Biology subjects

Zatz, M.

Publications and source records attributed to Zatz, M..

12 recordsLinked to original sources

bioRxiv2025

FOXM1 Inhibition Promotes Polyploidization and Metabolic Maturation in Human iPSC-Derived Hepatocytes by Modulating the Wnt/β-Catenin Pathway

Human induced pluripotent stem cell (iPSCs)-derived hepatocytes are widely used in regenerative medicine and disease modeling. However, existing protocols mainly produce fetal-like cells, limiting accurate modeling of liver functionality. Topoisomerase II (TOP2) and its transcription factor, forkhead box M1 (FOXM1), are silenced during late liver embryonic development; however, their roles in hepatocyte differentiation remain unclear. Here, we examined the effects of TOP2 and FOXM1 inhibition on the terminal differentiation of hepatocytes. We found that subtoxic TOP2 inhibition reduced nuclear chromatin condensation without causing DNA damage. RNA-seq analysis showed that TOP2 inhibition induced cell cycle arrest in a TOP2A-selective manner, with FOXM1 downregulation. ATAC-seq validation demonstrated that TOP2A inhibition decreases chromatin accessibility and modulates the Wnt/{beta}-catenin pathway. Proteomic analysis revealed that FOXM1 inhibition modulated TOP2A expression, replicated TOP2A-mediated cell cycle arrest, and reduced the levels of fetal hepatocyte proteins (HBG1/2, UGT2B7, and AFP). Prolonged FOXM1 inhibition is correlated with increased hepatocyte polyploidization, enhanced CYP450 activity, and improved lipid metabolism, suggesting a potential role in these processes. Overall, our findings suggest that FOXM1 inhibition significantly promotes the terminal differentiation of human iPSC-derived hepatocytes, indicating a potential role for FOXM1 and TOP2A in liver development, regeneration, and disease.

cell biology↗

bioRxiv2025

iPSC-derived skeletal muscle spheroids for Duchenne Muscular Dystrophy modeling

BackgroundThe progressive skeletal muscle degeneration observed in Duchenne Muscular Dystrophy (DMD) patients requires multiple cycles of satellite cells (SCs) activation to promote tissue regeneration. Dystrophic SCs present intrinsic defects, and the disrupting fibrotic niche hinders appropriate muscle recovery. Traditional 2D culture systems face challenges in modeling the DMD muscle niche and SCs behavior. Our aim was to validate a 3D culture of skeletal muscle spheroids (iSMS) for DMD modeling, as compared to the traditional 2D culture, while investigating the pathophysiological mechanisms of dystrophin deficiency in vitro. MethodsTo compare iSMS with traditional 2D myogenic differentiation, we differentiated PAX7 reporter wild-type (WT), dystrophic (DMD) isogenic induced pluripotent stem cells (iPSCs), and patients iPSCs, characterized myogenic markers levels and assessed differences in proliferation and differentiation using RT-qPCR, immunofluorescence, and flow cytometry. ResultsOur data showed that although both 2D and iSMS culture systems generated myogenic progenitors positive for MYOD, MYOG, MYF5, and MYH3, iSMS improved PAX7 expression in vitro. Moreover, we identified a differential regulation of canonical Notch signaling genes between iSMS and 2D, which may influence the comparison between WT and DMD. We also characterized the differentiation of myogenic progenitors derived from 2D and iSMS towards elongated myofibers, providing a valuable comparison with muscle fibers differentiated from human primary myoblasts. Additionally, DMD iSMS derived progenitors proliferated at reduced levels compared with WT iSMS, a characteristic not observed in progenitors derived from 2D cultures. Finally, we performed iSMS and 2D myogenic differentiation of iPSC lines from three patients with DMD, thus validating the iSMS protocol for DMD modeling. ConclusionOur results highlight the important advantages of using the iSMS differentiation platform over 2D for disease modeling. Exploring these 3D systems may help to gain a deeper understanding of SCs behavior to advance in novel treatments for DMD, which might be applicable to other forms of muscular disorders.

cell biology↗

bioRxiv2024

Multiomics analysis reveals key immunogenic signatures induced by oncolytic Zika virus infection of paediatric brain tumour cells

Brain tumours disproportionately affect children and are the largest cause of paediatric cancer-related death. Despite decades of research, paediatric standard-of-care therapy still predominantly relies on surgery, radiotherapy, and systemic use of cytotoxic chemotherapeutic agents, all of which can result in debilitating acute and late effects. Novel therapies that engage the immune system, such as oncolytic viruses (OVs), hold great promise and are desperately needed. Zika virus (ZIKV) infects and destroys aggressive cells from paediatric medulloblastoma, atypical teratoid rhabdoid tumour (ATRT), diffuse midline glioma (DMG), ependymoma and neuroblastoma. Despite this, the molecular mechanisms underpinning this therapeutic response are grossly unknown. By profiling the transcriptome across a time-course, we comprehensively investigated the response of paediatric medulloblastoma and ATRT brain tumour cells to ZIKV infection at the transcriptome level for the first time. We observed conserved TNF signalling pathway and cytokine signalling-related signatures following ZIKV infection. We demonstrated that the canonical TNF-alpha signalling pathway is implicated in oncolysis by reducing the viability of ZIKV-infected brain tumour cells and is a likely contributor to the anti-tumoural immune response through TNF-alpha secretion. Our findings have highlighted TNF-alpha as a potential prognostic marker for oncolytic ZIKV virotherapy. Performing a 49-plex ELISA, we generated the most comprehensive ZIKV-infected cancer cell secretome to date. We demonstrated that ZIKV infection induces a clinically relevant and diverse pro-inflammatory brain tumour cell secretome, thus circumventing the need for transgene modification to boost efficacy. We assessed publicly available scRNA-Seq data to model how the ZIKV-induced secretome may (i) interact with medulloblastoma tumour microenvironment (TME) cells via paracrine signalling and (ii) polarise lymph node immune cells via endocrine signalling. Our modelling has provided significant insight into the cytokine response that orchestrates the diverse anti-tumoural immune response during oncolytic ZIKV infection of brain tumours. Our findings have significantly contributed to understanding the molecular mechanisms governing oncolytic ZIKV infection and will help pave the way towards ZIKV-based virotherapy.

cancer biology↗

bioRxiv2024

Mitochondrial dysfunction heightens the integrated stress response to drive ALS pathogenesis

Vesicle-associated membrane protein-associated protein-B (VAPB) is an ER membrane bound protein. VAPB P56S causes a dominant, familial form of amyotrophic lateral sclerosis (ALS), however, the mechanism through which this mutation causes motor neuron (MN) disease remains unknown. Using inducible wild type (WT) and VAPB P56S expressing iPSC-derived MNs we show that VAPB P56S, but not WT, protein decreased neuronal firing and mitochondrial-ER contact (MERC) with an associated age-dependent decrease in mitochondrial membrane potential (MMP); all typical characteristics of MN-disease. We further show that VAPB P56S expressing iPSC-derived MNs have enhanced age-dependent sensitivity to ER stress. We identified elevated expression of the master regulator of the Integrated Stress Response (ISR) marker ATF4 and decreased protein synthesis in the VAPB P56S iPSC-derived MNs. Chemical inhibition of ISR with the compound, ISRIB, rescued all MN disease phenotype in VAPB P56S MNs. Thus, our results not only support ISR inhibition as a potential therapeutic target for ALS patients, but also provides evidence to pathogenesis.

neuroscience↗

bioRxiv2023

Optimal HLA imputation of admixed population with dimension reduction

Human genomics has quickly evolved, powering genome-wide association studies (GWASs). SNP-based GWASs cannot capture the intense polymorphism of HLA genes, highly associated with disease susceptibility. There are methods to statistically impute HLA genotypes from SNP-genotypes data, but lack of diversity in reference panels hinders their performance. We evaluated the accuracy of the 1,000 Genomes data as a reference panel for imputing HLA from admixed individuals of African and European ancestries, focusing on (a) the full dataset, (b) 10 replications from 6 populations, (c) 19 conditions for the custom reference panels. The full dataset outperformed smaller models, with a good F1-score of 0.66 for HLA-B. However, custom models outperformed the multiethnic or population models of similar size (F1-scores up to 0.53, against up to 0.42). We demonstrated the importance of using genetically specific models for imputing admixed populations, which are currently underrepresented in public datasets, opening the door to HLA imputation for every genetic population.

genomics↗

bioRxiv2022

IFNγ protects motor neurons from oxidative stress via enhanced global protein synthesis in FUS-associated Amyotrophic Lateral Sclerosis

Amyotrophic lateral sclerosis type 6 (ALS6) is a familial subtype of ALS linked to Fused in Sarcoma (FUS) gene mutation. FUS mutations lead to decreased global protein synthesis, but the mechanism that drives this has not been established. Here, we used ALS6 patient-derived induced pluripotent stem cells (hIPSCs) to study the effect of the ALS6 FUSR521H mutation on the translation machinery in motor neurons (MNs). We find, in agreement with findings of others, that protein synthesis is decreased in ALS6 MNs. Furthermore, ALS6 MNs are more sensitive to oxidative stress and display reduced expression of TGF-{beta} and mTORC gene pathways when stressed. Finally, we show that IFN{gamma} treatment reduces apoptosis of ALS6 MNs exposed to oxidative stress and partially restores the translation rates in ALS6 MNs. Overall, these findings suggest that a functional IFN{gamma} response is important for FUS-mediated protein synthesis, possibly by FUS nuclear translocation in ALS6. Highlights and eTOC blurbO_LIALS6 patient-derived motor neurons show decreased viability and reduced production of innate immune cytokines following oxidative stress C_LIO_LIFUS cytoplasmic localization coincides with decreased protein synthesis rates C_LIO_LIIFN{gamma} treatment of ALS6 patient-derived motor neurons reduces apoptosis and ameliorates translation rates resulting from oxidative stress C_LI

neuroscience↗

bioRxiv2021

MS-driven metabolic alterations are recapitulated in iPSC-derived astrocytes

ObjectiveAstrocytes play a significant role in the pathology of Multiple Sclerosis (MS). Nevertheless, for ethical reasons, most of the studies in these cells were performed on the Experimental Autoimmune Encephalomyelitis model. As there are significant differences between human and mouse cells, we aimed here to better characterize astrocytes from patients with MS (PwMS), focusing mainly on mitochondrial function and cell metabolism. MethodsWe obtained and characterized induced pluripotent stem cell (iPSC)-derived astrocytes from three PwMS and three unaffected controls and performed functional assays including electron microscopy, flow cytometry, cytokine measurement, gene expression, in situ respiration, and metabolomics. ResultsWe detected several differences in MS astrocytes including: (i) enrichment of genes associated with mitophagy and neurodegeneration, (ii) increased mitochondrial fission and decreased mitochondrial to nuclear DNA ratio, indicating disruption of mitochondrial content, (iii) increased production of superoxide and MS-related proinflammatory chemokines, (iv) increased electron transport capacity and proton leak, in line with the increased oxidative stress, and (v) a distinct metabolic profile, with a deficiency in amino acid catabolism and increased sphingolipid metabolism, which have already been linked to MS. InterpretationTo our knowledge, this is the first study thoroughly describing the metabolic profile of iPSC-derived astrocytes from PwMS, and validating this model as a powerful tool to study disease mechanisms and to perform non-invasive drug targeting assays in vitro. Our findings recapitulate several disease features described in patients and provide new mechanistic insights into the metabolic rewiring of astrocytes in MS, which could be targeted in future therapeutic studies.

neuroscience↗

bioRxiv2020

Genetic variability in COVID-19-related genes in the Brazilian population

SARS-CoV-2 employs the angiotensin-converting enzyme 2 (ACE2) receptor and the transmembrane serine protease (TMPRSS2) to infect human lung cells. Previous studies have suggested that different host genetic backgrounds in ACE2 and TMPRSS2 could contribute to differences in the rate of infection or severity of COVID-19. Recent studies also showed that variants in 15 genes related to type I interferon immunity to influenza virus could predispose to life-threatening COVID-19 pneumonia. Additional genes (SLC6A20, LZTFL1, CCR9, FYCO1, CXCR6, XCR1, IL6, CTSL, ABO, and FURIN) and HLA alleles have also been implicated in response to infection with SARS-CoV-2. Currently, Brazil has recorded the third-highest number of COVID-19 patients worldwide. We aim to investigate the genetic variation present in COVID-19-related genes in the Brazilian population. We analysed 27 candidate genes and HLA alleles in 954 admixed Brazilian exomes. We used the information available in two public databases (http://www.bipmed.org and http://abraom.ib.usp.br/), and additional exomes from individuals born in southeast Brazil, the region with the highest number of COVID-19 patients in the country. Variant allele frequencies were compared with the 1000 Genomes Project phase 3 (1KGP) and the gnomAD databases. We found 395 non-synonymous variants; of these, 325 were also found in the 1000 Genome Project phase 3 (1KGP) and/or gnomAD. Six of these variants were previously reported as putatively influencing the rate of infection or clinical prognosis for COVID-19. The remaining 70 variants were identified exclusively in the Brazilian sample, with a mean allele frequency of 0.0025. In silico prediction of the impact in protein function revealed that three of these rare variants were pathogenic. Furthermore, we identified HLA alleles that were previously associated with COVID-19 response at loci DQB1 and DRB1. Our results showed genetic variability common to other populations, but also rare and ultra-rare variants exclusively found in the Brazilian population. These findings could potentially lead to differences in the rate of infection or response to infection by SARS-CoV-2 and should be further investigated in patients with the disease.

genetics↗

bioRxiv2020

Whole-genome sequencing of 1,171 elderly admixed individuals from the largest Latin American metropolis (Sao Paulo, Brazil)

As whole-genome sequencing (WGS) becomes the gold standard tool for studying population genomics and medical applications, data on diverse non-European and admixed individuals are still scarce. Here, we present a high-coverage WGS dataset of 1,171 highly admixed elderly Brazilians from a census-based cohort, providing over 76 million variants, of which ~2 million are absent from large public databases. WGS enabled identifying ~2,000 novel mobile element insertions, nearly 5Mb of genomic segments absent from human genome reference, and over 140 novel alleles from HLA genes. We reclassified and curated nearly four hundred variant's pathogenicity assertions in genes associated with dominantly inherited Mendelian disorders and calculated the incidence for selected recessive disorders, demonstrating the clinical usefulness of the present study. Finally, we observed that whole-genome and HLA imputation could be significantly improved compared to available datasets since rare variation represents the largest proportion of input from WGS. These results demonstrate that even smaller sample sizes of underrepresented populations bring relevant data for genomic studies, especially when exploring analyses allowed only by WGS.

genomics↗

bioRxiv2019

Differential gene expression elicited by ZIKV infection in trophoblasts from congenital Zika syndrome discordant twins

Zika virus (ZIKV) causes congenital Zika syndrome (CZS), which is characterized by fetal demise, microcephaly and other abnormalities. ZIKV in the pregnant woman circulation must cross the placental barrier that includes fetal endothelial cells and trophoblasts, in order to reach the fetus. CZS occurs in [~]1-40% of cases of pregnant women infected by ZIKV, suggesting that mothers infection by ZIKV during pregnancy is not deterministic for CZS phenotype in the fetus. Therefore, other susceptibility factors might be involved, including the host genetic background. We have previously shown that in three pairs of dizygotic twins discordant for CZS, neural progenitor cells (NPCs) from the CZS-affected twins presented differential in vitro ZIKV susceptibility compared with NPCs from the non-affected. Here, we analyzed human-induced-pluripotent-stem-cell-derived (hiPSC-derived) trophoblasts from these twins and compared by RNA-Seq the trophoblasts from CZS-affected and non-affected twins. Following in vitro exposure to a Brazilian ZIKV strain (ZIKVBR), trophoblasts from CZS-affected twins were significantly more susceptible to ZIKVBR infection when compared with trophoblasts from the non-affected. Transcriptome profiling revealed no differences in gene expression levels of ZIKV candidate attachment factors, IFN receptors and IFN in the trophoblasts, either before or after ZIKVBR infection. Most importantly, ZIKVBR infection caused, only in the trophoblasts from CZS-affected twins, the downregulation of genes related to extracellular matrix organization and to leukocyte activation, which are important for trophoblast adhesion and immune response activation. In addition, only trophoblasts from non-affected twins secreted significantly increased amounts of chemokines RANTES/CCL5 and IP10 after infection with ZIKVBR. Overall, our results showed that trophoblasts from non-affected twins have the ability to more efficiently activate genes that are known to play important roles in cell adhesion and in triggering the immune response to ZIKV infection in the placenta, and this may contribute to predict protection from ZIKV dissemination into fetuses tissues. Author summaryThe Zika virus (ZIKV) infection in adults is usually characterized by mild flu-like symptoms, with most cases remaining asymptomatic. However, in the last years, widespread ZIKV infection was shown for the first time to be associated with congenital Zika syndrome (CZS) and death of neonates. CZS is a very debilitating condition that includes microcephaly and mental retardation, leading to a strong social and health impact. This dramatic condition calls for a careful evaluation of the molecular mechanisms involved in ZIKV infection in the maternal-fetal interface. It is estimated that CZS occurs in [~]1-40% of cases of pregnant women infected by ZIKV, which suggests that different susceptibility factors might be involved, including the host genetic background. By analyzing trophoblast cells that recapitulate the placenta from three pairs of dizygotic twins discordant for CZS, we were able to show that trophoblasts from CZS-affected twins were significantly more susceptible to ZIKV infection when compared with trophoblasts from the non-affected twins. We also provide a detailed picture of genes differentially expressed by trophoblasts from the discordant twins after infection with ZIKV. These genes can be further investigated as possible therapeutic targets to avoid viral dissemination into developing fetus tissues.

microbiology↗

bioRxiv2019

Admixture/fine-mapping in Brazilians reveals a West African associated potential regulatory variant (rs114066381) with a strong female-specific effect on body mass- and fat mass-indexes

Admixed populations are a resource to study the global genetic architecture of complex phenotypes, which is critical, considering that non-European populations are severely under-represented in genomic studies. Leveraging admixture in Brazilians, whose chromosomes are mosaics of fragments of Native American, European and African origins, we used genome-wide data to perform admixture mapping/fine-mapping of Body Mass Index (BMI) in three population-based cohorts from Northeast (Salvador), Southeast (Bambui) and South (Pelotas) of the country. We found significant associations with African-associated alleles in children from Salvador (PALD1 and ZMIZ1 genes), and in young adults from Pelotas (NOD2 and MTUS2 genes). More importantly, in Pelotas, rs114066381, mapped in a potential regulatory region, is significantly associated only in females (p= 2.76 e-06). This variant is very rare in Europeans but with frequencies of ~3% in West Africa, and has a strong female-specific effect (95%CI: 2.32-5.65 kg/m2 per each A allele). We confirmed this sex-specific association and replicated its strong effect for an adjusted fat-mass index in the same Pelotas cohort, and for BMI in another Brazilian cohort from Sao Paulo (Southeast Brazil). A meta-analysis confirmed the significant association. Remarkably, we observed that while the frequency of rs114066381-A allele ranges from 0.8 to 2.1% in the studied populations, it attains ~9% among morbidly obese women from Pelotas, Sao Paulo, and Bambui. The effect size of rs114066381 is at least five-times the effect size of the FTO SNPs rs9939609 and rs1558902, already emblematic for their high effects, and for which we replicated associations in Pelotas. We demonstrate how, after a decade of GWAS mostly performed in European-ancestry populations, non-European and admixed populations are a source of new relevant phenotype-associated genetic variants.

genetics↗

bioRxiv2019

Structural variation of the malaria-associated human glycophorin A-B-E region

Approximately 5% of the human genome consists of structural variants, which are enriched for genes involved in the immune response and cell-cell interactions. A well-established region of extensive structural variation is the glycophorin gene cluster, comprising three tandemly-repeated regions about 120kb in length, carrying the highly homologous genes GYPA, GYPB and GYPE. Glycophorin A and glycophorin B are glycoproteins present at high levels on the surface of erythrocytes, and they have been suggested to act as decoy receptors for viral pathogens. They act as receptors for invasion of a causative agent of malaria, Plasmodium falciparum. A particular complex structural variant (DUP4) that creates a GYPB/GYPA fusion gene is known to confer resistance to malaria. Many other structural variants exist, and remain poorly characterised. Here, we analyse sequences from 6466 genomes from across the world for structural variation at the glycophorin locus, confirming 15 variants in the 1000 Genomes project cohort, discovering 9 new variants, and characterising a selection using fibre-FISH and breakpoint mapping. We identify variants predicted to create novel fusion genes and a common inversion duplication variant at appreciable frequencies in West Africans. We show that almost all variants can be explained by unequal cross over events (non-allelic homologous recombination, NAHR) and. by comparing the structural variant breakpoints with recombination hotspot maps, show the importance of a particular meiotic recombination hotspot on structural variant formation in this region.

genetics↗