Effects of the ion-channel blocker quinine on human sperm volume, kinematics and mucus penetration, and the involvement of potassium channels

Sperm defects in the infertile c-ros knockout mouse model have recently highlighted the importance of volume regulation in sperm function. In this study, washed human spermatozoa were shown to change size and shape, as detected by flow cytometry and light microscopy, in response to the ion-channel blocker quinine (minimum effective doses at 20 and 125 micromol/l respectively). The increase in sperm volume was accompanied by reduced straight-line velocity (VSL) and linearity (LIN) of the swim-path but increased lateral head displacement and curvilinear velocity, while percentage motility was unaffected. Spermatozoa in semen and in artificial cervical mucus were similarly affected at 0.2 and 0.5 mmol/l quinine, resulting in marked reduction of mucus penetration and migration. The effects of quinine on sperm volume and kinematics were reduced or abolished by the K(+)-ionophores valinomycin (1 and 5 micromol/l) and gramicidin (0.5 and 1 micromol/l). In Ca(2+)-free medium; however, the quinine effects largely persisted. The K(+)-channel blocker, 4-aminopyridine (1 and 4 mmol/l), mimicked the quinine effects in the reduction of VSL and LIN, while the K(+)-channel blocker, tetraethylammonium chloride (TEA, 2.5-10 mmol/l), did not affect kinematics. The K(+)-channel (Kv1.3)-specific inhibitor, margatoxin, and the Ca(2+)-dependent K(+)-channel blocker, charybdotoxin, also had no effects. This study suggests that volume regulation in human spermatozoa and the linear trajectory of their motion may rely on quinine-sensitive and TEA-insensitive, largely calcium-independent, potassium channels, and possibly volume-sensitive organic anion channels. These channels could be targets for contraception.     

Genetic typing and HIV-1 diagnosis by using 96 capillary array electrophoresis and ultraviolet absorption detection

Current high-throughput approaches to the analysis of PCR products are based primarily on electrophoretic separation and laser-excited fluorescence detection. We show that capillary array electrophoresis can be applied to HIV-1 diagnosis and D1S80 VNTR genetic typing based simply on UV absorption detection. The additive contribution of each base pair to the total absorption signal provides adequate detection sensitivity for analyzing most PCR products. Not only is the use of specialized and potentially toxic fluorescent labels eliminated, but also the complexity and cost of the instrumentation are greatly reduced.     

Release of lymphotoxin by control and chemical carcinogen-treated lymphocyte cultures derived from black healthy subjects and cancer patients

In an age-adjusted comparison with white men, black men have a significantly higher increase in esophageal and other types of cancer associated with environmental causes. The basis of this increase in cancer rates in blacks over the last two decades is unknown. Since cancer patients generally show an impairment in cell-mediated immune (CMI) functions, we measured certain CMI reactions in cultured lymphocytes derived from black healthy subjects and cancer patients. We also determined the levels of aryl hydrocarbon hydroxylase (AHH) induced in these lymphocytes. AHH catalyzes the activation of polycyclic aromatic hydrocarbons (PAH) to intermediates which might alter CMI functions.     

Functional expression and properties of the human skeletal muscle sodium channel

Full-length deoxyribonucleic acid, complementary (cDNA) constructs encoding the-subunit of the adult human skeletal muscle Na⁺ channel, hSkM1, were prepared. Functional expression was studied by electrophysiological recordings from cRNA-injectedXenopus oocytes and from transiently transfected tsA201 cells. The Na⁺ currents of hSkM1 had abnormally slow inactivation kinetics in oocytes, but relatively normal kinetics when expressed in the mammalian cell line. The inactivation kinetics of Na⁺ currents in oocytes, during a depolarization, were fitted by a weighted sum of two decaying exponentials. The time constant of the fast component was comparable to that of the single component observed in mammalian cells. The block of hSkM1 Na⁺ currents by the extracellular toxins tetrodotoxin (TTX) and -conotoxin (CTX) was measured. The IC₅₀ values were 25 nM (TTX) and 1.2 M (CTX) in oocytes. The potency of TTX is similar to that observed for the rat homolog rSkM1, but the potency of CTX is 22-fold lower in hSkM1, primarily due to a higher rate of toxin dissociation in hSkM1. Single-channel recordings were obtained from outside-out patches of oocytes expressing hSkM1. The single-channel conductance, 24.9 pS, is similar to that observed for rSkM1 expressed in oocytes.     

Follicular fluid renin concentration and IVF outcome

Total renin protein concentration (TRC) was measured in stored follicular fluid (FF) samples from 42 women. Samples were selected according to their origin from follicles either without recovered ova ('empty', n = 38) or fertilized but with failed implantation ('failed', n = 36) or successful deliveries ('deliveries', n = 71). Ratios of number of embryos transferred to number of infants delivered were 2:1, 3:1 or 4:2 but 1:1 was not available. Non-parametric testing was applied to FF-TRC, volume and outcome. TRC was significantly higher in the delivery than the failed (P = 0.001) or empty (P = 0.002) categories. Assuming that the range of renin in failed follicles can identify the sub-population of unsuccessful follicles in the delivery category, then elevated FF-TRC was clearly associated with successful outcome. For individual women, the odds of infant delivery increased 17-fold as a function of average FF-TRC between 10,000 and 25,000 microIU/ml. For failed and delivery but not empty follicles, higher renin levels occurred in the smaller follicles, consistent with a burst of renin synthesis associated with the presence of an oocyte. The results suggest that FF-TRC relates to ovum viability with ovarian hyperstimulation and may have predictive use in IVF programmes.     

Effect of statins combined with estradiol on the proliferation of human receptor-positive and receptor-negative breast cancer cells

OBJECTIVE: Combination of a statin plus estrogen may reveal benefits on the cardiovascular system in postmenopausal women by additively ameliorating both the lipid profile and vascular function. Long-term therapy with estrogens, however, is associated with an increase of breast cancer risk. In contrast, evidence is accumulating that statins may inhibit carcinogenesis because of their central action on important cellular functions. It is of special clinical interest whether a statin/estrogen combination may reduce the most undesired side effect of estrogen therapy, that is, an increase in breast cancer risk. Therefore, in the present in vitro study, for the first time we have compared the effect of five statins on the proliferation of human breast cancer cells alone and in the presence of stimulatory estradiol (E(2)). DESIGN: As cell models, the receptor-positive cell line MCF-7 and the receptor-negative cell line MDA-MB 231 were used. The statins atorvastatin, fluvastatin, lovastatin, pravastatin, and simvastatin were tested in the concentration range of 1.6 microm to 50 microm alone and in the range of 0.01 nm to 10 microm in combination with E(2). Cell proliferation was measured after 4 days by the adenosinetriphosphate-chemosensitivity test. RESULTS: All statins except pravastatin were able to significantly inhibit dose dependently the cell proliferation of both cell lines. The inhibitory values were between 10% and 90%, whereby the potency was greater in the case of receptor-negative cancer cells. A significant difference in the efficacy of the statins was observed for MCF-7 cells, in which atorvastatin was less effective than the other statins. In contrast, in the presence of E(2), the statins showed similar antiproliferative actions in MCF-7 cells when tested in the concentration range of 0.01 nm to 10 microm. A reduction of cell proliferation of less than 10% was observed at the lower concentrations and between 15% and 25% at the highest concentration of 10 microm. CONCLUSIONS: The present data indicate that statins can inhibit the proliferation of receptor-positive and -negative human breast cancer cells but failed to completely abrogate the E(2)-induced proliferation of receptor-positive breast cancer cells. Clinical trials, however, are necessary to prove this anticarcinogenic action of statins.     

Alternate Pathway Activation in Sickle Cell Disease and ��-Thalassemia Major

Total hemolytic complement activity (CH50), immuno-electrophoretic conversion of Factor B (C3PA), and of C3 were studied in 16 patients with sickle cell disease in a steady state, eight patients in crisis, and ten patients with β-thalassemia major anemia maintained on a constant transfusion regimen. Patients with sickle cell disease in a steady state have moderatley 56 (percent) depressed conversion of Factor B in addition to markedly decreased conversion of C3 in four of ten patients. One of the three sickle cell patients and two of the four thalassemia patients with low C3 conversion levels have died subsequent to the studies. The combination of chronically decreased Factor B conversion in the face of markedly decreased C3 conversion may make these patients occasionally vulnerable to overwhelming infection analagous to the situation seen in postsplenectomy cases.