Characterization of human platelet glycoprotein antigens giving rise to individual immunoprecipitates in crossed-immunoelectrophoresis

Washed human platelets were labeled with 125I by the lactoperoxidase- catalyzed method and solubilized in 1% Triton X-100. The soluble proteins were analyzed by crossed-immunoelectrophoresis in 1% agarose, employing a rabbit antibody raised against whole human platelets. Analysis of autoradiograms developed from dried agarose gels led to the establishment of a normal reference pattern that was consistent for platelets obtained from more than 50 normal individuals. Six platelet membrane glycoprotein antigens contained in four distinguishable precipitates were identified. Each identification was based on direct sodium dodecyl sulfate-polyacrylamide gel electrophoretic analysis of 125I-antigens contained in individually excised precipitates. These platelet antigens include major membrane glycoproteins previously designated la, lb, lla, llb, llla, and lllb. Glycoproteins llb and llla were shown to be contained in a single immunoprecipitate, while glycoproteins la and lla were routinely detected in a single different immunoprecipitate. Analysis of soluble proteins from platelets of five patients with Glanzmann's thrombasthenia demonstrated either a complete absence or a marked reduction of only one radiolabeled precipitate, that containing membrane glycoproteins llb and llla. Platelet samples from two patients with Bernard-Soulier syndrome were devoid of a different precipitate, that containing membrane glycoprotein lb.     

Current Outcomes of Emergency Large Bowel Surgery

Introduction

Emergency large bowel surgery (ELBS) is known to carry an increased risk of morbidity and mortality. Previous studies have reported morbidity and mortality rates up to 14.3%. However, there has not been a recent study to document the outcomes of ELBS following several major changes in surgical training and provision of emergency surgery. The aim of this study was therefore to explore the current outcomes of ELBS.

Methods

A retrospective review was performed of a prospectively maintained database of the clinical records of all patients who had ELBS between 2006 and 2013. Data pertaining to patient demographics, ASA (American Society of Anesthesiologists) grade, diagnosis, surgical procedure performed, grade of operating surgeon and assistant, length of hospital stay, postoperative complications and in-hospital mortality were analysed.

Results

A total of 202 patients underwent ELBS during the study period. The mean patient age was 62 years and the most common cause was colonic carcinoma (n=67, 33%). There were 32 patients (15.8%) who presented with obstruction and 64 (31.7%) had bowel perforation. The overall in-hospital mortality rate was 14.8% (n=30). A consultant surgeon was involved in 187 cases (92.6%) as either first operator, assistant or available in theatre.

Conclusions

ELBS continues to carry a high risk despite several major changes in the provision of emergency surgery. Further developments are needed to improve postoperative outcomes in these patients.     

Effects of luteal phase administration of mifepristone (RU486) and prostaglandin analogue or inhibitor on endometrium in the rhesus monkey

Early luteal phase administration of a potent anti-progestin like mifepristone (RU486) inhibits blastocyst implantation and the establishment of pregnancy without marked changes in menstrual cyclicity and ovarian steroid hormone profiles; however, the underlying mechanism is not very clear. In the present study, a hypothesis that prostaglandins (PG) are involved in the anti-gestatory action of luteal phase mifepristone was tested. Endometrial changes in rhesus monkeys were examined following luteal phase administration of mifepristone, a prostaglandin synthesis inhibitor (diclofenac) and a prostaglandin analogue (misoprostol) either alone or in combination. Twenty-five monkeys were randomly assigned to six groups: group 1 (n = 4), normal control group; group 2 (n = 4), mifepristone (2 mg, daily, s.c.) treated group; group 3 (n = 4), diclofenac (25 mg, daily, i.m.) treated group; group 4 (n = 4), misoprostol (100 microg, daily, oral) treated group; group 5 (n = 5), mifepristone and diclofenac (same dosages as for groups 2 and 3) treated group; group 6 (n = 4), mifepristone and misoprostol (same dosages as for groups 2 and 4) treated group. All treatments were given to monkeys on days 16-18 of mated cycles and endometrial tissue samples were collected on day 20. With diclofenac alone (group 3), marginal changes were observed in glandular, stromal and vascular compartments, and there were few apoptotic bodies in gland cells; partial inhibition and delay in implantation was earlier reported. Significantly higher oestrogen receptor expression in glandular epithelial cells as compared with all other treatment groups was found after treatment with misoprostol alone (group 4) and was associated with normal fecundity. The anti-nidatory action of luteal phase antiprogestin treatment alone or in combination with diclofenac or misoprostol was associated with altered endometrial histometric features characterized by glandular apoptosis, regression in secretory functions, decreased oedema, extravasation and a higher degree of stromal leukocytic infiltration. In these three groups (groups 2, 5 and 6) receptors for oestrogen and progesterone receptors were significantly higher in stromal cells, and lower in vascular cells, while glandular cells showed significantly higher progesterone receptors compared with the control group. The anti-nidatory activity of mifepristone and associated endometrial changes could not be accentuated or attenuated with co-administration of PGE or diclofenac, nor could these be mimicked by these agents alone.      

A study of subunit selectivity,mechanism and site of action of the delta selective compound 2 (DS2) at human recombinant and rodent native GABAA receptors

Background and Purpose

Most GABA_A receptor subtypes comprise 2α, 2β and 1γ subunit, although for some isoforms, a δ replaces a γ-subunit. Extrasynaptic δ-GABA_A receptors are important therapeutic targets, but there are few suitable pharmacological tools. We profiled DS2, the purported positive allosteric modulator (PAM) of δ-GABA_A receptors to better understand subtype selectivity, mechanism/site of action and activity at native δ-GABA_A receptors.

Experimental Approach

Subunit specificity of DS2 was determined using electrophysiological recordings of Xenopus laevis oocytes expressing human recombinant GABA_A receptor isoforms. Effects of DS2 on GABA concentration–response curves were assessed to define mechanisms of action. Radioligand binding and electrophysiology utilising mutant receptors and pharmacology were used to define site of action. Using brain-slice electrophysiology, we assessed the influence of DS2 on thalamic inhibition in wild-type and δ^0/0 mice.

Key Results

Actions of DS2 were primarily determined by the δ-subunit but were additionally influenced by the α, but not the β, subunit (α4/6βxδ > α1βxδ >> γ2-GABA_A receptors > α4β3). For δ-GABA_A receptors, DS2 enhanced maximum responses to GABA, with minimal influence on GABA potency. (iii) DS2 did not act via the orthosteric, or known modulatory sites on GABA_A receptors. (iv) DS2 enhanced tonic currents of thalamocortical neurones from wild-type but not δ^0/0 mice.

Conclusions and Implications

DS2 is the first PAM selective for α4/6βxδ receptors, providing a novel tool to investigate extrasynaptic δ-GABA_A receptors. The effects of DS2 are mediated by an unknown site leading to GABA_A receptor isoform selectivity.