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Telomere Length in Peripheral Blood Mononuclear Cells Is Associated with Folate Status in Men

Human chromosomes are capped by telomeres, which consist of tandem repeats of DNA and associated proteins. The length of the telomeres is reduced with increasing cell divisions except when the enzyme telomerase is active, as in stem cells and germ cells. Telomere dysfunction has been associated with development of age-related pathologies, including cancer, cardiovascular disease, Alzheimers disease, and Parkinsons disease. DNA damage in the telomeric region causes attrition of telomeres. Because folate provides precursors for nucleotide synthesis and thus affects the integrity of DNA, including that of the telomeric region, folate status has the potential to influence telomere length. Telomere length is epigenetically regulated by DNA methylation, which in turn could be modulated by folate status. In this study, we determined whether folate status and the 677C > T polymorphism of the methylene tetrahydrofolate reductase (MTHFR) gene are associated with the telomere length of peripheral blood mononuclear cells in healthy men. The results of our study showed that plasma concentration of folate was associated with telomere length of peripheral blood mononuclear cells in a nonlinear manner. When plasma folate concentration was above the median, there was a positive relationship between folate and telomere length. In contrast, there was an inverse relationship between folate and telomere length when plasma folate concentration was below the median. The MTHFR 677C > T polymorphism was weakly associated (P = 0.065) with increased telomere length at below-median folate status. We propose that folate status influences telomere length by affecting DNA integrity and the epigenetic regulation of telomere length through DNA methylation.

Expression of endothelial protein C receptor and thrombomodulin in the intestinal tissue of patients with inflammatory bowel disease

Objective:Inflammatory bowel diseases are characterized by disorders of immunity, thrombosis of large vessels, and microthrombosis of mucosal vessels. The expression of endothelial protein C receptor (EPCR) and thrombomodulin—two receptors of the protein C pathway involved in thrombin scavenging and inflammation—was studied in intestinal resection specimens or mucosal biopsies from patients with inflammatory bowel disease and from controls. The soluble forms of the receptors in plasma were measured. Data Source:This study involved patients from two large university hospitals. After surgery or biopsy, tissue samples were either frozen or fixed in formalin and embedded in paraffin. Sections for immunohistochemistry examination were cut and tested with the specific antibodies to EPCR and thrombomodulin. RNA was extracted from frozen tissue for amplification via reverse-transcriptase polymerase chain reaction. Normal intestinal and diverticulitis tissue was used as a control. Resection samples from 36 patients with ulcerative colitis, 38 with Crohn’s disease, 38 with colonic cancer, and 32 with diverticulitis were studied by immunohistochemistry, and frozen sections from the same patients were studied by immunofluorescence. Twelve biopsy specimens of adjacent intestinal areas from six patients with inflammatory bowel disease were included in the study for reverse-transcriptase polymerase chain reaction. Soluble receptors were measured in the plasma of 52 inflammatory bowel disease patients and 52 controls. Data Summary:EPCR and thrombomodulin were expressed on the mucosal endothelium of controls, and the intensity of the signal decreased in inflammatory bowel disease patients. EPCR was expressed by dendritic-like cells in controls, which also stained positive for CD21. The EPCR+/CD21+ dendritic-like cells were not as commonly observed in sections from ulcerative colitis patients as they were in sections from control patients (12.0 ± 3.6 cells per high-power field vs. 23.8 ± 10.4 cells per high-power field, p = .03), and this decrease was less evident in sections from Crohn’s disease patients. Levels of messenger RNA for EPCR paralleled protein expression. Soluble thrombomodulin and EPCR levels were both higher in patients than in controls: 41.5 vs. 26.0 ng/mL (p <.0001) and 141 vs. 130 ng/mL (p <.05), respectively. Conclusions:EPCR expression on dendritic-like cells that bear the key complement receptor CD21 suggests a role for EPCR in innate immunity. The reduced expression of thrombomodulin and EPCR in the mucosal vessels in inflammatory bowel disease impairs protein C activation, favoring microthrombosis.

Determination of vitamin K1 in plasma by solid phase extraction and HPLC with fluorescence detection.

We describe a procedure for quantification of vitamin K(1) in human plasma by HPLC. Samples, enriched with a vitamin K derivative as internal standard, were deproteinized, purified on polymeric RP-SPE cartridges and injected into HPLC equipped with a post-column on-line zinc metal reactor and a fluorometric detector. Median level in blood donors (n=87) was 1.967 nmol/L (0.93-4.01, 5th-95th percentiles), with a significant correlation between plasma levels and age (r=0.276, p=0.00958) and a lower (not significant) value in women than in men. This method, easy-to-handle and with a high throughput, can be used to identify covert states of vitamin K intake deficiency in patients thus at risk of alterations in blood clotting or bone mineralization.

Haploinsufficiency of the platelet P2Y12 gene in a family with congenital bleeding diathesis

The platelet P2Y12 receptor plays a very important role both in thrombosis and hemostasis. This report describes haploinsufficiency of the platelet P2Y12 gene in a family with congenital bleeding diathesis. Two sisters with inherited, severe platelet dysfunction associated with P2Y12 deficiency displayed a single base pair deletion in their P2Y12 genes (378delC), resulting in a frame-shift and premature truncation of the protein. GL, the son of one of them, displayed mild platelet dysfunction and normal P2Y12 sequence. We hypothesized that the abnormal platelet phenotype of GL is due to haploinsufficiency of his P2Y12 gene. We analyzed genomic DNA from the family by Southern Blotting and real-time (RT) PCR. Southern Blotting results demonstrated that GL has a single P2Y12 allele, inherited from his father. RT-PCR revealed that GL, his mother and aunt have one single intact P2Y12 allele, while his father has two P2Y12 alleles. The single GL P2Y12 allele contains normal sequence, while his mother and aunt have the 378delC allele. The results of this study support our hypothesis and illustrate the platelet phenotype associated with P2Y12 haploinsufficiency.

Levels of soluble endothelial protein C receptor are associated with CD4+ changes in Maraviroc-treated HIV-infected patients.

Background Inflammation is a key feature of HIV infection and is correlated with long-term negative cardiovascular outcomes. Therapy-induced increases in CD4+ cell counts can control inflammation, as shown by decreases of coagulation and inflammation markers during efficacious therapy. Maraviroc, a CCR5-antagonist, has resulted in larger increases in CD4+ counts both in naïve and experienced subjects compared to traditional antiretroviral therapy. Objectives and Methods To examine if a member of the protein C anticoagulant and anti-inflammatory pathway, and marker of coagulation and inflammation, the soluble endothelial protein C receptor, is modified by infection and therapy-related variables in patients treated with Maraviroc. Endothelial protein C receptor, together with other established markers of inflammation and coagulation (CRP, IL-6, D-dimer and soluble thrombomodulin) was studied in 43 patients on traditional antiretroviral therapy and in 45 on Maraviroc during 48 weeks of follow-up. Results Soluble endothelial protein C receptor was the only marker that could discriminate at least partially between patients with a good response to Maraviroc and patients who did not respond with an adequate increase in CD4+ cell counts (more than 500 cells/µL by week 48). Conclusions Elevated levels of soluble endothelial protein C receptor, a sensitive marker of endothelial damage, indicated a low level of inflammation and coagulation activation in Maraviroc treated patients not picked up by other widely used markers. Persistent elevated levels of this marker at 48 weeks from beginning of treatment with Maraviroc were related to a poor increase in CD4+ cells.

Instability of cytosolic phospholipase A2α variant upon cellular expression as a basis for its clinical presentation

Instability of cytosolic phospholipase A2α variant upon cellular expression as a basis for its clinical presentation -