Impact of the Balanced School Day on Glycemic Control in Children with Type 1 Diabetes.

OBJECTIVE: The balanced school day (BSD) is an alternative elementary school schedule whereby children have 2 20-minute eating periods instead of 1 20-minute lunch, as is found in the traditional schedule (TS). We assessed the glycated hemoglobin (A1C) levels of children with type 1 diabetes in the TS vs. the BSD because 2 eating periods have the potential to impact blood glucose control. METHODS: A1C levels representative of the summer months (SumA1C) and A1C levels occurring at least 3 months after the start of the school year (SchA1C) were obtained retrospectively. A parental survey of perceptions of lunch planning, activity levels and diabetes management at school was also completed. RESULTS: Our sample included 97 students (TS=42, BSD=55). The mean age ± SD was 10.9±2.6 and 10.1±2.8 years in the TS and BSD, respectively (p=0.12). Sex distribution was not statistically different; 54% were female in TS vs. 36% in BSD; p=0.08. SumA1C was similar in the 2 groups (TS: 8.3±1.1% vs. BSD: 8.0±0.8%; p=0.08). There was a significant within-group increase from SumA1C to SchA1C in the BSD group only (p=0.001), with mean A1C values increasing from 8.0%±0.8% to 8.5%±1.0% in the BSD group compared to no significant increase in the TS group. Parental perceptions of lunch planning, physical activity and diabetes management were similar, regardless of school schedule. CONCLUSIONS: Children with type 1 diabetes in the BSD appear to have worse diabetes control during the school year compared to the summer, which is not evident in children in the TS. Additional school supports may assist students in the BSD.

Meta-iodobenzylguanidine, an inhibitor of arginine-dependent mono(ADP-ribosyl)ation, prevents neointimal hyperplasia.

The association of ADP-ribosylation with cell proliferation and ischemia-reperfusion injury suggests that it may be a suitable target for therapeutic control of revascularization-induced injury. The purpose of this study was to investigate the inhibitory actions of ADP-ribosylation inhibitors on restenosis. In organ culture, the poly(ADP-ribose) polymerase (PARP) inhibitor 3,4-dihydro-5-methylisoquinolinone (PD128763) was unable to prevent neointimal hyperplasia, whereas the arginine-dependent mono(ADP-ribosyl)transferase (ART) inhibitor meta-iodobenzylguanidine (MIBG) was highly effective (EC(50) 21 microM). Treatment with 3-aminobenzamide (3AB), a less potent ART inhibitor, also produced a significant reduction in neointimal hyperplasia. Single doses (25 mM) of MIBG and 3AB were also applied within a fibrin coagulum directly to the adventitial surface of the porcine femoral artery after balloon catheter injury in vivo. MIBG reduced the neointimal index, measured 14 days after angioplasty, by 82%, whereas 3AB was ineffective. However, when extended to 45 days, the neointimal index was not significantly decreased by MIBG treatment relative to control. Assessment of MIBG release from the fibrin glue showed that the bulk of the compound was eluted within 3 days, suggesting that the vehicle was not suitable for long-term delivery. On the other hand, direct infusion of MIBG into vessels was able to reduce neointimal hyperplasia over 14 days in organ culture. These data support the conclusion that the cellular retention characteristics of MIBG contribute significantly to the efficacy of this compound. Based on these results, ART, but not PARP, may be a credible target for therapeutic treatment of restenosis.

Injury-induced expression of cytokeratins 8 and 18 by vascular smooth muscle cells requires concurrent activation of cytoskeletal and growth factor receptors.

Cytokeratins are not present in the vascular smooth muscle cells (VSMCs) of normal arteries, but they are detectable in the VSMCs of atherosclerotic lesions. A correlation between cytokeratin expression and VSMC phenotype is proposed, but an examination of VSMCs after mechanical injury has yet to be performed. Immunohistochemistry was used to monitor proteins in arterial sections. Western blotting enabled quantification of protein levels. Angioplasty of porcine femoral artery in vivo and porcine coronary artery in vitro served as models of vascular injury. Cytokeratins 8 and 18 were expressed by VSMCs in porcine femoral artery lesions 14 days after balloon angioplasty. Cytokeratins were also present in the neointima of porcine coronary artery segments placed into organ culture for 4 days. Cytokeratin expression was decreased in the presence of inhibitors that affect MAP kinase, PI3 kinase, Src kinase, and G protein, but not in the presence of an AT1 receptor antagonist. Cytokeratin expression also occurred when VSMCs were plated onto collagen in the presence of serum. We conclude that mechanical injury induces expression of cytokeratin 8 and 18 both in vitro and in vivo by synthetic VSMCs that migrate into the neointima. Furthermore, cytokeratin expression requires cellular attachment to extracellular matrix proteins in conjunction with mitogenic stimulation.

MIBG, an inhibitor of arginine-dependent mono(ADP-ribosyl)ation, prevents differentiation of L6 skeletal myoblasts by inhibiting expression of myogenin and p21(cip1).

The development of skeletal muscle is controlled by a highly synchronized series of cellular events, and various signals from both inside and outside the cell play a role in the switch from multipotential mesodermal stem cells to muscle fibers. Meta-iodobenzylguanidine (MIBG), an inhibitor of mono(ADP-ribosyl)ation, has been shown to prevent terminal differentiation of skeletal myoblasts; however, its mechanism of action has not been established. We recently reported that MIBG is capable of preventing phenotypic modulation of smooth muscle cells by interfering with specific trans-acting factors [L. Yau, B. Litchie, S. Thomas, B. Storie, N. Yurkova, P. Zahradka, Endogenous mono-ADP-ribosylation mediates smooth muscle cell proliferation and migration via protein kinase N-dependent induction of c-fos expression. Eur. J. Biochem. 270 (2003) 101-110.]. We therefore examined the effect of MIBG on select myogenic regulatory factors known to control terminal differentiation. It was confirmed that MIBG, but not inhibitors of poly-ADP-ribose polymerase (3-aminobenzamide, PD128763), inhibits fusion of L6 skeletal myoblasts in a concentration-dependent manner. Moreover, inhibition by MIBG correlated with a failure to induce expression of myogenin and p21(cip1), while levels of MyoD and MEF2 were unaffected. Time-of-addition studies revealed that MIBG also affected a late event possibly linked to cell fusion. Finally, arginine-dependent mono(ADP-ribosyl)transferase activity increased over the first 24 h of the differentiation period. These data support a role for arginine-dependent mono(ADP-ribosyl)transferase as an essential positive regulator of differentiation in skeletal muscle cells that operates by modulating the expression of specific myogenic factors.

Perivascular delivery of losartan with surgical fibrin glue prevents neointimal hyperplasia after arterial injury.

OBJECTIVE: Long-term success of revascularization procedures is limited by recurrent stenosis, a reduction in vascular lumen area that results from neointimal hyperplasia. Inhibitors of the renin-angiotensin system, such as losartan, have potential to prevent recurrent stenosis; however, to date, efficacy has not been demonstrated in either animal models or human beings. While we have previously reported that treatment with a satisfactory dose may be an important element in obtaining efficacy, oral delivery cannot achieve the required concentration. We therefore tested the ability of losartan to restrict neointimal hyperplasia after local delivery of an elevated dose in a fibrin glue. METHODS: The porcine saphenous artery was subjected to balloon angioplasty. Losartan (25 micromol/mL) was applied directly to the adventitial surface of the injured vessel after mixing with 1.0 mL of Tisseel. Neointimal formation was quantified after 14 days with morphometry, and immunologic staining was used to monitor expression of proteins associated with cell proliferation, migration, and phenotypic modulation. RESULTS: A statistically significant decrease of 82% (n = 5) in neointimal area was obtained with losartan, and cell proliferation, as defined by proliferating cell nuclear antigen (PCNA) expression, was inhibited by 97%. Reduced cyclin A expression in losartan-treated vessels confirmed that cell cycle progression was blocked; however, the presence of cytokeratin 8 and tenascin in the media and neointima of injured vessels, regardless of treatment, suggested that losartan does not restrict phenotypic modulation. Inhibition of MT1-MMP (MMP-14) expression by losartan indicated that this inhibitor likely suppresses cell migration as well. CONCLUSIONS: These data demonstrate that losartan can effectively prevent recurrent stenosis when delivered locally with a fibrin glue such as Tisseel. Our results also indicate that losartan may operate by interfering with the expression of proteins required for cell cycle progression and migration. CLINICAL RELEVANCE: Release of angiotensin II in response to vascular injury may promote neointimal hyperplasia, because this hormone can stimulate smooth muscle cell proliferation and migration. This study demonstrates that local application of an angiotensin receptor antagonist, losartan, to the site of injury can effectively prevent neointimal hyperplasia after balloon angioplasty. Application of losartan to the perivascular surface of the injured vessel in a surgical fibrin glue enabled delivery of a dose that exceeds the maximum attainable, via a systemic delivery route. The glue also served as a depot from which the drug was slowly released over time. Treatment with losartan may be a viable approach for controlling neointimal hyperplasia at locations (eg, grafts) that are accessible during a surgical procedure.

PGE(2) stimulates vascular smooth muscle cell proliferation via the EP2 receptor.

Prostaglandins released by injured vascular tissue can modulate smooth muscle cell (SMC) proliferation. The mechanism of action of PGE(2) was investigated with porcine coronary artery SMCs obtained by explant culture. DNA and RNA syntheses exhibited a concentration-dependent increase following treatment of quiescent SMCs with PGE(2), while PGI(2) had no effect. PGE(2) also elevated PCNA expression, bromodeoxyuridine incorporation, and cell number, indicative of a hyperplastic growth response. Furthermore, induction of c-fos expression required activation of both phosphatidylinositol 3-kinase and mitogen-activated protein kinase. Contrary to these data, treatment of proliferating cells with PGE(2) caused a reduction in DNA synthesis. A role for PKA in either growth stimulation or inhibition was excluded. Interestingly, only quiescent SMCs expressed EP2 receptors, and the selective EP2 receptor agonist butaprost confirmed that this receptor was essential for growth stimulation and possibly inhibition. These data suggest that the growth state-dependent actions of PGE(2) on SMC proliferation may be mediated via the EP2 receptor.

Endogenous mono-ADP-ribosylation mediates smooth muscle cell proliferation and migration via protein kinase N-dependent induction of c-fos expression.

ADP-ribosylation has been coupled to intracellular events associated with smooth muscle cell vasoreactivity, cytoskeletal integrity and free radical damage. Additionally, there is evidence that ADP-ribosylation is required for smooth muscle cell proliferation. Our investigation employed selective inhibitors to establish that mono-ADP-ribosylation and not poly(ADP-ribosyl)ation was necessary for the stimulation of DNA synthesis by mitogens. Mitogen treatment increased concomitantly the activity of both soluble and particulate mono-ADP-ribosyltransferase, as well as the number of modified proteins. Inclusion of meta-iodobenzylguanidine (MIBG), a selective decoy substrate of arginine-dependent mono-ADP-ribosylation, prevented the modification of these proteins. MIBG also blocked the stimulation of DNA and RNA synthesis, prevented smooth muscle cell migration and suppressed the induction of c-fos and c-myc gene expression. An examination of relevant signal transduction pathways showed that MIBG did not interfere with MAP kinase and phosphatidylinositol 3-kinase stimulation; however, it did inhibit phosphorylation of the Rho effector, PRK1/2. This novel observation suggests that mono-ADP-ribosylation participates in a Rho- dependent signalling pathway that is required for immediate early gene expression.