Müller glial dysfunction during diabetic retinopathy in rats is linked to accumulation of advanced glycation end-products and advanced lipoxidation end-products.

Aims/hypothesis: The impact of AGEs and advanced lipoxidation end-products (ALEs) on neuronal and Müller glial dysfunction in the diabetic retina is not well understood. We therefore sought to identify dysfunction of the retinal Müller glia during diabetes and to determine whether inhibition of AGEs/ALEs can prevent it.

Methods: Sprague-Dawley rats were divided into three groups: (1) non-diabetic; (2) untreated streptozotocin-induced diabetic; and (3) diabetic treated with the AGE/ALE inhibitor pyridoxamine for the duration of diabetes. Rats were killed and their retinas were evaluated for neuroglial pathology. Results: AGEs and ALEs accumulated at higher levels in diabetic retinas than in controls (p<0.001). AGE/ALE immunoreactivity was significantly diminished by pyridoxamine treatment of diabetic rats. Diabetes was also associated with the up-regulation of the oxidative stress marker haemoxygenase-1 and the induction of glial fibrillary acidic protein production in Müller glia (p<0.001). Pyridoxamine treatment of diabetic rats had a significant beneficial effect on both variables (p<0.001). Diabetes also significantly altered the normal localisation of the potassium inwardly rectifying channel Kir4.1 and the water channel aquaporin 4 to the Müller glia end-feet interacting with retinal capillaries. These abnormalities were prevented by pyridoxamine treatment.

Conclusions/interpretation: While it is established that AGE/ALE formation in the retina during diabetes is linked to microvascular dysfunction, this study suggests that these pathogenic adducts also play a role in Müller glial dysfunction.

Barriers to increasing fruit and vegetable intakes in the older population of Northern Ireland: low levels of liking and low awareness of current recommendations

Objective: To investigate barriers to increasing fruit and vegetable (f + v) intakes in a large sample of the older population of Northern Ireland (NI), in relation to current intakes.

p63 maintains keratinocyte proliferative capacity through regulation of Skp2-p130 levels

p63 is a master regulator of proliferation and differentiation in stratifying epithelia, and its expression is frequently altered in carcinogenesis. However, its role in maintaining proliferative capacity remains unclear. Here, we demonstrate that hypoproliferation and loss of differentiation in organotypic raft cultures of primary neonatal human foreskin keratinocytes (HFKs) depleted of the a and ß isoforms of p63 result from p53-p21-mediated accumulation of retinoblastoma (Rb) family member p130. Hypoproliferation in p63-depleted HFKs can be rescued by depletion of p53, p21(CIP1) or p130. Furthermore, we identified the gene encoding S-phase kinase-associated protein 2 (Skp2), the recognition component of the SCF(Skp2) E3 ubiquitin ligase, as a novel target of p63, potentially influencing p130 levels. Expression of Skp2 is maintained by p63 binding to a site in intron 2 and mRNA levels are downregulated in p63-depleted cells. Hypoproliferation in p63-depleted cells can be restored by re-expression of Skp2. Taken together, these results indicate that p63 plays a multifaceted role in maintaining proliferation in the mature regenerating epidermis, in addition to being required for differentiation.

GPVI levels in platelets: relationship to platelet function at high shear

We have investigated the density of the collagen receptors glycoprotein VI (GPVI) and alpha(2)beta(1) on human platelets and their relationship to polymorphisms within the GPVI gene. GPVI levels varied 1.5-fold and showed a weak correlation (r = 0.35) with the levels of alpha(2)beta(1), which varied 3-fold. GPVI genotype had a significant effect on receptor levels with carriers of the proline 219 allele (approximately 22% of the population) having 10% lower GPVI levels than the more common serine homozygotes. GPVI and alpha(2)beta(1) levels were found to be significantly decreased on platelets from patients with myeloproliferative disorders (MPDs). In both the MPD and the control group, GPVI levels were found not to affect platelet function under high shear in whole blood. Similarly murine platelets that express up to 5-fold lower levels of GPVI showed no significant difference than controls in thrombus formation on a high-density collagen-coated surface. However platelets lacking the GPVI/Fc receptor gamma-chain (FcR gamma-chain) complex or a functional FcR gamma-chain (immunoreceptor tyrosine-based activation motif [ITAM] point mutant) exhibited severely abrogated thrombus formation at 800 s(-1) and 1500 s(-1). These results demonstrate that GPVI levels are tightly controlled and play a critical role in thrombus formation on collagen; nevertheless, a range of receptor densities can support platelet function under high shear. (C) 2003 by The American Society of Hematology.

Differential effects of reduced glycoprotein VI levels on activation of murine platelets by glycoprotein VI ligands

We have investigated the effects of decreased levels of the complex between glycoprotein VI (GPVI) and the Fc receptor gamma-chain (FcRgamma) on responses to collagen and GPVI-specific ligands in murine platelets. We show that levels of GPVI-FcRgamma of the order of 50 % and 20 % of wild-type levels caused 2- and 5-fold shifts to the right respectively in the dose-response curve for aggregation in response to collagen, the snake toxin convulxin and the monoclonal antibody JAQ1. In addition, there is a delay in the onset of aggregation in response to collagen. In contrast, the stimulation of protein tyrosine phosphorylation by collagen (as measured after 150 s) and adhesion to a collagen-coated surface under static conditions were unaffected in platelets with 50 % and 20 % of wild-type levels of GPVI. In contrast, responses to a collagen-related peptide (CRP), made up of repeat glycine-proline-hydroxyproline motifs, were markedly inhibited and abolished in platelets expressing 50 % and 20 % of wild-type levels of GPVI respectively. We suggest that the marked effect of a reduction in GPVI levels on the CRP-induced activation of platelets is due to the multivalent nature of CRP and the fact that GPVI is its sole receptor on platelets. Thus it appears that the interaction of CRP with GPVI is determined by a combination of affinity and avidity. The observation that collagen does not behave like CRP in platelets expressing reduced levels of GPVI, even in the combined presence of blocking antibodies against integrin alpha2beta1 and GPV, suggests that collagen has a greater affinity than CRP for GPVI, and/or that other receptors are involved in its binding to platelets. The clinical significance of these results is discussed.

Levels of Neonatal Thyroid Hormone in Preterm Infants and Neurodevelopmental Outcome at 51/2 Years: Millennium Cohort Study

Context: Transient hypothyroxinemia is the commonest thyroid dysfunction of premature infants, and recent studies have found adverse associations with neurodevelopment. The validity of these associations is unclear because the studies adjusted for a differing range of factors likely to influence neurodevelopment. Objective: The aim was to describe the association of transient hypothyroxinemia with neurodevelopment at 5.5 yr corrected age. Design: We conducted a follow-up study of a cohort of infants born in Scotland from 1999 to 2001 =34 wk gestation. Main Outcome Measures: We measured scores on the McCarthy scale adjusted for 26 influences of neurodevelopment including parental intellect, home environment, breast or formula fed, growth retardation, and use of postnatal drugs. Results: A total of 442 infants =34 wk gestation who had serum T4 measurements on postnatal d 7, 14, or 28 and 100 term infants who had serum T4 measured in cord blood were followed up at 5.5 yr. Infants with hypothyroxinemia (T4 level = 10th percentile on d 7, 14, or 28 corrected for gestational age) scored significantly lower than euthyroid infants (T4 level greater than the 10th percentile and less than the 90th percentile on all days) on all McCarthy scales, except the quantitative. After adjustment for confounders of neurodevelopment, hypothyroxinemic infants scored significantly lower than euthyroid infants on the general cognitive and verbal scales. Conclusions: Our findings do not support the view that the hypothyroxinemic state, in the context of this analysis, is harmless in preterm infants. Many factors contribute both to the etiology of hypothyroxinemia and neurodevelopment; strategies for correction of hypothyroxinemia should acknowledge its complex etiology and not rely solely on one approach.

Holothurians – potential biomonitors for metal levels in deep-sea sediments

Levels of cadmium, copper, aluminium, iron, manganese and zinc (Cd, Cu, Al, Fe, Mn and Zn) in deep-sea sediments and sediment-feeding holothurians are reported. Profiles of metals were found to vary with depth in the sediment. Holothurian foregut metal values are generally closer to surficial sediment levels, whereas body wall levels, with the exception of Cd and Cu, were generally lower. Holothurians are presented as potential biomonitors of metal concentrations in the deep-sea floor environment.

Altered stress stimulation of inward rectifier potassium channels in Andersen-Tawil syndrome

Inward rectifier potassium channels of the Kir2 subfamily are important determinants of the electrical activity of brain and muscle cells. Genetic mutations in Kir2.1 associate with Andersen-Tawil syndrome (ATS), a familial disorder leading to stress-triggered periodic paralysis and ventricular arrhythmia. To identify the molecular mechanisms of this stress trigger, we analyze Kir channel function and localization electrophysiologically and by time-resolved confocal microscopy. Furthermore, we employ a mathematical model of muscular membrane potential. We identify a novel corticoid signaling pathway that, when activated by glucocorticoids, leads to enrichment of Kir2 channels in the plasma membranes of mammalian cell lines and isolated cardiac and skeletal muscle cells. We further demonstrate that activation of this pathway can either partly restore (40% of cases) or further impair (20% of cases) the function of mutant ATS channels, depending on the particular Kir2.1 mutation. This means that glucocorticoid treatment might either alleviate or deteriorate symptoms of ATS depending on the patient's individual Kir2.1 genotype. Thus, our findings provide a possible explanation for the contradictory effects of glucocorticoid treatment on symptoms in patients with ATS and may open new pathways for the design of personalized medicines in ATS therapy. © FASEB.