Microemulsions strongly promoted the activity of α-bisabolol against different Leishmania species and its skin permeation.

This study aimed to develop microemulsions (MEs) containing α-bisabolol for the topical treatment of cutaneous leishmaniasis (CL). Initially, pseudoternary phase diagrams were developed using α-bisabolol as the oil phase, Eumulgin® CO 40 as the surfactant, Polymol® HE as the co-surfactant, and distilled water as the aqueous phase. Two transparent liquid systems (TLS) containing 5% of α-bisabolol were selected and characterized (F5E25 and F5EP25). Next, skin permeation and retention assays were performed using Franz cells. The interaction of the formulation with the stratum corneum (SC) was evaluated using the FTIR technique. The cytotoxicity was evaluated in murine peritoneal macrophages. Finally, the antileishmanial activity of microemulsions was determined in promastigotes and amastigotes of L. amazonensis (strain MHOM/BR/77/LTB 0016). As a result, the selected formulations showed isotropy, nanometric size (below 25 nm), Newtonian behavior and pH ranging from 6.5 to 6.9. The MEs achieved a 2.5-fold increase in the flux and skin-permeated amount of α-bisabolol. ATR-FTIR results showed that microemulsions promoted fluidization and extraction of lipids and proteins of the stratum corneum, increasing the diffusion coefficient and partition coefficient of the drug in the skin. Additionally, F5E25 and F5EP25 showed higher activity against promastigotes (IC50 13.27 and 18.29, respectively) compared to unencapsulated α-bisabolol (IC50 53.8). Furthermore, F5E25 and F5EP25 also showed antileishmanial activity against intracellular amastigotes of L. amazonensis, with IC50 50 times lower than free α-bisabolol and high selectivity index (up to 15). Therefore, the systems obtained are favorable to topical administration, with significant antileishmanial activity against L. amazonensis promastigotes and amastigotes, being a promising system for future in vivo trials.

In vitro antileishmanial activity of thioridazine on amphotericin B unresponsive/ sensitive Leishmania donovani promastigotes and intracellular amastigotes.

The recent increase in the drug (liposomal amphotericin-B) unresponsive cases becomes hostile for the visceral leishmaniasis (VL) elimination target. The quest for new antileishmanial drugs is on the way and may demand more time. Meanwhile, drug repurposing is a quite promising option to explore further. We made such an attempt with thioridazine (TRZ), a first-line antipsychotic drug, which was reported for antimicrobial activity. In this study, we evaluated the drug activity of TRZ against amphotericin-B (Amp-B) sensitive and unresponsive Leishmania donovani promastigotes, as well as intracellular amastigotes (drug sensitive). We observed a potent antileishmanial activity of TRZ with significantly low half maximal inhibitory concentrations (IC50) on both the variants of promastigotes (0.61 ± 0.15 µM). These concentrations are comparable to the previously reported IC50 concentration of the current antileishmanial drug (Amp-B) against L. donovani. Light microscopy reveals the perturbations in promastigote morphology upon TRZ treatment. The in vitro studies on human macrophage cell lines determine the 50% cytotoxicity concentration (CC50) of TRZ on host cells as 20.046 µM and a half maximal effective concentration (EC50) as 0.91 µM during L. donovani infection, in turn selectivity index (SI) was calculated as 22.03 µM. Altogether, the results demonstrate that TRZ has the potential for drug repurposing and further studies on animal models could provide better insights for VL treatment.

Unilateral keratitis secondary to Leishmania spp. infection in a horse: Clinical signs and successful topical therapy.

Dermatological clinical signs have been seldom reported in the literature secondary to equine leishmaniasis. This case depicts the clinical signs, treatment, and outcome of a young horse with a pink, elevated lesion on the ventromedial quadrant of the cornea. A corneal cytology was performed and revealed the presence of leishmania amastigotes reaching the diagnosis of keratitis secondary to leishmania. Surgical resection was recommended but the owner declined the procedure, and the lesion was treated with a topical antimonial for 6 weeks. The lesion reduced remarkably during the first weeks of treatment. The patient had not shown recurrence of the lesion for 2 years since the treatment was started. Leishmania spp. can be responsible for ocular surface abnormalities such as keratitis. Corneal cytology is an inexpensive diagnostic method that should be considered when ocular surface abnormalities are identified in horses in endemic areas.

Design and synthesis of N-acyl and dimeric N-Arylpiperazine derivatives as potential antileishmanial agents.

The current regime for leishmaniasis is associated with several adverse effects, expensive, parenteral treatment for longer periods and the emergence of drug resistance. To develop affordable and potent antileishmanial agents, a series of N-acyl and homodimeric aryl piperazines were synthesized with high purity, predicted druggable properties by in silico methods and investigated their antileishmanial activity. The in vitro biological activity of synthesized compounds against clinically validated intracellular amastigote and extracellular promastigote form of Leishmania donovani parasite showed eight compounds inhibited 50% amastigotes growth below 25 µM. The half maximal inhibitory concentration (IC50) and cytotoxicity assessment of eight active compounds, 4a, 4d and 4e demonstrated activity with an IC50 2.0 - 9.1 µM and selectivity index 10 - 42. Compound 4d (IC50 2.0 µM, SI = 42) found to be the best among them with four-folds more potent and eight-folds less toxic than the control drug miltefosine. Overall, results demonstrated that compound 4d is a promising lead candidate for further development as antileishmanial drug.

Protozoal meningoencephalitis and myelitis in 4 dogs associated with Trypanosoma cruzi infection.

American trypanosomiasis is caused by the zoonotic protozoa Trypanosoma cruzi and primarily results in heart disease. Organisms also infect the central nervous system (CNS). The Texas A&M University veterinary teaching hospital archive was searched for dogs with CNS disease with intralesional protozoal amastigotes. This study summarizes 4 cases of dogs with disseminated trypanosomiasis and CNS involvement confirmed by quantitative polymerase chain reaction (qPCR) with T. cruzi primers. Clinical signs included lethargy, respiratory distress, tetraparesis, and seizures. Central nervous system lesions included meningeal congestion (1/4), necrosis with hemorrhage in the spinal cord gray and white matter (2/4), and histiocytic meningoencephalitis (4/4), and meningomyelitis (2/4) with intralesional and intracellular protozoal. Genotyping identified 1 case of T. cruzi discrete typing unit (DTU) TcI and 2 cases as TcIV, both are common variants in the United States. Trypanosomiasis should be considered a differential diagnosis for dogs with CNS signs in T. cruzi-endemic areas.

The cytostome-cytopharynx complex of intracellular and extracellular amastigotes of Trypanosoma cruzi exhibit structural and functional differences.

Endocytosis in Trypanosoma cruzi is mainly performed through a specialised membrane domain called cytostome-cytopharynx complex. Its ultrastructure and dynamics in endocytosis are well characterized in epimastigotes, being absent in trypomastigotes, that lack endocytic activity. Intracellular amastigotes also possess a cytostome-cytopharynx but participation in endocytosis of these forms is not clear. Extracellular amastigotes can be obtained from the supernatant of infected cells or in vitro amastigogenesis. These amastigotes share biochemical and morphological features with intracellular amastigotes but retain trypomastigote's ability to establish infection. We analysed and compared the ultrastructure of the cytostome-cytopharynx complex of intracellular amastigotes and extracellular amastigotes using high-resolution tridimensional electron microscopy techniques. We compared the endocytic ability of intracellular amastigotes, obtained through host cell lysis, with that of extracellular amastigotes. Intracellular amastigotes showed a cytostome-cytopharynx complex similar to epimastigotes'. However, after isolation, the complex undergoes ultrastructural modifications that progressively took to an impairment of endocytosis. Extracellular amastigotes do not possess a cytostome-cytopharynx complex nor the ability to endocytose. Those observations highlight morpho functional differences between intra and extracellular amastigotes regarding an important structure related to cell metabolism. TAKE AWAYS: T. cruzi intracellular amastigotes endocytose through the cytostome-cytopharynx complex. The cytostome-cytopharynx complex of intracellular amastigotes is ultrastructurally similar to the epimastigote. Intracellular amastigotes, once outside the host cell, disassembles the cytostome-cytopharynx membrane domain. Extracellular amastigotes do not possess a cytostome-cytopharynx either the ability to endocytose.

Quantitative monitoring of experimental and human leishmaniasis employing amastigote-specific genes.

The gold standard for diagnosis of leishmaniasis is the microscopic detection of amastigotes/Leishman Donovan (LD) bodies, but its moderate sensitivity necessitates the development of molecular approaches. This study aimed to quantify in experimental animal models and human leishmaniasis the expression of amastigote-specific virulence genes, A2 and amastin by droplet digital polymerase chain reaction (ddPCR). Total RNA was isolated from L. donovani-infected hamsters or murine peritoneal macrophages and lesional biopsies from patients with post kala-azar dermal leishmaniasis (PKDL). Following cDNA conversion, EvaGreen-based ddPCR was performed using specific primers for A2 or amastin and parasite load expressed in copies per µL. Assay was optimized and the specificity of amastigote-specific A2 and amastin was confirmed. In hepatic and splenic tissues of L. donovani-infected hamsters and peritoneal macrophages, ddPCR demonstrated a greater abundance of A2 than amastin. Treatment of L. donovani-infected peritoneal macrophages with conventional anti-leishmanials, miltefosine and amphotericin B translated into a dose-dependent reduction in copies per µL of A2 and amastin, and the extrapolated IC50 was comparable with results obtained by counting LD bodies in Giemsa-stained macrophages. Similarly, in dermal biopsies of patients with PKDL, A2 and amastin were detected. Overall, monitoring of A2 by ddPCR can be an objective measure of parasite burden and potentially adaptable into a high throughput approach necessary for drug development and monitoring disease progression when the causative species is L. donovani.

Direct In Vitro Comparison of the Anti-Leishmanial Activity of Different Olive Oil Total Polyphenolic Fractions and Assessment of Their Combined Effects with Miltefosine.

The bioactive compounds present in the edible products of the olive tree have been extensively studied and their favorable effects on various disease risk factors have been demonstrated. The aim of this study was to perform a comparative analysis of the anti-leishmanial effects of total phenolic fractions (TPFs) derived from extra virgin olive oil with different phenolic contents and diverse quantitative patterns. Moreover, the present study investigated their association with miltefosine, a standard anti-leishmanial drug, against both extracellular promastigotes and intracellular amastigotes of a viscerotropic and a dermotropic Leishmania strain. The chemical compositions of TPFs were determined by high performance liquid chromatography with diode array detection (HPLC-DAD). Analysis of parasite growth kinetics, reactive oxygen species production and apoptotic events were determined by microscopy and flow cytometry. Our results revealed that the presence of oleacein (OLEA) and oleocanthal (OLEO) secoiridoids enhances the anti-leishmanial effect of TPF. The association between TPFs and miltefosine was suggested as being additive in Leishmania infantum and Leishmania major promastigotes, and as antagonistic in intracellular amastigotes, as was evaluated with the modified isobologram method. The obtained data verified that TPFs are bioactive dietary extracts with a strong anti-leishmanial activity and highlighted that fractions that are richer in OLEA and OLEO phenolic compounds possess stronger inhibitory effects against parasites. This study may contribute to improving the therapeutic approaches against leishmaniasis.

In vivo reflectance confocal microscopic imaging of Leishmania amastigotes (Leishman bodies): A case report.

Cutaneous leishmaniasis (CL) is an intracellular parasitic infectious skin disease with a chronic self-limited course. In vivo reflectance confocal microscopy (RCM) findings in CL have been described in only two cases of CL. We report another case with RCM findings; however to our knowledge, this is the first demonstration of Leishmania amastigotes in RCM imaging. A centrally eroded reddish nodular lesion with a diameter of 12 mm was observed on the leg of a 36-years-old male with a 1-month history. On dermoscopy, a central yellowish crust, and irregularly distributed whitish opaque structures ranging in size and shape (round to polygonal) were observed. There were also irregular vessels mostly at the center and dotted/glomerular vessels at the periphery. On RCM, mild epidermal disarray with some scattered bright cells at the basal layer was observed. At the dermis, dense infiltration of polymorphic/roundish cells with heterogeneous reflectivity was seen. These large, mildly reflecting cells with fine granular structures in their cytoplasm were compatible with macrophages. Histopathology was concordant with CL. The Leishmania amastigotes seen as cytoplasmic granularity on RCM were the clue feature for the initial diagnosis.

Novel phenanthridine amide analogs as potential anti-leishmanial agents: In vitro and in silico insights.

In the current work, sixteen novel amide derivatives of phenanthridine were designed and synthesized using 9-fluorenone, 4-Methoxy benzyl amine, and alkyl/aryl acids. The characterization of the title compounds was performed using LCMS, elemental analysis, 1HNMR, 13CNMR and single crystal XRD pattern was also developed for compounds A8. All the final analogs were screened in vitro for anti-leishmanial activity against promastigote form of L. infantum strain. Among the tested analogs, four compounds (A-06, A-11, A-12, and A-15) exhibited significant anti-leishmanial activity with EC50 value ranges from 8.9 to 21.96 µM against amastigote forms of tested L. infantum strain with SI ranges of 1.0 to 4.3. From the activity results it was found that A-11 was the most active compound in both promastigote and amastigotes forms with EC50 values 8.53 and 8.90 µM respectively. In-silico ADME prediction studies depicted that the titled compounds obeyed Lipinski's rule of five as that of the approved marketed drugs. The predicted in-silico toxicity profile also confirmed that the tested compounds were non-toxic. Finally, molecular docking and molecular dynamics study was also performed for significantly active compound (A-11) in order to study it's putative binding pattern at the active site of the selected leishmanial trypanothione reductase target as well as to understand the stability pattern of target-ligand complex for 100 ns. Single crystal XRD of compound A-08 revealed that the compound crystallizes in monoclinic C2/c space group and showed interesting packing arrangements.

Pyrrolidine-based 3-deoxysphingosylphosphorylcholine analogs as possible candidates against neglected tropical diseases (NTDs): identification of hit compounds towards development of potential treatment of Leishmania donovani.

A rational-based process was adopted for repurposing pyrrolidine-based 3-deoxysphingosylphosphorylcholine analogs bearing variable acyl chains, different stereochemical configuration and/or positional relationships. Structural features were highly influential on activity. Amongst, enantiomer 1e having 1,2-vicinal relationship for the -CH2O- and the N-acyl moieties, a saturated palmitoyl chain and an opposite stereochemical configuration to natural sphingolipids was the most potent hit compound against promastigotes showing IC50 value of 28.32 µM. The corresponding enantiomer 1a was 2-fold less potent showing a eudismic ratio of 0.54 in promastigotes. Compounds 1a and 1e inhibited the growth of amastigotes more potently relative to promastigotes. Amongst, enantiomer 1a as the more selective and safer. In silico docking study using a homology model of Leishmania donovani inositol phosphoceramide synthase (IPCS) provided plausible reasoning for the molecular factors underlying the found activity. Collectively, this study suggests compounds 1a and 1e as potential hit compounds for further development of new antileishmanial agents.

Preparation and characterization of itraconazole loaded nanomicelles based on dextran-behenic acid for cutaneous leishmaniasis treatment.

Cutaneous leishmaniasis is known as the most prevalent clinical form of leishmaniasis. It needs the development of new therapies due to the serious side-effects promoted by taking the current drugs. In the present study, dextran-behenic acid (DEX-BA) based nanomicelles were developed to direct the delivery of itraconazole (ITZ) to the macrophages and enhance its toxic effects against Leishmania parasites. DEX-BA was synthesized through the esterification of dextran with behenic acid. The critical micelle concentration of the newly developed conjugate was evaluated using pyrene as the fluorescent probe. The nanomicelles were generated by the dialysis method; then they were optimized by applying a Box-Behnken design. The effects of the dialysis temperature, polymer content, and sonication time on the characteristics of micelles were subsequently studied. Furthermore, in vitro efficacy against Leishmania major promastigotes and parasite-infected macrophages was evaluated. The optimized formulation showed the particle size of 195.16 ± 3.06 nm, the polydispersity index of 0.39 ± 0.01, the zeta potential of -16.29 ± 0.89 mV, the encapsulation efficiency % of 56.11 ± 4.9, and the release efficiency % of 51.29 ± 1.97. According to scanning electron microscopy, the nanomicelles were found to be nearly spherical in shape. ITZ-loaded nanomicelles showed the strongest anti-leishmanial activities when compared with the free ITZ and drug-free nanomicelles. It could be, therefore, concluded that ITZ-loaded nanomicelles might be useful as an alternative therapy for the treatment of cutaneous leishmania.

In Vitro Antileishmanial and Antischistosomal Activities of Anemonin Isolated from the Fresh Leaves of Ranunculus multifidus Forsk.

Leishmaniasis and schistosomiasis are neglected tropical diseases (NTDs) infecting the world's poorest populations. Effectiveness of the current antileishmanial and antischistosomal therapies are significantly declining, which calls for an urgent need of new effective and safe drugs. In Ethiopia fresh leaves of Ranunculus multifidus Forsk. are traditionally used for the treatment of various ailments including leishmaniasis and eradication of intestinal worms. In the current study, anemonin isolated from the fresh leaves of R. multifidus was assessed for its in vitro antileishmanial and antischistosomal activities. Anemonin was isolated from the hydro-distilled extract of the leaves of R. multifidus. Antileishmanial activity was assessed on clinical isolates of the promastigote and amastigote forms of Leishmania aethiopica and L. donovani clinical isolates. Resazurin reduction assay was used to determine antipromastigote activity, while macrophages were employed for antiamastigote and cytotoxicity assays. Antischistosomal assays were performed against adult Schistosoma mansoni and newly transformed schistosomules (NTS). Anemonin displayed significant antileishmanial activity with IC50 values of 1.33 nM and 1.58 nM against promastigotes and 1.24 nM and 1.91 nM against amastigotes of L. aethiopica and L. donovani, respectively. It also showed moderate activity against adult S. mansoni and NTS (49% activity against adult S. mansoni at 10 µM and 41% activity against NTS at 1 µM). The results obtained in this investigation indicate that anemonin has the potential to be used as a template for designing novel antileishmanial and antischistosomal pharmacophores.

Purified Splenic amastigotes of Leishmania donovani-Immunoproteomic approach for exploring Th1 stimulatory polyproteins.

Visceral leishmaniasis (VL) represents one of the most challenging infectious diseases worldwide. The reason that once infected, patient develops immunity against Leishmania parasite has paved way to develop prophylactic vaccines against disease, but only some of these have moved ahead for clinical trials. Herein, the study to explore novel and potential vaccine candidates was extended to pathogenic form of parasite, that is, amastigote form which is less explored due to complexity of its purification process. Methods and results. Classical protocol of purification of splenic amastigotes was modified to obtain highly pure amastigotes which was confirmed by Western blotting in support with proteomics studies. Fractionation and sub-fractionation of purified splenic amastigotes revealed four sub-fractions, belonging to 97 to 68 kDa and 68 to 43 kDa ranges, which showed long-lasting protection with remarkable Th1-type cellular responses in hamsters vaccinated with these sub-fractions (LTT, NO, QRT-PCR). Further proteomics analysis, to identify and understand the precise nature and function of these protective protein sub-fractions, identified a total of 47 proteins including twenty-five hypothetical proteins/unknowns. Amastigote stage has potential Th1-stimulatory vaccine candidates, notably, among identified proteins, major were uncharacterized proteins/hypothetical proteins, which once characterized may serve as novel and potential vaccine candidates/drug targets.

Nucleotides and AHCC Enhance Th1 Responses In Vitro in Leishmania-Stimulated/Infected Murine Cells.

A stronger Th1 (cellular) immune response in canine leishmaniosis (CanL) leads to a better prognosis. Dietary nucleotides plus AHCC® have shown beneficial effects in dogs with clinical leishmaniosis and in clinically healthy Leishmania-infected dogs. The potential leishmanicidal activity of nucleotides and AHCC was assessed by quantifying nitric oxide (NO) production and replication of parasites. Their effects on lymphocyte proliferation were studied with and without soluble Leishmania infantum antigen (SLA) stimulation. Cytokine level variations were assessed using naïve and L. infantum-infected macrophages/lymphocytes cocultures. Promastigotes and amastigotes proliferation and NO macrophage production were not directly affected. Lymphocyte proliferation was significantly enhanced by nucleotides, AHCC, and their combinations only after SLA stimulation. Nucleotides and AHCC significantly increased the production of IL-1ß, IL-2, IL-5, IL-9, IL-10, and IL-12 by naïve immune cells. In naïve and L. infantum-infected macrophage/lymphocyte cocultures, nucleotides with or without AHCC led to significant increases in IFN-γ and TNF-α. Given that these cytokines are involved in the effective Th1 immune response against Leishmania parasites, these mechanism of action could explain the previously reported in vivo clinical efficacy of such combination and further support the use of nucleotides with or without AHCC in the management of CanL patients.

Biological evaluation and structure activity relationship of 9-methyl-1-phenyl-9H-pyrido[3,4-b]indole derivatives as anti-leishmanial agents.

A series of piperazinyl-ß-carboline-3-carboxamide derivatives were designed through a molecular hybridization approach. Designed analogues were synthesized, characterized and evaluated for anti-leishmanial activity against Leishmania infantum and Leishmania donovani. In L. infantum inhibition assay, compounds 7d, 7g and 7c displayed potent inhibition of promastigotes (EC50 1.59, 1.47 and 3.73 µM respectively) and amastigotes (EC50 1.4, 1.9 and 2.6 µM respectively). SAR studies revealed that, para substitution of methoxy, chloro groups and methyl group on ortho position favored anti-leishmanial activity against L. infantum. Among these analogues 7d, 7h, 7n and 7g exhibited potent inhibition against L. donovani promastigotes (EC50 0.91, 4.0, 4.57 and 5.02 µM respectively), axenic amastigotes (EC50 0.9, 3.5, 2.2 and 3.8 µM respectively) and intracellular amastigotes (EC50 1.3, 7.8, 5.6 and 6.3 µM respectively). SAR studies suggested that, para substitution of methoxy group, para and meta substitution of chloro groups and benzyl replacement recommended for significant anti-leishmanial against L. donovani.

The activity of encapsulated meglumine antimoniate in stearylamine-bearing liposomes against cutaneous leishmaniasis in BALB/c mice.

Topical treatment of cutaneous leishmaniasis has demonstrated appropriate alternative for reducing toxicity of conventional treatments, improving patients' compliance and reducing treatment costs. Furthermore, outbreak of cutaneous leishmaniasis in war and conflict zones emerges finding an effective, economical and user-friendly treatment. In the context of liposomal topical drug delivery, we developed and characterized meglumine antimoniate (MA) loaded liposomes and investigated their effectiveness in topical treatment of cutaneous leishmaniasis. Previously, we showed the promising use of liposomal formulation of MA in treatment of cutaneous leishmaniasis in BALB/c mice. Here, we included Stearylamine (SA) in liposomes' structure which has antileishmanial activity by itself. . Size and encapsulation efficiency of liposomes were measured and in vitro permeation was performed using mice model. In vitro toxicity of liposomes was measured against leishmania promastigotes and amastigotes. Liposomes were used topically twice a day for 4 weeks to treat leishmania lesions in BALB\c mice model. In vitro permeation study showed liposomal formulations improved the percent of MA permeation compared with MA-cream. Promastigotes and amastigotes assay results showed significant enhanced toxicity in Liposomal-MA containing SA compared to Lip-MA. In BALB\c mice model of cutaneous leishmaniasis, liposomal groups exhibited significantly smaller lesion size compared to control groups (p < 0.01). In addition, the spleen parasite burden was significantly (P < 0.01) lower in mice treated with selected liposomal MA than in mice treated with PBS, control liposomes and MA cream. The results of this study showed that SA-liposomes encapsulated with MA might be useful as a candidate for the topical treatment of CL which merit further investigation.

The antidepressant clomipramine induces programmed cell death in Leishmania amazonensis through a mitochondrial pathway.

Despite many efforts, the currently available treatments for leishmaniasis are not fully effective. To discover new medications, drug repurposing arises as a promising strategy. We present data that supports the use of the antidepressant clomipramine against Leishmania amazonensis. The drug presented selective activity at micromolar range against both the parasite forms and stimulated nitric oxide production in host macrophages. Regarding the mechanism of action, clomipramine led parasites do mitochondrial depolarization, which coupled with the inhibition of trypanothione reductase induced strong oxidative stress in the parasites. The effects observed in promastigotes included lipoperoxidation, plasma membrane permeabilization, and apoptosis hallmarks (i.e., DNA fragmentation, phosphatidylserine exposure, and cell shrinkage). The mechanism of action in both parasitic forms was quite similar, but amastigotes also exhibited energetic stress, reflected by a reduction of adenosine triphosphate levels. Such differential effects might be attributable to the metabolic particularities of each form of the parasitic. Ultrastructural alterations of the endomembrane system and autophagy were also observed, possibly indicating an adaptive response to oxidative stress. Our results suggest that clomipramine interferes with the redox metabolism of L. amazonensis. In spite of the cellular responses to recover the cellular homeostasis, parasites underwent programmed cell death.

Hybrids of Cinchona Alkaloids and Bile Acids as Antiparasitic Agents Against Trypanosoma cruzi.

The current chemotherapy of Chagas disease needs to be urgently improved. With this aim, a series of 16 hybrids of Cinchona alkaloids and bile acids were prepared by functionalization at position C-2 of the quinoline nucleus by a radical attack of a norcholane substituent via a Barton-Zard decarboxylation reaction. The antitrypanosomal activity of the hybrids was tested on different stages and strains of T. cruzi. In particular, eight out of 16 hybrids presented an IC50 ≤1 µg/mL against trypomastigotes of the CL Brener strain and/or a selectivity index higher than 10. These promising hybrids yielded similar results when tested on trypomastigotes from the RA strain of T. cruzi (discrete typing unit-DTU-VI). Surprisingly, trypomastigotes of the Y strain (DTU II) were more resistant to benznidazole and to most of the hybrids than those of the CL Brener and RA strains. However, the peracetylated and non-acetylated forms of the cinchonine/chenodeoxycholic bile acid conjugate 4f and 5f were the most trypanocidal hybrids against Y strain trypomastigotes, with IC50 values of 0.5 and 0.65 µg/mL, respectively. More importantly, promising results were observed in invasion assays using the Y strain, where hybrids 5f and 4f induced a significant reduction in intracellular amastigotes and on the release of trypomastigotes from infected cells.

Protective effect of aspirin treatment on mouse behavior in the acute phase of experimental infection with Trypanosoma cruzi.

Chagas disease is a potentially fatal disease caused by the parasite Trypanosoma cruzi, which can in some cases affect the central nervous system. The objective was to evaluate the effect of aspirin (ASA) in the behavior of mice infected with T. cruzi during the acute phase. This was an experimental study with random assignation. Twenty four BALB/c mice were divided into four groups of six animals each as follows: only ASA (OA), ASA before infection (BI), ASA after infection (AI) and only infection (OI). The strain used for infection was M/HOM/Bra/53/Y. An ASA dose of 100 mg/kg per day was administered 72 h before infection to BI group and the same dose 48 h after infection to AI group. Mice behavior in the open field test, mortality, and brain histopathology was evaluated. Data were analyzed using ANOVA, chi square test, and Kaplan-Meier with long-rank for survival analysis. In the open field test, the OA group has similar results with the BI group, in the variables of immobility and escape. Also, the OA group displayed significantly higher rates of micturition (p < 0.001) and defecation (p < 0.001) compared to infected groups. Mortality was higher in BI group (p = 0.02). The presence of T. cruzi amastigotes were higher in brain tissues of the AI and OI groups (p = 0.008). In conclusion, the administration of ASA before infection seemed to prevent behavioral changes induced by the acute infection, but it led to accelerated mortality. The study highlighted the potential importance of the pathways inhibited by ASA in the early hours of acute infection with T. cruzi.