Functional development of the visual system in normal and protein-deprived rats. VIM. Post-natal development of optic nerve axons

Protein deprivation results in a persistent impairment of transmission of impulses from the specific cortical projection within the visual cortex. In order to evaluate changes in subcortical structures during pre-weaning development, a study was made on number, calibre and myelination of optic nerve axons in control (C) and protein-deprived (PD) rats 5, 12, 20 and 30 days of age. Protein deprivation was induced by giving rat mothers a diet containing 7% protein by weight (control diet 14%, during gestation and lactation. The cross-sectional area of the nerve was measured on a digitizer. Between 3000 and 4000 axons collected from 7–13 field areas sampled by a random, systematic procedure from a cross-shaped area of the nerve were counted and measured. Between 5 and 12 days after birth, the number of axons was reduced by 50% in C rats. The total number of optic nerve axons was not significantly different in PD compared to C rats, indicating that protein deprivation does not affect the formation or naturally occurring nerve cell death of retinal ganglion cells. At all ages examined there were significant reductions in the number of axons larger than 0.52μm as well as the number of myelinated axons. The rates of growth/maturation and myelination of axons in C rats and PD rats suggested that the reductions seen in PD rats up to 20 days of age may represent a developmental delay of approximately 4 days. At 30 days, a delay or a distortion of development may be present. The retarded development of optic nerve axons is discussed in relation to delays, distortions and deficits during visual system maturation in protein-deprived rats.     

Diabetes in pregnancy: a new decade of challenges ahead

Every 10 years, the Diabetic Pregnancy Study Group, a study group of the EASD, conducts an audit meeting to review the achievements of the preceding decade and to set the directions for research and clinical practice improvements for the next decade. The most recent meeting focused on the following areas: improving pregnancy outcomes for women with pregestational type 1 diabetes and type 2 diabetes; the influence of obesity and gestational diabetes on pregnancy outcomes; the determinants and assessment of fetal growth and development; and public health issues, including consideration of transgenerational consequences and economic burden. The audit meeting also considered the likely impact of ‘omics’ on research within the field and the potential of these technologies to enable precision-medicine approaches to management. Through sharing of the findings and ideas of audit meeting participants, the DPSG hopes to promote networking, research and advances in clinical care, to improve outcomes for all women and their offspring affected by diabetes and obesity in pregnancy.     

Occult incontinence in women with pelvic organ prolapse - does it matter?

Objective

Many surgeons perform an anti-incontinence procedure during prolapse surgery in women in whom occult stress urinary incontinence has been demonstrated. Others prefer a two-step approach. It was the aim of the study to find out how many women really need a second operation and if a positive cough stress test with the prolapse reduced is associated with the development of stress urinary incontinence after prolapse surgery.

Methods

233 women were operated for primary or recurrent prolapse without complaining of SUI. Preoperatively, 53/233 women had a full urogynecological workup with the prolapse reduced. Postoperatively, if the patient suffered from stress urinary incontinence, an anti-incontinence surgery was performed.

Results

19/53 (35.8%) women who had a stress test with the prolapse reduced before surgery were defined as occult stress incontinent. Only 3 women (15.8%) of these 19 women developed symptoms of incontinence after prolapse surgery and had to be operated because of that. 18/233 (7.7%) complained of SUI 6 weeks to 6 months after surgery and received a TVT-tape.

Conclusion

The incidence of stress urinary incontinence manifesting after prolapse surgery is low in this study with 7.7%. This fact and the possible severe side effects of an incontinence operation justify a two-step approach if the patient is counseled and agrees. However, there is a small subgroup of women (3/19, 15.8%) with preoperative OSUI and SUI after surgery, who would benefit from a one-step approach. Further research is required to identify these women before surgical intervention.     

Improving outcome of malignant brain tumours in very young children: a population-based study in Finland during 1975-93

Abstract Sixty-four children with malignant brain tumours diagnosed at less than 3 years of age were reported to the Finnish Cancer Registry from 1975 to 1993. The survival rate has improved significantly: the 5-year survival rate was 26% for all children, 13% for children diagnosed during 1975-85 ( n = 30) and 40% for those diagnosed during 1986-93 ( n = 4). Of the surviving children in 1986-93, 43% were categorized in Bloom's group I or II and could lead active lives without major disabilities. The remaining children had severe neurologic late complications, such as hemiplegia, intractable seizures, and mental retardation.     

CD63 is a component of Weibel-Palade bodies of human endothelial cells

Weibel-Palade bodies are secretory granules of vascular endothelial cells specialized in the storage of von Willebrand factor (vWF) and P- selectin, two adhesion proteins that can be rapidly mobilized to the cell surface by exocytosis in response to thrombin or other agonists. In this study, we attempted to identify additional components of Weibel- Palade bodies by raising monoclonal antibodies to these granules, purified by cell fractionation. One antibody, 2C6, was found to be specific for CD63, a membrane glycoprotein previously described in the lysosomes of platelets and other cell types. The immunopurified 2C6 antigen was recognized by an anti-CD63 reference antibody, 2.28, by Western blotting. Also, the biosynthetic profile of the 2C6 antigen in endothelial cells showed a nascent molecular mass and a glycosylation pattern identical to that of CD63. Immunofluorescence staining with 2C6 showed the lysosomes, and also elongated structures identified as Weibel-Palade bodies by their shape, distribution, and positive staining with anti-vWF antibodies, CD63 was also found by Western blotting of subcellular fractions highly enriched in Weibel-Palade bodies. Our results indicate that CD63 colocalizes with vWF and P- selectin in the Weibel-Palade bodies of endothelial cells, and together with these adhesion proteins it could be rapidly expressed on the cell surface in areas of vascular injury and inflammation.     

Regulation of platelet production and function by megakaryocyte growth and development factor in nonhuman primates

The primary physiologic regulator of platelet production, Mpl ligand, has recently been cloned and characterized. To define the regulatory role of Mpl ligand on platelet production and function we measured the effects of a recombinant truncated human Mpl ligand, megakaryocyte growth and development factor (rHu-MGDF) on megakaryocytopoiesis, platelet function, and thrombogenesis in nonhuman primates. rHu-MGDF was administered to 10 baboons for 28 days while performing pharmacokinetics and repeated measurements of the following: (1) platelet count, volume, turnover, and function ex vivo and in vitro; (2) marrow megakaryocyte number, volume, and ploidy; and (3) platelet deposition and fibrin accumulation on segments of vascular graft and endarterectomized aorta in vivo. Daily subcutaneous injections of rHu- MGDF (5 microgram/kg/d) attained plasma concentrations averaging 1,300 +/- 300 pg/mL 2 hours after injection with trough levels of 300 +/- 65 pg/mL before the next dose. These levels of rHu-MGDF incrementally increased the peripheral platelet concentration threefold by day 7 and fivefold by day 28 (P < 10(-4)) associated with a reciprocal decrease of 25% in mean platelet volumes (P < 10(-3)). Platelet mass turnover, a steady-state measure of platelet production, increased fivefold (P < 10(-4)). Platelet morphology, life span, and recovery were normal. No significant change occurred in peripheral leukocyte, neutrophil, or erythrocyte counts (P > .1 in all cases). The platelet count gradually returned to baseline within 2 weeks after discontinuing rHu-MGDF infections. Marrow megakaryocyte volume doubled (P < 10(-3)) three days after initiating rHu-MGDF therapy and the modal ploidy shifted from 16N to 64N (P < 10(-4)). Marrow megakaryocyte number increased twofold by day 7, and nearly fourfold by day 28 (P < 10(-4)), resulting in a 6.5- fold increase in marrow megakaryocyte mass (P < 10(-3)). The effects of rHu-MGDF on thrombosis were determined by comparing baseline, day 5, and day 28 rHu-MGDF-treatment measurements of 111In-platelet deposition and 125I-fibrin accumulation on segments of homologous endarterectomized aorta (EA) and vascular graft (VG) interposed in arteriovenous femoral shunts. rHu-MGDF increased 111In-platelet deposition in direct proportion to the circulating concentration of platelets for both EA and VG (r=.98 in both cases), without significant changes in fibrin accumulation (P > .5 in both cases). During the first week of rHu-MGDF treatment ex vivo platelet aggregatory responsiveness was enhanced to physiologic agonists (adenosine diphosphate, collagen, and thrombin receptor agonist peptide, TRAP1-6) (P < .05 in all cases). Although in vitro platelet aggregation was not induced by any concentration of rHu-MGDF tested (P > .5), rHu-MGDF enhanced aggregatory responses to low doses of physiologic agonists, effects that were maximal at 10 ng/mL for baboon platelets and 100 ng/mL for human platelets, and were blocked by excess soluble c-Mpl receptor. Flow cytometric expression of platelet activation epitopes was not increased on resting platelets (ligand-induced binding sites, P- selectin, or Annexin V binding sites; P > .1 in all cases). Megakaryocyte growth and development factor regulates platelet production and function by stimulating endoreduplication and megakaryocyte formation from marrow progenitor cells, and transiently enhancing platelet functional responses ex vivo. rHu-MGDF has the potential for achieving platelet hemostatic protection with minimal thrombo-occlusive risk.     

Demonstration of single chain urokinase-type plasminogen activator on human platelet membrane

Fibrinolytic activity was found to be associated with sonicated platelet membranes after separation from cytosol by differential centrifugation. This fibrinolytic activity was attributed to the presence of a plasminogen activator, which was immunochemically identified as urinary-type plasminogen activator (uPA) by antibody neutralization assay, immunoblotting, and immunofluorescence. The molecular weight (mol wt) of this uPA was 54,000 and was present as the single chain form, although a small amount was detected in a higher mol wt complex indicative of a uPA-inhibitor complex. Treatment of membrane preparations with Triton X-100, 3 mol/L KCl, and 0.1 mol/L glycine, (pH 2.3), but not 10 mmol/L ethylenediamine tetraacetic acid (EDTA), removed the uPA from the membrane. This suggests that uPA is a peripheral membrane protein and that metal ions do not mediate protein- membrane association. Immunofluorescent staining revealed the presence of uPA on the outer surface of the platelet in preparations of intact unstimulated platelets. Thus, uPA is associated with the outer leaflet of the platelet membrane and may be involved with the acceleration of thrombus degradation observed with platelet-rich thrombi.