Penile vibratory stimulation yields increased spermatozoa and accessory gland production compared with rectal electroejaculation in a neurologically intact primate (Saimiri boliviensis)

Assisted reproductive techniques require an efficient semen collection procedure in cases of ejaculatory dysfunction. Anejaculation may be of psychogenic or neurogenic origin but can be overcome with stimulatory techniques. Penile vibratory stimulation (PVS) therapy for anejaculation has recently emerged as an alternative to rectal probe electroejaculation (RPE) and more invasive testicular procedures. Comparison of the stimulatory procedures in neurologically intact subjects is not ethically possible due to the discomfort involved with electroejaculation, and comparison in spinal cord injured men may be compromised due to the intricate effects of chronic denervation on semen quality. We have previously shown the efficacy of PVS in a non-human primate, the squirrel monkey. A cross-over study design comparing semen collected by PVS and RPE was employed during the breeding season in which 15 donor males were divided into two groups. One group received PVS and the other RPE, then, three days later, treatments were reversed. Twelve of 15 animals responded to PVS (80%), all with spermatozoa in the ejaculate. Mean volume (436 +/- 90 microl), motility (80.6 +/- 4.3%), and total spermatozoa (32.8 +/- 10.2 x 10(6)) were significantly higher than in the semen after RPE. RPE resulted in ejaculation in all 15 animals with a semen volume of 205 +/- 25 microl, but fewer samples contained spermatozoa (9/15) resulting in a low total count (0.5 +/- 0.3 x 10(6)). The motility was reduced in those samples with spermatozoa (n = 9; 44.1 +/- 11.4%). Additionally, accessory gland activity was measured via the seminal vesicle and prostrate markers, fructose and citric acid, respectively. The PVS specimens had significantly more fructose (2.9 +/- 0.7 mg/ejaculate) and citric acid (0.46 +/- 0.14 mg/ejaculate) compared to RPE collected specimens (1.2 +/- 0.3 mg/ejaculate and 0.24 +/- 0.04 mg/ejaculate, respectively). In conclusion, PVS produces a much greater sperm yield and increased accessory gland secretion compared to RPE in our neurologically intact squirrel monkey model.     

Increased urinary free cortisol: a potential intermediate phenotype of essential hypertension

We evaluated urinary cortisol excretion as a potential intermediate phenotype of essential hypertension in 153 white patients with essential hypertension and 18 normotensive white control subjects. Analyses were controlled for dietary sodium and gender to adjust for potential confounding effects of these variables on cortisol excretion. Urinary cortisol excretion measured on both high- and low-salt diets was significantly related to hypertension by repeated measures ANCOVA (P=.02). Additional determinants of urinary free cortisol included dietary sodium intake and gender; cortisol excretion was significantly higher in men (P=.0006) and during a high-sodium diet (P=.0001). Maximum likelihood analysis showed urinary cortisol to have a bimodal distribution on both 200-mmol (P<.01) and 10-mmol (P<.002) sodium diets in hypertensive subjects. On the low-salt diet, the mean urinary cortisol in normotensive subjects (108.7+/-44.7 nmol/d) was similar to the mean of hypertensive subjects in the low mode (127.2+/-43.0 nmol/d). The high mode comprised 31.2% of the hypertensive population and had a mean urinary cortisol of 224.3+/-93.8 nmol/d. Subjects with the highest urinary free cortisol showed the least sensitivity of blood pressure to dietary sodium loading (P<.05). These data suggest that there is an association between salt-resistant hypertension and high urine cortisol levels. This association may have a genetic basis.     

An intersection of the cAMP/PKA and two-component signal transduction systems in Dictyostelium

Terminal differentiation of both stalk and spore cells in Dictyostelium can be triggered by activation of cAMP-dependent protein kinase (PKA). A screen for mutants where stalk and spore cells mature in isolation produced three genes which may act as negative regulators of PKA: rdeC (encoding the PKA regulatory subunit), regA and rdeA. The biochemical properties of RegA were studied in detail. One domain is a cAMP phosphodiesterase (Km approximately 5 microM); the other is homologous to response regulators (RRs) of two-component signal transduction systems. It can accept phosphate from acetyl phosphate in a reaction typical of RRs, with transfer dependent on Asp212, the predicted phosphoacceptor. RegA phosphodiesterase activity is stimulated up to 8-fold by the phosphodonor phosphoramidate, with stimulation again dependent on Asp212. This indicates that phosphorylation of the RR domain activates the phosphodiesterase domain. Overexpression of the RR domain in wild-type cells phenocopies a regA null. We interpret this dominant-negative effect as due to a diversion of the normal flow of phosphates from RegA, thus preventing its activation. Mutation of rdeA is known to produce elevated cAMP levels. We propose that cAMP breakdown is controlled by a phosphorelay system which activates RegA, and may include RdeA. Cell maturation should be triggered when this system is inhibited.     

STAR MR angiography for rapid detection of vascular abnormalities in patients with acute cerebrovascular disease

BACKGROUND AND PURPOSE: We undertook to investigate the usefulness of signal targeting with alternating radiofrequency magnetic resonance angiography (STAR MRA) in the diagnosis of acute cerebrovascular disease. The potential advantage of the technique is that angiographic images can be acquired in less than 1 minute. METHODS: We studied 19 patients (11 men and 8 women, ranging in age from 36 to 84 years [mean age, 66 years]) presenting with signs and symptoms of acute stroke. Patients underwent STAR MRA and three-dimensional fast imaging with steady-state precession (3D FISP) MRA. The MRAs were analyzed as to image quality and vascular abnormalities in the vascular territory of stroke as defined by diffusion-weighted imaging abnormalities and compared using a Wilcoxon signed-rank test. RESULTS: STAR MRAs had slightly inferior image quality compared with 3D FISP MRA (P < .05). STAR MRA and 3D FISP MRA agreed in 18 of 19 cases regarding vascular abnormalities in the territory of the infarct (occlusion, n = 8; stenosis, n = 4; no abnormality, n = 6). In one patient, the techniques disagreed, when 3D FISP MRA was normal and STAR MRA demonstrated a vessel occlusion in the vascular territory of a stroke as defined by diffusion-weighted imaging abnormalities (P > .05). CONCLUSIONS: Despite slightly inferior image quality compared with 3D FISP MRA, STAR MRA is comparable with 3D FISP MRA in depicting abnormalities in the proximal parts of the cerebral arteries corresponding to ischemic regions on diffusion-weighted imaging, in a strikingly shorter acquisition time. Further studies are necessary to confirm that the smaller branches are better shown by using longer inversion times.     

Th1-like cytokine production profile and individual specific alterations in TCRBV-gene usage of T cells from newly diagnosed type 1 diabetes patients after stimulation with beta-cell antigens

Thirty complete coding sequences of human major histocompatibility complex (Mhc) class II DRB alleles, spanning 237 codons, were analyzed for phylogenetic information using distance, parsimony, and likelihood approaches. Allelic genealogies derived from different parts of the coding sequence (exon 2, the 5' and 3' ends of exon 2, respectively, and exons 3-6) were compared. Contrary to prior assertions, a rigorous analysis of allelic genealogies in this gene family cannot be used to justify the claim that the lineage leading to modern humans contained on average at least 100,000 individuals. Phylogenetic inferences based upon the exon 2 region of the DRB loci are complicated by selection and recombination, so this part of the gene does not provide a complete and accurate view of allelic relationships. Attempts to reconstruct human history from genetic data must use realistic models which consider the complicating factors of nonequilibrium populations, recombination, and different patterns of selection.     

The G2/M DNA damage checkpoint inhibits mitosis through Tyr15 phosphorylation of p34cdc2 in Aspergillus nidulans

It is possible to cause G2 arrest in Aspergillus nidulans by inactivating either p34cdc2 or NIMA. We therefore investigated the negative control of these two mitosis-promoting kinases after DNA damage. DNA damage caused rapid Tyr15 phosphorylation of p34cdc2 and transient cell cycle arrest but had little effect on the activity of NIMA. Dividing cells deficient in Tyr15 phosphorylation of p34cdc2 were sensitive to both MMS and UV irradiation and entered lethal premature mitosis with damaged DNA. However, non-dividing quiescent conidiospores of the Tyr15 mutant strain were not sensitive to DNA damage. The UV and MMS sensitivity of cells unable to tyrosine phosphorylate p34cdc2 is therefore caused by defects in DNA damage checkpoint regulation over mitosis. Both the nimA5 and nimT23 temperature-sensitive mutations cause an arrest in G2 at 42 degrees C. Addition of MMS to nimT23 G2-arrested cells caused a marked delay in their entry into mitosis upon downshift to 32 degrees C and this delay was correlated with a long delay in the dephosphorylation and activation of p34cdc2. Addition of MMS to nimA5 G2-arrested cells caused inactivation of the H1 kinase activity of p34cdc2 due to an increase in its Tyr15 phosphorylation level and delayed entry into mitosis upon return to 32 degrees C. However, if Tyr15 phosphorylation of p34cdc2 was prevented then its H1 kinase activity was not inactivated upon MMS addition to nimA5 G2-arrested cells and they rapidly progressed into a lethal mitosis upon release to 32 degrees C. Thus, Tyr15 phosphorylation of p34cdc2 in G2 arrests initiation of mitosis after DNA damage in A. nidulans.     

The use of amplified variable number of tandem repeats (VNTR) in the detection of chimerism following bone marrow transplantation. A comparison with restriction fragment length polymorphism (RFLP) by Southern blotting

A 49-year-old woman patient with atypical myelodysplastic syndrome (MDS) showing a der(3)t(3;12)(q21;p13), and der(12)t(3;12)(q21;p13)inv(3)(q21q26) as an acquired chromosomal abnormality in the bone marrow is described. The chromosomal breakpoints of the presented complex aberration with combination of the inv(3)(q21q26) and t(3;12)(q21;p13) were defined by fluorescence in situ hybridization (FISH) with yeast artificial chromosomes (YACs). The inv(3) is a relatively frequent chromosomal rearrangement in patients with myeloid malignancies and dysmegakaryopoiesis and t(3;12)(q26;p13) has also been reported as a recurrent abnormality in MDS and in blast crisis of chronic myelogenous leukemia (CML). Whereas the t(3;12), inv(3), and t(3;3) are associated with a very poor prognosis, our patient surprisingly had a mild clinical course.     

Expression of gtfS is essential for normal insoluble glucan synthesis by Streptococcus downei

The gtfI and gtfS genes of Streptococcus downei were investigated to determine the contribution of the respective enzymes to glucan production in the presence and absence of other glucosyltransferases. Extracts of Escherichia coli expressing cloned gtfS produced a short linear dextran from sucrose which could act as a primer for insoluble glucan synthesis when mixed with extracts of a strain expressing recombinant gtfI. To elucidate the contribution of gtfS to glucan production by S. downei, a mutant was constructed by insertionally inactivating gtfS. S. downei (gtfS mutant) colonies exhibited a marked phenotypic change on sucrose-containing media and a decreased ability to adhere to glass and produced no detectable water-insoluble glucan. These experiments confirm that expression of gtfS is essential for normal insoluble glucan synthesis by S. downei.     

Microperoxidases catalytically degrade reactive oxygen species and may be anti-cataract agents

1,25-(OH)2D3 and 24,25-(OH)2D3 mediate their effects on chondrocytes through the classic vitamin D receptor (VDR) as well as through rapid membrane-mediated mechanisms which result in both nongenomic and genomic effects. In intact cells, it is difficult to distinguish between genomic responses via the VDR and genomic and nongenomic responses via membrane-mediated pathways. In this study, we used two hybrid analogues of 1,25-(OH)2D3 which have been modified on the A-ring and C,D-ring side chain (1 alpha-(hydroxymethyl)-3 beta-hydroxy-20-epi-22-oxa-26,27-dihomo vitamin D3 (analogue MCW-YA = 3a) and 1 beta-(hydroxymethyl)-3 alpha-hydroxy-20-epi-22-oxa-26,27-dihomo vitamin D3 (analogue MCW-YB = 3b) to examine the role of the VDR in response of rat costochondral resting zone (RC) and growth zone (GC) chondrocytes to 1,25-(OH)2D3 and 24,25-(OH)2D3. These hybrid analogues are only 0.1% as effective in binding to the VDR from calf thymus as 1,25-(OH)2D3. Chondrocyte proliferation ([3H]-thymidine incorporation), proteoglycan production ([35S]-sulfate incorporation), and activity of protein kinase C (PKC) were measured after treatment with 1,25-(OH)2D3, 24,25-(OH)2D3, or the analogues. Both analogues inhibited proliferation of both cell types, as did 1,25-(OH)2D3 and 24,25-(OH)2D3. Analogue 3a had no effect on proteoglycan production by GCs but increased that by RCs. Analogue 3b increased proteoglycan production in both GC and RC cultures. Both analogues stimulated PKC in GC cells; however, neither 3a nor 3b had an effect on PKC activity in RC cells. 1,25-(OH)2D3 and 3a decreased PKC in matrix vesicles from GC cultures, whereas plasma membrane PKC activity was increased, with 1,25-(OH)2D3 having a greater effect. 24,25-(OH)2D3 caused a significant decrease in PKC activity in matrix vesicles from RC cultures; 24,25-(OH)2D3, 3a, and 3b increased PKC activity in the plasma membrane fraction, however. Thus, with little or no binding to calf thymus VDR, 3a and 3b can affect cell proliferation, proteoglycan production, and PKC activity. The direct membrane effect is analogue-specific and cell maturation-dependent. By studying analogues with greatly reduced affinity for the VDR, we have provided further evidence for the existence of a membrane receptor(s) involved in mediating nongenomic effects of vitamin D metabolites.