Elsevier

International Journal of Cardiology

Volume 299, 15 January 2020, Pages 243-248
International Journal of Cardiology

Combination therapy with benznidazole and doxycycline shows no additive effect to monotherapy with benznidazole in mice infected with the VL-10 strain of the Trypanosoma cruzi

https://doi.org/10.1016/j.ijcard.2019.07.047Get rights and content

Highlights

  • Dox was unable to improve ejection fraction in T. cruzi (VL-10 strain)-infected mice.

  • Dox did not ameliorate immune-parasitological parameters in T. cruzi-infected mice.

  • Benznidazole ameliorated ejection fraction in T. cruzi (VL-10 strain)-infected mice.

  • Benznidazole reduced parasitemia, CCL2/CCL5 plasma levels and cardiac infiltration.

  • Combination therapy with Dox does not potentiate the effects of benznidazole.

Abstract

Background

Chagas heart disease is the most important clinical manifestation of Trypanosoma cruzi infection. Pharmacological therapies have been proposed aiming to reduce inflammatory response and cardiac damage in infected hosts. In this study, we investigated the use of doxycycline (Dox), in a sub-antimicrobial dose, in monotherapy and in combination with benznidazole (Bz) during the acute phase of infection with the VL-10 strain of T. cruzi, evaluating the therapeutic effect during the acute and chronic phases of the infection.

Methods and results

C57BL/6 mice were treated for 20 days with Dox (30 mg/kg), Bz (100 mg/kg), or both drugs in combination starting 9 days after infection. Parasitemia was measured during the acute phase and the animals were monitored for 12 months, after which echocardiography analysis was performed. Blood samples were obtained from euthanized mice for CCL2, CCL5, IL-10 analysis, and cardiac fragments were collected for histopathological evaluation. Dox treatment did not ameliorate parasitological/inflammatory parameters but reduced the cardiac collagen neoformation (CN) in 35%. In contrast, Bz administration reduced parasitemia, plasma levels of CCL2 and CCL5, and cardiac infiltration during acute infection, and reduced the level of IL-10 and CN (95%) at 12 months. Dox was unable to improve ejection fraction, while Bz treatment ameliorated the ejection fraction. No additive effect was observed in combination therapy.

Conclusion

Dox monotherapy is not effective in the acute or chronic phases of experimental cardiomyopathy induced by the VL-10 strain of T. cruzi. Furthermore, combination therapy with Dox does not potentiate the effects of Bz monotherapy.

Introduction

Trypanosoma cruzi infection represents a relevant social problem that still occurs in many countries, especially in Latin America. Cardiac involvement is present in many cases of Chagas disease and can be considered as the more severe clinical form of T. cruzi infection [1,2]. This cardiac disease results in a remodeling of the collagen matrix and subsequent fibrosis, leading to myocarditis, increased tissue rigidity, diastolic and systolic dysfunction, functional loss and, in some cases, severe dilated cardiomyopathy associated with ventricular arrhythmias [3,4].

Benznidazole (Bz) is the most commonly used drug against T. cruzi in Brazil and is used to reduce or eliminate the parasites that trigger these cardiac inflammatory processes [[5], [6], [7], [8]]. Unfortunately, Bz has low efficacy and leads to undesired adverse reactions. For this reason, new anti-T. cruzi compounds have been tested in the last decade, as well as anti-inflammatory drugs that could be useful in preserving cardiac morpho-functional structure. These drugs include angiotensin converting enzyme (ACE) inhibitors, beta-blockers, statins, and, more recently, Doxycycline (Dox) [[9], [10], [11], [12]]. Doxycycline is a broad-spectrum bacteriostatic agent invented and clinically developed in the early 1960s by Pfizer Inc., NY, USA [13]. Besides its original antibacterial role, Dox has emerged as a co-adjuvant therapy in non-bacterial illnesses such as malaria and tick-borne rickettsial diseases [14]. Dox has also stood out as a potent inhibitor of metalloproteinases and suppressor of the hydrolases (e.g., phospholipase A2 and alpha-amylase) when administered in low doses (<50 mg/kg), which can reflect, in part, its capacity to scavenge reactive oxygen species and suppress inflammatory cytokines involved in distinct inflammatory disorders [15,16].

Due to the inefficiency of current treatments against T. cruzi, new pharmacological strategies directed towards the modulation of systemic and/or tissue-specific inflammatory response have been pursued, with the aim of minimizing tissue damage caused by this protozoan. The purpose of this study was to identify whether Dox and Bz therapies could positively modify immune and cardiac parameters during acute (30 days) and long-term (12 months) infection in C57BL/6 mice infected with the VL-10 strain of T. cruzi.

Section snippets

Animals

C57BL/6 male mice, aged 6–8 weeks, were bred and maintained at the Center of Animal Science (CCA) at the Federal University of Ouro Preto (UFOP), Brazil. The procedures adopted are in accordance with the ethical principles of animal experimentation pre-established by the National Council for Control of Animal Experimentation (CONCEA). This research was previously approved by the Ethics Committee on Animal Research of UFOP-CEUA (protocol n° 2016/63).

Parasites and infection

The male C57BL/6 mice (60 days of birth) were

Parasitemia curve

Dox treatment did not inhibit replication of the VL-10 strain of T. cruzi, with no difference observed between the untreated and Dox-treated infected animals. On the other hand, Bz reduced the peak and profile of the parasitemia curve, maintaining activity both in monotherapy and in combination with Dox, as shown in Fig. 1. In this way, as the parasite load is closely related to the inflammatory response, any drug capable of reducing parasitemia (such as Bz) is likely to partially reduce the

Conclusions

Sub-therapeutic dose of Dox, in monotherapy, during the acute phase of T. cruzi infection (VL-10 strain) was not effective to ameliorate the murine cardiomyopathy in both, acute and chronic phases. Furthermore, combination therapy with Bz does not potentiate the effects of Bz monotherapy in chagasic cardiomyopathy.

Financial support

Universidade Federal de Ouro Preto (UFOP), Centro Nacional de Biologia Estrutural e Bioimagem (CENABIO/UFRJ). Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES), and Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq). The authors are grateful to the CAPES and FAPEMIG for the support of the graduate scholarship and AT and ACCC acknowledge CNPq and FAPERJ (ACCC) for a research productivity fellowship.

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