Polymeric Nanoparticles of Enoxaparin as a Delivery System: In Vivo Evaluation in Normal Rats and in a Venous Thrombosis Rat Model.

Enoxaparin is an anticoagulant widely used in the treatment and prophylaxis of deep vein thrombosis (DVT). The subcutaneous route of administration, sometimes in repeated doses during 24 hours, represents a limitation to its use. Thus, the development of a product that can be administered either subcutaneously, in a smaller number of applications becomes a major challenge, with interesting clinical applications. The use of a system for sustained release of drugs can help to meet that goal, by protecting and enabling a gradual released of the agent. This study consisted of the evaluation of in vivo anticoagulant and antithrombotic activity of biodegradable nanoparticles of poly (ε-caprolactone) (PCL) with enoxaparin after subcutaneous injection. The nanoparticles were prepared by the method of double emulsion (w/o/w) and solvent evaporation. Subcutaneous enoxaparin encapsulated in PCL nanoparticles (1000 IU/kg) showed a sustained release in vivo for up to 12 hours (Cmax 0.62 IU/mL) a significantly longer period (P < 0.01) when compared to free enoxaparin (1000 IU/Kg) that disappeared after 9 hours (Cmax 1.50 IU/mL), however with lower anti-Xa activity. The antithrombotic action of enoxaparin-nanoparticles was tested in a DVT model by stasis in rats. There were virtually no formation of venous thrombosis in any of the rats that received enoxaparin encapsulated in nanoparticles (0.03 mg), with a significant difference when compared to groups that received saline (17.2 mg, P < 0.001) and free enoxaparin (2.87 mg, P = 0.001). In summary, enoxaparin-encapsulated in polymeric nanoparticles showed a sustained release for a greater period than that of enoxaparin, and with excellent antithrombotic action. These results corroborate the promising use of pharmacological nanoparticles in clinical practice.

CD144, CD146 and VEGFR-2 properly identify circulating endothelial cell

Studies evaluating circulating endothelial cells by flow cytometry are faced by a lack of con- sensus about the best combination of monoclonal antibodies to be used. The rarity of these cells in peripheral blood, which represent 0.01% of mononuclear cells, drastically increases this challenge. Objective: The aim of this study is to suggest some combinations of markers that would safely and properly identify these cells. Methods: Flow cytometry analysis of circulating endothelial cells was performed applying three different panels composed of different combinations of the CD144, CD146, CD31, CD133, CD45 and anti-Vascular endothelial growth factor receptor-2 antibodies. Results: In spite of the rarity of the events, they were detectable and presented similar interperson numbers of circulating endothelial cells. Conclusion: The combination of markers successfully identified the circulating endothelial cells in healthy individuals, with the use of three different panels confirming the obtained data as reliable.

CD144, CD146 and VEGFR-2 properly identify circulating endothelial cell.

UNLABELLED: Studies evaluating circulating endothelial cells by flow cytometry are faced by a lack of consensus about the best combination of monoclonal antibodies to be used. The rarity of these cells in peripheral blood, which represent 0.01% of mononuclear cells, drastically increases this challenge. OBJECTIVE: The aim of this study is to suggest some combinations of markers that would safely and properly identify these cells. METHODS: Flow cytometry analysis of circulating endothelial cells was performed applying three different panels composed of different combinations of the CD144, CD146, CD31, CD133, CD45 and anti-Vascular endothelial growth factor receptor-2 antibodies. RESULTS: In spite of the rarity of the events, they were detectable and presented similar inter-person numbers of circulating endothelial cells. CONCLUSION: The combination of markers successfully identified the circulating endothelial cells in healthy individuals, with the use of three different panels confirming the obtained data as reliable.

Estrogen receptor alpha and beta gene polymorphisms are not risk factors for recurrent miscarriage in a Brazilian population.

The aim of this study was to determine the prevalence of alpha (ESR1: c.454-397T>C and c.454-351A>G) and beta (ESR2: 1082G>A and 1730G>A) estrogen receptor gene polymorphisms in 2 Brazilian ethnic groups (Caucasian, African Brazilian) and to investigate their association with recurrent miscarriage (RM) in 75 women with a history of 3 or more consecutive pregnancy losses and 139 controls with at least 2 live births and no history of pregnancy loss. Polymerase chain reaction and restriction fragment length polymorphism were used to identify gene polymorphisms. Coagulation methods were used to measure protein C, protein S, and fibrinogen, and a chromogenic method was used for antithrombin quantification. Significantly higher prevalences of 1082G>A and 1730G>A polymorphisms were seen in African Brazilian and Caucasian controls, respectively. There was no association between RM and ESR polymorphisms. There was a difference in the genotype prevalence in the c.454-39T>C polymorphism between RM and control Caucasians, but this finding was not associated with an increased risk of miscarriage. There was no synergistic or additive effect between ESR polymorphisms and thrombophilia in RM patients. A difference in the prevalence of ESR polymorphisms was observed, according to ethnic origin. ESR polymorphisms could not be considered a risk factor for RM.

Association between estrogen receptor alpha and beta gene polymorphisms and deep vein thrombosis.

BACKGROUND: Estrogen and the estrogen receptors alpha (ESR1) and beta (ESR2) play a role in regulating genes, including coagulation and fibrinolysis genes. OBJECTIVE: We investigated the association between ESR1 c.454-397T>C and c.454-351A>G and ESR2 1082A>G and 1730A>G polymorphisms and the risk of deep vein thrombosis (DVT), in 134 patients and 134 controls with acquired risk factors for thrombosis associated with estrogen alterations, such as pregnancy, puerperium, oral contraceptives (OC), and hormone replacement therapy (HRT). We also analysed 134 men with DVT. We investigated the relationship of these polymorphisms and the levels of fibrinogen, protein C (PC), protein S (PS), and antithrombin (AT) activity. METHODS: Gene polymorphisms were identified by using PCR and RFLP. Coagulation methods were used to measure PC, PS, and fibrinogen. Chromogenic methods were used to quantify AT. RESULTS AND CONCLUSIONS: The presence of the AA genotype of the 1730G>A polymorphism (OR=0.18; 95%CI=0.05-0.62) suggests a protective effect for DVT in women using OC. As the GG genotype of the 1730G>A polymorphism is associated with increased PS activity in all control women and women using OC, this suggested that a protective effect must occur by another pathway not related to PS. The AA and AG genotypes of the c.454-351A>G and GG genotype of the 1082G>A polymorphisms are associated with increased fibrinogen concentration in pregnant women. The GG haplotype in the ESR2 gene (P<0.001) was related to factor V Leiden or G20210A mutation in the prothrombin gene, or both, as predictive factors of DVT.