Quantitative Time-Resolved Fluorescence Imaging of Androgen Receptor and Prostate-Specific Antigen in Prostate Tissue Sections

Activins are multifunctional growth and differentiation factors and stimulate FSH-β gene expression and FSH secretion by the pituitary gonadotropes. Follistatins bind activin, resulting in the neutralization of activin bioactivity. The activin/follistatin system is present in the prostate tissue. Prostate-specific antigen (PSA) plays an important role in male reproductive physiology as well as being very important as a tumor marker for prostate cancer. Thus the regulation of PSA has important clinical implications. Previous studies showed that PSA is primarily regulated by androgens. In the present study, we evaluated the direct effects of activin A on the proliferation and PSA production of prostate cancer LNCaP cells, which express functional activin receptors and androgen receptor and PSA. LNCaP cells were treated with activin A and 5α-dihydrotestosterone (DHT) with or without their antagonists (follistatin or the nonsteroidal anti-androgen bicalutamide). Activin A decreased cell growth of LNCaP cells in a dose-dependent manner, whereas DHT increased it in a biphasic manner. In contrast to their opposing actions on cell growth, both activin A and DHT upregulated PSA gene expression and increased PSA secretion by LNCaP cells. The effects of activin A and DHT to increase PSA production were synergistic or additive. Follistatin or bicalutamide was without effect on cell growth or PSA production. The effects of activin A on LNCaP cells were blocked by follistatin, not by bicalutamide, whereas effects of DHT were prevented by bicalutamide, not by follistatin. Activin A upregulates PSA production, and the effect is through an androgen receptor-independent pathway. The activin/follistatin system can be a physiological modulator of PSA gene transcription and secretion in the prostate tissue, and activins may cooperate with androgen to upregulate PSA in vivo.

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Europium Nanoparticles and Time-resolved Fluorescence for Ultrasensitive Detection of Prostate-specific Antigen

Clinical Chemistry ◽

10.1093/clinchem/47.3.561 ◽

2001 ◽

Vol 47 (3) ◽

pp. 561-568 ◽

Cited By ~ 179

Author(s):

Harri Härmä ◽

Tero Soukka ◽

Timo Lövgren

Keyword(s):

Detection Limit ◽

Prostate Specific Antigen ◽

Specific Antigen ◽

High Specific Activity ◽

Microtiter Plate ◽

Detection Sensitivity ◽

Time Resolved Fluorescence ◽

Time Resolved ◽

Biochemical Assays ◽

Microtiter Plate Format

Abstract Background: Nanoparticle-based detection technologies have the potential to improve detection sensitivity in miniature as well as in conventional biochemical assays. We introduce a detection technology that relies on the use of europium(III) nanoparticles and time-resolved fluorometry to improve the detection limit of biochemical assays and to visualize individual molecules in a microtiter plate format. Methods: Streptavidin was covalently coated on 107-nm nanoparticles containing >30 000 europium molecules entrapped with β-diketones. In a model assay system, these nanoparticles were used to trace biotinylated prostate-specific antigen (PSA) in a microtiter plate format. Results: The detection limit (mean + 3 SD of the zero calibrator) of biotinylated PSA was 0.38 ng/L, corresponding to 10 fmol/L or 60 zeptomoles (60 × 10−21 moles) of PSA. Moreover, single nanoparticles, representing individual PSA molecules, were visualized in the same microtiter wells with a time-resolved fluorescence microscope using a ×10 objective. Single nanoparticles, possessing high specific activity, were also detected in solution by a standard time-resolved plate fluorometer. Conclusions: The universal streptavidin-coated europium(III) nanoparticle label is suitable for detection of any biotinylated molecule either in solution or on a solid phase. The europium(III) nanoparticle labeling technology is applicable to many areas of modern biochemical analysis, such as immunochemical and multianalyte DNA-chip assays as well as histo- and cytochemistry to improve detection sensitivities.

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