S100A alarmins and thymic stromal lymphopoietin (TSLP) regulation in severe asthma following bronchial thermoplasty.

RATIONALE: Severe asthma affects a small proportion of asthmatics but represents a significant healthcare challenge. Bronchial thermoplasty (BT) is an interventional treatment approach preconized for uncontrolled severe asthma after considering biologics therapy. It was showed that BT long-lastingly improves asthma control. These improvements seem to be related to the ability of BT to reduce airway smooth muscle remodeling, reduce the number of nerve fibers and to modulate bronchial epithelium integrity and behavior. Current evidence suggest that BT downregulates epithelial mucins expression, cytokine production and metabolic profile. Despite these observations, biological mechanisms explaining asthma control improvement post-BT are still not well understood. OBJECTIVES: To assess whether BT affects gene signatures in bronchial epithelial cells (BECs). METHODS: In this study we evaluated the transcriptome of cultured bronchial epithelial cells (BECs) of severe asthmatics obtained pre- and post-BT treatment using microarrays. We further validated gene and protein expressions in BECs and in bronchial biopsies with immunohistochemistry pre- and post-BT treatment. MEASUREMENTS AND MAIN RESULTS: Transcriptomics analysis revealed that a large portion of differentially expressed genes (DEG) was involved in anti-viral response, anti-microbial response and pathogen induced cytokine storm signaling pathway. S100A gene family stood out as five members of this family where consistently downregulated post-BT. Further validation revealed that S100A7, S100A8, S100A9 and their receptor (RAGE, TLR4, CD36) expressions were highly enriched in severe asthmatic BECs. Further, these S100A family members were downregulated at the gene and protein levels in BECs and in bronchial biopsies of severe asthmatics post-BT. TLR4 and CD36 protein expression were also reduced in BECs post-BT. Thymic stromal lymphopoietin (TSLP) and human ß-defensin 2 (hBD2) were significantly decreased while no significant change was observed in IL-25 and IL-33. CONCLUSIONS: These data suggest that BT might improve asthma control by downregulating epithelial derived S100A family expression and related downstream signaling pathways.

A management model in blood, tissue and cell establishments to ensure rapid and sustainable patient access to advanced therapy medicinal products in Europe.

Blood, tissue and cell establishments (BTCs) stand out in the management of donor selection, procurement and processing of all types of substances of human origin (SoHO). In the last decades, the framework created around BTCs, including hospitals and national health system networks, and their links to research, development and innovation organizations and agencies have spurred their involvement in the study of groundbreaking advanced therapy medicinal products (ATMP). To further improve strategic synergies in the development of ATMPs, it will be required to promote intra- and inter-European collaborations by creating an international network involving BTCs and major stakeholders (i.e., research organizations, hospitals, universities, patient associations, public agencies). This vision is already shared with the European Blood Alliance, the association of non-profit blood establishments, with 26 member states throughout the European Union and European Free Trade Association states. Herein we present and analyze the "BTC for ATMP Development And Manufacture" (BADAM) model, an ethically responsible business model based on the values and missions of BTCs and their commitment to health equity, patient access and education (based on voluntary donation of SoHO to address unmet clinical needs, while contributing to training professionals and scientific literacy of our Society).

CRISPR/Cas9-mediated gene knockout and interallelic gene conversion in human induced pluripotent stem cells using non-integrative bacteriophage-chimeric retrovirus-like particles.

BACKGROUND: The application of CRISPR/Cas9 technology in human induced pluripotent stem cells (hiPSC) holds tremendous potential for basic research and cell-based gene therapy. However, the fulfillment of these promises relies on the capacity to efficiently deliver exogenous nucleic acids and harness the repair mechanisms induced by the nuclease activity in order to knock-out or repair targeted genes. Moreover, transient delivery should be preferred to avoid persistent nuclease activity and to decrease the risk of off-target events. We recently developed bacteriophage-chimeric retrovirus-like particles that exploit the properties of bacteriophage coat proteins to package exogenous RNA, and the benefits of lentiviral transduction to achieve highly efficient, non-integrative RNA delivery in human cells. Here, we investigated the potential of bacteriophage-chimeric retrovirus-like particles for the non-integrative delivery of RNA molecules in hiPSC for CRISPR/Cas9 applications. RESULTS: We found that these particles efficiently convey RNA molecules for transient expression in hiPSC, with minimal toxicity and without affecting the cell pluripotency and subsequent differentiation. We then used this system to transiently deliver in a single step the CRISPR-Cas9 components (Cas9 mRNA and sgRNA) to generate gene knockout with high indel rate (up to 85%) at multiple loci. Strikingly, when using an allele-specific sgRNA at a locus harboring compound heterozygous mutations, the targeted allele was not altered by NHEJ/MMEJ, but was repaired at high frequency using the homologous wild type allele, i.e., by interallelic gene conversion. CONCLUSIONS: Our results highlight the potential of bacteriophage-chimeric retrovirus-like particles to efficiently and safely deliver RNA molecules in hiPSC, and describe for the first time genome engineering by gene conversion in hiPSC. Harnessing this DNA repair mechanism could facilitate the therapeutic correction of human genetic disorders in hiPSC.

The Transcriptome Landscape of the In Vitro Human Airway Epithelium Response to SARS-CoV-2.

Airway-liquid interface cultures of primary epithelial cells and of induced pluripotent stem-cell-derived airway epithelial cells (ALI and iALI, respectively) are physiologically relevant models for respiratory virus infection studies because they can mimic the in vivo human bronchial epithelium. Here, we investigated gene expression profiles in human airway cultures (ALI and iALI models), infected or not with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), using our own and publicly available bulk and single-cell transcriptome datasets. SARS-CoV-2 infection significantly increased the expression of interferon-stimulated genes (IFI44, IFIT1, IFIT3, IFI35, IRF9, MX1, OAS1, OAS3 and ISG15) and inflammatory genes (NFKBIA, CSF1, FOSL1, IL32 and CXCL10) by day 4 post-infection, indicating activation of the interferon and immune responses to the virus. Extracellular matrix genes (ITGB6, ITGB1 and GJA1) were also altered in infected cells. Single-cell RNA sequencing data revealed that SARS-CoV-2 infection damaged the respiratory epithelium, particularly mature ciliated cells. The expression of genes encoding intercellular communication and adhesion proteins was also deregulated, suggesting a mechanism to promote shedding of infected epithelial cells. These data demonstrate that ALI/iALI models help to explain the airway epithelium response to SARS-CoV-2 infection and are a key tool for developing COVID-19 treatments.

Homo erectus at Trinil on Java used shells for tool production and engraving.

The manufacture of geometric engravings is generally interpreted as indicative of modern cognition and behaviour. Key questions in the debate on the origin of such behaviour are whether this innovation is restricted to Homo sapiens, and whether it has a uniquely African origin. Here we report on a fossil freshwater shell assemblage from the Hauptknochenschicht ('main bone layer') of Trinil (Java, Indonesia), the type locality of Homo erectus discovered by Eugène Dubois in 1891 (refs 2 and 3). In the Dubois collection (in the Naturalis museum, Leiden, The Netherlands) we found evidence for freshwater shellfish consumption by hominins, one unambiguous shell tool, and a shell with a geometric engraving. We dated sediment contained in the shells with (40)Ar/(39)Ar and luminescence dating methods, obtaining a maximum age of 0.54 ± 0.10 million years and a minimum age of 0.43 ± 0.05 million years. This implies that the Trinil Hauptknochenschicht is younger than previously estimated. Together, our data indicate that the engraving was made by Homo erectus, and that it is considerably older than the oldest geometric engravings described so far. Although it is at present not possible to assess the function or meaning of the engraved shell, this discovery suggests that engraving abstract patterns was in the realm of Asian Homo erectus cognition and neuromotor control.

Concise Review: Assessing the Genome Integrity of Human Induced Pluripotent Stem Cells: What Quality Control Metrics?

Human induced pluripotent stem cells (hiPSCs) have the potential to differentiate virtually into any cell type in unlimited quantities. Therefore, they are ideal for in vitro tissue modeling or to produce cells for clinical use. Importantly, and differently from immortalized and cancer cell lines, the hiPSC genome scrupulously reproduces that of the cell from which they were derived. However, hiPSCs can develop genetic abnormalities during reprogramming or prolonged cell culture, such as aneuploidies or oncogenic mutations (e.g., in TP53). Therefore, hiPSC genome integrity must be routinely monitored because serious genome alterations would greatly compromise their usefulness or safety of use. Here, we reviewed hiPSC genome quality control monitoring methods and laboratory practice. Indeed, due to their frequency and functional consequences, recurrent genetic defects found in cultured hiPSCs are inacceptable and their appearance should be monitored by routine screening. Hence, for research purposes, we propose that the genome of hiPSC lines should be systematically screened at derivation, at least by karyotyping, and then regularly (every 12 weeks) during experiments, for instance with polymerase chain reaction-based techniques. For some specific applications, such as research on aging, cell cycle, apoptosis or cancer, other tests (e.g., TP53 mutation detection) should also be included. For clinical use, in addition to karyotyping, we advise exome sequencing. Stem Cells 2018;36:814-821.

Rejuvenating senescent and centenarian human cells by reprogramming through the pluripotent state.

Direct reprogramming of somatic cells into induced pluripotent stem cells (iPSCs) provides a unique opportunity to derive patient-specific stem cells with potential applications in tissue replacement therapies and without the ethical concerns of human embryonic stem cells (hESCs). However, cellular senescence, which contributes to aging and restricted longevity, has been described as a barrier to the derivation of iPSCs. Here we demonstrate, using an optimized protocol, that cellular senescence is not a limit to reprogramming and that age-related cellular physiology is reversible. Thus, we show that our iPSCs generated from senescent and centenarian cells have reset telomere size, gene expression profiles, oxidative stress, and mitochondrial metabolism, and are indistinguishable from hESCs. Finally, we show that senescent and centenarian-derived pluripotent stem cells are able to redifferentiate into fully rejuvenated cells. These results provide new insights into iPSC technology and pave the way for regenerative medicine for aged patients.

Induced Pluripotent Stem Cells for Primary Ciliary Dyskinesia Modeling and Personalized Medicine.

Primary ciliary dyskinesia (PCD) is a rare and heterogeneous genetic disorder that affects the structure and function of motile cilia. In the airway epithelium, impaired ciliary motion results in reduced or absent mucociliary clearance that leads to the appearance of chronic airway infection, sinusitis, and bronchiectasis. Currently, there is no effective treatment for PCD, and research is limited by the lack of convenient models to study this disease and investigate innovative therapies. Furthermore, the high heterogeneity of PCD genotypes is likely to hinder the development of a single therapy for all patients. The generation of patient-derived, induced pluripotent stem cells, and their differentiation into airway epithelium, as well as genome editing technologies, could represent major tools for in vitro PCD modeling and for developing personalized therapies. Here, we review PCD pathogenesis and then discuss how human induced pluripotent stem cells could be used to model this disease for the development of innovative, patient-specific biotherapies.

Combining DGE and RNA-sequencing data to identify new polyA+ non-coding transcripts in the human genome.

Recent sequencing technologies that allow massive parallel production of short reads are the method of choice for transcriptome analysis. Particularly, digital gene expression (DGE) technologies produce a large dynamic range of expression data by generating short tag signatures for each cell transcript. These tags can be mapped back to a reference genome to identify new transcribed regions that can be further covered by RNA-sequencing (RNA-Seq) reads. Here, we applied an integrated bioinformatics approach that combines DGE tags, RNA-Seq, tiling array expression data and species-comparison to explore new transcriptional regions and their specific biological features, particularly tissue expression or conservation. We analysed tags from a large DGE data set (designated as 'TranscriRef'). We then annotated 750,000 tags that were uniquely mapped to the human genome according to Ensembl. We retained transcripts originating from both DNA strands and categorized tags corresponding to protein-coding genes, antisense, intronic- or intergenic-transcribed regions and computed their overlap with annotated non-coding transcripts. Using this bioinformatics approach, we identified ∼34,000 novel transcribed regions located outside the boundaries of known protein-coding genes. As demonstrated using sequencing data from human pluripotent stem cells for biological validation, the method could be easily applied for the selection of tissue-specific candidate transcripts. DigitagCT is available at http://cractools.gforge.inria.fr/softwares/digitagct.

Growth in fossil and extant deer and implications for body size and life history evolution.

BACKGROUND: Body size variation within clades of mammals is widespread, but the developmental and life-history mechanisms by which this variation is achieved are poorly understood, especially in extinct forms. An illustrative case study is that of the dwarfed morphotypes of Candiacervus from the Pleistocene of Crete versus the giant deer Megaloceros giganteus, both in a clade together with Dama dama among extant species. Histological analyses of long bones and teeth in a phylogenetic context have been shown to provide reliable estimates of growth and life history patterns in extant and extinct mammals. RESULTS: Similarity of bone tissue types across the eight species examined indicates a comparable mode of growth in deer, with long bones mainly possessing primary plexiform fibrolamellar bone. Low absolute growth rates characterize dwarf Candiacervus sp. II and C. ropalophorus compared to Megaloceros giganteus displaying high rates, whereas Dama dama is characterized by intermediate to low growth rates. The lowest recorded rates are those of the Miocene small stem cervid Procervulus praelucidus. Skeletal maturity estimates indicate late attainment in sampled Candiacervus and Procervulus praelucidus. Tooth cementum analysis of first molars of two senile Megaloceros giganteus specimens revealed ages of 16 and 19 years whereas two old dwarf Candiacervus specimens gave ages of 12 and 18 years. CONCLUSIONS: There is a rich histological record of growth across deer species recorded in long bones and teeth, which can be used to understand ontogenetic patterns within species and phylogenetic ones across species. Growth rates sensu Sander & Tückmantel plotted against the anteroposterior bone diameter as a proxy for body mass indicate three groups: one with high growth rates including Megaloceros, Cervus, Alces, and Dama; an intermediate group with Capreolus and Muntiacus; and a group showing low growth rates, including dwarf Candiacervus and Procervulus. Dwarf Candiacervus, in an allometric context, show an extended lifespan compared to other deer of similar body size such as Mazama which has a maximum longevity of 12 years in the wild. Comparison with other clades of mammals reveals that changes in size and life history in evolution have occurred in parallel, with various modes of skeletal tissue modification.

Mesenchymal stromal cells orchestrate follicular lymphoma cell niche through the CCL2-dependent recruitment and polarization of monocytes.

Accumulating evidence indicates that infiltrating stromal cells contribute directly and indirectly to tumor growth in a wide range of cancers. In follicular lymphoma (FL), malignant B cells are found admixed with heterogeneous lymphoid-like stromal cells within invaded lymph nodes and BM. In addition, mesenchymal stromal cells (MSCs) support in vitro FL B-cell survival, in particular after their engagement toward lymphoid differentiation. We show here that BM-MSCs obtained from patients with FL (FL-MSCs) display a specific gene expression profile compared with MSCs obtained from healthy age-matched donors (HD-MSCs). This FL-MSC signature is significantly enriched for genes associated with a lymphoid-like commitment. Interestingly, CCL2 could be detected at a high level within the FL-cell niche, is up-regulated in HD-MSCs by coculture with malignant B cells, and is overexpressed by FL-MSCs, in agreement with their capacity to recruit monocytes more efficiently than HD-MSCs. Moreover, FL-MSCs and macrophages cooperate to sustain malignant B-cell growth, whereas FL-MSCs drive monocyte differentiation toward a proangiogenic and lipopolysaccharide-unresponsive phenotype close to that of tumor-associated macrophages. Altogether, these results highlight the complex role of FL stromal cells that promote direct tumor B-cell growth and orchestrate FL-cell niche, thus emerging as a potential therapeutic target in this disease.

Generation of a healthy heavy smoker patient-derived induced pluripotent stem cell line UHOMi007-A from peripheral blood mononuclear cells.

Human pluripotent stem cells (hiPSC) represent a unique opportunity to model lung development and chronic bronchial diseases. We generated a hiPSC line from a highly characterized healthy heavy smoker male donor free from emphysema or tobacco related disease. Peripheral blood mononuclear cells (PBMCs) were reprogrammed using integration-free Sendai virus. The cell line had normal karyotype, expressed pluripotency hallmarks, and differentiated into the three primary germ layers. The reported UHOMi007-A iPSC line may be used as a control to model lung development, study human chronic bronchial diseases and drug testing.

Oxygen isotopes in orangutan teeth reveal recent and ancient climate variation.

Studies of climate variation commonly rely on chemical and isotopic changes recorded in sequentially produced growth layers, such as in corals, shells, and tree rings, as well as in accretionary deposits-ice and sediment cores, and speleothems. Oxygen isotopic compositions (δ18O) of tooth enamel are a direct method of reconstructing environmental variation experienced by an individual animal. Here, we utilize long-forming orangutan dentitions (Pongo spp.) to probe recent and ancient rainfall trends on a weekly basis over ~3-11 years per individual. We first demonstrate the lack of any consistent isotopic enrichment effect during exclusive nursing, supporting the use of primate first molar teeth as environmental proxies. Comparisons of δ18O values (n=2016) in twelve molars from six modern Bornean and Sumatran orangutans reveal a high degree of overlap, with more consistent annual and bimodal rainfall patterns in the Sumatran individuals. Comparisons with fossil orangutan δ18O values (n=955 measurements from six molars) reveal similarities between modern and late Pleistocene fossil Sumatran individuals, but differences between modern and late Pleistocene/early Holocene Bornean orangutans. These suggest drier and more open environments with reduced monsoon intensity during this earlier period in northern Borneo, consistent with other Niah Caves studies and long-term speleothem δ18O records in the broader region. This approach can be extended to test hypotheses about the paleoenvironments that early humans encountered in southeast Asia.

When an animal drinks water, two naturally occurring variants of oxygen ­ known as oxygen-18 and oxygen-16 ­ are incorporated into its growing teeth. The ratio of these variants in water changes with temperature, rainfall and other environmental conditions and therefore can provide a record of the climate during an animal's life. Teeth tend to be well preserved as fossils, which makes it possible to gain insights into this climate record even millions of years after an animal's death. Orangutans are highly endangered great apes that today live in rainforests on the islands of Borneo and Sumatra. During a period of time known as the Pleistocene (around 2.6 million years to 12,000 years ago), these apes were more widely spread across Southeast Asia. Climate records from this area in the time before human-induced climate change are somewhat limited. Therefore, fossilized orangutan teeth offer a possible way to investigate past seasonal rainfall patterns and gain insight into the kind of environments early humans would have encountered. To address this question, Smith et al. measured oxygen-18 and oxygen-16 variants in thin slices of modern-day orangutan teeth using a specialized analytical system. This established that the teeth showed seasonal patterns consistent with recent rainfall trends, and that the ratio of these oxygen variants did not appear to be impacted by milk intake in young orangutans. These findings indicated that the oxygen variants could be a useful proxy for predicting prehistoric weather patterns from orangutan teeth. Further measurements of teeth from fossilized Sumatran orangutans showed broadly similar rainfall patterns to those of teeth from modern-day orangutans. On the other hand, fossilized teeth from Borneo suggested that the environment used to be drier, with less intense wet seasons. The approach developed by Smith et al. provides an opportunity for scientists to leverage new fossil discoveries as well as existing collections to investigate past environments. This could allow future research into how climate variation may have influenced the spread of early humans through the region, as well as the evolution of orangutans and other endangered animals.

Pharmacometabolomics applied to low-dose interleukin-2 treatment in amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) is a devastating motor neuron disease. The immunosuppressive functions of regulatory T lymphocytes (Tregs) are impaired in ALS, and correlate to disease progression. The phase 2a IMODALS trial reported an increase in Treg number in ALS patients following the administration of low-dose (ld) interleukin-2 (IL-2). We propose a pharmacometabolomics approach to decipher metabolic modifications occurring in patients treated with ld-IL-2 and its relationship with Treg response. Blood metabolomic profiles were determined on days D1, D64, and D85 from patients receiving 2 MIU of IL-2 (n = 12) and patients receiving a placebo (n = 12). We discriminated the three time points for the treatment group (average error rate of 42%). Among the important metabolites, kynurenine increased between D1 and D64, followed by a reduction at D85. The percentage increase of Treg number from D1 to D64, as predicted by the metabolome at D1, was highly correlated with the observed value. This study provided a proof of concept for metabolic characterization of the effect of ld-IL-2 in ALS. These data could present advances toward a personalized medicine approach and present pharmacometabolomics as a key tool to complement genomic and transcriptional data for drug characterization, leading to systems pharmacology.

U-series and radiocarbon analyses of human and faunal remains from Wajak, Indonesia.

Laser ablation U-series dating results on human and faunal bone fragments from Wajak, Indonesia, indicate a minimum age of between 37.4 and 28.5 ka (thousands of years ago) for the whole assemblage. These are significantly older than previously published radiocarbon estimates on bone carbonate, which suggested a Holocene age for a human bone fragment and a late Pleistocene age for a faunal bone. The analysis of the organic components in the faunal material show severe degradation and a positive δ(13)C ratio indicate a high degree of secondary carbonatisation. This may explain why the thermal release method used for the original age assessments yielded such young ages. While the older U-series ages are not in contradiction with the morphology of the Wajak human fossils or Javanese biostratigraphy, they will require a reassessment of the evolutionary relationships of modern human remains in Southeast Asia and Oceania. It can be expected that systematic direct dating of human fossils from this area will lead to further revisions of our understanding of modern human evolution.

Characterization of a transitional preplasmablast population in the process of human B cell to plasma cell differentiation.

The early steps of differentiation of human B cells into plasma cells are poorly known. We report a transitional population of CD20(low/-)CD38(-) preplasmablasts along differentiation of human memory B cells into plasma cells in vitro. Preplasmablasts lack documented B cell or plasma cell (CD20, CD38, and CD138) markers, express CD30 and IL-6R, and secrete Igs at a weaker level than do plasmablasts or plasma cells. These preplasmablasts further differentiate into CD20(-)CD38(high)CD138(-) plasmablasts and then CD20(-)CD38(high)CD138(+) plasma cells. Preplasmablasts were fully characterized in terms of whole genome transcriptome profiling and phenotype. Preplasmablasts coexpress B and plasma cell transcription factors, but at a reduced level compared with B cells, plasmablasts, or plasma cells. They express the unspliced form of XBP1 mRNA mainly, whereas plasmablasts and plasma cells express essentially the spliced form. An in vivo counterpart (CD19(+)CD20(low/-)CD38(-)IL-6R(+) cells) of in vitro-generated preplasmablasts could be detected in human lymph nodes (0.06% of CD19(+) cells) and tonsils (0.05% of CD19(+) cells). An open access "B to Plasma Cell Atlas," which makes it possible to interrogate gene expression in the process of B cell to plasma cell differentiation, is provided. Taken together, our findings show the existence of a transitional preplasmablast population using an in vitro model of plasma cell generation and of its in vivo counterpart in various lymphoid tissues.

[Reprogramming equals gambling?]. / La reprogrammation, un jeu de hasard ?

While somatic cell reprogramming is now part of our text books, we ignore most of the mechanisms governing this cellular transformation. The most enigmatic question is why only rare cells undergo reprogramming, and whether this is governed by stochastc or deterministic events. In the late 2012, several major studies have addressed this question through a clonal analysis of the reprogramming process in murine MEF. In this mini-review, we describe these results and discuss future perspectives based on these date to optimize and secure the derivation of iPSC.

Comprehensive quantum chemical analysis of the (ro)vibrational spectrum of thiirane and its deuterated isotopologue.

The (ro)vibrational spectra of thiirane, c-C2H4S, and its fully deuterated isotopologue, c-C2D4S, have been studied by means of vibrational configuration interaction theory, VCI, its incremental variant, iVCI, and subsequent variational rovibrational calculations, RVCI, which rely on multidimensional potential energy surfaces of coupled-cluster quality including up to four-mode coupling terms. Accurate geometrical parameters, fundamental vibrational transitions and first overtones, rovibrational spectra and rotational spectroscopic constants have been determined from these calculations and were compared with experimental results whenever available. A number of tentative misassignments in the vibrational spectra could be resolved and most results for the deuterated thiirane are high-level predictions, which may guide experiments to come. Besides this, a new implementation of infrared intensities within the iVCI framework has been tested for the transitions of the title compounds and are compared with results obtained from standard VCI calculations.

[New models to study human embryonic development]. / Du nouveau dans les modèles d'étude de l'embryon humain.

Since 2021, assisted reproductive technologies (ART) are available to infertile couples, but also to single women and female couples. The process of in vitro fertilization (IVF) has allowed to cross the threshold of 5 million births worldwide, between 1978 and 2013. However, the failure rate per each IVF cycle is estimated to be around 75%. Therefore, there is a need to better understand human embryonic development in order to improve the success rate of IVF. Study models have evolved significantly in recent years: development of embryo culture, sequencing of the transcriptome of individualized cells, discovery of culture conditions for naive pluripotent stem cells and generation of blastoids. Here, we review these recent advances in human embryo modeling that establish a new knowledge base for improving ART.

Title: Du nouveau dans les modèles d'étude de l'embryon humain. Abstract: Depuis 2021, l'assistance médicale à la procréation (AMP) est accessible aux couples infertiles, mais aussi aux femmes seules et aux couples de femmes. Le processus de fécondation in vitro (FIV) a permis de franchir le seuil de cinq millions de naissances dans le monde, entre 1978 et 2013. Cependant, le taux d'échec à chaque cycle est évalué à environ 75 %. Il est donc nécessaire de mieux comprendre le développement embryonnaire humain afin d'améliorer le taux de succès des FIV. Les modèles d'étude ont beaucoup évolué ces dernières années : mise au point de la culture embryonnaire, séquençage du transcriptome de cellules individualisées, découverte des conditions de culture de cellules souches pluripotentes naïves et génération de blastoïdes. Nous revenons dans cette revue sur ces avancées récentes concernant la modélisation de l'embryon humain, qui établissent un nouveau socle de connaissances pour améliorer l'AMP.

Brief report: benchmarking human pluripotent stem cell markers during differentiation into the three germ layers unveils a striking heterogeneity: all markers are not equal.

Pluripotent stem cells (PSC) are functionally characterized by their capacity to differentiate into all the cell types from the three germ layers. A wide range of markers, the expression of which is associated with pluripotency, has been used as surrogate evidence of PSC pluripotency, but their respective relevance is poorly documented. Here, we compared by polychromatic flow cytometry the kinetics of loss of expression of eight widely used pluripotency markers (SSEA3, SSEA4, TRA-1-60, TRA-1-81, CD24, OCT4, NANOG, and alkaline phosphatase [AP]) at days 0, 5, 7, and 9 after induction of PSC differentiation into cells representative of the three germ layers. Strikingly, each marker showed a different and specific kinetics of disappearance that was similar in all the PSC lines used and for all the induced differentiation pathways. OCT4, SSEA3, and TRA-1-60 were repeatedly the first markers to be downregulated, and their expression was completely lost at day 9. By contrast, AP activity, CD24, and NANOG proteins were still detectable at day 9. In addition, we show that differentiation markers are coexpressed with pluripotency markers before the latter begin to disappear. These results suggest that OCT4, SSEA3, and TRA-1-60 might be better to trace in vitro the emergence of pluripotent cells during reprogramming.