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V X Farias,
F H P Macêdo,
M B Oquendo,
A R Tomé,
S N Báo,
D O S Cintra,
C F Santos,
A A C Albuquerque, D B Heimark,
J Larner,
M C Fonteles,
J H Leal-Cardoso,
N R F Nascimento
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ABSTRACT: D-chiro-inositol (DCI) has been shown to prevent and reverse endothelial dysfunction in diabetic rats and rabbits. The present study evaluates the preventive effect of DCI on experimental diabetic neuropathy (DN).
Streptozotocin-induced (STZ) diabetic mice were treated by oral gavage for 60 days with DCI (20 mg/kg/12 h) or saline (NaCl 0.9%; 0.1 ml/10 g/12 h; Diab) and compared with euglycaemic groups treated with saline (0.1 ml/10 g/12 h; Eugly). We compared the response of the isolated sciatic nerve, corpora cavernosa or vas deferens to electrical stimulation.
The electrically evoked compound action potential of the sciatic nerve was greatly blunted by diabetes. The peak-to-peak amplitude (PPA) was decreased from 3.24 ± 0.7 to 0.9 ± 0.2 mV (p < 0.05), the conduction velocity (CV) of the first component was reduced from 46.78 ± 4.5 to 26.69 ± 3.8 ms (p < 0.05) and chronaxy was increased from 60.43 ± 1.9 to 69.67 ± 1.4 ms (p < 0.05). These parameters were improved in nerves from DCI-treated mice (p < 0.05). PPA in the DCI group was 5.79 ± 0.8 mV (vs. 0.9 ± 0.2 mV-Diab; p < 0.05) and CV was 45.91 ± 3.6 ms (vs. 26.69 ± 3.8 ms-Diab; p < 0.05). Maximal relaxation of the corpus cavernosum evoked by electrical stimulation (2-64 Hz) in the Diab group was 36.4 ± 3.8% compared to 65.4 ± 2.8% in Eugly and 59.3 ± 5.5% in the DCI group (p < 0.05). Maximal contraction obtained in the vas deferens was 38.0 ± 9.2% in Eugly and 11.5 ± 2.6% in Diab (decrease of 69.7%; p < 0.05), compared to 25.2 ± 2.3% in the DCI group (p < 0.05 vs. diabetic). Electron microscopy of the sciatic nerves showed prevention of neuronal damage.
DCI has a neuroprotective action in both autonomic and somatic nerves in STZ-induced DN.
Diabetes Obesity and Metabolism 03/2011; 13(3):243-50. · 3.38 Impact Factor
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N R F Nascimento,
L M A Lessa,
M R Kerntopf,
C M Sousa,
R S Alves,
M G R Queiroz,
J Price, D B Heimark,
J Larner,
X Du,
M Brownlee,
A Gow,
C Davis,
M C Fonteles
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ABSTRACT: Endothelial dysfunction (ED) is an early feature of cardiovascular risk and diabetes. Hyperglycemia and hyperlipidemia are causative factors. Excessive endothelial mitochondrial superoxide (ROS) production with hyperglycemia and hyperlipidemia is a key mechanism. Inositol components of an insulin inositol glycan mediator, d-chiro-inositol (DCI) and 3-O-methyl DCI (pinitol), decrease hyperglycemia and hyperlipidemia. We tested whether these, myoinositol and dibutyryl DCI (db-DCI), would prevent or reverse ED in diabetic rats and rabbits. Oral inositols reduced hyperglycemia and hypertriglyceridemia with different potencies and prevented ED in rat aortic rings and mesenteric beds. Inositols added in vitro to five diabetic tissues reversed ED. Relaxation by Ach, NO, and electrical field stimulation was potentiated by inositols in vitro in rabbit penile corpus cavernosa. Inositols in vitro restored impaired contraction by the eNOS inhibitor l-NAME and increased NO effectiveness. DCI and db-DCI decreased elevated ROS in endothelial cells in high glucose and db-DCI reduced PKC activation, hexosamine pathway activity, and advanced glycation end products to basal levels. Xanthine/xanthine oxidase generated superoxide was reduced by superoxide dismutase or inositols, with db-DCI efficacious in a mechanism requiring chelated Fe(3+). Histochemical examination of rat aortic rings for protein SNO demonstrated a decrease in diabetic rings with restoration by inositols. In summary, inositols prevented and reversed ED in rat and rabbit vessels, reduced elevated ROS in endothelial cells, potentiated nitrergic or vasculo-myogenic relaxations, and preserved NO signaling. These effects are related to their metabolic actions, direct superoxide scavenging, and enhancing and protecting NO signaling. Of the inositols tested, db-DCI was most effective.
Proceedings of the National Academy of Sciences 02/2006; 103(1):218-23. · 9.68 Impact Factor
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D L Brautigan,
M Brown,
S Grindrod,
G Chinigo,
A Kruszewski,
S M Lukasik,
J H Bushweller,
M Horal,
S Keller,
S Tamura, D B Heimark,
J Price,
A N Larner,
J Larner
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ABSTRACT: Insulin-stimulated glucose disposal in skeletal muscle proceeds predominantly through a nonoxidative pathway with glycogen synthase as a rate-limiting enzyme, yet the mechanisms for insulin activation of glycogen synthase are not understood despite years of investigation. Isolation of putative insulin second messengers from beef liver yielded a pseudo-disaccharide consisting of pinitol (3-O-methyl-d-chiro-inositol) beta-1,4 linked to galactosamine chelated with Mn(2+) (called INS2). Here we show that chemically synthesized INS2 has biological activity that significantly enhances insulin reduction of hyperglycemia in streptozotocin diabetic rats. We used computer modeling to dock INS2 onto the known three-dimensional crystal structure of protein phosphatase 2C (PP2C). Modeling and FlexX/CScore energy minimization predicted a unique favorable site on PP2C for INS2 in a surface cleft adjacent to the catalytic center. Binding of INS2 is predicted to involve formation of multiple H-bonds, including one with residue Asp163. Wild-type PP2C activity assayed with a phosphopeptide substrate was potently stimulated in a dose-dependent manner by INS2. In contrast, the D163A mutant of PP2C was not activated by INS2. The D163A mutant and wild-type PP2C in the absence of INS2 had the same Mn(2+)-dependent phosphatase activity with p-nitrophenyl phosphate as a substrate, showing that this mutation did not disrupt the catalytic site. We propose that INS2 allosterically activates PP2C, fulfilling the role of a putative mediator mimetic of insulin signaling to promote protein dephosphorylation and metabolic responses.
Biochemistry 09/2005; 44(33):11067-73. · 3.42 Impact Factor
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ABSTRACT: Insulin signaling to generate inositol phosphoglycans (IPGs) was demonstrated to occur via the participation of the heterotrimeric G-proteins G(q/11). IPGs were measured as two specific inositol markers, myo-inositol and chiro-inositol after strong acid hydrolysis. Insulin and Pasteurella multocida toxin (PMT) generated both myo-inositol and chiro-inositol IPGs in a dose-dependent manner. PMT has been shown to activate G(q) specifically. Insulin action was abrogated by pre-treatment with anti G(q/11) antibody. Western blotting demonstrated the enrichment of both insulin receptor beta subunit and G(q/11) in the liver membrane vesicles. Vesicles also contained clathrin, caveolin PLC beta 1 and PLC Delta. Immunogold staining revealed the co-localization of both insulin receptor beta subunit and G(q/11) in an approximate stochiometric ratio of 1:3. No vesicles were detected with either component alone. The present and considerable published data provide strong evidence for insulin signaling both via a tyrosine kinase cascade mechanism and via heterotrimeric G-protein interactions.
Biochemical and Biophysical Research Communications 08/2002; 295(2):561-9. · 2.48 Impact Factor
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ABSTRACT: Adherence of Entamoeba histolytica trophozoites to colonic mucins and resistance to lysis by the membrane attack complex of complement are mediated by a galactose- and N-acetyl-D-galactosamine-specific cell-surface lectin. This lectin is a heterodimeric glycoprotein of heavy (170 kDa) and light (35/31 kDa) subunits. In this work, the amino acid sequence and membrane anchor of the light subunit were analyzed. The light subunit cDNA encoded a protein with a calculated molecular mass of 32 kDa containing two potential sites for N-linked glycosylation and putative amino- and carboxyl-terminal signal sequences characteristic of glycosylphosphatidylinositol (GPI)-anchored proteins. No classical carbohydrate-binding domains common to C- or S-type eukaryotic lectins were detected by sequence analysis of either the heavy or light subunits, leaving the location of the ligand-binding site of the lectin unknown. Analysis of restriction enzyme-digested E. histolytica DNA by Southern blotting was consistent with the presence of more than one light subunit gene. Two light subunit isoforms of 31 and 35 kDa were identified by SDS-polyacrylamide gel electrophoresis analysis of affinity-purified lectin, and the isoforms were shown on two-dimensional gel analysis to form distinct 170/35- and 170/31-kDa heterodimers. The amino acid compositions and cyanogen bromide peptide patterns of the two light subunit isoforms were nearly identical. The 35-kDa isoform labeled more efficiently than the 31-kDa isoform with [3H]glucosamine, while only the 31-kDa isoform labeled with [3H]myristate and [3H]palmitate. Nitrous acid deamination released lipid from the 31-kDa isoform, which co-migrated on thin layer chromatography with acylphosphatidylinositol, a component of some GPI anchors. Gas chromatography and mass spectrometry of the deamination product from the 31-kDa subunit identified both myo- and chiro-inositols, supporting the presence of a GPI membrane anchor. The covalent association of a transmembrane protein with a GPI-anchored protein, as suggested by the cDNA sequences of the lectin heavy and light subunits, is novel and suggests unique roles for the two subunits in the pathogenesis of amebiasis.
Journal of Biological Chemistry 12/1993; 268(32):24223-31. · 4.77 Impact Factor
