Posaconazole (POS) is an inhibitor of ergosterol biosynthesis in clinical use for treating invasive fungal infections. POS has potent and selective anti-Trypanosoma cruzi activity and has been evaluated as a possible treatment for Chagas disease. Microtissues are a 3D culture system that has been shown to reproduce better tissue architecture and functionality than cell cultures in monolayer (2D). It has been used to evaluate chemotropic response as in vitro disease models. We previously developed an in vitro model that reproduces aspects of cardiac fibrosis observed in Chagas cardiomyopathy, using microtissues formed by primary cardiac cells infected by the T. cruzi, here called T. cruzi fibrotic cardiac microtissue (TCFCM). We also showed that the treatment of TCFCM with a TGF-β pathway inhibitor reduces fibrosis. Here, we aimed to evaluate the effect of POS in TCFCM, observing parasite load and molecules involved in fibrosis. To choose the concentration of POS to be used in TCFCM we first performed experiments in a monolayer of primary cardiac cell cultures and, based on the results, TCFCM was treated with 5 nM of POS for 96 h, starting at 144 h post-infection. Our previous studies showed that at this time the TCFCM had established fibrosis, resulting from T. cruzi infection. Treatment with POS of TCFCM reduced 50 % of parasite load as observed by real-time PCR and reduced markedly the fibrosis as observed by western blot and immunofluorescence, associated with a strong reduction in the expression of fibronectin and laminin (45 % and 54 %, respectively). POS treatment also changed the expression of proteins involved in the regulation of extracellular matrix proteins (TGF-β and TIMP-4, increased by 50 % and decreased by 58 %, respectively) in TCFCM. In conclusion, POS presented a potent trypanocidal effect both in 2D and in TCFCM, and the reduction of the parasite load was associated with a reduction of fibrosis in the absence of external immunological effectors.

泊沙康唑（POS）是临床上用于治疗侵袭性真菌感染的麦角固醇生物合成抑制剂。POS具有强效和选择性的抗克氏锥虫活性，已被评估为可治疗南美锥虫病的一种方法。微组织是一种3D培养系统，已显示比单层（2D）中的细胞培养可复制更好的组织结构和功能。它已用于评估趋化反应作为体外疾病模型。我们以前开发了一种体外模型，心肌纤维化的抄录方面的南美锥虫病观察，使用由感染主要心脏细胞形成的微组织锥虫，这里被称为克氏锥虫纤维化心脏微组织（TCFCM）。我们还显示，用TGF-β途径抑制剂治疗TCFCM可减少纤维化。在这里，我们旨在评估POS在TCFCM中的作用，观察寄生虫负荷和参与纤维化的分子。为了选择用于TCFCM的POS浓度，我们首先在原代心脏细胞培养的单层中进行了实验，根据结果，从感染后144 h开始，用5 nM POS处理TCFCM 96小时。我们以前的研究表明，此时TCFCM已确定了由克鲁氏锥虫引起的纤维化。感染。实时PCR观察到，TCFCM的POS处理可减少50％的寄生虫负荷，而蛋白质印迹和免疫荧光检测可显着减少纤维化，与纤连蛋白和层粘连蛋白的表达显着降低有关（分别为45％和54％，分别）。POS处理还改变了TCFCM中参与细胞外基质蛋白调节的蛋白表达（TGF-β和TIMP-4，分别增加了50％和减少了58％）。总之，POS在2D和TCFCM中均表现出强大的锥虫杀伤作用，并且在没有外部免疫效应子的情况下，寄生虫负荷的减少与纤维化的减少有关。