Therapeutic ultrasound as a potential male contraceptive: power, frequency and temperature required to deplete rat testes of meiotic cells and epididymides of sperm determined using a commercially available system.

BACKGROUND: Studies published in the 1970s by Mostafa S. Fahim and colleagues showed that a short treatment with ultrasound caused the depletion of germ cells and infertility. The goal of the current study was to determine if a commercially available therapeutic ultrasound generator and transducer could be used as the basis for a male contraceptive. METHODS: Sprague-Dawley rats were anesthetized and their testes were treated with 1 MHz or 3 MHz ultrasound while varying power, duration and temperature of treatment. RESULTS: We found that 3 MHz ultrasound delivered with 2.2 Watt per square cm power for fifteen minutes was necessary to deplete spermatocytes and spermatids from the testis and that this treatment significantly reduced epididymal sperm reserves. 3 MHz ultrasound treatment reduced total epididymal sperm count 10-fold lower than the wet-heat control and decreased motile sperm counts 1,000-fold lower than wet-heat alone. The current treatment regimen provided nominally more energy to the treatment chamber than Fahim's originally reported conditions of 1 MHz ultrasound delivered at 1 Watt per square cm for ten minutes. However, the true spatial average intensity, effective radiating area and power output of the transducers used by Fahim were not reported, making a direct comparison impossible. We found that germ cell depletion was most uniform and effective when we rotated the therapeutic transducer to mitigate non-uniformity of the beam field. The lowest sperm count was achieved when the coupling medium (3% saline) was held at 37 degrees C and two consecutive 15-minute treatments of 3 MHz ultrasound at 2.2 Watt per square cm were separated by 2 days. CONCLUSIONS: The non-invasive nature of ultrasound and its efficacy in reducing sperm count make therapeutic ultrasound a promising candidate for a male contraceptive. However, further studies must be conducted to confirm its efficacy in providing a contraceptive effect, to test the result of repeated use, to verify that the contraceptive effect is reversible and to demonstrate that there are no detrimental, long-term effects from using ultrasound as a method of male contraception.

Furan-induced dose-response relationships for liver cytotoxicity, cell proliferation, and tumorigenicity (furan-induced liver tumorigenicity).

Rodent studies of furan are associated with liver cell necrosis, release of liver-associated enzymes, increased hepatocyte proliferation, and hepatocarcinogenesis. For carcinogens whose proposed mode of action is cytolethality, it is hypothesized that the dose-response curve for tumor development would parallel the dose-response curve for cell death with compensatory proliferation in the target organ. To prospectively test this hypothesis, female B6C3F(1) mice were exposed to furan at carcinogenic doses and lower for 3 weeks or 2 years. At 3 weeks and in the 2-year study, there were dose-dependent and significant increases in hepatic cytotoxicity at 1.0, 2.0, 4.0, and 8.0mg furan/kg. For cell proliferation as measured by 5-bromo-2'-deoxyuridine (BrdU) labeling index (LI), there was a statistically significant trend with increasing dose levels of furan and increased LI at 8.0mg/kg. There was an increased incidence of foci of altered hepatocytes, hepatocellular adenomas, and adenomas or carcinomas at 4.0 and 8.0mg/kg and carcinomas at 8.0mg/kg. The multiplicity of microscopic tumors was increased and latency was decreased in mice exposed to 8.0mg/kg. Prevalence of hepatic nodules at necropsy was increased in mice exposed to 4.0 and 8.0mg/kg. Data demonstrate an association among furan-induced hepatic cytotoxicity, compensatory cell replication, and liver tumor formation in mice; at high doses >or=4.0mg/kg, furan induced hepatotoxicity, compensatory cell replication and tumorigenesis in a dose-related manner, while furan did not produce tumors at cytotoxic doses of 1.0 and 2.0mg/kg.

An investigation of the effects of late-onset dietary restriction on prostate cancer development in the TRAMP mouse.

In our previous work we showed that dietary restriction initiated at puberty reduced prostate cancer development in the TRAMP mouse model. The current study was conducted to ascertain whether a dietary restriction regime would similarly reduce lesion development if imposed once tumor development was well established. Male TRAMP mice were maintained on an ad libitum diet until 20 weeks of age when proliferative prostate lesions are clearly evident. Mice were then subjected to a 20% restriction in dietary calories compared to matched controls, which were continued on ad libitum feeding. Mice were sacrificed at 20, 24, 32, and 39 weeks of age and proliferative epithelial lesions of the prostate were assessed using an established grading scheme. In this study, although dietary restriction reduced mean sex pluck weight (prostate and seminal vesicles), and mean grade of epithelial proliferative lesions in the dorsal and lateral lobes of the prostate, the effect was not as pronounced as was the case with dietary restriction from puberty. There was no relationship between serum insulin like growth factor (IGF-1) and prostate lesion grade. Additionally, we also report the relationship between lobe specific lesion development and SV40 immunostaining and, the occurance of neuroendocrine tumors (NETs) in the ventral prostate and urethra of the TRAMP mouse. NETs stained with high specificity and sensitivity for the neuroendocrine markers, synaptophysin and neuron-specific enolase (NSE), less for serotonin, but not for chromogranin A. NETs did not stain for cyclo-oxygenase-2 (COX-2) nor androgen receptor (AR). SV40 positive tubulo-acinar tumors seen occasionally in the kidney, did not stain for synaptophysin nor NSE.

Boron supplementation inhibits the growth and local expression of IGF-1 in human prostate adenocarcinoma (LNCaP) tumors in nude mice.

Prostate-specific antigen (PSA) is a serine protease and one of the most abundant proteins secreted by the human prostate epithelium. PSA is used as a well-established marker of prostate cancer. The involvement of PSA in several early events leading to the development of malignant prostate tumors has made it a target for prevention and intervention. It is thought that PSA cleaves insulin-like growth factor binding protein-3 (IGFBP-3), providing increased local levels of IGF-1, leading to tumor growth. Separately, there are data that suggest an enzymatic regulatory role for dietary boron, which is a serine protease inhibitor. In this study we have addressed the use of boric acid as a PSA inhibitor in an animal study. We have previously reported that low concentrations (6 ug/mL) of boric acid can partially inhibit the proteolytic activity of purified PSA towards a synthetic fluorogenic substrate. Also, by Western blot we have followed the degradation of fibronectin by enzymatically active PSA and have found significant inhibition in the presence of boric acid. We proposed that dietary supplementation with boric acid would inhibit PSA and reduce the development and proliferation of prostate carcinomas in an animal model. We tested this hypothesis using nude mice implanted subcutaneously with LNCaP cells in Matrigel. Two groups (10 animals/group) were dosed with boric acid solutions (1.7, 9.0 mgB/kg/day) by gavage. Control group received only water. Tumor sizes were measured weekly for 8 weeks. Serum PSA and IGF-1 levels were determined at terminal sacrifice. The size of tumors was decreased in mice exposed to the low and high dose of boric acid by 38% and 25%, respectively. Serum PSA levels decreased by 88.6% and 86.4%, respectively, as compared to the control group. There were morphological differences between the tumors in control and boron-dosed animals, including a significantly lower incidence of mitotic figures in the boron-supplemented groups. Circulating IGF-1 levels were not different among groups, though expression of IGF-1 in the tumors was markedly reduced by boron treatment, which we have shown by immunohistochemistry. These data indicate that low-level dietary boron supplementation reduced tumor size and content of a tumor trophic factor, IGF-1. This promising model is being evaluated in further studies.

A grading scheme for the assessment of proliferative lesions of the mouse prostate in the TRAMP model.

To improve the precision and consistency of experimental results, we have developed a scoring system for proliferative epithelial lesions in the mouse prostate based on histological growth patterns observed in individual lobes. Severity of proliferative lesions was divided into 6 categories; the grade of the most advanced lesion was identified for each lobe and its distribution estimated semiquantitatively. A numerical score combining grade and distribution of the most advanced lesion in each lobe was assigned and termed the "distribution-adjusted lesion grade"; the mean of these scores was calculated for each treatment group. Using this grading scheme, we assessed lesion development in ad libitum-fed and 20%-diet-restricted groups of TRAMP (Transgenic Adenocarcinoma of Mouse Prostate) mice that were started on study at 7 weeks of age and sacrificed when 11 and 20 weeks old. The anterior, dorsal, lateral, and ventral prostate lobes showed clear reductions in lesion severity in diet-restricted TRAMPS at 11 and 20 weeks. This method for scoring the epithelial pathology of the prostate in the TRAMP model with minimal to severe proliferative lesions utilizes the natural history of lesion development for assessing the effects of chemical and dietary interventions.