ST8Sia2 polysialyltransferase protects against infection by Trypanosoma cruzi.

Glycosylation is one of the most structurally and functionally diverse co- and post-translational modifications in a cell. Addition and removal of glycans, especially to proteins and lipids, characterize this process which has important implications in several biological processes. In mammals, the repeated enzymatic addition of a sialic acid unit to underlying sialic acids (Sia) by polysialyltransferases, including ST8Sia2, leads to the formation of a sugar polymer called polysialic acid (polySia). The functional relevance of polySia has been extensively demonstrated in the nervous system. However, the role of polysialylation in infection is still poorly explored. Previous reports have shown that Trypanosoma cruzi (T. cruzi), a flagellated parasite that causes Chagas disease (CD), changes host sialylation of glycoproteins. To understand the role of host polySia during T. cruzi infection, we used a combination of in silico and experimental tools. We observed that T. cruzi reduces both the expression of the ST8Sia2 and the polysialylation of target substrates. We also found that chemical and genetic inhibition of host ST8Sia2 increased the parasite load in mammalian cells. We found that modulating host polysialylation may induce oxidative stress, creating a microenvironment that favors T. cruzi survival and infection. These findings suggest a novel approach to interfere with parasite infections through modulation of host polysialylation.

Generation of genetically modified human induced pluripotent stem cell lines harboring haploin sufficient or dominant negative variants in the FBN1 gene.

Marfan Syndrome (MFS) is an autosomal dominant connective tissue disorder caused by mutations in the FBN1 gene. To investigate the molecular mechanisms of pathogenesis for the syndrome, we genetically modified the FBN1 gene in a line of induced pluripotent stem cells (hiPSCs) derived from a healthy donor using the CRISPR/Cas9 gene editing technology. The sublines described here were characterized according to established criteria and were shown to maintain pluripotency, three germ layer differentiation potential and genomic integrity. These clones can now be used to better understand the pathogenesis of MFS in different cell types.

Generation of 6 lines of human pluripotent stem cells from hypertensive patients and 3 lines of human pluripotent stem cell from normotensive patients.

Hypertension is a complex multifactorial disease characterized by a chronic increase of arterial pressure. Ninety percent of the cases are idiopathic and thus classified as essential hypertension. Uncontrolled arterial pressure has devasting consequences including cardiac insufficiency, stroke, dementia, chronic renal disease, ischemic heart disease and death. The hiPSC lines described here from six hypertensive patients and three controls were characterized according to established criteria and were shown to maintain pluripotency, differentiation into the three germ layers and genomic integrity. These cell lines can contribute to the understanding of the molecular mechanisms involved in hypertension in different cell types.

Generation of two human induced pluripotent stem cell (hiPSC) lines derived from unrelated Marfan Syndrome patients.

Marfan Syndrome (MFS) is a pleiotropic and autosomal dominant condition caused by pathogenic variants in FBN1. Although fully penetrant, clinical variability is frequently observed among patients and there are only few genotype-phenotype correlations described so far. Here, we describe the generation and characterization of hiPSC lines derived from two unrelated MFS patients harboring heterozygous variants in FBN1. Human iPSCs were obtained from erythroblasts reprogrammed with episomal vectors carrying the reprogramming factors OCT4, SOX2, KLF4, c-MYC and LIN-28, and characterized according to established criteria. Differentiated cells demonstrated different patterns of fibrillin-1 expression suggesting different molecular mechanisms between the two patients.

Human induced pluripotent stem cells as a tool for disease modeling and drug screening for COVID-19.

The emergence of the new corona virus (SARS-CoV-2) and the resulting COVID-19 pandemic requires fast development of novel prevention and therapeutic strategies. These rely on understanding the biology of the virus and its interaction with the host, and on agnostic phenotypic screening for compounds that prevent viral infection. In vitro screenings of compounds are usually performed in human or animal-derived tumor or immortalized cell lines due to their ease of culturing. However, these platforms may not represent the tissues affected by the disease in vivo, and therefore better models are needed to validate and expedite drug development, especially in face of the COVID-19 pandemic. In this scenario, human induced pluripotent stem cells (hiPSCs) are a powerful research tool due to their ability to generate normal differentiated cell types relevant for the disease. Here we discuss the different ways hiPSCs can contribute to COVID-19 related research, including modeling the disease in vitro and serving as a platform for drug screening.

Trypanosoma cruzi infection of human induced pluripotent stem cell-derived cardiomyocytes: an in vitro model for drug screening for Chagas disease.

Chagas disease, caused by Trypanosoma cruzi, is an important global public health problem which, despite partial efficacy of benznidazole (Bz) in acute phase, urgently needs an effective treatment. Cardiotoxicity is a major safety concern for conduction of more accurate preclinical drug screening platforms. Human induced pluripotent stem cells derived cardiomyocytes (hiPSC-CM) are a reliable model to study genetic and infectious cardiac alterations and may improve drug development. Herein, we introduce hiPSC-CM as a suitable model to study T. cruzi heart infection and to predict the safety and efficacy of anti-T. cruzi drugs.

Monitoring cell line identity in collections of human induced pluripotent stem cells.

The ability to reprogram somatic cells into induced pluripotent stem cells (hiPSCs) has led to the generation of large collections of cell lines from thousands of individuals with specific phenotypes, many of which will be shared among different research groups as invaluable tools for biomedical research. As hiPSC-based research involves extensive culture of many cell lines, the issue periodic cell line identification is particularly important to ensure that cell line identity remains accurate. Here we analyzed the different commercially available genotyping methods considering ease of in-house genotyping, cost and informativeness, and applied one of them in our workflow for hiPSC generation. We show that the chosen STR method was able to establish a unique DNA profile for each of the 35 individuals/hiPSC lines at the examined sites, as well as identify two discrepancies resulting from inadvertently exchanged samples. Our results highlight the importance of hiPSC line genotyping by an in-house method that allows periodic cell line identification and demonstrate that STR is a useful approach to supplement less frequent karyotyping and epigenetic evaluations.