Reduced thrombin formation and excessive fibrinolysis are associated with bleeding complications in patients with dengue fever: a case-control study comparing dengue fever patients with and without bleeding manifestations

Abstract

Background: Dengue cases have been classified according to disease severity into dengue fever (DF) and dengue hemorrhagic fever (DHF). Although DF is considered a non-severe manifestation of dengue, it has been recently demonstrated that DF represents a heterogeneous group of patients with varied clinical complications and grades of severity. Particularly, bleeding complications, commonly associated to DHF, can be detected in half of the patients with DF. Although a frequent complication, the causes of bleedings in DF have not been fully addressed. Thus, the aim of this study was to perform a comprehensive evaluation of possible pathophysiological mechanisms that could contribute to the bleeding tendency observed in patients with DF.

Methods: This is a case-control study that enrolled adults with DF without bleeding and adults with DF and bleeding complications during the defervescence period. Healthy controls were also included. Peripheral blood counts, inflammatory, fibrinolysis and endothelial cell activation markers, and thrombin generation were evaluated in patients and controls.

Results: We included 33 adults with DF without complications, 26 adults with DF and bleeding and 67 healthy controls. Bleeding episodes were mild in 15 (57.6%) and moderate in 11 (42.4%) patients, 8 (30.7%) patients had bleedings in multiple sites. Patients with DF and bleedings had lower platelet counts than DF without bleeding (median = 19,500 vs. 203,500/mm3, P < 0,0001). Levels of TNF-α, thrombomodulin and VWF were significantly increased in the two dengue groups than in healthy controls, but similar between patients with and without bleedings. Plasma levels of tPA and D-dimer were significantly increased in patients with bleedings (median tPA levels were 4.5, 5.2, 11.7 ng/ml, P < 0.0001 and median D-dimer levels were 515.5, 1028 and 1927 ng/ml, P < 0.0001). The thrombin generation test showed that patients with bleeding complications had reduced thrombin formation (total thrombin generated were 3753.4 in controls, 3367.5 in non-bleeding and 2274.5nM in bleeding patients, P < 0.002).

Conclusions: DF can manifest with spontaneous bleedings, which are associated with specific coagulation and fibrinolysis profiles that are not significantly present in DF without this complication. Particularly, thrombocytopenia, excessive fibrinolysis and reduced thrombin formation may contribute to the bleeding manifestations in DF.

Figure 1

Flow chart of the study.

Figure 2

Peripheral blood counts. This figure illustrates the presence of thrombocytopenia (a) and monocytosis (b) in patients with bleeding complications. These parameters were performed in all patients. P values were calculated using the Wilcoxon-Mann–Whitney test. CT = controls.

Figure 3

Inflammatory and endothelial activation markers. This figure illustrates the activation of the inflammatory response and the endothelial cells in dengue patients. Levels of inflammatory and endothelial cells activation markers were similar between DF groups. a. TNF-α, b. plasma TM levels. c. VWF antigen. These parameters were performed in all patients. P values were calculated using the Kruskal-Wallis analysis, followed by the Tukey post-test. CT = controls . * Place of differences detected by Tukey post-test.

Figure 4

Fibrinolysis parameters. This figure illustrates the presence of hyperfibrinolysis in dengue patients. Fibrinolysis activity markers, such as tPA and D-dimer, were particularly elevated in patients with DF and bleeding complications. a. tPA levels; b. D-dimer levels. These parameters were performed in all patients. P values were calculated using the Kruskal-Wallis analysis, followed by the Tukey post-test. CT = controls. * Place of differences detected by Tukey post-test.