Fertility of male adult rats submitted to forced swimming stress

We investigated whether stress interferes with fertility during adulthood. Male Wistar rats (weighing 220 g in the beginning of the experiment) were forced to swim for 3 min in water at 32ºC daily for 15 days. Stress was assessed by the hot-plate test after the last stressing session. To assess fertility, control and stressed males (N = 15 per group) were mated with sexually mature normal females. Males were sacrificed after copulation. Stress caused by forced swimming was demonstrated by a significant increase in the latency of the pain response in the hot-plate test (14.6 ± 1.25 s for control males vs 26.0 ± 1.53 s for stressed males, P = 0.0004). No changes were observed in body weight, testicular weight, seminal vesicle weight, ventral prostate weight or gross histological features of the testes of stressed males. Similarly, no changes were observed in fertility rate, measured by counting live fetuses in the uterus of normal females mated with control and stressed males; no dead or incompletely developed fetuses were observed in the uterus of either group. In contrast, there was a statistically significant decrease in spermatid production demonstrated by histometric evaluation (154.96 ± 5.41 vs 127.02 ± 3.95 spermatids per tubular section for control and stressed rats, respectively, P = 0.001). These data demonstrate that 15 days of forced swimming stress applied to adult male rats did not impair fertility, but significantly decreased spermatid production. This suggests that the effect of stress on fertility should not be assessed before at least the time required for one cycle of spermatogenesis.

Desquamation Is A Novel Phenomenon For Collective Prostate Epithelial Cell Deletion After Castration.

The mechanism underlying castration-induced prostate regression, which is a classical physiological concept translated into the therapeutic treatment of advanced prostate cancer, involves epithelial cell apoptosis. In searching for events and mechanisms contributing to prostate regression in response to androgen modulation, we have frequently observed the collective deletion of epithelial cells. This work was undertaken to characterize this phenomenon hereafter named desquamation and to verify its presence after 17β-estradiol (E2) administration. Electron microscopy revealed that the desquamating cells had preserved cell-cell junctions and collapsed nuclear contents. The TUNEL reaction was negative for these cells, which were also negative for cleaved caspases-8, -9, -3 and nuclear apoptosis-inducing factor. Detailed analyses revealed that the condensed chromatin was first affected detaching from the nuclear lamina, which was observable after lamin A immunohistochemistry, suggesting the lack of lamin A degradation. A search in animals treated with supraphysiological E2 employed as an alternative anti-androgen treatment revealed no desquamation. The combined treatment (Cas + E2 group) caused changes particular to each treatment, including desquamation. In conclusion, desquamation appeared as a novel phenomenon contributing to collective prostate epithelial cell deletion, distinct from the classical castration-induced apoptosis and particular to the androgen deprivation resulting from surgical castration, and should be considered as part of the mechanisms promoting organ regression.

Proteomic analysis of normal and malignant prostate tissue to identify novel proteins lost in cancer

BACKGROUND. Alterations of important protein pathways, including loss of prostate secretory granules, and disruption of the prostatic secretory pathway have been identified as early events in malignancy. In this study, proteomics was used to map the differences in protein expression between normal and malignant prostate tissues and to identify and analyze differentially expressed proteins in human prostate tissue with particular regard to the proteins lost in malignancy. METHODS. Small quantities of normal and malignant prostate tissue were taken fresh from 34 radical prostatectomy cases. After histological examination, proteins were solubilized from selected tissues and separated using two-dimensional electrophoresis. Using image analysis, the proteome of normal and malignant tissues were mapped and differentially expressed proteins (present in normal and absent in malignant tissue) were identified and subsequently analyzed using peptide mass finger printing and N-terminal sequencing. Western blotting and immunohistochemistry were performed to examine expression profiles and tissue localization of candidate proteins. RESULTS. Comparison of protein maps of normal and malignant prostate were used to identify 20 proteins which were lost in malignant transformation, including prostate specific antigen (PSA), alpha-l antichymotrypsin (ACT), haptoglobin, and lactoylglutathione lyase. Three of the 20 had not previously been reported in human prostate tissue (Ubiquitin-like NEDD8, calponin, and a follistatin-related protein). Western blotting confirmed differences in the expression profiles of NEDD8 and calponin, and immunohistochemistry demonstrated differences in the cellular localization of these two proteins in normal and malignant prostate glands. CONCLUSIONS. The expression of NEDD8, calponin, and the follistatin-related protein in normal prostate tissues is a novel finding and the role of these important functional proteins in normal prostate and their loss or reduced expression in prostate malignancy warrants further investigations. (C) 2002 Wiley-Liss, Inc.

Effects of medical therapy, alcohol, smoking, and endocrine disruptors on male infertility

Infertility affects up to 15% of the sexually active population, and in 50% of cases, a male factor is involved, either as a primary problem or in combination with a problem in the female partner. Because many commonly encountered drugs and medications can have a detrimental effect on male fertility, the medical evaluation should include a discussion regarding the use of recreational and illicit drugs, medications, and other substances that may impair fertility. With the knowledge of which drugs and medications may be detrimental to fertility, it may be possible to modify medication regimens or convince a patient to modify habits to decrease adverse effects on fertility and improve the chances of achieving a successful pregnancy. Concern is growing that male sexual development and reproduction have changed for the worse over the past 30 to 50 years. Although some reports find no changes, others suggest that sperm counts appear to be decreasing and that the incidence of developmental abnormalities such as hypospadias and cryptorchidism appears to be increasing, as is the incidence of testicular cancer. These concerns center around the possibility that our environment is contaminated with chemicals - both natural and synthetic - that can interact with the endocrine system.

Is hospital discharge administrative data an appropriate source of information for cancer registries purposes? Some insights from four Spanish registries

Background. The use of hospital discharge administrative data (HDAD) has been recommended for automating, improving, even substituting, population-based cancer registries. The frequency of false positive and false negative cases recommends local validation. Methods. The aim of this study was to detect newly diagnosed, false positive and false negative cases of cancer from hospital discharge claims, using four Spanish population-based cancer registries as the gold standard. Prostate cancer was used as a case study. Results. A total of 2286 incident cases of prostate cancer registered in 2000 were used for validation. In the most sensitive algorithm (that using five diagnostic codes), estimates for Sensitivity ranged from 14.5% (CI95% 10.3-19.6) to 45.7% (CI95% 41.4-50.1). In the most predictive algorithm (that using five diagnostic and five surgical codes) Positive Predictive Value estimates ranged from 55.9% (CI95% 42.4-68.8) to 74.3% (CI95% 67.0-80.6). The most frequent reason for false positive cases was the number of prevalent cases inadequately considered as newly diagnosed cancers, ranging from 61.1% to 82.3% of false positive cases. The most frequent reason for false negative cases was related to the number of cases not attended in hospital settings. In this case, figures ranged from 34.4% to 69.7% of false negative cases, in the most predictive algorithm. Conclusions. HDAD might be a helpful tool for cancer registries to reach their goals. The findings suggest that, for automating cancer registries, algorithms combining diagnoses and procedures are the best option. However, for cancer surveillance purposes, in those cancers like prostate cancer in which care is not only hospital-based, combining inpatient and outpatient information will be required.

Testostérone et prostate [Testosterone and prostate].

Age related testosterone deficiency syndrome may occur with other diseases of the elderly men, as prostate diseases. The relationship between testosterone and prostate has been widely studied the last 10 years, with the increased use of testosterone replacement therapy. The traditional belief that testosterone administration causes prostate cancer growth has been challenged by recent studies. To date, nothing has been found to support the evidence that restoring testosterone levels within physiological range increases the incidence of prostate cancer in hypogonadic patients. In these patients, testosterone replacement therapy does not seem to worsen lower urinary tract symptoms.

Substrate specificity of human kallikrein 2 (hK2) as determined by phage display technology.

Human glandular kallikrein 2 (hK2) is a trypsin-like serine protease expressed predominantly in the prostate epithelium. Recently, hK2 has proven to be a useful marker that can be used in combination with prostate specific antigen for screening and diagnosis of prostate cancer. The cleavage by hK2 of certain substrates in the proteolytic cascade suggest that the kallikrein may be involved in prostate cancer development; however, there has been very little other progress toward its biochemical characterization or elucidation of its true physiological role. In the present work, we adapt phage substrate technology to study the substrate specificity of hK2. A phage-displayed random pentapeptide library with exhaustive diversity was generated and then screened with purified hK2. Phages displaying peptides susceptible to hK2 cleavage were amplified in eight rounds of selection and genes encoding substrates were transferred from the phage to a fluorescent system using cyan fluorescent protein (derived from green fluorescent protein) that enables rapid determination of specificity constants. This study shows that hK2 has a strict preference for Arg in the P1 position, which is further enhanced by a Ser in P'1 position. The scissile bonds identified by phage display substrate selection correspond to those of the natural biological substrates of hK2, which include protein C inhibitor, semenogelins, and fibronectin. Moreover, three new putative hK2 protein substrates, shown elsewhere to be involved in the biology of the cancer, have been identified thus reinforcing the importance of hK2 in prostate cancer development.

Continuing declines in cancer mortality in the European Union

BACKGROUND: From 1988 to 1997 age-standardised total cancer mortality rates in the European Union (EU) fell by around 9% in both sexes. Available cancer mortality data in Europe up to 2002 allow a first check of the forecast of further declines in cancer mortality. PATIENTS AND METHODS: We considered trends in age-standardised mortality from major cancer sites in the EU during the period 1980-2002. RESULTS: For men, total cancer mortality, after a peak of 191.1/100,000 in 1987 declined to 177.8 in 1997 (-7%), and to 166.5 in 2002. Corresponding figures for females were 107.9/100,000, 100.5 and 95.2, corresponding to falls of 7% from 1987 to 1997, and to 5% from 1997 to 2002. Over the last 5 years, lung cancer declined by 1.9% per year in men, to reach 44.4/100,000, but increased by 1.7% in women, to reach 11.4. In 2002, for the first year, lung cancer mortality in women was higher than that for intestinal cancer (11.1/100,000), and lung cancer became the second site of cancer deaths in women after breast (17.9/100,000). From 1997 to 2002, appreciable declines were observed in mortality from intestinal cancer in men (-1.6% per year, to reach 18.8/100,000), and in women (-2.5%), as well as for breast (-1.7% per year) and prostate cancer (-1.4%). CONCLUSIONS: Despite the persisting rises in female lung cancer, the recent trends in cancer mortality in the EU are encouraging and indicate that an 11% reduction in total cancer mortality from 2000 to 2015 is realistic and possible.

Foxm1 expression in prostate epithelial cells is essential for prostate carcinogenesis.

The treatment of advanced prostate cancer (PCa) remains a challenge. Identification of new molecular mechanisms that regulate PCa initiation and progression would provide targets for the development of new cancer treatments. The Foxm1 transcription factor is highly up-regulated in tumor cells, inflammatory cells, and cells of tumor microenvironment. However, its functions in different cell populations of PCa lesions are unknown. To determine the role of Foxm1 in tumor cells during PCa development, we generated two novel transgenic mouse models, one exhibiting Foxm1 gain-of-function and one exhibiting Foxm1 loss-of-function under control of the prostate epithelial-specific Probasin promoter. In the transgenic adenocarcinoma mouse prostate (TRAMP) model of PCa that uses SV40 large T antigen to induce PCa, loss of Foxm1 decreased tumor growth and metastasis. Decreased prostate tumorigenesis was associated with a decrease in tumor cell proliferation and the down-regulation of genes critical for cell proliferation and tumor metastasis, including Cdc25b, Cyclin B1, Plk-1, Lox, and Versican. In addition, tumor-associated angiogenesis was decreased, coinciding with reduced Vegf-A expression. The mRNA and protein levels of 11β-Hsd2, an enzyme playing an important role in tumor cell proliferation, were down-regulated in Foxm1-deficient PCa tumors in vivo and in Foxm1-depleted TRAMP C2 cells in vitro. Foxm1 bound to, and increased transcriptional activity of, the mouse 11β-Hsd2 promoter through the -892/-879 region, indicating that 11β-Hsd2 was a direct transcriptional target of Foxm1. Without TRAMP, overexpression of Foxm1 either alone or in combination with inhibition of a p19(ARF) tumor suppressor caused a robust epithelial hyperplasia, but was insufficient to induce progression from hyperplasia to PCa. Foxm1 expression in prostate epithelial cells is critical for prostate carcinogenesis, suggesting that inhibition of Foxm1 is a promising therapeutic approach for prostate cancer chemotherapy.

The use of telomerase activity for the detection of prostatic cancer cells after prostatic massage.

PURPOSE: Prostate cancer is the most commonly diagnosed cancer in the United States. The diagnosis or followup of prostate cancer in men older than 50 years is based on digital rectal examination, measurement of the free-to-total prostatic specific antigen ratio and transrectal ultrasound assisted needle biopsy of the prostate. We developed and evaluated a noninvasive method for diagnosing prostate cancer based on the measurement of telomerase activity after prostatic massage in fresh voided urine or after urethral washing. MATERIALS AND METHODS: We obtained 36 specimens of cells after prostatic massage in the fresh voided urine of 16 patients who subsequently underwent radical prostatectomy and after urethral washing in 20 who underwent prostate needle biopsies. Ethylenediaminetetraacetic acid was immediately added to the collected urine or washing to a final concentration of 20 mM. After protein extraction by CHAPS buffer each specimen was tested for telomerase activity in a 2-step modified telomeric repeat amplification protocol assay. The 2 prostate cancer cell lines PC-3 and LNCaP with high telomerase activity were used as a positive control. RESULTS: Telomerase activity was detected in 14 of 24 samples with known prostate cancer (sensitivity 58%). In contrast, no telomerase activity was found in the 12 cases without histological evidence of prostate tumor (specificity 100%). Eight of 9 poorly differentiated cancers expressed telomerase activity (89%), while only 6 of 15 well and moderately differentiated cancers showed telomerase activity (40%). CONCLUSIONS: Our data illustrate that telomerase activity may be detected in voided urine or washing after prostatic massage in patients with prostate cancer. Sensitivity was higher for poorly differentiated tumors. This approach is not currently available for detecting prostate cancer in clinical practice. However, these results are promising and further studies are ongoing.

The Effects of Fibroblast Growth Factor 8b on Reproductive Organs and Prostate Tumorigenesis

Fibroblast growth factors (FGFs) are involved in the development and homeostasis of the prostate and other reproductive organs. FGF signaling is altered in prostate cancer. Fibroblast growth factor 8 (FGF8) is a mitogenic growth factor and its expression is elevated in prostate cancer and in premalignant prostatic intraepithelial neoplasia (PIN) lesions. FGF8b is the most transforming isoform of FGF8. Experimental models show that FGF8b promotes several phases of prostate tumorigenesis - including cancer initiation, tumor growth, angiogenesis, invasion and development of bone metastasis. The mechanisms activated by FGF8b in the prostate are unclear. In the present study, to examine the tumorigenic effects of FGF8b on the prostate and other FGF8b expressing organs, an FGF8b transgenic (TG) mouse model was generated. The effect of estrogen receptor beta (ERβ) deficiency on FGF8binduced prostate tumorigenesis was studied by breeding FGF8b-TG mice with ERβ knockout mice (BERKOFVB). Overexpression of FGF8b caused progressive histological and morphological changes in the prostate, epididymis and testis of FGF8b-TG-mice. In the prostate, hyperplastic, preneoplastic and neoplastic changes, including mouse PIN (mPIN) lesions, adenocarcinomas, sarcomas and carcinosarcomas were present in the epithelium and stroma. In the epididymis, a highly cancer-resistant tissue, the epithelium contained dysplasias and the stroma had neoplasias and hyperplasias with atypical cells. Besides similar histological changes in the prostate and epididymis, overexpression of FGF8b induced similar changes in the expression of genes such as osteopontin (Spp1), connective tissue growth factor (Ctgf) and FGF receptors (Fgfrs) in these two tissues. In the testes of the FGF8b-TG mice, the seminiferous epithelium was frequently degenerative and the number of spermatids was decreased. A portion of the FGF8b-TG male mice was infertile. Deficiency of ERβ did not accelerate prostate tumorigenesis in the FGF8b-TG mice, but increased significantly the frequency of mucinous metaplasia and slightly the frequency of inflammation in the prostate. This suggests putative differentiation promoting and anti-inflammatory roles for ERβ. In summary, these results underscore the importance of FGF signaling in male reproductive organs and provide novel evidence for a role of FGF8b in stromal activation and prostate tumorigenesis.

Sexual behavior and fertility of male rats submitted to prolonged immobilization-induced stress

In order to investigate whether prolonged stress interferes with the onset of sexual behavior at puberty and with fertility at adulthood, prepubertal male Wistar rats (40 days of age) were immobilized 6 h a day for 15 days (up to early puberty) or for 60 days (until sexual maturity). Pubertal stressed rats showed a two-fold increase in the latency for the first mount (probably due to repeated aversive experience in which a change of environment was always followed by immobilization) and a 2.5-fold increase in the frequency of thrusting (indicative of enhanced sexual performance). The apparently stimulatory effect of prolonged stress on the onset of sexual behavior is discussed in terms of increased testosterone level and interference with the complex interchanges between the neurotransmitters/neuropeptides involved in the central control of male sexual activity. Adult stressed animals were mated with normal females, which became pregnant but exhibited a more than two-fold increase in both pre-implantation and post-implantation loss, probably due to a smaller rate of fertilization and/or fertilization with damaged spermatozoa.

Rôle de GPR40 dans la survie et la prolifération cellulaires induites par l’oléate dans les cellules de cancer du sein MDA-MB-231 et de cancer de la prostate DU145

La relation entre l’obésité et le cancer, bien qu’établie par des études épidémiologiques, est peu connue. Pourtant, environ 25 % des cancers pourraient y être attribuables. Parmi les cancers reliés à l’obésité, les cancers du côlon, du sein chez les femmes ménopausées et de la prostate sont les plus fréquents. Des études sur modèles animaux ont suggéré une association positive entre une diète riche en gras et le développement du cancer mammaire et de la prostate. Nous avons étudié les mécanismes moléculaires par lesquels les acides gras influencent le devenir de lignées de cellules cancéreuses du sein et de la prostate. Ces travaux ont montré que les acides gras insaturés, dont l’oléate, induisent la prolifération cellulaire tandis que les acides gras saturés, dont le palmitate, diminuent la prolifération. Un traitement à l’oléate stimule la formation de gouttelettes lipidiques dans le cytoplasme des cellules de cancer du sein MDA-MB-231 et de la prostate DU145 alors qu’un traitement au palmitate entraîne l’apoptose. Le mécanisme d’action de l’oléate sur la prolifération a été étudié de façon plus approfondie. L’utilisation d’inhibiteurs pharmacologiques nous a permis de déterminer que l’effet prolifératif de l’oléate implique la voie PI3K/Akt, la voie ERK1/2 et l’activation d’un ou de plusieurs récepteur(s) couplé(s) aux protéines G (GPCR). L’oléate induit la phosphorylation rapide des protéines Akt et ERK1/2 dans les cellules de cancer du sein MDA-MB-231 et de la prostate DU145. Au cours des dernières années, deux GPCRs ont été identifiés comme étant activables par des acides gras à moyennes et à longues chaînes, GPR40 et GPR120. GPR40 étant exprimé dans plusieurs lignées cellulaires de cancer du sein et de la prostate contrairement à l’expression de GPR120 qui était inexistante dans la plupart des lignées, nous avons étudié l’implication de GPR40 dans l’effet prolifératif de l’oléate. Ces deux récepteurs n’étant pas exprimés dans les cellules épithéliales mammaires humaines en culture primaire, ces cellules ne répondent pas aux effets de l’oléate sur la prolifération et l’activation des voies de signalisation. L’activation des voies Akt et ERK1/2 par l’oléate dans les cellules MDA-MB-231 et DU145 est potentialisée par la surexpression du récepteur GPR40 et inhibée par l’utilisation d’un siRNA dirigé contre ce récepteur. Cependant, la prolifération induite par l’oléate ne semble pas affectée par la présence d’un siRNA dirigé contre GPR40. L’oléate étant un acide gras, il est capable d’entrer librement dans les cellules et une partie de ses effets sur la prolifération pourrait être attribuée à sa métabolisation. Un agoniste de GPR40, le GW9508, est en mesure d’activer GPR40 sans toutefois entrer dans les cellules ni activer le métabolisme de l’oléate. Le GW9508 stimule la phosphorylation des protéines Akt et ERK1/2 dans les cellules du cancer du sein MDA-MB-231 et de la prostate DU145, mais il n’est pas en mesure d’induire la prolifération cellulaire comme le fait l’oléate. Ces résultats nous permettent de mieux comprendre le mécanisme d’action de l’oléate sur les cellules de cancer du sein et de la prostate. L’oléate induit la signalisation de GPR40 qui est impliquée dans l’activation rapide des voies de signalisation Akt et ERK1/2. De son côté, l’effet prolifératif induit par l’oléate s’effectue par un mécanisme GPR40-indépendant, possiblement lié au métabolisme de l’oléate.

Implication de la voie alternative NF-kappa B dans le cancer de la prostate

Le cancer de la prostate (CaP) est le plus diagnostiqué chez les hommes au Canada et représente le troisième cancer le plus meurtrier au sein de cette population. Malgré l’efficacité des traitements de première ligne, de nombreux patients finiront par développer une résistance et, le cas échéant, verront leur CaP progresser vers une forme plus agressive. Plusieurs paramètres, essentiellement cliniques, permettent de prédire la progression du CaP mais leur sensibilité, encore limitée, implique la nécessité de nouveaux biomarqueurs afin de combler cette lacune. Dans cette optique nous nous intéressons au facteur de transcription NF-κB. Des études réalisées au laboratoire et ailleurs, associent RelA(p65) à un potentiel clinique dans le CaP, soulignant ainsi l’importance de la voie classique NF-κB. L’implication de la voie alternative NF-κB dans la progression du CaP a aussi été suggérée dans une de nos études illustrant la corrélation entre la distribution nucléaire de RelB et le score de Gleason. Alors que la voie classique est largement documentée et son implication dans la progression du CaP établie, la voie alternative, elle, reste à explorer. La présente thèse vise à clarifier l’implication de la voie alternative NF-κB dans le CaP et répond à deux objectifs fixés dans ce but. Le premier objectif fut d’évaluer l’impact de l'activation de la voie alternative NF-κB sur la biologie des cellules cancéreuses prostatiques. L’étude de la surexpression de RelB a souligné les effets de la voie alternative NF-κB sur la prolifération et l'autophagie. Étant ainsi impliquée tant dans la croissance tumorale que dans un processus de plus en plus associée à la progression tumorale, quoique potentiellement létal pour les cellules cancéreuses, son impact sur la tumorigénèse du CaP reste encore difficile à définir. Il n'existe, à ce jour, aucune étude permettant de comparer le potentiel clinique des voies classique et alternative NF-κB. Le second objectif de ce projet fut donc l'analyse conjointe de RelA(p65) et RelB au sein de mêmes tissus de patients atteints de CaP afin de déterminer l'importance clinique des deux signalisations NF-κB, l'une par rapport à l'autre. Le marquage immunofluorescent de RelA(p65) et RelB en a permis l'analyse quantitative et objective par un logiciel d'imagerie. Nos travaux ont confirmé le potentiel clinique associé à RelA(p65). La variable RelA(p65)/RelB s’est, elle, avérée moins informative que RelA(p65). Par contre, aucune corrélation entre RelB et les paramètres cliniques inclus dans l'étude n’est ressortie. En définitive, mon projet de thèse aura permis de préciser l'implication de la voie alternative NF-κB sur la biologie du CaP. Son impact sur la croissance des cellules cancéreuses prostatiques ainsi que sur l'autophagie, dénote l’ambivalence de la voie alternative NF-κB face à la tumorigénèse du CaP. L’étude exhaustive de la signalisation NF-κB souligne davantage l'importance de la voie classique dont l’intérêt clinique est principalement associé au statut de RelA(p65). Ainsi, bien que RelB n’affiche aucun potentiel en tant que biomarqueur exploitable en clinique, l’analyse de l’intervention de la voie alternative NF-κB sur la biologie des cellules cancéreuses prostatiques reste d’intérêt pour la compréhension de son rôle exact dans la progression du CaP.