Luis Fernando Bittar

Long-term increased factor VIII levels are associated to interleukin-6 levels but not to post-thrombotic syndrome in patients with deep venous thrombosis.

INTRODUCTION Increased FVIII levels are a well established risk factor for deep venous thrombosis (DVT), whose etiopathogenesis is not yet well understood. In this study, we aimed to evaluate the possibility that inflammatory markers and post-thrombotic syndrome (PTS) could contribute to FVIII levels in patients with a history of DVT. DESIGN AND METHODS It is a case-control study that included 68 patients with DVT of the lower limbs 32 months after the acute episode, and 67 healthy adults as controls. We evaluated plasma levels of FVIII, VWF, D-dimer and serum levels of CRP, IL-6, IL-8, TNF-α in patients and controls. The presence of PTS was evaluated by the Villalta scale. RESULTS Patients with DVT presented higher levels of FVIII, VWF and D-dimer when compared to controls (P ≤ 0.001). Almost 50% of patients presented FVIII levels above 90th percentile. Furthermore, IL-6 (1.19 vs. 0.98 pg/mL, P = 0.01) and TNF-α (2.27 vs. 1.57 pg/mL, P ≤ 0.001) were also higher in patients when compared to controls. In a linear regression multivariate model, VWF and IL-6 levels were independent factors associated with FVIII levels (P ≤ 0.001). FVIII levels were not increased in patients with PTS. Patients with PTS showed higher levels of IL-8 when compared to patients without PTS (23.03 vs. 18.20 pg/mL, P = 0.04). CONCLUSIONS In conclusion, we demonstrated that DVT is associated with increased levels of inflammatory and coagulation markers, including FVIII, even a long time after the acute episode. Moreover, IL-6 levels were an independent factor associated with FVIII levels. Finally, PTS seems to be related to inflammatory cytokine IL-8, a proinflammatory and proangiogenic chemokine, but not to FVIII levels.

Folate, Vitamin B12 and Homocysteine status in the post-folic acid fortification era in different subgroups of the Brazilian population attended to at a public health care center

BackgroundFolate and vitamin B12 are essential nutrients, whose deficiencies are considerable public health problems worldwide, affecting all age groups. Low levels of these vitamins have been associated with high concentrations of homocysteine (Hcy) and can lead to health complications. Several genetic polymorphisms affect the metabolism of these vitamins. The aims of this study were to assess folate, vitamin B12 and homocysteine status in distinct Brazilian individuals after the initiation of folic acid fortification by Brazilian authorities and to investigate the effects of RFC1 A80G, GCPII C1561T and MTHFR C677T polymorphisms on folate, vitamin B12 and Hcy levels in these populations.MethodsA total of 719 individuals including the elderly, children, as well as pregnant and lactating women were recruited from our health care center. Folate, vitamin B12 and Hcy levels were measured by conventional methods. Genotype analyses of RFC1 A80G, GCPII C1561T and MTHFR C677T polymorphisms were performed by PCR-RFLP.ResultsThe overall prevalence of folate and vitamin B12 deficiencies were 0.3% and 4.9%, respectively. Folate deficiency was observed only in the elderly (0.4%) and pregnant women (0.3%), whereas vitamin B12 deficiency was observed mainly in pregnant women (7.9%) and the elderly (4.2%). Plasma Hcy concentrations were significantly higher in the elderly (33.6%). Pregnant women carrying the MTHFR 677TT genotype showed lower serum folate levels (p = 0.042) and higher Hcy levels (p = 0.003). RFC1 A80G and GCPII C1561T polymorphisms did not affect folate and Hcy levels in the study group. After a multivariate analysis, Hcy levels were predicted by variables such as folate, vitamin B12, gender, age and RFC1 A80G polymorphism, according to the groups studied.ConclusionOur results suggest that folate deficiency is practically nonexistent in the post-folic acid fortification era in the subgroups evaluated. However, screening for vitamin B12 deficiency may be particularly relevant in our population, especially in the elderly.

Severe Post-thrombotic Syndrome is Associated With Higher Levels of Factor VIII

Increased levels of factor VIII (FVIII) are a prevalent and independent risk factor for deep venous thrombosis (DVT). After a median of 10 years of the first DVT, we evaluated FVIII coagulation levels in 55 patients with DVT of the lower limbs and previous high levels of FVIII and in 74 controls. Subsequently, we analyzed the presence of post-thrombotic syndrome (PTS) in patients and its relationship with FVIII levels. After a median of 10 years of the first DVT, the FVIII levels were still significantly higher in patients when compared to controls (P < .001). Patients with severe PTS showed increased levels of FVIII when compared to patients with moderate or absent PTS (P < .001). We demonstrated a persistent increase in FVIII levels in a subset of patients with DVT, but in a lower magnitude after 10 years of the first DVT episode. Moreover, we observed a significant association between increased FVIII levels and severe PTS.

Polymorphisms and mutations in vWF and ADAMTS13 genes and their correlation with plasma levels of FVIII and vWF in patients with deep venous thrombosis.

Background: Increased levels of factor VIII (FVIII) are a prevalent and independent risk factor for deep venous thrombosis (DVT) and are affected by von Willebrand factor (vWF) levels. Design and Methods: ADAMTS13 contributes to vWF levels, and we investigated genetic polymorphisms previously described to be associated with decreased levels of these proteins in 435 patients with DVT (126 M and 309 F; median age 37 years, range 18-68 years) and 580 controls (163 M and 417 F; median age 35 years, range 18-68 years). Subsequently, we investigated the relationship between the genotypes and plasma levels of FVIII, vWF, and DVT risk. Results: Patients with DVT showed higher plasma levels of FVIII:C, FVIII:Ag, and vWF:Ag (P < .001) when compared to controls. Patients and controls heterozygous for the 4751A>G polymorphism in the vWF gene presented decreased levels of vWF:Ag, FVIII:Ag, and FVIII:C (P < .001), but this was not a protective factor for DVT. Individuals heterozygous for 1852C>G polymorphism in ADAMTS13 gene, which is associated with reduced levels of ADAMTS13, had significantly elevated levels of vWF:Ag (P = .001), FVIII:Ag (P = .01), and FVIII:C (P = .02). However, this polymorphism was not a risk factor for DVT in our study. Heterozygosis for a new polymorphism identified in ADAMTS13 gene, 1787-26G>A, was significantly associated with elevated levels of FVIII:C (P = .02) when compared to wild type. Conclusions: Despite the tempting assumption that genetic factors that change ADAMTS13 activity might modulate the risk of DVT by altering vWF and FVIII levels, the polymorphisms analyzed in this study did not correlate with DVT risk among patients investigated.

Circulating Progenitor and Mature Endothelial Cells in Deep Vein Thrombosis

Introduction: Mature circulating endothelial cells (CEC) and circulating endothelial progenitor cells (EPC) have been described in several conditions associated with endothelial injury. Their role in deep vein thrombosis (DVT) has not been previously evaluated. Patients and Methods: In this pilot study we evaluated the time course of CEC and EPC release after vena cava experimental DVT in mice, using the FeCl3 model. We also evaluated their presence in patients with DVT at different phases of the disease (acute and chronic phase). CEC and EPC were evaluated by Flow Cytometry. Results: In mice, both CEC and EPC were increased 24 hours after DVT induction, peaking 48 hours thereafter. After 72 hours, CEC counts decreased sharply, whereas EPC counts decreased less substantially. In DVT patients we observed a significant increase in CEC counts immediately after DVT compared to healthy individuals. Patients with chronic disease also presented a significant elevation of these cell count. In a subgroup of patients for whom serial samples were available, CEC counts decreased significantly after 9-15 months of the acute event. Conclusions: Our results suggest the participation of these cells in the reparative processes that follows DVT, both at immediate and late time-points. The different kinetics of CEC and EPC release in experimental DVT suggests a heterogeneous role for these cells in the reparative events after DVT.

Increased adhesive properties of neutrophils and inflammatory markers in venous thromboembolism patients with residual vein occlusion and high D-dimer levels

BACKGROUND Venous thromboembolism (VTE) develops via a multicellular process on the endothelial surface. Although widely recognized, the relationship between inflammation and thrombosis, this relationship has been mostly explored in clinical studies by measuring circulating levels of inflammatory cytokines. However, the role of inflammatory cells, such as neutrophils, in the pathogenesis of VTE is not clear in humans. AIMS To evaluate the adhesive properties of neutrophils, erythrocytes and platelets in VTE patients and to correlate findings with inflammatory and hypercoagulability marker levels. METHODS Study group consisted of twenty-nine VTE patients and controls matched according to age, gender and ethnic background. Adhesive properties of neutrophils, erythrocytes and platelets were determined using a static adhesion assay. Neutrophil adhesion molecules expressions were evaluated by flow cytometry. Inflammatory and hypercoagulability marker levels were evaluated by standard methods. Residual vein occlusion (RVO) was evaluated by Doppler ultrasound. RESULTS No significant difference could be observed in platelet and erythrocyte adhesion between VTE patients and controls. Interestingly, VTE patients with high levels of D-dimer and RVO, demonstrated a significant increase in neutrophil adhesion, compared to controls and remaining patients. Inflammatory markers (IL-6, IL-8, TNF-α) were also significantly elevated in this subgroup, compared to other VTE patients. Adhesive properties of neutrophils correlated with IL-6 and D-dimer levels. Neutrophils adhesion molecules (CD11a, CD11b and CD18) were not altered in any of the groups. CONCLUSION These findings not only support the hypothesis of an association between inflammation and hypercoagulability, but more importantly, highlight the role of neutrophils in this process.

Severe postthrombotic syndrome is associated with characteristic sonographic pattern of the residual thrombosis.

Postthrombotic syndrome (PTS) may affect 50% of patients with deep venous thrombosis, 5–10% of them may present severe manifestations. The causes for PTS development and severity have not been well established. This study evaluated whether PTS may be associated with the presence, and echogenicity, of the residual vein thrombosis (RVT). We included patients with a history of deep venous thrombosis in the past 58 months. These patients were further evaluated for PTS diagnosis, clinical comorbidities, plasma levels of D-dimer, serum levels of C-reactive protein and for the presence of RVT. Particularly, RVT was detected by ultrasound examination and the residual thrombi echogenicity was determined by grayscale median (GSM). Fifty-six patients were included, of which 41 presented PTS. Mild PTS was detected in 23 patients, moderate PTS in 11 and severe PTS in seven patients. Patients with severe PTS showed higher body mass index, higher abdominal circumference and higher C-reactive protein levels when compared with the other patients (P = 0.007, P = 0.002, P = 0.02, respectively). The ultrasound-generated GSM was significantly lower in patients with severe PTS compared with patients with mild–moderate PTS or no PTS (median = 24, 35 and 41, respectively; P = 0.04). A GSM value less than 25, which was consistent with a hypoechoic RVT, was the best cut-off value to discriminate patients with severe PTS from those with mild or moderate PTS and those without PTS. RVT is a common finding among patients with PTS and the echogenicity of the RVT may impact the severity of PTS.

Genetic variations in sites of affinity between FVIII and LRP1 are not associated with high FVIII levels in venous thromboembolism

Increased factor VIII (FVIII) levels are a prevalent and independent risk factor for venous thromboembolism (VTE). The low density lipoprotein receptor-related protein 1 (LRP1) has been associated with FVIII catabolism. After a median of 10 years of the first thrombotic episode, we evaluated FVIII activity levels in 75 patients with VTE and high FVIII levels and in 74 healthy controls. Subsequently, we evaluated the regions of F8 and LRP1 genes coding sites of affinity between these proteins, with the objective of determining genetic alterations associated with plasma FVIII levels. After a median time of 10 years after the VTE episode, FVIII levels were significantly higher in patients when compared to controls (158.6 IU/dL vs. 125.8 IU/dL; P ≤ 0.001]. Despite the fact that we found 14 genetic variations in F8 and LRP1 genes, no relationship was found between FVIII levels with these variations. We demonstrated a persistent increase of FVIII levels in patients with VTE, but in a much lower magnitude after 10 years when compared to 3-years after the episode. Moreover, we observed no relationship of genetic variations in the gene regions coding affinity sites between LRP1 and FVIII with FVIII levels.

Residual Vein Thrombosis Echogenicity Is Associated to the Risk of DVT Recurrence: A Cohort Study

Although deep vein thrombosis (DVT) recurrence is a common late complication of the disease, there are few predictive markers to risk-stratify patients long-term after the thrombotic event. The accuracy of residual vein thrombosis (RVT) in this context is controversial, possibly due to a lack of a standardized methodology. The objective of the study was to evaluate the accuracy of RVT echogenicity as a predictive marker of late DVT recurrence. To evaluate the accuracy of RVT echogenicity as a predictive marker of late DVT recurrence. This prospective study included patients with history of DVT in the past 33 months. Ultrasound examination was performed to detect the presence of RVT, and its echogenicity was determined by calculating the grayscale median (GSM) of the images. Blood samplings were taken for plasma D-dimer levels. Patients were followed-up for 28 months and the primary end point was DVT recurrence. Deep vein thrombosis recurrence was confirmed or excluded by ultrasound during the follow-up. Fifty-six patients were included, of which 10 presented DVT recurrence during the follow-up. D-dimer levels above 630 ng/mL conferred higher risk for recurrence with a negative predictive value of 94%. The absence of RVT was a protective marker for recurrence with a negative predictive value of 100%. Also, the presence of hypoechoic RVT, determined by GSM values below 24, positively predicted 75% of DVT recurrences. Our results suggest that the persistence of RVT and, particularly, the presence of hypoechoic thrombi (GSM < 24) are predictive markers of the risk of DVT recurrence. Residual vein thrombosis echogenicity, by GSM analysis, could represent a new strategy for the evaluation of recurrence risk in patients with DVT.