Long-term type 1 diabetes influences haematopoietic stem cells by reducing vascular repair potential and increasing inflammatory monocyte generation in a murine model.

AIMS/HYPOTHESIS: We sought to determine the impact of long-standing type 1 diabetes on haematopoietic stem/progenitor cell (HSC) number and function and to examine the impact of modulating glycoprotein (GP)130 receptor in these cells. METHODS: Wild-type, gp130(-/-) and GFP chimeric mice were treated with streptozotocin to induce type 1 diabetes. Bone marrow (BM)-derived cells were used for colony-formation assay, quantification of side population (SP) cells, examination of gene expression, nitric oxide measurement and migration studies. Endothelial progenitor cells (EPCs), a population of vascular precursors derived from HSCs, were compared in diabetic and control mice. Cytokines were measured in BM supernatant fractions by ELISA and protein array. Flow cytometry was performed on enzymatically dissociated retina from gfp(+) chimeric mice and used to assess BM cell recruitment to the retina, kidney and blood. RESULTS: BM cells from the 12-month-diabetic mice showed reduced colony-forming ability, depletion of SP-HSCs with a proportional increase in SP-HSCs residing in hypoxic regions of BM, decreased EPC numbers, and reduced eNos (also known as Nos3) but increased iNos (also known as Nos2) and oxidative stress-related genes. BM supernatant fraction showed increased cytokines, GP130 ligands and monocyte/macrophage stimulating factor. Retina, kidney and peripheral blood showed increased numbers of CD11b(+)/CD45(hi)/ CCR2(+)/Ly6C(hi) inflammatory monocytes. Diabetic gp130(-/-) mice were protected from development of diabetes-induced changes in their HSCs. CONCLUSIONS/INTERPRETATION: The BM microenvironment of type 1 diabetic mice can lead to changes in haematopoiesis, with generation of more monocytes and fewer EPCs contributing to development of microvascular complications. Inhibition of GP130 activation may serve as a therapeutic strategy to improve the key aspects of this dysfunction.

Retinal and choroidal microangiopathies: therapeutic opportunities.

Pathological angiogenesis in the retina and underlying choroid is a major cause of visual impairment in all age groups. The last decade has seen an explosion in the clinical availability of antiangiogenic compounds. Emphasis has been placed on inhibitors of the VEGF signaling pathway and considerable success has been achieved with aptamers and antibodies that bind VEGF. However, regression of neovascularization is rarely permanent and the regrowth of new vessels, often within a few months, requires multiple applications of drug. A number of antiangiogenic factors such as IGFBP3, SDF-1 blockers, PEDF, gamma-secretase, Delta-like ligand 4, and integrin antagonists have been identified, which act either indirectly on the VEGF system or independent of it. The importance of other candidates such as HIF-1alpha and protein kinase CK2, which act as "master" regulators of angiogenesis, offer realistic alternative targets for pharmacological intervention. The concept of combination therapy is rapidly gaining interest in the eye field and co-administration of two angiogenic agents (e.g., a CK2 inhibitor with a somatostatin analog, octreotide) are often significantly more effective at inhibiting retinal angiogenesis than either drug alone. The following review will discuss the current therapies available for aberrant ocular angiogenesis, consider new candidate targets for development of antiangiogenic compounds and emphasize the importance of combinatorial pharmacological agents in the treatment of such a dynamic cellular event as angiogenesis.

High sensitivity C-reactive protein in systemic lupus erythematosus: relation to disease activity, clinical presentation and implications for cardiovascular risk.

Measurement of high sensitivity C-reactive protein (hs-CRP), has been used in the assessment of disease activity in numerous rheumatic conditions including systemic lupus erythematosus (SLE). However, the utility of hs-CRP measurement in patients with lupus is uncertain. This study examined if hs-CRP can be used to assess disease activity, severity and cardiovascular risk in SLE. Serum samples from 601 visits of 213 SLE patients and 134 controls were analysed for hs-CRP by nephelometry. Detailed demographic data were obtained from all subjects and medication history and key laboratory parameters were collected. Disease activity was assessed using the SLEDAI. High sensitivity CRP was not associated with disease activity (SLEDAI), number of ACR SLE criteria or presence of any particular organ involvement. hs-CRP levels were significantly correlated with standard cardiovascular risk factors including body weight (P = 0.0002), hypertension (P = 0.001), and apolipoprotein A-I (P < 0.0001). Interestingly an inverse correlation was seen between hs-CRP levels and antimalarial use (P = 0.0018). Our results suggest that measurement of hs-CRP, though not valuable as marker of disease activity in SLE may be of some use in the assessment of cardiovascular risk. We speculate that antimalarials may help to reduce cardiovascular risk in patients with SLE.

Effect of pH, ionic charge, and osmolality on cytochrome c-mediated caspase-3 activity.

Cytochrome c-mediated activation of caspase-3 is the final common pathway for most signals that induce apoptosis. Before release of cytochrome c from mitochondria, K(+) and Cl(-) efflux and intracellular acidification must occur. We have utilized an in vitro assay to examine the role of pH, cations, anions, and uncharged molecules on the process of cytochrome c-mediated activation of procaspase-3. In this cell-free system, a pH above 7.4 severely suppressed the activation of procaspase-3 but not the activity of caspase-3. KCl, NaCl, and other salts all inhibited caspase activation, but uncharged molecules did not. Comparison of the inhibitory capacity of various salts suggests that the crucial element in causing suppression is the cation. The inhibition of alkaline pH could be overcome by increasing concentrations of cytochrome c, whereas the inhibition of ionic charge could not, suggesting that pH and salts affect the activation of caspase-3 by different mechanisms.

Disulfide bond formation during the folding of influenza virus hemagglutinin.

To study the importance of individual sulfhydryl residues during the folding and assembly in vivo of influenza virus hemagglutinin (HA), we have constructed and expressed a series of mutant HA proteins in which cysteines involved in three disulfide bonds have been substituted by serine residues. Investigations of the structure and intracellular transport of the mutant proteins indicate that (a) cysteine residues in the ectodomain are essential both for efficient folding of HA and for stabilization of the folded molecule; (b) cysteine residues in the globular portion of the ectodomain are likely to form native disulfide bonds rapidly and directly, without involvement of intermediate, nonnative linkages; and (c) cysteine residues in the stalk portion of the ectodomain also appear not to form intermediate disulfide bonds, even though they have the opportunity to do so, being separated from their correct partners by hundreds of amino acids including two or more other sulfhydryl residues. We propose a role for the cellular protein BiP in shielding the cysteine residues of the stalk domain during the folding process, thus preventing them from forming intermediate, nonnative disulfide bonds.

Clinical use of non-invasive method for determining oxygen saturation: ear oximetry.

Oxygen saturation can be estimated by measuring the absorption properties of the arterialized blood perfusing the ear. Simultaneous determination of oxygen saturation by blood gas and ear oximetry showed no statistical difference. This non-invasive method allows rapid and accurate monitoring of the oxygenation status of patients with respiratory insufficiency.

Gas exchange and arterial blood gas tensions to terbutaline sulfate in older patients with reversible airway obstruction.

Ten patients 55 years and older, known to have chronic obstructive airway disease with reversible component and who had required daily sympathomimetics for many years, were studied. At a clinically stable state in a sitting position, spirometry, ventilation, gas exchange, diffusion and arterial blood gas determinations were performed before and 10, 30, 60 and 120 minutes after administration of 0.5 mg of terbutaline sulfate by inhalation. Although changes were small, significant improvement in FEV1 remained even two hours after inhaling terbutaline. Increase of conducting airways reflected an increase of dead space for 120 minutes. Early rise of alveolar-arterial oxygen tension (PAO2-PaO2) with decreased arterial oxygen tension was observed. This was a transient phenomenon and not severe enough to warrant immediate therapy. No significant EKG or blood pressure changes were noted throughout the study. It appears that terbutaline, an effective long acting bronchodilator, can be safely administered to older patients with bronchospasms.

Sudden disappearance of electrocardiographic pattern of anteroseptal myocardial infarction. Result of superimposed acute posterior myocardial infarction.

In a 76-year-old man an electrocardiographic pattern of acute anteroseptal myocardial infarction disappeared suudenly. At necropsy, a more recent posterior myocardial infarct was found, in addition to an acute anteroseptal infarct. "Normalization" of the electrocardiogram from the pattern of anteroseptal myocardial infarction in this case resulted from the loss of opposing electromotive forces in the posterior wall because of posterior infarction.

Isoproterenol aerosols.

Notable acute improvement in pulmonary function in the majority of 32 patients with known bronchial asthma followed two inhalations of Isoproterenol. No deleterious clinical or cardiovascular effects of EKG changes (after one-half hour) were observed. A slight drop in blood pressure was noted in previously hypertensive patients. Isoproterenol's short duration of effects is ideal and one or two inhalations spaced four hours apart may be safely recommended for the relief of the acute asthma paroxysm.