Compliance with postpartum Rh isoimmunization prophylaxis in Alberta

A retrospective review of obstetric records for 1979 in two major Calgary hospitals was undertaken to determine the rate of compliance with postpartum Rh isoimmunization prophylaxis in Alberta. The charts of 4528 women ranging in age from 13 to 46 years were reviewed. The prevalence rate of Rh negativity was found to be 16%. Of the 710 Rh-negative women 490 (69%) were eligible to receive Rh immune globulin (RhIG); that is, they had no anti-D antibodies, and the baby/fetus was Rh-positive or Rh-unknown. RhIG had been administered to 93.6% of the eligible women; the compliance rate ranged from 66.7% for obstetric emergencies (i.e., spontaneous abortion, antepartum or early-pregnancy hemorrhage, or ectopic pregnancy) to 98.2% for postpartum diagnoses. In more than half (54.7%) of the women who underwent amniocentesis Rh type was not determined; the implications of this finding are discussed. Although poor compliance with postpartum RhIG administration is not a reason for withholding antepartum administration of RhIG, maximum compliance with the more cost-effective programs should be attained before antepartum programs are fully implemented.     

Changes in quality of stallion spermatozoa during cryopreservation: Plasma membrane integrity and motion characteristics

Better procedures for freezing and thawing equine sperm are needed since variable fertility is obtained when cryopreserved sperm are used. To evaluate current methods of freezing equine sperm, we examined spermatozoal quality by means of two new techniques. These measured the integrity of plasma-acrosomal membranes by immunofluorescent analyses of binding of an antibody specific to the acrosome and evaluated eight parameters of spermatozoal motion using a fully automated computerized system. Five ejaculates from each of eight stallions were processed for freezing in egg yolk-lactose extender with 4% glycerol. Spermatozoal quality was assessed at four different points: at less than 15 min after collecting and before processing (Step 1); after centrifugation and just before freezing (Step 2); immediately after thawing less than 3 h after freezing (Step 3); and immediately after thawing 10 to 20 d after freezing (Step 4). Acrosome-specific monoclonal antibody detected differences (P <0.05) among steps and ejaculates within stallions. All parameters of spermatozoal motion, including the percentage of motile sperm, percentage of progressively motile sperm, curvilinear velocity, straight line velocity, linearity, amplitude of lateral head displacement, and radius of the average path for circularly swimming sperm, differed (P <0.05) among steps, and most of these parameters differed among ejaculates within a stallion and among stallions. For Steps 2 and 3, 62 and 37% of the sperm were motile, and 56 and 23% of all motile sperm had a curvilinear velocity of >100 mum/sec. Most damage to sperm occurred as a result of freezing-thawing, whereas centrifugation of sperm caused only minor damage.     

Bone morphometrics and tetracycline marking patterns in young growing American alligators (Alligator mississippiensis)

Nine young American alligators (Alligator mississippiensis) were injected at monthly intervals with tetracycline to determine the bone apposition rate and the resorption patterns over a 3-mo period. The periosteal apposition rate increased progressively over the 3-mo period from 2.99 microns/day to 5.94 microns/day. Endosteal apposition rate was much slower with incomplete tetracycline lines being observed on the endosteum. This suggests that most modeling-resorptive activities occur on the endosteal envelope.     

Cyclic AMP induces rapid increases in gap junction permeability and changes in the cellular distribution of connexin43

The rapid effects of cAMP on gap junction-mediated intercellular communication were examined in several cell types which express different levels of the gap junction protein, connexin43 (Cx43), including immortalized rat hepatocyte and granulosa cells, bovine coronary venular endothelial cells, primary rat myometrial and equine uterine epithelial cells. Functional analysis of changes in junctional communication induced by 8-bromo-cAMP was monitored by a fluorescence recovery after photobleaching assay in subconfluent cultures in the presence or absence of 1.0 mm 1-octanol (an agent which uncouples cells by closing gap junction channels). Communicating cells treated with 1.0 mm 8-bromo-cAMP alone exhibited significant increases in the percent of fluorescence recovery which were detected within 1–3 min depending on cell type, and junctional communication remained significantly elevated for up to 24 hr. Addition of 1.0 mm 8-bromo-cAMP to cultured cells, which were uncoupled with 1.0 mm octanol for 1 min, exhibited partial restoration of gap junctional permeability beginning within 3–5 min. Identical treatments were performed on cultures that were subsequently processed for indirect immunofluorescence to monitor Cx43 distribution. The changes in junctional permeability of cells correlated with changes in the distribution of immunoreactive Cx43. Cells treated for 2 hr with 10 m monensin exhibited a reduced communication rate which was accompanied by increased vesicular cytoplasmic Cx43 staining and reduced punctate surface staining of junctional plaques. Addition of 1.0 mm 8-bromo-cAMP to these cultures had no effect on the rate of communication or the distribution of Cx43 compared to cultures treated with monensin alone. These data suggest that an effect of cyclic AMP on Cx43 gap junctions is to promote increases in gap junctional permeability by increasing trafficking and/or assembly of Cx43 to plasma membrane gap junctional plaques.We acknowledge the technical assistance of Richard Lewis and Meghan Abella. We thank Dr. Hugh Dookwah for contributions to the myometrial cell isolation protocol and Drs. Stephen H. Safe, Timothy D. Phillips, and Evelyn Tiffany-Castiglioni for helpful discussions. This work was funded by NIH (HD-26182, P42-ES04917, ES05871-01A1), the March of Dimes Birth Defects Foundation Basic Research grant #1-0796, and USDA 92-37203-7952.     

Social organization of confined male collared lemmings (Dicrostonyx groenlandicus Traill)

This study describes and quantifies the social organization and agonistic behaviour of adult male collared lemmings (Dicrostonyx groenlandicus) in an indoor enclosure. Five groups of four lemmings were observed during separate 8-day tests. Experimental design permitted an increase in group size while density remained constant. When in groups of two, males established stable dominant-subordinate relationships and, when in groups of four, they formed stable non-linear dominance hierarchies. For either group size, frequency of agonistic behaviour declined over time. Habituation, formation of dominance relationships and spatio-temporal partitioning of space are suggested as possible explanations of this decline. Percentage of initial body weight lost during the experiments varied inversely with social rank. Possible causes of this weight loss are discussed. Social relationships in this study are discussed in light of field observations of male lemming home range distribution.     

Interaction Energy Analysis of Nonclassical Antifolates with Human Dihydrofolate Reductase

The x-ray structure of the PTX:NADPH:L22F human mutant DHFR ternary complex was used as a structural template to generate structural models for the following wild type DHFR complexes: PTX:DHFR:NADPH, TMP:DHFR:NADPH, EPM:DHFR:NADPH, and TMQ:DHFR:NADPH. Each of these complexes were subsequently modeled in a 60 Å cube of explicit water and minimized to a rms gradient of from 1.0-3.0·10^-5 kcal·Å^-1. For each complex, interaction energies were calculated for the antifolate interaction with each of the following: the DHFR binding site residues, the entire DHFR protein, the solvated complex (containing DHFR, NADPH, and solvent water), water alone, and NADPH. Additionally, each antifolate was subdivided into distinct substructural regions and interaction energy calculations were performed in order to evaluate their contributions to overall antifolate interaction. Each antifolate showed its most stable interaction with the solvated complex. Substructural regions which consisted of a nitrogen containing aromatic ring system contributed most to the stability of the antifolate interactions, while the hydrocarbon aromatic rings, methoxy, and ethoxy groups showed much less stable interaction energies. Since the different substructural regions of nonclassical antifolates differ in their contributions to overall antifolate binding, those substructural regions which exhibit relatively unfavorable interaction energies may constitute important targets in the design of improved DHFR inhibitors.     

MHC gene configuration variation in geographically disparate populations of California sea lions (Zalophus californianus)

Major histocompatibility complex (MHC) class II DRB genotypes were examined in two geographically isolated populations of California sea lions (Zalophus californianus) (Gulf of California and California coastal Pacific Ocean). Genomic DNA from 227 California sea lions was examined using eight sequence-specific primer (SSP) pairs flanking the putative peptide-binding site. A total of 40 different Zaca-DRB genotype configurations were identified among the 227 individuals. Using SSP-PCR, significant differences were found between coastal California and Gulf of California Zalophus populations in numbers of DRB sequences per individual and configuration of sequences within individuals. Additionally, unique local patterns of MHC diversity were identified among the Midriff Island animals. These population differences are consistent with either ecologically distinct patterns of selection pressures and/or geographical isolation. The consequences of these partitioned MHC configurations at the population level are as yet unknown; however, the worldwide increase in emerging marine diseases lends urgency to their examination.     

Deficiency of PTP1B in POMC neurons leads to alterations in energy balance and homeostatic response to cold exposure

The adipose tissue-derived hormone leptin regulates energy balance through catabolic effects on central circuits, including proopiomelanocortin (POMC) neurons. Leptin activation of POMC neurons increases thermogenesis and locomotor activity. Protein tyrosine phosphatase 1B (PTP1B) is an important negative regulator of leptin signaling. POMC neuron-specific deletion of PTP1B in mice results in reduced high-fat diet-induced body weight and adiposity gain due to increased energy expenditure and greater leptin sensitivity. Mice lacking the leptin gene (ob/ob mice) are hypothermic and cold intolerant, whereas leptin delivery to ob/ob mice induces thermogenesis via increased sympathetic activity to brown adipose tissue (BAT). Here, we examined whether POMC PTP1B mediates the thermoregulatory response of CNS leptin signaling by evaluating food intake, body weight, core temperature (T(C)), and spontaneous physical activity (SPA) in response to either exogenous leptin or 4-day cold exposure (4°C) in male POMC-Ptp1b-deficient mice compared with wild-type controls. POMC-Ptp1b(-/-) mice were hypersensitive to leptin-induced food intake and body weight suppression compared with wild types, yet they displayed similar leptin-induced increases in T(C). Interestingly, POMC-Ptp1b(-/-) mice had increased BAT weight and elevated plasma triiodothyronine (T(3)) levels in response to a 4-day cold challenge, as well as reduced SPA 24 h after cold exposure, relative to controls. These data show that PTP1B in POMC neurons plays a role in short-term cold-induced reduction of SPA and may influence cold-induced thermogenesis via enhanced activation of the thyroid axis.