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Biomedical subjects

R Sullivan

Publications and source records attributed to R Sullivan.

At least 19 recordsLinked to original sources
PubMed2002

Effect of immunization of hamsters against recombinant P26h on fertility rates.

Despite the various contraceptive methods available, an effective and inexpensive method remains to be established. Immunocontraception may help to achieve this goal. P26h has been proposed as a candidate for the development of a male contraceptive vaccine. P26h, a hamster sperm protein, interacts with the zona pellucida. Furthermore, in vivo fertilization can be blocked completely by active immunization of male hamsters against P26h. Maltose binding protein (MBP)-P26 shares antigenic determinants with the native P26h present on cauda epididymal spermatozoa. The aim of the present study was to reproduce the immunocontraceptive properties of native P26h by immunizing male hamsters against a recombinant P26h fused with a maltose binding protein (MBP). Active immunization of male hamsters with the MBP-P26h showed that specific anti-P26h circulating IgGs could be generated. Mating of immunized male hamsters with superovulated females resulted in a significant decrease, 20-25%, in the fertilization rate. This result is in agreement with results from in vitro sperm-zona pellucida binding assays. Indeed, the anti-recombinant P26h IgGs showed lower inhibitory properties when compared with anti-native P26h IgG. Despite the high anti-P26h IgG titres generated in hamsters, histological studies showed that active immunization has no pathological sequelae to the reproductive tissues. The potential of P26h as a candidate for a contraceptive vaccine is discussed.

Alcohol Oxidoreductases↗
PubMed2001

Calmodulin binding to the C-terminus of the small-conductance Ca2+-activated K+ channel hSK1 is affected by alternative splicing.

We identified three splice variants of hSK1 whose C-terminal structures are determined by the independent deletion of two contiguous nucleotide sequences. The upstream sequence extends 25 bases in length, is initiated by a donor splice site within exon 8, and terminates at the end of the exon. The downstream sequence consists of nine bases that compose exon 9. When the upstream sequence (hSK1(-)(25b)) or both sequences (hSK1(-)(34b)) are deleted, truncated proteins are encoded in which the terminal 118 amino acids are absent. The binding of calmodulin to these variants is diminished, particularly in the absence of Ca2+ ions. The first 20 amino acids of the segment deleted from hSK1(-)(25b) and hSK1(-)(34b) contain a 1-8-14 Ca2+ calmodulin binding motif, and synthetic oligopeptides based on this region bind calmodulin better in the presence than absence of Ca2+ ions. When the downstream sequence (hSK1(-)(9b)) alone is deleted, only the three amino acids A452, Q453, and K454 are removed, and calmodulin binding is not reduced. On the basis of the relative abundance of mRNA encoding each of the four isoforms, the full-length variant appears to account for most hSK1 in the human hippocampus, while hSK1(-)(34b) predominates in reticulocytes, and hSK1(-)(9b) is especially abundant in human erythroleukemia cells in culture. We conclude that the binding of calmodulin by hSK1 can be modulated through alternative splicing.

Alternative Splicing↗
PubMed2001

The effect of heparin on motility parameters and protein phosphorylation during bovine sperm capacitation.

It is generally accepted that incubation with heparin is required to induce capacitation of ejaculated bovine spermatozoa in vitro. The capacitation process implicates many biochemical events, and is correlated with modified sperm motility and the phosphorylation of specific proteins on tyrosine residues. To better understand the molecular basis of heparin-induced capacitation, bovine spermatozoa were incorporated with a radioactive substrate of protein kinases [gamma32P]-ATP, to observe protein phosphorylation dynamics over time. Sperm motion parameters including the percentage of motile spermatozoa, amplitude of lateral head displacement (ALH) and flagellar beat cross frequency (BCF) were assessed to determine whether the protein phosphorylation patterns induced by heparin also promote changes in motility. Capacitation was confirmed using the chlortetracycline fluorescence assay and the appearance of 'pattern B' stained spermatozoa. Evaluation of the different motility parameters during capacitation reveal that heparin has a marked negative effect, over time, on the percentage of motile spermatozoa, consistent with hyperactivation. Indeed, the presence of heparin greatly increases the BCF of bull spermatozoa and induces a significant increase in the ALH compared to spermatozoa incubated without heparin. We detected several sperm proteins that are phosphorylated over time. A 45 kDa protein is the most intensely phosphorylated of the sperm proteins. However, it is visible regardless of the presence of heparin. Interestingly, a second phosphorylated protein of approximately 50 kDa undergoes more intense phosphorylation with heparin than without. In summary, the present study demonstrated that heparin induces physiological changes in several sperm motility parameters including ALH, BCF and the percentage of motile spermatozoa. Heparin also increases the intensity of phosphorylation of a 50 kDa sperm protein. These results suggest that capacitation of bovine spermatozoa and capacitation-associated motility changes may be regulated by a mechanism that includes protein phosphorylation, and that a presently unknown protein kinase is involved.

Adenosine Triphosphate↗
PubMed2001

Prostasome-like particles are involved in the transfer of P25b from the bovine epididymal fluid to the sperm surface.

In bulls, P25b is a sperm protein associated with the plasma membrane covering the acrosome. The amount of P25b bound to a constant number of spermatozoa varies from one individual to the other, low levels being associated with bull subfertility. In this study, we describe the epididymal origin of P25b using Western blot analysis. Whereas P25b was undetectable on caput spermatozoa, the amount of P25b associated to a constant number of spermatozoa increases from the corpus to the cauda. Prostasome-like particles were prepared by ultracentrifugation of epididymal fluid. P25b appears to be also associated with those membranous vesicles in increasing amounts along the epididymis. P25b is anchored to the plasma membrane of spermatozoa through glycosylphosphatidyl-inositol as shown by the ability of phospholipase C. but not of high salt treatment, to release P25b. Coincubation experiments revealed that prostasome-like particles are able to transfer P25b to spermatozoa, this process being more efficient at slightly acidic pH. P25b thus appears to be a marker of sperm epididymal maturation in bulls.

Animals↗
PubMed2001

Identification and characterization of P31m, a novel sperm protein in Cynomolgus monkey (Macaca fascicularis).

We have previously described a hamster sperm glycoprotein, P26h, which is implicated in the cascade of events occurring during the interaction between mature spermatozoa and the oocyte's zona pellucida. The P26h is acquired on the acrosomal cap of the spermatozoon during its maturation arising within the epididymis. Lately, using a polyclonal antiserum raised against P26h, a 34 kDa protein, P34H, has been identified on the acrosomal cap of the human spermatozoon. The cloning and sequencing of the cDNA encoding P34H has revealed a 65% similarity between the P34H and P26h amino acid sequences. Considering that P26h shows total immunocontraceptive properties in the hamster, it is of relevant importance to have an animal model phylogenetically closer to the human. Using the Cynomolgus monkey, we searched for a protein autologous to the human P34H. A 31 kDa protein, the P31m, localized on the acrosomal cap of the monkey spermatozoon has been identified by a Western blot analysis and by immunohistochemical techniques using an anti-hamster P26h antiserum. Northern blot analysis showed increasing high levels of the P31m mRNA through the epididymis and at lower levels in the testis. In situ hybridization showed the presence of the P31m mRNA in the principal cells of the epididymis. The cloning and sequencing of the cDNA encoding the P31m showed a high homology of 97% identity between the P31m and P34H nucleotidic sequences. This study clearly demonstrates that the monkey P31m is the homologous protein of the hamster P26h and of the human P34H. Mol. Reprod. Dev. 59: 431-441, 2001.

Acrosome↗
PubMed2001

Gender differences in the depressive effect of widowhood in later life.

OBJECTIVES: This study documented the stronger adverse effect of widowhood on the psychological well-being of men than that of women and explained why this gender difference in the effect of widowhood exists. METHODS: Data came from Wave 1 of the National Survey of Families and Households. Married and widowed people aged 65 and older were selected (n = 1,686). The dependent variable was the Center for Epidemiologic Studies--Depression scale (CES-D). RESULTS: Widowhood was indeed more depressing for men than women. However, this was due primarily to the fact that married men were much less depressed than married women; widowed men and women were comparably depressed. Other contributors to the stronger effect of widowhood for men included men's shorter average time since widowhood, lower frequency of church attendance, stronger dislike of domestic labor, and lessened ability to assist their children. DISCUSSION: Although widowhood has a strong depressive effect for older men, its effect for women is nonsignificant, and it explains a small proportion of the variation in depressive symptomatology. This suggests that most people, particularly women, adapt relatively well in the long run.

Adaptation, Psychological↗
PubMed2001

Effect of vasectomy on P34H messenger ribonucleic acid expression along the human excurrent duct: a reflection on the function of the human epididymis.

Sperm surface proteins involved in fertilization can be added or modified during epididymal transit. P34H, a human epididymal-sperm protein, appears on the sperm acrosomal cap in the distal caput-proximal corpus epididymis. In previous studies, it was shown that P34H is present on spermatozoa in men of proven fertility, is absent in 50% of men presenting with idiopathic infertility, and that a high proportion of men with normospermic vasovasectomy produce spermatozoa deficient in this sperm surface protein. P34H mRNA was expressed in the principal cells of the epididymis of normal men, predominantly in the corpus region. Recently, results coming from the assisted reproductive technologies have questioned the importance of the human epididymis in sperm maturation. In order to understand the effect of obstruction on the physiological state of the human epididymis and its function in sperm maturation, we have analyzed the expression of P34H mRNA at the level of the vas deferens and along the epididymis of normal and vasectomized men. In situ hybridization experiments showed that obstruction of the vas deferens alters the pattern of P34H mRNA expression compared with the tract of normal tissues. The P34H transcript was detected in the proximal caput epididymis of vasectomized men at a much higher intensity than that observed in the same region of normal tissues, being restricted to the principal cells of the epididymal epithelium. Compared with the normal duct, the lumen of vasectomized men was distended throughout the duct and the height of the epithelium was maximal in the caput. P34H mRNA was detectable in vas deferens, was not affected by vasectomy, and a 912-base pair P34H transcript was restricted to the epithelial cells of the vas deferens. Thus, using P34H as a marker, these results show that vasectomy alters the pattern of gene expression along the human epididymis, and suggest that the vas deferens can be a major contributor to sperm maturation in certain situations.

Adult↗
PubMed2001

Expression of the hamster sperm protein P26h during spermatogenesis.

P26h is a hamster sperm protein of 26 kDa that has been previously characterized as a surface protein covering the acrosome acquired during epididymal transit. P26h is involved in sperm-egg interactions. Recently, it has been shown that the P26h transcript is highly expressed in the testis, and the P26h cDNA has been cloned from a hamster testicular cDNA library. Herein we report the production of a fusion protein (maltose binding protein-P26h) with the whole P26h cDNA encoding sequence and the production of a polyclonal antiserum against it. In Western blots, this antiserum recognized both the P26h extracted from cauda epididymal spermatozoa and the MBP-P26h. We also determined the age of appearance of P26h and which germ cell types express P26h mRNA and its translational product. Northern blots and in situ hybridization analysis showed that P26h transcripts appear at 3 wk of age, within the first round of spermatogenesis in the golden hamster. In situ hybridization showed that P26h transcripts are expressed in spermatocytes and round spermatids, whereas immunostaining revealed the presence of P26h in the cytoplasm of round spermatids and elongated spermatids. P26h was undetectable in testicular spermatozoa. Both in situ hybridization and immunostaining showed P26h expression to be dependent of the testicular cell type and the epithelium cycle. The implications for P26h in sperm-egg interaction and the testicular origin of P26h are discussed.

Alcohol Oxidoreductases↗
PubMed2001

Gamma-aminobutyric acid type B receptors with specific heterodimer composition and postsynaptic actions in hippocampal neurons are targets of anticonvulsant gabapentin action.

Gamma-aminobutyric acid (GABA) activates two qualitatively different inhibitory mechanisms through ionotropic GABA(A) multisubunit chloride channel receptors and metabotropic GABA(B) G protein-coupled receptors. Evidence suggests that pharmacologically distinct GABA(B) receptor subtypes mediate presynaptic inhibition of neurotransmitter release by reducing Ca2+ conductance, and postsynaptic inhibition of neuronal excitability by activating inwardly rectifying K+ (Kir) conductance. However, the cloning of GABA(B) gb1 and gb2 receptor genes and identification of the functional GABA(B) gb1-gb2 receptor heterodimer have so far failed to substantiate the existence of pharmacologically distinct receptor subtypes. The anticonvulsant, antihyperalgesic, and anxiolytic agent gabapentin (Neurontin) is a 3-alkylated GABA analog with an unknown mechanism of action. Here we report that gabapentin is an agonist at the GABA(B) gb1a-gb2 heterodimer coupled to Kir 3.1/3.2 inwardly rectifying K+ channels in Xenopus laevis oocytes. Gabapentin was practically inactive at the human gb1b-gb2 heterodimer, a novel human gb1c-gb2 heterodimer and did not block GABA agonism at these heterodimer subtypes. Gabapentin was not an agonist at recombinant GABA(A) receptors as well. In CA1 pyramidal neurons of rat hippocampal slices, gabapentin activated postsynaptic K+ currents, probably via the gb1a-gb2 heterodimer coupled to inward rectifiers, but did not presynaptically depress monosynaptic GABA(A) inhibitory postsynaptic currents. Gabapentin is the first GABA(B) receptor subtype-selective agonist identified providing proof of pharmacologically and physiologically distinct receptor subtypes. This selective agonism of postsynaptic GABA(B) receptor subtypes by gabapentin in hippocampal neurons may be its key therapeutic advantage as an anticonvulsant.

Acetates↗
PubMed2000

Galileo at Io: results from high-resolution imaging.

During late 1999/early 2000, the solid state imaging experiment on the Galileo spacecraft returned more than 100 high-resolution (5 to 500 meters per pixel) images of volcanically active Io. We observed an active lava lake, an active curtain of lava, active lava flows, calderas, mountains, plateaus, and plains. Several of the sulfur dioxide-rich plumes are erupting from distal flows, rather than from the source of silicate lava (caldera or fissure, often with red pyroclastic deposits). Most of the active flows in equatorial regions are being emplaced slowly beneath insulated crust, but rapidly emplaced channelized flows are also found at all latitudes. There is no evidence for high-viscosity lava, but some bright flows may consist of sulfur rather than mafic silicates. The mountains, plateaus, and calderas are strongly influenced by tectonics and gravitational collapse. Sapping channels and scarps suggest that many portions of the upper approximately 1 kilometer are rich in volatiles.

Extraterrestrial Environment↗
PubMed2000

Inhibition of protein kinase C(alpha) by dequalinium analogues: dependence on linker length and geometry.

Analogues of a bipartite compound, dequalinium (DECA) (quinolinium, 1,1'-(1,10-decanediyl)bis(4-amino-2-methyl diiodide)), were tested for inhibition of protein kinase C(alpha) (PKC(alpha)). In vitro assays of monomeric and dimeric analogues support a model in which DECA inhibits PKC(alpha) by an obligatory two-point contact, a unique mechanism among PKC inhibitors. The presence of unsaturation in the center of the C(10)-alkyl linker produced geometric isomers with different inhibitory potencies: cis IC(50) = 52 +/- 12 microM and trans IC(50) = 12 +/- 3 microM, where the trans isomer was equipotent to that of the saturated C(10)-DECA. DECA analogues with longer, saturated linkers (C(12), C(14), or C(16)) exhibited enhanced inhibitory potencies which reached a plateau with the C(14)-linker (IC(50) = 2.6 +/- 0.2 microM). Metastatic melanoma cells treated with 250 nM C(12)-, C(14)-, or C(16)-DECA and irradiated with long-wave UV light (which causes irreversible inhibition of PKC(alpha) by DECA) confirmed the linker-dependent inhibition of intracellular PKC(alpha) activity.

Animals↗
PubMed2000

North-south geological differences between the residual polar caps on Mars.

Polar processes can be sensitive indicators of global climate, and the geological features associated with polar ice caps can therefore indicate evolution of climate with time. The polar regions on Mars have distinctive morphologic and climatologic features: thick layered deposits, seasonal CO2 frost caps extending to mid latitudes, and near-polar residual frost deposits that survive the summer. The relationship of the seasonal and residual frost caps to the layered deposits has been poorly constrained, mainly by the limited spatial resolution of the available data. In particular, it has not been known if the residual caps represent simple thin frost cover or substantial geologic features. Here we show that the residual cap on the south pole is a distinct geologic unit with striking collapse and erosional topography; this is very different from the residual cap on the north pole, which grades into the underlying layered materials. These findings indicate that the differences between the caps are substantial (rather than reflecting short-lived differences in frost cover), and so support the idea of long-term asymmetry in the polar climates of Mars.

Carbon Dioxide↗
PubMed2000

Reporting child abuse: pediatricians' experiences with the child protection system.

OBJECTIVE: To explore pediatricians' reluctance to adhere to a legal mandate to report suspected child abuse and to compare their perspectives with those described in other jurisdictions in studies informed by different theories and employing diverse methodologies. METHOD: An anonymous survey questionnaire was distributed to all pediatricians with admitting privileges to B.C. Children's Hospital which has a specialized Child Protection Services Unit. Physicians were asked about their own prior reporting experience and the influence of that experience upon future reporting decisions. They were also asked why they think other physicians might be reluctant to report (N = 26). RESULTS: While respondents were generally positive about their experience in filing a report to Child Protective Services, they were less positive about the dearth of feedback they received and they were undecided as to whether there was a positive outcome to their report. Their perceptions of the reasons colleagues might fail to report emphasized dissatisfaction with CPS, concern with loss of relationship with the child's parents, and a desire to avoid court proceedings. CONCLUSIONS: Physicians' noncompliance or at least dissatisfaction with their child abuse reporting mandate is an international problem that can not be addressed by institutional protocols alone. Confusions about the limitations on confidentiality and information sharing might be better addressed through more shared training and opportunities for collaboration than by legal mandates and institutional protocols alone. Organizational theory and exchange theory might inform future research about the conditions under which collaboration is enhanced in the service of protecting children.

Adult↗
PubMed2000

A comparison of 12- and 15-lead ECGS in ED chest pain patients: impact on diagnosis, therapy, and disposition.

The objective of this study was to investigate the diagnostic and therapeutic impact of the 15-lead electrocardiogram (15ECG) on the emergency department (ED) management of chest pain (CP) patients. The design was prospective use of 15ECG with real-time physician survey and retrospective comparison to 12-lead ECG (12ECG). The study took place in a University hospital ED. Adult CP patients participated. During the 15ECG period (June 1996 to July 1996), 595 patients (92% of CP patients) had 15ECG analysis. Diagnoses of acute coronary ischemic syndromes (ACIS) were as follows: 13 acute myocardial infarction (AMI, 7 anterior [ANT], 5 inferior [INF], 1 lateral [LAT], 2 posterior [POST], 1 right ventricular [RV]) and 136 unstable angina (USA) with 47% exhibiting ECG abnormality; the 2 POST and 1 RV AMI occurred in the setting of coexisting INF AMI. The following management strategies were used: 6 fibrinolytic therapy (TT), 4 primary angioplasty (PTCA), 67 rule-out myocardial infarction (ROMI), and 144 admission to critical care unit (CCU). During the 12ECG period (June 1995 to July 1995), 599 patients were encountered. The diagnoses of ACIS were as follows: 11 AMI (5 ANT, 4 INF, 2 LAT) and 146 USA with 51% exhibiting ECG abnormality (P = NS for diagnostic comparisons to 15ECG). The following management strategies were used: 5 TT, 5 PTCA, 59 ROMI, and 137 admission to CCU (P = NS for all treatment comparisons to 15ECG). Of 15ECG cases 81% had completed real-time physician survey, showing that the diagnosis and management ACIS were not altered by the 15ECG; physicians felt, however, that the 15ECG provided a more complete anatomic picture of the ACIS. No false-positive cases of additional lead STE were noted in this investigation except in cases involving abnormal intraventricular conduction such as the bundle branch block scenario. The 15ECG provided a more complete description of myocardial injury without altering the ED diagnosis, ED-based therapy, or hospital disposition in adult CP patients. Further study is required to identify patient subset(s) which may benefit from the 15ECG.

Angina, Unstable↗