of electrostatic separation and differential scanning calorimetry for microplastic analysis in river sediments

A method with the potential for comprehensive microplastic monitoring in river sediments is presented in this study. We introduce a novel combination of electrostatic separation, density separation, and differential scanning calorimetry (DSC). Currently, microplastic analysis in sediments is limited in terms of sample masses, processing time, and analytical robustness. This work evaluated a method to process large sample masses efficiently and still obtain robust results. Four particulate matrices, including commercial sands and river sediments, were spiked with PCL, LD-PE, and PET microplastic particles (63–200 µm). Samples with a mass of 100 g and 1,000 g (sand only) contained 75 mg of each microplastic. After electrostatic separation, the mass of sand samples was reduced by 98%. Sediment samples showed a mass reduction of 70–78%. After density separation, the total mass reduction of sediment samples was above 99%. The increased concentration of total organic carbon seems to have the highest impact on mass reduction by electrostatic separation. Nevertheless, the recovery of microplastic was independent of the particulate matrix and was polymer-specific. In 100 g samples, the average recovery rates for PCL, LD-PE, and PET were 74 ± 9%, 93 ± 9%, and 120 ± 18%, respectively. The recoveries of microplastic from 1,000 g samples were 50 ± 8%, 114 ± 9%, and 82 ± 11%, respectively. In scale up experiments, high recoveries of all microplastics were observed with a decrease in standard deviation. Moreover, the biodegradable polymer PCL could be used as an internal standard to provide quality assurance of the process. This method can overcome the current limitations of routine microplastic analysis in particulate matrices. We conclude that this method can be applied for comprehensive microplastic monitoring in highly polluted sediments. More studies on electrostatic separation and polymer-specific recovery rates in complex matrices are proposed

Application of electrostatic separation and differential scanning calorimetry for microplastic analysis in river sediments

A method with the potential for comprehensive microplastic monitoring in river sediments is presented in this study. We introduce a novel combination of electrostatic separation, density separation, and differential scanning calorimetry (DSC). Currently, microplastic analysis in sediments is limited in terms of sample masses, processing time, and analytical robustness. This work evaluated a method to process large sample masses efficiently and still obtain robust results. Four particulate matrices, including commercial sands and river sediments, were spiked with PCL, LD-PE, and PET microplastic particles (63–200 µm). Samples with a mass of 100 g and 1,000 g (sand only) contained 75 mg of each microplastic. After electrostatic separation, the mass of sand samples was reduced by 98%. Sediment samples showed a mass reduction of 70–78%. After density separation, the total mass reduction of sediment samples was above 99%. The increased concentration of total organic carbon seems to have the highest impact on mass reduction by electrostatic separation. Nevertheless, the recovery of microplastic was independent of the particulate matrix and was polymer-specific. In 100 g samples, the average recovery rates for PCL, LD-PE, and PET were 74 ± 9%, 93 ± 9%, and 120 ± 18%, respectively. The recoveries of microplastic from 1,000 g samples were 50 ± 8%, 114 ± 9%, and 82 ± 11%, respectively. In scale up experiments, high recoveries of all microplastics were observed with a decrease in standard deviation. Moreover, the biodegradable polymer PCL could be used as an internal standard to provide quality assurance of the process. This method can overcome the current limitations of routine microplastic analysis in particulate matrices. We conclude that this method can be applied for comprehensive microplastic monitoring in highly polluted sediments. More studies on electrostatic separation and polymer-specific recovery rates in complex matrices are proposed

Steroidogenesis in Peripheral and Transition Zones of Human Prostate Cancer Tissue

The peripheral zone (PZ) and transition zone (TZ) represent about 70% of the human prostate gland with each zone having differential ability to develop prostate cancer. Androgens and their receptor are the primary driving cause of prostate cancer growth and eventually castration-resistant prostate cancer (CRPC). De novo steroidogenesis has been identified as a key mechanism that develops during CRPC. Currently, there is very limited information available on human prostate tissue steroidogenesis. The purpose of the present study was to investigate steroid metabolism in human prostate cancer tissues with comparison between PZ and TZ. Human prostate cancer tumors were procured from the patients who underwent radical prostatectomy without any neoadjuvant therapy. Human prostate homogenates were used to quantify steroid levels intrinsically present in the tissues as well as formed after incubation with 2 µg/mL of 17-hydroxypregnenolone (17-OH-pregnenolone) or progesterone. A Waters Acquity ultraperformance liquid chromatography coupled to a Quattro Premier XE tandem quadrupole mass spectrometer using a C18 column was used to measure thirteen steroids from the classical and backdoor steroidogenesis pathways. The intrinsic prostate tissue steroid levels were similar between PZ and TZ with dehydroepiandrosterone (DHEA), dihydrotestosterone (DHT), pregnenolone and 17-OH-pregnenolone levels higher than the other steroids measured. Interestingly, 5-pregnan-3,20-dione, 5-pregnan-3-ol-20-one, and 5-pregnan-17-ol-3,20-dione formation was significantly higher in both the zones of prostate tissues, whereas, androstenedione, testosterone, DHT, and progesterone levels were significantly lower after 60 min incubation compared to the 0 min control incubations. The incubations with progesterone had a similar outcome with 5-pregnan-3,20-dione and 5-pregnan-3-ol-20-one levels were elevated and the levels of DHT were lower in both PZ and TZ tissues. The net changes in steroid formation after the incubation were more observable with 17-OH-pregnenolone than with progesterone. In our knowledge, this is the first report of comprehensive analyses of intrinsic prostate tissue steroids and precursor-driven steroid metabolism using a sensitive liquid chromatography-mass spectrometry assay. In summary, the PZ and TZ of human prostate exhibited similar steroidogenic ability with distinction in the manner each zone utilizes the steroid precursors to divert the activity towards backdoor pathway through a complex matrix of steroidogenic mechanisms.AlumniNon UBCMedicine, Faculty ofUrologic Sciences, Department ofReviewedFacult

Androgen levels increase by intratumoral de novo steroidogenesis during progression of castration-resistant prostate cancer

Although systemic androgen deprivation prolongs life in advanced prostate cancer, remissions are temporary because patients almost uniformly progress to a state of a castration-resistant prostate cancer (CRPC) as indicated by recurring PSA. This complex process of progression does not seem to be stochastic as the timing and phenotype are highly predictable, including the observation that most androgen-regulated genes are reactivated despite castrate levels of serum androgens. Recent evidence indicates that intraprostatic levels of androgens remain moderately high following systemic androgen deprivation therapy, whereas the androgen receptor (AR) remains functional, and silencing the AR expression following castration suppresses tumor growth and blocks the expression of genes known to be regulated by androgens. From these observations, we hypothesized that CRPC progression is not independent of androgen-driven activity and that androgens may be synthesized de novo in CRPC tumors leading to AR activation. Using the LNCaP xenograft model, we showed that tumor androgens increase during CRPC progression in correlation to PSA up-regulation. We show here that all enzymes necessary for androgen synthesis are expressed in prostate cancer tumors and some seem to be up-regulated during CRPC progression. Using an ex vivo radiotracing assays coupled to high-performance liquid chromatography-radiometric/mass spectrometry detection, we show that tumor explants isolated from CRPC progression are capable of de novo conversion of [(14)C]acetic acid to dihydrotestosterone and uptake of [(3)H]progesterone allows detection of the production of six other steroids upstream of dihydrotestosterone. This evidence suggests that de novo androgen synthesis may be a driving mechanism leading to CRPC progression following castration

Insulin increases De Novo Steroidogenesis in prostate cancer cells

Androgen-dependent pathways regulate maintenance and growth of normal and malignant prostate tissues. Androgen deprivation therapy (ADT) exploits this dependence and is used to treat metastatic prostate cancer; however, regression initially seen with ADT gives way to development of incurable castration-resistant prostate cancer (CRPC). Although ADT generates a therapeutic response, it is also associated with a pattern of metabolic alterations consistent with metabolic syndrome including elevated circulating insulin. Because CRPC cells are capable of synthesizing androgens de novo, we hypothesized that insulin may also influence steroidogenesis in CRPC. In this study, we examined this hypothesis by evaluating the effect of insulin on steroid synthesis in prostate cancer cell lines. Treatment with 10 nmol/L insulin increased mRNA and protein expression of steroidogenesis enzymes and upregulated the insulin receptor substrate insulin receptor substrate 2 (IRS-2). Similarly, insulin treatment upregulated intracellular testosterone levels and secreted androgens, with the concentrations of steroids observed similar to the levels reported in prostate cancer patients. With similar potency to dihydrotestosterone, insulin treatment resulted in increased mRNA expression of prostate-specific antigen. CRPC progression also correlated with increased expression of IRS-2 and insulin receptor in vivo. Taken together, our findings support the hypothesis that the elevated insulin levels associated with therapeutic castration may exacerbate progression of prostate cancer to incurable CRPC in part by enhancing steroidogenesis