No abstract

Ejaculated mammalian sperm require several hours exposure to secretions in female reproductive tracts, or incubation in appropriate culture medium in vitro, before acquiring the capacity to fertilize eggs. Arachidonylethanolamide (AEA), also known as anandamide, is a novel lipid-signal molecule that is an endogenous agonist (endocannabinoid) for cannabinoid receptors. We now report that AEA is present in human seminal plasma, mid-cycle oviductal fluid, and follicular fluid analyzed by high-performance liquid chromatography/mass spectrometry. Sperm are sequentially exposed to these reproductive fluids as they move from the vagina to the site of fertilization in the oviduct. Specific binding of the potent cannabinoid agonist [3 H]CP-55,940 to human sperm was saturable (K D 9.71 AE 1.04 nM), suggesting that they express cannabinoid receptors. R-methanandamide , a potent and metabolically stable AEA analog, and (À)D 9 tetrahydrocannabinol (THC), the major psychoactive constituent of Cannabis, modulated capacitation and fertilizing potential of human sperm in vitro. AM-356 elicited biphasic effects on the incidence of hyperactivated sperm motility (HA) between 1 and 6 hr of incubation: at (2.5 nM) it inhibited HA, while at (0.25 nM) it stimulated HA. Both AM-356 and THC inhibited morphological alterations over acrosomal caps between 2 and 6 hr (IC 50 5.9 AE 0.6 pM and 3.5 AE 1.5 nM, respectively). Sperm fertilizing capacity, measured in the Hemizona Assay, was reduced 50% by (1 nM) AM-356. These findings suggest that AEA-signaling may regulate sperm functions required for fertilization in human reproductive tracts, and imply that smoking of marijuana could impact these processes. This study has potential medical and public policy ramifications because of the incidence of marijuana abuse by adults in our society, previously documented reproductive effects of marijuana, and the ongoing debate about medicinal use of marijuana and cannabinoids.

No abstract

Tumor angiogenesis, a major requirement for tumor outgrowth and metastasis formation, is regulated by pro-and anti-angiogenic factors. We have studied the expression of a panel of angiogenic factors, and of the angiogenesis inhibitor angiostatin, in a panel of human melanoma cell lines giving rise to xenografts with different vascular densities. Angiogenic-factor expression was analyzed in vitro (cell lines) and in vivo (xenografts), both at mRNA (RT-PCR and Northern blot) and at protein level (ELISA and Western blot). In vitro angiostatin generation was assessed by Western-blot analysis. Expression of bFGF and VEGF was clearly correlated with a high degree of vascularization, confirming the importance of these factors for tumor angiogenesis. In addition, there was exclusive or elevated in vitro expression of angiogenic factors IL-8, PDGF-AB, and, to a lesser extent, midkine in cell lines that formed highly vascularized tumors. A similar angiogenic-factor-expression pattern was found in the corresponding xenografts, with the exception of VEGF. In most cell lines, this factor had low expression in vitro which was strongly enhanced in vivo. Although all 8 melanoma cell lines were able to excise the angiostatin fragment from the plasminogen parent molecule in vitro, cell lines BLM and M14 showed the most potent angiostatin generation. In vitro angiostatin generation by cell lysates prepared from melanoma xenografts was comparable in all xenograft types. Thus, in our model system we found no correlation between angiostatin generation and vascular density. Our study has limited the number of pro-angiogenic factors that may be involved in melanoma angiogenesis, and provides evidence for the notion that regulation of tumor angiogenesis is dependent on multiple factors. Inhibition of angiogenesis for therapeutic purposes, therefore, should preferably not concentrate on a single factor. Int.

A B S T R A C T Daily administration of estrogen antagonists to premenopausal women has been incorporated into the adjuvant treatment ofbreast cancer. We have studied the changes in reproductive hormones, pituitary responses to hypothalamic-releasing hormones, and endometrial histology during treatment with the antiestrogen tamoxifen in five healthy, premenopausal women. These studies were carried out during one menstrual cycle before and during two cycles ofantiestrogen treatment. All subjects continued to have regular menses with biphasic basal body temperature records. During treatment, estradiol (E2) levels were increased but followed the usual pattern reflecting follicular maturation and corpus luteum formation. The mean E2 concentration at the midcycle peak and during the luteal phase was twice that observed during the non-treatment cycle. By contrast, the concentrations and secretory patterns of luteinizing hormone and follicle-stimulating hormone were not greatly changed, and the gonadotropin responses to gonadotropin-releasing hormone were not suppressed.Endometrial biopsies obtained during the follicular phase of control and tamoxifen treatment cycles showed no differences whereas biopsies obtained during the luteal phase of tamoxifen cycles uniformly showed a lack of changes attributed to progesterone action with no progression of histologic changes beyond those expected on day 7-8 of the luteal phase.These observations are consistent with maturation of multiple ovarian follicles, a surprising finding considering the normal gonadotropin concentrations. The retarded development of the endometrium in the presence of supranormal serum E2 and progesterone concentrations is a morphologic demonstration of the antiprogestational effect of antiestrogens. The lack of gonadotropin suppression in the presence of hyper-