Introduction and ObjectiveDiabetes is characterized by chronic inflammation, endothelial dysfunction, increased risk of infections and early cardiovascular disease. By releasing neutrophil extracellular traps (NETs), neutrophils kill bacteria and exert pro-inflammatory and pro-thrombotic activities. Increased NETosis has been found in cross-sectional studies including treated type 2 diabetes mellitus (T2DM) patients. In this study, we determined whether the ability of neutrophils to form NETs differs in diabetic patients pre- and post-hyperglycemic control versus healthy donors (HD), and the relationship between NETosis with pro-thrombotic, pro-inflammatory biomarkers and thrombotic clinical events.MethodsDiabetic patients recently diagnosed and after 6 and 12 months of treatment (N = 25) and HD (N = 25) were included. NET formation was studied by microscopy and fluorometry. Nucleosomes, HNE-DNA complexes, von Willebrand factor (vWF), IL6 and TNFα plasma levels were measured by ELISA and P-selectin on the platelet surface was assessed by cytometry.ResultsBasal levels of NETs in recently diagnosed T2DM patients were higher compared to HD. While TNFα stimulation of control neutrophils resulted in DNA release, patient neutrophils were not responsive. Although glycemia decreased after 6 months of metformin treatment, basal and TNFα and PMA-stimulated NETs reached normal values after 12 months. Compared to controls, nucleosomes, HNE-DNA complexes, IL-6 and TNFα levels were increased in recently diagnosed patients and decreased after 12 months of treatment. P-selectin and vWF levels were similar in both populations.ConclusionOur data suggest that NETs could represent a biomarker for T2DM. Increased NETosis in T2DM patients does not appear to be the consequence of impaired glycemic control but rather due to pro-inflammatory cytokines and is not related to thrombotic events.

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Two siblings (HSN and AcSN) with congenital goitrous hypothyroidism were investigated in terms of clinical, biochemical, and molecular biology. Diagnosis of defective thyroglobulin (Tg) was based on findings of low serum T4, low normal or normal serum T3, a negative percholate discharge test, and the virtual absence of the serum Tg response to challenge by bovine TSH. Only minute amounts of Tg-related antigens were detected by RIA in the goitrous tissue (HSN, 0.82 mg/g, compared to 70-90 mg/g in normal thyroid tissue), as confirmed by sodium dodecyl sulfate-agarose gel electrophoresis that indicated the virtual absence of Tg. The Tg messenger ribonucleic acids (mRNAs) from controls and HSN thyroid tissue were first reverse transcribed and then divided into several portions from positions 57-8448; the resulting complementary DNAs were, in turn, amplified by reverse polymerase chain reaction. The amplification of nucleotides 5165-6048 from control thyroid tissue Tg mRNA showed a fragment of 884 base pairs (bp). In contrast, the fragment present in the HSN was +/- 750 bp and lacked the normal fragment. The sequencing of the smaller fragment revealed that 138 bp were missing between positions 5590-5727 of the HSN Tg mRNA. This deletion does not affect the reading frame of the resulting mRNA and is potentially fully translatable into a Tg polypeptide chain that is shorter by 46 residues. A cysteine residue is maintained by the junction between the proximal T from leucine 1831 and the distal GT from cysteine 1877. DNA genomic polymerase chain reaction amplification excludes a deletion in the Tg gene and indicates that the deleted 138-nucleotide sequences lie in the same exon. The functional consequences of the deletion are not entirely clear, but it is conceivable that the excision of this segment of the Tg molecule could affect the protein structure, resulting in its premature degradation, very low colloid storage, and diminished thyroid hormone production rate.