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.     

Identification of HLA-B44 subtypes associated with extended MHC haplotypes

The HLA class I antigen B44 is found in each of two different extended major histocompatibility haplotypes (allele combinations of HLA-B, HLA-DR, and complement genes BF, C2, C4A, and C4B in linkage disequilibrium). Using isoelectric focusing, two variants of HLA-B44 were identified. The basic variant was found in all cell lines with the extended haplotype HLA-B44, DR7, FC31, and the acidic variant in all cell lines with the extended haplotype HLA-B44, DR4, SC30. The occurrence of each antigen variant with a unique extended haplotype explains previous observations concerning the nonrandom association of B44 variants with DR antigens.     

Affinity labeling of delta-opiate receptors using [D-Ala2,Leu5,Cys6]enkephalin. Covalent attachment via thiol-disulfide exchange

[D-Ala2,Leu5,Cys6]Enkephalin (DALCE) is a synthetic enkephalin analog which contains a sulfhydryl group. DALCE binds with high affinity to delta-receptors, with moderate affinity to mu-receptors, and with negligible affinity to kappa-receptors. Pretreatment of rat brain membranes with DALCE resulted in concentration-dependent loss of delta-binding sites. Using 2 nM [3H][D-Pen2,D-Pen5]enkephalin (where Pen represents penicillamine) to label delta-sites, 50% loss of sites occurred at about 3 microM DALCE. Loss of sites was not reversed by subsequent incubation in buffer containing 250 mM NaCl and 100 microM guanyl-5'-yl imidodiphosphate (Gpp(NH)p), conditions which cause dissociation of opiate agonists. By contrast, the enkephalin analogs [D-Ala2,D-Leu5]enkephalin, [D-Ser2,Leu5,Thr6]enkephalin, [D-Pen2,D-Pen5]enkephalin, and [D-Ala2,D-Leu5,Lys6]enkephalin were readily dissociated by NaCl and Gpp(NH)p, producing negligible loss at 3 microM. This suggests that DALCE binds covalently to the receptors. Pretreatment of membranes with the reducing agents dithiothreitol and beta-mercaptoethanol had no effect on opiate binding. Thus, loss of sites required both specific recognition by opiate receptors and a thiol group. The irreversible effect of DALCE was completely selective for delta-receptors. Pretreatment with DALCE had no effect on binding of ligands to mu- or kappa-receptors. The effect of DALCE on delta-binding was: 1) markedly attenuated by inclusion of dithiothreitol in the preincubation buffer, 2) partially reversed by subsequent incubation with dithiothreitol, 3) slightly enhanced when converted to the disulfide-linked dimer, and 4) prevented by blocking the DALCE sulfhydryl group with N-ethylmaleimide or iodoacetamide. These results indicate that DALCE binds covalently to delta-receptors by forming a disulfide bond with a sulfhydryl group in the binding site. The mechanism may involve a thiol-disulfide exchange reaction.     

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.     

Diffusional dynamics of an active rhodamine-labeled 1,4-dihydropyridine in sarcolemmal lipid multibilayers.

A "membrane bilayer pathway" model, involving ligand partition into the bilayer, lateral diffusion, and receptor binding has been invoked to describe the 1,4-dihydropyridine (DHP) calcium channel antagonist receptor binding mechanism. In an earlier study (Chester et al. 1987. Biophys. J. 52:1021-1030), the diffusional component of this model was examined using an active fluorescence labeled DHP calcium channel antagonist, nisoldipine-lissamine rhodamine B (Ns-R), in purified cardiac sarcolemmal (CSL) lipid multibilayers. Diffusion coefficient measurements on membrane-bound drug and phospholipid at maximum bilayer hydration yielded similar values (3.8 x 10(-8) cm2/s). However, decreases in bilayer hydration resulted in dramatically reduced diffusion coefficient values for both probes with substantially greater impact on Ns-R diffusion. These data suggested that hydration dependent diffusional differences could be a function of relative probe location along the bilayer normal. In this communication, we have addressed the relative effect of the rhodamine substituent on Ns-R diffusion complex by examining the diffusional dynamics of free rhodamine B under the same conditions used to evaluate Ns-R complex and phospholipid diffusion. X-ray diffraction studies were performed to determine the Ns-R location in the membrane and model the CSL lipid bilayer profile structure to give a rationale for the differences in probe diffusional dynamics as a function of interbilayer water space.     

Antiviral T cell competence and restriction specificity of mixed allogeneic (P1 + P2----P1) irradiation chimeras

Mixed irradiation bone marrow chimeras were prepared by reconstituting lethally irradiated C57BL/10 (B10) or B10.D2 mice with T cell-depleted bone marrow cells of B10 plus B10.D2 origin. These chimeras were healthy and survived well under conventional housing conditions and after experimental laboratory infections. Of a total of 17 chimeras tested, 2 died spontaneously or from the injected virus. Twelve of fifteen chimeras mounted a measurable cytotoxic T cell response to virus. Despite approximately equal percentages of B10 and B10.D2 lymphocytes in chimeras, cytotoxic T cell responses to vaccinia virus and lymphocytic choriomeningitis virus were mediated variably by either syngeneic or allogeneic donor lymphocytes; thus the H-2 type of effector T cells frequently did not correspond to the 50:50 distribution of spleen or peripheral blood lymphocytes. Cytotoxic responses were restricted exclusively to recipient H-2 type. All mixed chimeras examined were able to mount a good IgG response to vesicular stomatitis virus. These results confirm previous data suggesting that such mixed chimeras are healthy and immunocompetent and demonstrate strict recipient-determined restriction specificity of effector T cells; they also suggest that if T help is necessary for induction of virus-specific cytotoxic T cells, it does not require host-restricted interactions between helper T cells and precursor cytotoxic T cells.