Glycation does not alter LDL-induced secretion of tissue plasminogen activator and plasminogen activator inhibitor-1 from human aortic endothelial cells

Diabetes may induce both quantitative and qualitative changes in lipoproteins, especially low-density lipoprotein (LDL). Effects of LDL glycation on endothelial cell secretion of tissue plasminogen activator (tPA) and plasminogen activator inhibitor-1 (PAI-1) have not been fully elucidated. Human aortic endothelial cell (HAEC) tPA and PAI-1 production were determined after incubation with LDL (50 to 500 microg/mL protein, 24 h) from three sources: (1) nondiabetic LDL (N-LDL) modified in vitro to form six preparations: native, nonmodified (N); glycated (G); minimally oxidized (MO); minimally oxidized and glycated (MOG); heavily oxidized (HO); and heavily oxidized and glycated (HOG); (2) in vivo glycated and relatively nonglycated LDL subfractions from type 1 diabetic patients; (3) LDL from type 1 diabetic patients and matched controls, which was subfractionated using density gradient ultracentrifugation. In experiments using LDL modified in vitro, the rate of tPA release by HAECs incubated with N-LDL (83 +/- 4 ng/mg cell protein/24 h) did not differ significantly from those incubated with G-LDL (73 +/- 7), MO-LDL (74 +/- 13), or MOG-LDL (66 +/- 15) and was not influenced by LDL concentration. The rate of PAI-1 release was similar in HAECs incubated with N-LDL (5.7 +/- 0.6 mug/mg cell protein/24 h), G-LDL (5.7 +/- 0.7), MO-LDL (5.5 +/- 0.8), or MOG-LDL (5.7 +/- 0.9) and was not influenced by LDL concentration. In contrast, tPA release was significantly decreased in cells incubated with LDL (10 microg/mL) modified extensively by oxidation, and averaged 45.2 +/- 5.0 and 43.7 +/- 9.9 ng/mg/24 h for HO-LDL and HOG-LDL, respectively, and was further decreased with increasing concentrations of the heavily oxidized LDL preparations. PAI-1 release was not significantly decreased relative to N-LDL in cells incubated with low concentrations (5 to 50 microg/mL) of HO-LDL and HOG-LDL, but was decreased to 3.2 +/- 0.5 and 3.1 +/- 0.7 microg/mg/24 h for HO-LDL and HOG-LDL at 200 microg/mL, respectively. Results using in vivo glycated versus nonglycated LDL showed that tPA and PAI-1 release did not differ between subfractions. Release of tPA averaged 5.11 +/- 0.6 and 5.12 +/- 0.7 ng/mg/24 h, whereas release of PAI-1 averaged 666 +/- 27 ng/mg/24 h and 705 +/- 30 ng/mg/24 h for nonglycated and glycated LDL subfractions, respectively. Using LDL of different density subclasses, tPA and PAI-1 release in response to LDL from diabetic patients compared with control subjects did not differ when HAECs were incubated with LDLs of increasing density isolated from each subject pair. We conclude that oxidation of LDL, but not glycation, may contribute to the altered fibrinolysis observed in diabetes.

Impact of different health systems on the burden of type 2 diabetes in two populations

Objectives: Health policy directs the management of patients with chronic disease in a country, but evaluating nationwide policies is difficult, not least because of the absence of suitable comparators. This paper examines the management of patients with type 2 diabetes in two demographically comparable populations with different health care systems to see if this represents a viable approach to evaluation.

Methods: A secondary analysis of centralized prescribing databases for 2010 was undertaken to compare the levels and costs of care of patients with type 2 diabetes in Northern Ireland’s National Health Service (NHS) (NI, n = 1.8 million) which has structured care, financial incentives related to diabetes care and an emphasis on generic prescribing, with that of the Republic of Ireland (ROI, n = 4.3 million) where management of diabetes care is guided solely by clinical and other guidelines.

Results: The prevalence of treated type 2 diabetes was 3.59% in NI and 3.09% in ROI, but there were similar and high levels of prescribing of secondary cardiovascular medications. Medication costs per person for anti-diabetic, anti-obesity and cardiovascular medication were 46% higher in ROI than NI, due to differences in levels of generic prescribing.

Conclusions: These different health care systems appear to be producing similar levels of care for patients with type 2 diabetes, although at different levels of cost. The findings question the need for financial incentives in NI and highlight the large cost savings potentially accruing from a greater shift to generic prescribing in ROI. Cross-country comparison, though not without difficulties, may prove a useful adjunct to within-country analysis of policy impact.

AcT-2: A Novel Myotropic and Antimicrobial Type 2 Tryptophyllin from the Skin Secretion of the Central American Red-Eyed Leaf Frog, Agalychnis callidryas

Tryptophyllins are a diverse family of amphibian peptides originally found in extracts of phyllomedusine frog skin by chemical means. Their biological activities remain obscure. Here we describe the isolation and preliminary pharmacological characterization of a novel type 2 tryptophyllin, named AcT-2, from the skin secretion of the red-eyed leaf frog, Agalychnis callidryas. The peptide was initially identified during smooth muscle pharmacological screening of skin secretion HPLC fractions and the unique primary structure—GMRPPWF-NH2—was established by both Edman degradation and electrospray MS/MS fragmentation sequencing. A. cDNA encoding the biosynthetic precursor of AcT-2 was successfully cloned from a skin secretion-derived cDNA library by means of RACE PCR and this contained an open-reading frame consisting of 62 amino acid residues with a single AcT-2 encoding sequence located towards the C-terminus. A synthetic replicate of AcT-2 was found to relax arterial smooth muscle (EC50 = 5.1 nM) and to contract rat urinary bladder smooth muscle (EC50 = 9.3 μM). The peptide could also inhibit the growth of the microorganisms, Staphylococcus aureus, (MIC = 256 mg/L) Escherichia coli (MIC = 512 mg/L), and Candida albicans (128 mg/L). AcT-2 is thus the first amphibian skin tryptophyllin found to possess both myotropic and antimicrobial activities.

Excess molar volumes and excess molar enthalpies in binary systems N-alkyl-triethylammonium bis(trifluoromethylsulfonyl)imide plus methanol

In the present paper, a study on the influence of the alkyl chain length in N-alkyl-triethylammonium bis(trifluoromethylsulfonyl)imide ionic liquids, [N_R,222][Tf₂N] (R   = 6, 8 or 12), on the excess molar enthalpy at 303.15 K and excess molar volume within the temperature interval (283.15–338.15 K) of ionic liquid + methanol mixtures is carried out. Small excess molar volumes with highly asymmetric curves (i.e. S-shape) as a function of mole fraction composition were obtained, with negative values showing in the methanol-rich regions. The excess molar volumes increase with the increase of the alkyl-chain length of the ammonium cation of the ionic liquid and decrease with temperature. The excess enthalpies of selected binary mixtures are positive over the whole composition range and increase slightly with the length of the alkyl side-chain of the cation on the ionic liquid. Both excess properties were subsequently correlated using a Redlich–Kister-type equation, as well as by using the ERAS model. From this semipredictive model the studied excess quantities could be obtained from its chemical and physical contribution. Finally, the COSMOThermX software has been used to evaluate its prediction capability on the excess enthalpy for investigated mixtures at 303.15 K and 0.1 MPa. From this work, it appears that COSMOThermX method predicts this property with good accuracy of approx. 10%, providing at the same time the correct order of magnitude of the partial molar excess enthalpies at infinite dilution for the studied ILs,

pLR-HL: a novel amphibian Bowman-Birk-type trypsin inhibitor from the skin secretion of the broad-folded frog, Hylarana latouchii

In this study, we report a novel heptadecapeptide (LIGGCWTKSIPPKPCLV) of the pLR/ranacyclin family, named pLR-HL, whose structure was deduced from its biosynthetic precursor-encoding cDNA cloned from the skin secretion-derived cDNA library of the broad-folded frog, Hylarana latouchii, by employing a "shotgun" cloning technique. It contains a disulphide loop between Cys5 and Cys15 which is consistent with Bowman-Birk-type protease inhibitors. The primary structure of pLR-HL deduced from the cDNA sequence was confirmed by fractionating the skin secretion using reverse phase HPLC and subsequent analysis using MALDI-TOF mass spectrometry and LC/MS/MS fragmentation sequencing. On the basis of the establishment of unequivocal amino acid sequence, a synthetic replicate was synthesised by solid-phase Fmoc chemistry, and it displayed a moderately potent trypsin inhibition with a Ki of 143 nM. The substitution of Lys-8 by Phe (Phe8 -pLR-HL) resulted in abolition of trypsin inhibition but generation of modest inhibition on chymotrypsin with a Ki of 2.141 μM. Additionally, both the disulphide loops of pLR-HL and Phe8 -pLR-HL were synthesised and tested. Both of the catalytic loops retained similar inhibitory potencies towards trypsin or chymotrypsin in comparison with the original intact molecules. Thus, the replacement of reactive site residues could alter the specificity of these protease inhibitors, while the canonical reactive loop alone can independently constitute biologically-active moiety.

PESSTO monitoring of SN 2012hn: further heterogeneity among faint Type I supernovae

We present optical and infrared monitoring data of SN 2012hn collectedby the Public European Southern Observatory Spectroscopic Survey forTransient Objects. We show that SN 2012hn has a faint peak magnitude(MR ˜ -15.65) and shows no hydrogen and no clearevidence for helium in its spectral evolution. Instead, we detectprominent Ca II lines at all epochs, which relates this transient topreviously described `Ca-rich' or `gap' transients. However, thephotospheric spectra (from -3 to +32 d with respect to peak) of SN2012hn show a series of absorption lines which are unique and a redcontinuum that is likely intrinsic rather than due to extinction. Linesof Ti II and Cr II are visible. This may be a temperature effect, whichcould also explain the red photospheric colour. A nebular spectrum at+150 d shows prominent Ca II, O I, C I and possibly Mg I lines whichappear similar in strength to those displayed by core-collapsesupernovae (SNe). To add to the puzzle, SN 2012hn is located at aprojected distance of 6 kpc from an E/S0 host and is not close to anyobvious star-forming region. Overall SN 2012hn resembles a group offaint H-poor SNe that have been discovered recently and for which aconvincing and consistent physical explanation is still missing. Theyall appear to explode preferentially in remote locations offset from amassive host galaxy with deep limits on any dwarf host galaxies,favouring old progenitor systems. SN 2012hn adds heterogeneity to thissample of objects. We discuss potential explosion channels includingHe-shell detonations and double detonations of white dwarfs as well aspeculiar core-collapse SNe.

Ultrastructure of head organs (anterior adhesive apparatus) and posterior secretory systems of Caballeria liewi Lim, 1995 (Monogenea, Ancyrocephalidae)

Caballeria liewi Lim, 1995, uses adhesive secretions from the head organs and posterior secretory systems to assist in locomotion and attachment. Ultrastructural investigations show that the head organs of C. liewi consist of three pairs of antero-lateral pit-like openings bearing microvilli and ducts leading from two types of uninucleated gland cells (located lateral to the pharynx), one type producing rod-like (S1) bodies with an electron-dense matrix containing less electron-dense vesicles and the second type producing oval (S2) bodies with a homogeneous electron-dense matrix. Interlinking band-like structures are observed between S1 bodies and between S2 bodies. S1 body is synthesised in the granular endoplasmic reticulum, transported to a Golgi complex to be packaged into vesicles and routed into ducts for exudation. The synthesis of the S2 body is unresolved. Haptoral secretions manifested externally as net-like structures are derived from dual electron-dense (DED) secretory body produced in the peduncular gland cells. The DED body consists of a less electron-dense oval core in a homogeneous electron-dense matrix. On exocytosis into the pyriform haptoral reservoir, DED bodies are transformed into a secretion with two types of inclusions (less electron-dense oval and electron-dense spherical inclusions) in an electron-dense matrix. The secretions are further transformed (as small, oval, electron-dense bodies) when transported to the superficial anchor grooves, and on exudation into the gill tissues, the secretions become an electron-dense matrix. Secretory bodies associated with uniciliated structures, anchor sleeves and marginal hooks are also observed.