alina perrone

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Papers by alina perrone

Trypanosoma cruzi Secreted Cyclophilin TcCyP19 as an Early Marker for Trypanocidal Treatment Efficiency

International Journal of Molecular Sciences

Cyclophilins (CyPs) are a family of enzymes involved in protein folding. Trypanosoma cruzi, the c... more Cyclophilins (CyPs) are a family of enzymes involved in protein folding. Trypanosoma cruzi, the causative agent of Chagas disease, has a 19-kDa cyclophilin, TcCyP19, that was found to be secreted in parasite stages of the CL Brener clone and recognized by sera from T. cruzi-infected mice and patients. The levels of specific antibodies against TcCyP19 in T. cruzi-infected mice and subjects before and after drug treatment were measured by an in-house enzyme linked immunosorbent assay (ELISA). Mice in the acute and chronic phase of infection, with successful trypanocidal treatments, showed significantly lower anti-TcCyP19 antibody levels than untreated mice. In children and adults chronically infected with T. cruzi, a significant decrease in the anti-TcCyP19 titers was observed after 12 months of etiological treatment. This decrease was maintained in adult chronic patients followed-up 30–38 months post-treatment. These results encourage further studies on TcCyP19 as an early biomarker ...

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A homolog of cyclophilin D is expressed in Trypanosoma cruzi and is involved in the oxidative stress-damage response

Cell death discovery, 2017

Mitochondria have an important role in energy production, homeostasis and cell death. The opening... more Mitochondria have an important role in energy production, homeostasis and cell death. The opening of the mitochondrial permeability transition pore (mPTP) is considered one of the key events in apoptosis and necrosis, modulated by cyclophilin D (CyPD), a crucial component of this protein complex. In Trypanosoma cruzi, the protozoan parasite that causes Chagas disease, we have previously described that mitochondrial permeability transition occurs after oxidative stress induction in a cyclosporin A-dependent manner, a well-known cyclophilin inhibitor. In the present work, a mitochondrial parasite cyclophilin, named TcCyP22, which is homolog to the mammalian CyPD was identified. TcCyP22-overexpressing parasites showed an enhanced loss of mitochondrial membrane potential and loss of cell viability when exposed to a hydrogen peroxide stimulus compared with control parasites. Our results describe for the first time in a protozoan parasite that a mitochondrial cyclophilin is a component of...

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Improved Immuno-Detection of a Low-Abundance Cyclophilin Allows the Confirmation of its Expression in a Protozoan Parasite

Immunochemistry & Immunopathology, 2015

Protein samples can be challenging to analyze due to the presence of high-abundance proteins mask... more Protein samples can be challenging to analyze due to the presence of high-abundance proteins masking lowabundance proteins of interest, such as biomarkers and novel physiological mediators. Cyclophilins are chaperones involved in the cis/trans isomerization of peptidyl-prolyl bonds in peptides or proteins and have been found in every organism sequenced to date. Although considerable progress has been made in the characterization of some cyclophilins expressed in diverse parasites invading humans, the main aspects of low-abundance members of this family remain unknown. In the present work, we present that the combined strategy of using more specific antibodies and increasing the presence of subcellular proteins in the sample, allowed us to confirm the expression of a 21.1 kDa cyclophilin for the first time in Trypanosoma cruzi.

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Oxidative stress damage in the protozoan parasite Trypanosoma cruzi is inhibited by Cyclosporin A

Parasitology, Jan 31, 2015

Cyclosporin A (CsA) specifically inhibits the mitochondrial permeability transition pore (mPTP). ... more Cyclosporin A (CsA) specifically inhibits the mitochondrial permeability transition pore (mPTP). Opening of the mPTP, which is triggered by high levels of matrix [Ca2+] and/or oxidative stress, leads to mitochondrial dysfunction and thus to cell death by either apoptosis or necrosis. In the present study, we analysed the response of Trypanosoma cruzi epimastigote parasites to oxidative stress with 5 mm H2O2, by studying several features related to programmed cell death and the effects of pre-incubation with 1 μ m of CsA. We evaluated TcPARP cleavage, DNA integrity, cytochrome c translocation, Annexin V/propidium iodide staining, reactive oxygen species production. CsA prevented parasite oxidative stress damage as it significantly inhibited DNA degradation, cytochrome c translocation to cytosol and TcPARP cleavage. The calcein-AM/CoCl2 assay, used as a selective indicator of mPTP opening in mammals, was also performed in T. cruzi parasites. H2O2 treatment decreased calcein fluorescen...

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How to Improve the Early Diagnosis of Trypanosoma cruzi Infection: Relationship between Validated Conventional Diagnosis and Quantitative DNA Amplification in Congenitally Infected Children

PLoS Neglected Tropical Diseases, 2013

Background: According to the Chagas congenital transmission guides, the diagnosis of infants, bor... more Background: According to the Chagas congenital transmission guides, the diagnosis of infants, born to Trypanosoma cruzi infected mothers, relies on the detection of parasites by INP micromethod, and/or the persistence of T. cruzi specific antibody titers at 10-12 months of age. Methodology and Principal Findings: Parasitemia levels were quantified by PCR in T. cruzi-infected children, grouped according to the results of one-year follow-up diagnosis: A) Neonates that were diagnosed in the first month after delivery by microscopic blood examination (INP micromethod) (n = 19) had a median parasitemia of 1,700 Pe/mL (equivalent amounts of parasite DNA per mL); B) Infants that required a second parasitological diagnosis at six months of age (n = 10) showed a median parasitemia of around 20 Pe/mL and 500 Pe/mL at 1 and 6 months old, respectively, and C) babies with undetectable parasitemia by three blood microscopic observations but diagnosed by specific anti-T. cruzi serology at around 1 year old, (n = 22), exhibited a parasitemia of around 5 Pe/mL, 800 Pe/mL and 20 Pe/mL 1, 6 and 12 month after delivery, respectively. T. cruzi parasites were isolated by hemoculture from 19 congenitally infected children, 18 of which were genotypified as DTU TcV, (former lineage TcIId) and only one as TcI. Significance: This report is the first to quantify parasitemia levels in more than 50 children congenitally infected with T. cruzi, at three different diagnostic controls during one-year follow-up after delivery. Our results show that the parasite burden in some children (22 out of 51) is below the detection limit of the INP micromethod. As the current trypanocidal treatment proved to be very effective to cure T. cruzi-infected children, more sensitive parasitological methods should be developed to assure an early T. cruzi congenital diagnosis.

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Morphological and biological characterization of cell line developed from bovine Echinococcus granulosus

In Vitro Cellular & Developmental Biology - Animal, 2010

The taeniid tapeworm Echinococcus granulosus is the causative agent of echinococcal disease, a ma... more The taeniid tapeworm Echinococcus granulosus is the causative agent of echinococcal disease, a major zoonosis with worldwide distribution. Several efforts to establish an in vitro model of E. granulosus have been undertaken; however, many of them have been designed for Echinococcus multilocularis. In the present study, we have described and characterized a stable cell line obtained from E. granulosus bovine protoscoleces maintained 3 yr in vitro. Growth characterization, morphology by light, fluorescent and electronic microscopy, and karyotyping were carried out. Cell culture origin was confirmed by immunofluorescent detection of AgB4 antigen and by PCR for the mitochondrial cytochrome c-oxidase subunit 1 (DCO1) gene. Cells seeded in agarose biphasic culture resembled a cystic structure, similar to the one formed in secondary hosts. This cell line could be a useful tool to research equinococcal behavior, allowing additional physiological and pharmacological studies, such as the effect of growth factors, nutrients, and antiparasitic drugs on cell viability and growth and on cyst formation.

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Biological markers for evaluating therapeutic efficacy in Chagas disease, a systematic review

Expert Review of Anti-infective Therapy, 2014

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Trypanosoma cruzi: Biological characterization of a isolate from an endemic area and its susceptibility to conventional drugs

Experimental Parasitology, 2010

We describe some biological and molecular characteristics of a Trypanosoma cruzi isolate derived ... more We describe some biological and molecular characteristics of a Trypanosoma cruzi isolate derived from a Triatomine captured in Nicaragua. PCR based typification showed that this isolate, named Nicaragua, belonged to the lineage Tc I. Nicaragua infected culture cells were treated with allopurinol, showing different behaviour according to the cellular compartment, being cardiomyocyte primary cultures more resistant to this drug. The course of the infection in a mice experimental model and its susceptibility to benznidazole and allopurinol was analysed. In benznidazole treatment, mice reverted the high lethal effect of parasites during the acute infection, however, a few parasites were detected in the heart of 88 % of mice one year post-infection. Since Trypanosoma cruzi is a heterogeneous species population it is important to study and characterize different parasites actually circulating in humans in endemic areas. In this work we show that T. cruzi Nicaragua isolate, is sensitive to early benznidazole treatment.

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Estudios funcionales de la ciclofilina de 19 kDa de Trypanosoma cruzi con células y proteínas de mamífero

Circulating cytokine and chemokine profiles of Trypanosoma cruzi-infected women during pregnancy and its association with congenital transmission

The Journal of Infectious Diseases

Background Trypanosoma cruzi, the causative agent of Chagas disease, can be transmitted to the of... more Background Trypanosoma cruzi, the causative agent of Chagas disease, can be transmitted to the offspring of infected women, which constitutes an epidemiologically significant parasite transmission route in non-endemic areas. It is relevant to evaluate differentially expressed factors in T. cruzi-infected pregnant women as potential markers of Chagas congenital transmission. Methods Circulating levels of twelve cytokines and chemokines were measured by ELISA or cytometric bead array in T. cruzi-infected and uninfected pregnant women in their second trimester of pregnancy, and control groups of T. cruzi-infected and uninfected non-pregnant women. Results T. cruzi-infected women showed a pro-inflammatory Th1-biased profile, with increased levels of TNF-α, IL-12p70, IL-15 and MIG. Uninfected pregnant women presented a biased response towards Th2/Th17/Treg profiles, with increased plasma levels of IL-5, IL-6, IL-1β, IL-17A and IL-10. Finally, we identified that high parasitemia together ...

Trypanosoma cruzi infection in Cyclophilin D deficient mice

Experimental Parasitology

Transmigration of Trypanosoma cruzi trypomastigotes through 3D cultures resembling a physiological environment

Cellular Microbiology

Trypanosoma cruzi Infection at the Maternal-Fetal Interface: Implications of Parasite Load in the Congenital Transmission and Challenges in the Diagnosis of Infected Newborns

Frontiers in Microbiology

Trypanosoma cruzi is the protozoan unicellular parasite that causes Chagas disease. It can be tra... more Trypanosoma cruzi is the protozoan unicellular parasite that causes Chagas disease. It can be transmitted from infected mothers to their babies via the connatal route, thus being able to perpetuate even in the absence of Triatomine insect vectors. Chagas disease was originally endemic in Central and South America, but migration of infected women of childbearing age has spread the T. cruzi congenital infection to non-endemic areas like North America, Europe, Japan, and Australia. Currently, 7 million people are affected by this infection worldwide. This review focuses on the relevance of the T. cruzi parasite levels in different aspects of the congenital T. cruzi infection such as the mother-to-child transmission rate, the maternal and fetal immune response, and its impact on the diagnosis of infected newborns. Improvements in detection of this parasite, with tools that can be easily adapted to be used in remote rural areas, will make the early diagnosis of infected children possible, allowing a prompt trypanocidal treatment and avoiding the current loss of opportunities for the diagnosis of 100% of T. cruzi congenitally infected infants.

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Phenotypic diversity and drug susceptibility of Trypanosoma cruzi TcV clinical isolates

PloS one, 2018

Trypanosoma cruzi is a genetically heterogeneous group of organisms that cause Chagas disease. It... more Trypanosoma cruzi is a genetically heterogeneous group of organisms that cause Chagas disease. It has been long suspected that the clinical outcome of the disease and response to therapeutic agents are, at least in part, related to the genetic characteristics of the parasite. Herein, we sought to validate the significance of the genotype of T. cruzi isolates recovered from patients with different clinical forms of Chagas disease living in Argentina on their biological behaviour and susceptibility to drugs. Genotype identification of the newly established isolates confirmed the reported predominance of TcV, with a minor frequency of TcI. Epimastigote sensitivity assays demonstrated marked dissimilar responses to benznidazole, nifurtimox, pentamidine and dihydroartemisinin in vitro. Two TcV isolates exhibiting divergent response to benznidazole in epimastigote assays were further tested for the expression of anti-oxidant proteins. Benznidazole-resistant BOL-FC10A epimastigotes had dec...

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A Functional Analysis of the Cyclophilin Repertoire in the Protozoan Parasite Trypanosoma Cruzi

Biomolecules

Trypanosoma cruzi is the etiological agent of Chagas disease. It affects eight million people wor... more Trypanosoma cruzi is the etiological agent of Chagas disease. It affects eight million people worldwide and can be spread by several routes, such as vectorborne transmission in endemic areas and congenitally, and is also important in non-endemic regions such as the United States and Europe due to migration from Latin America. Cyclophilins (CyPs) are proteins with enzymatic peptidyl-prolyl isomerase activity (PPIase), essential for protein folding in vivo. Cyclosporin A (CsA) has a high binding affinity for CyPs and inhibits their PPIase activity. CsA has proved to be a parasiticidal drug on some protozoa, including T. cruzi. In this review, we describe the T. cruzi cyclophilin gene family, that comprises 15 paralogues. Among the proteins isolated by CsA-affinity chromatography, we found orthologues of mammalian CyPs. TcCyP19, as the human CyPA, is secreted to the extracellular environment by all parasite stages and could be part of a complex interplay involving the parasite and the ...

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Some Limitations for Early Diagnosis of Congenital Chagas Infection by PCR

Pediatrics

Trypanosoma cruzi is the etiological agent of Chagas disease. This hemoflagellate protozoan paras... more Trypanosoma cruzi is the etiological agent of Chagas disease. This hemoflagellate protozoan parasite is genetically diverse and classified into 6 discrete typing units (DTUs), T cruzi discrete typing unit I (TcI) through T cruzi discrete typing unit VI (TcVI). Regarding the T cruzi DTUs infecting humans, TcI is found mainly in the north of the Amazon basin but is usually associated with the sylvatic cycle throughout the American continent. T cruzi DTU II is found mainly in the Atlantic and central Brazil, whereas T cruzi discrete typing unit V (TcV) and TcVI are found in the American Southern Cone, in the Gran Chaco area. 1, 2

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