Early Stem Research Articles

Effects of Microgravity on Early Embryonic Development and Embryonic Stem Cell Differentiation: Phenotypic Characterization and Potential Mechanisms

With the development of science and technology, mankind’s exploration of outer space has increased tremendously. Settling in outer space or on other planets could help solve the Earth’s resource crisis, but such settlement will first face the problem of reproduction. There are considerable differences between outer space and the Earth’s environment, with the effects of gravity being one of the most significant. Studying the possible effects and underlying mechanisms of microgravity on embryonic stem cell (ESC) differentiation and embryonic development could help provide solutions to healthy living and reproduction in deep space. This article summarizes recent research progress on the effects of microgravity on ESCs and early embryonic development and proposes hypotheses regarding the potential mechanisms. In addition, we discuss the controversies and key questions in the field and indicate directions for future research.

Novel evolution of a hyper-elongated tongue in a Cretaceous enantiornithine from China and the evolution of the hyolingual apparatus and feeding in birds.

The globally distributed extinct clade Enantiornithes comprises the most diverse early radiation of birds in the Mesozoic with species exhibiting a wide range of body sizes, morphologies, and ecologies. The fossil of a new enantiornithine bird, Brevirostruavis macrohyoideus gen. et sp. nov., from the Lower Cretaceous Jiufotang Formation in Liaoning Province, northeastern China, preserves a few important skeletal features previously unknown among early stem and extant birds, including an extremely elongate bony hyoid element (only slightly shorter than the skull), combined with a short cranial rostrum. The long hyoid provides direct evidence for the evolution of specialized feeding in this extinct species, and appears similar to the highly mobile tongue that is mobilized by the paired epibranchials present in living hummingbirds, honeyeaters, and woodpeckers. The likely linkage between food acquisition and tongue protrusion might have been a key factor in the independent evolution of particularly elongate hyobranchials in early birds.

GENDER GAPS IN ENTREPRENEURSHIP AND EDUCATION LEVELS FROM THE PERSPECTIVE OF CLUSTERS OF EUROPEAN COUNTRIES

In European countries, the gender gap in entrepreneurship is persistently observed because females make up on average 30 percent of entrepreneurs. The aim of the paper is to investigate the impact of gender gaps in education at different levels (youth, higher education, adult learning and STEM education) on the gender gap in entrepreneurship, both in all analyzed European countries and in identified clusters of countries. The research is based on the cluster analysis and regression modelling of yearly panel data for 31 European countries for the period 2013–2018. Generally speaking, to the same extent, the gender gap in entrepreneurship can be explained by the gender gap in educational outcomes at different levels, mostly in early leavers and in STEM education. However, European countries are not monolithic in these aspects, as four clusters of countries are identified. Relationships between the gender gaps strongly depend on the country’s characteristics. In countries with a relatively lower level of female entrepreneurship and their education outcomes, the gender gap in entrepreneurship is affected the most significant in comparison to other clusters, by gender gaps in early leavers, higher education and STEM education. However, in clusters of countries with relatively moderate or high levels of female entrepreneurship and education outcomes, their relationships are rather limited.

Differential Effects of Low and High Radiation Dose Rates on Mouse Spermatogenesis.

The adverse effects of radiation are proportional to the total dose and dose rate. We aimed to investigate the effects of radiation dose rate on different organs in mice. The mice were subjected to low dose rate (LDR, ~3.4 mGy/h) and high dose rate (HDR, ~51 Gy/h) radiation. LDR radiation caused severe tissue toxicity, as observed in the histological analysis of testis. It adversely influenced sperm production, including sperm count and motility, and induced greater sperm abnormalities. The expression of markers of early stage spermatogonial stem cells, such as Plzf, c-Kit, and Oct4, decreased significantly after LDR irradiation, compared to that following exposure of HDR radiation, in qPCR analysis. The compositional ratios of all stages of spermatogonia and meiotic cells, except round spermatid, were considerably reduced by LDR in FACS analysis. Therefore, LDR radiation caused more adverse testicular damage than that by HDR radiation, contrary to the response observed in other organs. Therefore, the dose rate of radiation may have differential effects, depending on the organ; it is necessary to evaluate the effect of radiation in terms of radiation dose, dose rate, organ type, and other conditions.

Overlapping functions of RBBP4 and RBBP7 in regulating cell proliferation and histone H3.3 deposition during mouse preimplantation development

ABSTRACT Preimplantation development is critical for reproductive successes in mammals. Thus, it is important to understand how preimplantation embryogenesis is regulated. As a key event of preimplantation development, epigenetic reprogramming has been widely studied, yet how epigenetic complexes regulate preimplantation development remains largely unknown. Retinoblastoma binding protein 4 (RBBP4) and 7 (RBBP7) are integral components of epigenetic complexes including SIN3A, NuRD, and CoREST. Here, we demonstrate that double knockdown of Rbbp4 and 7, but not individually, causes embryonic lethality during the morula-to-blastocyst transition. Mechanistically, depletion of RBBP4 and 7 results in dysregulation of genes related to cell cycle, lineage development, and regulation of transcription, which is accompanied by cell cycle block, disrupted lineage specification and chromatin structure. Interestingly, RBBP4/7 depletion leads to a dramatic increase in H3.3 and H3K27ac abundance during morula-to-blastocyst transition. ChIP-seq analysis in early embryos and embryonic stem cells reveals enrichment of H3.3 at the promoter regions of RBBP4/7 target genes. In summary, our studies demonstrate the compensatory role of RBBP4/7 and reveal its potential mechanisms in preimplantation development. Summary sentence: RBBP4 and RBBP7 play a compensatory role in regulating cell proliferation, apoptosis, and histone H3.3 deposition during preimplantation development.

Dissecting Richter's Syndrome in a Multiplexed CRISPR-Based Mouse Model Reveals Key Changes in MYC, Interferon and BCR Signaling Underlying Transformation

Richter's syndrome (RS) represents one of the foremost challenges in CLL management, and its pathogenesis remains largely undefined. We recently leveraged CRISPR-Cas9 in vivo gene editing to develop mouse models of RS by engineering multiplexed loss-of-function lesions typical of CLL (Atm, Trp53, Chd2, Birc3, Mga, Samhd1) in early stem and progenitor cells [Lineage - Sca-1 + c-kit + (LSK)] from MDR-Cd19Cas9 donor mice. These animals express Cas9-GFP in a B-cell restricted fashion and the leukemogenic MDR lesion, which mimics del(13q) when the sgRNA-transduced LSK cells are transplanted in CD45.1 immunocompetent recipients. Through these methods, we observed not only development of CLL, but also transformation into RS, and even captured a stage where CLL and RS were co-existing in the same animal (CLL/RS).We hypothesized that the molecular events underlying RS development would be markedly distinct from those of CLL and performed transcriptome analysis of FACS-sorted CLL and/or RS cells (5 CLL, 4 CLL/RS, 10 RS) and normal B cell controls from 4 age-matched wild type MDR-Cd19Cas9 mice. We identified a unique transcriptional profile of RS (ANOVA, FDR<0.1), characterized by upregulation of pathways involved in cell survival and proliferation (E2F/MYC targets, G2-M checkpoint, mitotic spindle). In contrast, genes involved in interferon gamma response, JAK-STAT and BCR signaling were predominantly downregulated.We asked whether these oncogenic circuitries would be recapitulated in human RS. By correlating the differentially expressed genes in murine RS with those of 7 human RS cases (compared to matched CLL), we identified similar pathway dysregulations with >100 commonly altered genes including upregulated cell cycle regulators (CDK1, CCNA2) and downregulated signaling adapters (ITPKB, MAP3K9).To further dissect gene regulatory networks driving transformation in the mouse, we profiled one CLL and one RS case by single cell ATAC sequencing (scATAC-seq). Consistent with the RNA-seq profiles, we detected increased chromatin accessibility of MYC-family associated transcription factor motifs (MAX, MYCN), and reduced accessibility of the pro-inflammatory STAT2 motif in RS (-log10adjP>50). Functionally, decreased interferon gamma responses were confirmed by the reduced ability of RS cells to phosphorylate STAT1 and STAT3 at 5‘ and 15‘ after IFN-gamma stimulation, compared to CLL and normal B cells (Western Blot).To define the genetic landscape underlying these changes, we performed whole genome sequencing analysis, and identified loss of chr12 and chr16 as recurrent events in RS (6/8 cases) and CLL/RS (2/2), but not in CLL cases (0/5). Among the genes encoded by these chromosomes, we identified several epigenetic drivers (Dnmt3a, Crebbp, Setd3/4), MAP kinase family members (Map4k5, Mapk1), cytoskeletal regulators (Hcls1, Rhoj), and interferon family receptors (Ifnar1/2, Ifngr2), suggesting that broad epigenetic modifications together with loss of BCR and interferon signaling molecules represent key events of transforming disease. RS cases were also characterized by a significantly higher number of full chromosome amplifications or deletions (median=6; range: 2-9), as compared to CLL or CLL/RS (1; 0-5, P=0.0008), consistent with the high degree of genomic instability observed in human disease.Finally, we asked whether the observed changes would impact RS therapeutic vulnerabilities, and exposed 15 primary murine RS splenocyte samples to 20 drugs in vitro for 24 hours, followed by CellTiter-Glo assessment of cellular viability. We observed strong sensitivity to the BRD4 inhibitor JQ1 and the mTOR inhibitor everolimus (both reported to interfere with MYC signaling, P<0.0001), and to CDK inhibitors (e.g. the CDK4/6 inhibitor palbociclib, P=0.0007), modest activity of the JAK1/2 inhibitor ruxolitinib (P=0.05), and minimal, if any, response to ibrutinib, venetoclax and fludarabine.In conclusion, we define the evolutionary trajectories and therapeutic vulnerabilities of RS in a mouse model, with unique transcriptional, genetic, and epigenetic features, indicative of broad changes in MYC, IFN and BCR signaling pathways and remarkable similarities with human disease. In-depth analyses of BCR signaling and in vivo treatment studies are underway and will refine mechanistic insights into the biology of RS. DisclosuresDavids: Surface Oncology: Research Funding; Eli Lilly and Company: Consultancy; Genentech: Consultancy, Research Funding; Takeda: Consultancy; MEI Pharma: Consultancy; Janssen: Consultancy; Verastem: Consultancy, Research Funding; Ascentage Pharma: Consultancy, Research Funding; Pharmacyclics: Consultancy, Research Funding; TG Therapeutics: Consultancy, Research Funding; Astra-Zeneca: Consultancy, Research Funding; Merck: Consultancy; Adaptive Biotechnologies: Consultancy; Research to Practice: Consultancy; AbbVie: Consultancy; MEI Pharma: Consultancy, Research Funding; Novartis: Consultancy, Research Funding; BMS: Consultancy, Research Funding; Celgene: Consultancy; BeiGene: Consultancy. Letai: Dialectic Therapeutics: Other: equity holding member of the scientific advisory board; Flash Therapeutics: Other: equity holding member of the scientific advisory board; Zentalis Pharmaceuticals: Other: equity holding member of the scientific advisory board. Neuberg: Madrigal Pharmaceuticals: Other: Stock ownership; Pharmacyclics: Research Funding. Wu: Pharmacyclics: Research Funding; BioNTech: Current equity holder in publicly-traded company.

Nodal/SMAD2 and Bone Morphogenic Protein 4 RNA Expression in Microgravity. Implications for Spacial Orientation Signaling in Hematopoesis and the Implications for the Hematosuppressive Effects of Space Flight

Introduction: Nodal proteins are members of the TGF alpha family which direct right left orientation in the developing embryo. Nodal/SMAD2 signaling controls BMP4 expression and with canonical Wnt signaling, regulates the posteriorization of the primitive streak, development of the mesoderm, development of early hematopoetic stem cells, and the property of self renewal in adult hematopoiesis. This study investigated the extent that microgravity alters expression of these spatially orienting proteins of BMP4, Nodal, Wnt, and SMAD2 thus influencing the anemia, thrombocytopenia, and immune alteration associated with space flight.Method: The Gene Expression Omnibus Database (GEO) of the National Institutes of Health was accessed to define RNA expression of Nodal, SMAD2, Wnt and BMP4 in hematopoetic cells in microgravity. Initial queries for “Low Gravity” studies resulted in 565 investigations within the GEO Omnibus. Of the 565 studies, the GSE136939, GSE101102, and the GSE101309 datasets were explored which assessed the lymphoid and macrophage cell lines TK6, U937, and Jurkat. RNA expression with normal and microgravity environments were compared. In the U937 cells hypergravity environments were also studied. Nodal, Lefty, Wnt, BMP4, as well as the NODAL signaling cascade proteins of SMAD 1/5, SMAD2, SMAD3, and the inhibitory SMAD 7. Affymetrix Expression Arrays were utilized for the TK^ and Jurkat cells assays. The U937 cell lines were assayed with NimbleGen Human Gene Assay Arrays.Results: Significant decreases in BMP4 expression were seen in microgravity environments (P value, <0.005) where SMAD2 expression was significantly elevated (P value <0.05) in microgravity. Conversely, hypergravity effects on U937 cells resulted in a reverse expression pattern. (BMP4 expression increased while SMAD2 decreased (P value <0.01) ). Comparisons of 14, 12, and 4 expression assays were made in U937, Jurkat, and TK6 cells respectively. LEFTY, NODAL, Wnt, SMA1/5/3/7 expression assays all showed changes in expression, but the small sample numbers resulted in poor statistical power and insignificant P values to detect differences in the RNA expression assessed. Comparisons of the gravitational pressures and time spent in these environments for the cellular cultures resulted in insignificant statistical differences.Conclusions: Microgravity induces decreases in BMP4 RNA expression and increases in SMAD2 RNA expression in TK6, U937, and Jurkat cell as defined by the GSE101309, GSE101102 and GSE136939 datasets. BMP4 is required for stem cell posteriorization, HSC number, and adult hematopoetic self renewal. This diminishment in BMP4 expression in microgravity may result in diminished HSC number and self renewal capacity. This qualitative change in gene expression in microgravity and may be a significant component of the anemia, thrombocytopenia, and immune alterations of space flight. Moreover such changes in RNA expression in microgravity implicates the importance of hematopoetic spacial orientation signals in normal hematopoiesis. Therapeutic targeting of the BMP/SMAD2 pathway in refractory multilineage hematopoetic diseases may prove to be fruitful. Statistical interpretation of the data is hindered by the small number of samples, the cell type, hematopoetic maturation stage of the samples, and microenvironmental differences in the studies utilized in these datasets. If this result is confirmed in future and larger studies, it would have significant biologic and therapeutic implications for long term space flight and the developing concept of spacial hematopoiesis. DisclosuresNo relevant conflicts of interest to declare.

Spectrum of Molecular Modes of Immune Escape in Idiopathic Aplastic Anemia and Paroxysmal Nocturnal Hemoglobinuria

The pathogenesis of idiopathic aplastic anemia (IAA) involves a human leukocyte antigen (HLA)-restricted T-cell autoreactivity against unknown antigens preferentially distributed on early hematopoietic stem and progenitor cells (HSPCs). Genetically acquired GPI-anchor and HLA deficiency have been both linked to clonal immune evasion from T-cell pressure. We hypothesized that, in analogy to anti-tumor adaptive immune evasion, pathophysiology of immune escape in IAA originates together with a broader dysfunction of antigen presentation/processing machinery and immune regulatory proteins, beyond HLA molecules, as an effect of immune pressure under T-cell attack. This initial immune reaction would produce up-modulation of these pathways, ultimately promoting the acquisition of mutations and expansion of immune resistant clones.To test this hypothesis, we first performed single-cell RNAseq analysis in HSPCs in IAA patients at disease manifestation, 1 which showed signatures of dysfunction of antigen presentation machinery, with up-regulation of most of the HLA molecules, proteasome subunits and endoplasmic reticulum related organelle transporters. Strikingly, DRB1 was among the top 3 genes upregulated in IAA patients compared to controls (q-values 1.23E-35; Fig.1A), underscoring the etiological impact that antigen presentation via this locus has in the initiation of autoimmune process. Mild upregulation was also seen in DQB1 and B loci (q-values 4.7E-07 and 2.1E-10, respectively).We then studied molecular escape mechanisms by genotyping 204 IAA and PNH patients, with either a targeted or whole genome sequencing (WGS) platform. By application of a newly in-house developed bioinformatic pipeline, we detected somatic aberrations in HLA region involving both class I and II alleles in 36% of IAA patients including point mutations, frameshift insertions or deletions and copy number variations inducing allelic loss. B*14:02 and A*02:01 emerged as the most commonly mutated class I alleles with a few hotspot mutations identified, particularly in exon 1 (c.19C>T, p.R7X, Fig.1B,C), confirming previous reports. 2,3 In class II, DQB1 and DPA1 loci were frequently targeted by fine mutational events, while more complex allelic loss phenomena interested prevalently DRB1 and DQB1 loci. Those aberrations were identified at diagnosis (35%), during disease follow-up (33%) or at the time of malignant evolution (27%), with higher clonal size in specimens collected during the course of the disease (median VAF 3% [2-27%] at diagnosis, 8% [2-98%] at follow-up, and 2.2% [2.0-6.1%] at evolution). Of 41 patients with at least one HLA aberration and characterized with an extended genotypic study, only 6 harbored also >1 somatic myeloid mutation (14%), versus 30/90 (33%) not affected by somatic hits in HLA (p=.026; Fig.1D). HLA aberrant cases also showed lower number of somatic myeloid mutations (OR=0.44; p=.0262) with driver hits rarely present (Fig.1E). In terms of PIGA mutations, an increased PIGA mosaicism was observed in the HLA mutant group, underlying that both processes have similar pathophysiologic origin as a product of the immune selection pressure (OR: 1.55 [95%CI 1.1-4.2], p=.0201). We then investigated, through WGS of 53 patients, the presence of somatic mutations in other immune genes which could be triggered by immune pressure. Hence, in 47% of the cases we were able to find pathogenic or likely pathogenic hits in genes encompassing proteasome complex, vesicle trafficking, transactivators and interferon regulatory factors, including CREBBP, TAP1, CIITA, PSMC5, PSMB4 and IRF9 (Fig.1F), whose pathogenicity was computationally assessed through recently implemented somatic classifiers. 4 Those hits were not mutually exclusive neither with HLA nor with PIGA mutations, however their VAF was significantly lower compared to concurrent HLA and PIGA lesions, underscoring their lower driver potential within the immune escape environment compared to PNH and HLA-lacking clones.Altogether our results describe the diversity of molecular and immune events taking place in IAA and PNH. Our study suggests that following initial immune insult, clonal architecture of residual hematopoiesis can be dominated by multiple modes of immune escape, agonistically participating to a mechanism of “adaptive” clonal recovery, likely in opposition to the “maladaptive” malignant progression. [Display omitted] DisclosuresMaciejewski: Alexion: Consultancy; Regeneron: Consultancy; Novartis: Consultancy; Bristol Myers Squibb/Celgene: Consultancy.

Early Hematopoietic Stem Cell Transplantation in the Treatment of High-Risk Infant Leukemia

INTRODUCTIONAcute lymphoblastic leukemia (ALL) and acute myeloid leukemia (AML) are rare diseases in children less than one and two years old respectively. Infant ALL and AML demonstrates unique biological characteristics associated with aggressive clinical features, i.e. high frequency of KMT2A gene rearrangements, poor response to standard chemotherapy, and thus worse prognosis compared to older children. These patients (pts) are at high risk (HR). Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is a potentially curative therapy for HR infant leukemia especially in 1 st CR, but proportion of these patients unable to undergo allo-HSCT due to early events, mainly relapse or treatment-related toxicity during initial therapy. We investigated the role of early allo-HSCT from different types of donors. The primary end points were overall survival (OS), transplant-relate mortality (TRM), graft-versus-host disease (GVHD) free, relapse-free survival (GRFS). Secondary end points included neutrophil engraftment and acute and chronic GVHD. MethodsWe included 50 infants with HR ALL (n=10, of with MLLr+ n=6) and AML (n=40, of with MLLr+ n=14) in CR 1. Median age at allo-HSCT 12,5 months old (range from 4 m.o. till 35 m.o.). Median time from diagnosis to allo-HSCT was 9 months (range from 3 till 21). Donor of SCT was: MSD-10% (n=5), MUD-46% (n=23), Haplo-44% (n=22). Majority of patients, 86% received MAC based on busulfan 12-16mg/kg (n=43, including IV busulfan-20pts), 14% (n=7) received RIC based on melphalan 140 mg/m 2. GVHD prophylaxis consisted of PTCy±CNI and m-TOR inhibitor in 62% (n=31), seroprophylaxis±CNI in 38% (n=19). Median follow-up was 64,3 months (range 1-220). Patient data were censored at the time of death or last follow-up. Probabilities of OS and GRFS were estimated using Kaplan-Meier curves. TRM was defined as any death that occurred in the absence of disease relapse; relapse was a competing risk for this event. ResultsEngraftment rate was 82%. Cumulative incidences by day 100 of TRM in Haplo were 2% vs 4% in MSD+MUD (р=0,9). Ten-year rates OS in Haplo and MSD+MUD were 77,3% (±4,7%) vs 78,6% (±9%) respectively (p=0,73). Ten-year GRFS did not differ between two study cohorts being 50% vs 57% for Haplo and MSD+MUD (p=0,76). Clinically significant aGVHD (grades 2-4) after allo-HSCT was observed in 27% (n=6) Haplo and 32% (n=9) MSD+MUD (p=0,77). A total of 12% (n = 6) patients developed severe cGVHD. ConclusionsEarly allogeneic HSCT in 1 st CR from different types of donors, including haploidentical is associated with favorable outcomes in infant acute leukemias. This approach should be evaluated in further controlled clinical studies. DisclosuresKulagin: X4 Pharmaceuticals, Alexion, Apellis, Biocad: Research Funding; Novartis, Generium, Sanofi, Roche, Johnson & Johnson, Pfizer: Speakers Bureau.

Advancing Early STEM Identity Development: Insights into Early Childhood Mathematics Education

Advancing Early STEM Identity Development: Insights into Early Childhood Mathematics Education

Deficiencies in the DNA Binding Protein ARID3a Alter Chromatin Structures Important for Early Human Erythropoiesis.

ARID3a is a DNA-binding protein important for normal hematopoiesis in mice and for in vitro lymphocyte development in human cultures. ARID3a knockout mice die in utero with defects in both early hematopoietic stem cell populations and erythropoiesis. Recent transcriptome analyses in human erythropoietic systems revealed increases in ARID3a transcripts implicating potential roles for ARID3a in human erythrocyte development. However, ARID3a transcript levels do not faithfully reflect protein levels in many cells, and the functions and requirements for ARID3a protein in those systems have not been explored. We used the erythroleukemic cell line K562 as a model to elucidate functions of ARID3a protein in early human erythropoiesis. ARID3a knockdown of hemin-stimulated K562 cells resulted in lack of fetal globin production and modifications in gene expression. Temporal RNA sequencing data link ARID3a expression with the important erythroid regulators Gata1, Gata2, and Klf1 Ablation of ARID3a using CRISPR-Cas9 further demonstrated it is required to maintain chromatin structures associated with erythropoietic differentiation potential. These data demonstrate that the ARID3a protein is required for early erythropoietic events and provide evidence for the requirement of ARID3a functions for proper maintenance of appropriate chromatin structures.

Biomarkers of Early Liver Graft Damage in Circulatory Death and Brain Death Donors: A Propensity Score Matching Analysis

Donation after circulatory death (DCD) is related to a warm ischemia time and more complications compared with traditional donors (donation after brain death [DBD]). This study included biopsy samples retrospectively collected from November 2014 to December 2018 to compare histologic and biological markers of DCD and DBD liver grafts. The analysis includes marker of early apoptosis (p21), senescence (telomerase reverse transcriptase [TERT]), cell damage (caspase-3 active), endothelial damage (vascular endothelial growth factor), stem cell (CD90), hypoxia (HIF1A), inflammatory activation (COX-2), and cross-organ allograft rejection (CD44). A propensity score matching (PSM) was used to match patients receiving DCD livers to those receiving DBD livers. We analyzed the immunohistochemical initial liver damage-related warm ischemia time. Positive staining expression of liver damage biomarkers (COX-2, CD44, TERT, HIF1A, and CD90) was found, but no significant differences were found between DCD and DBD and with ischemic cholangiopathy. After PSM, there was a significant relationship between CD90 and male donors (odds ratio [OR], 0.26; 95% confidence interval [CI], 0.07-0.91), TERT with donor sodium (OR, 1.11; 95% CI, 1.02-1.2), HIF1A with steatosis (OR, 0.33; 95% CI, 0.13-0.83), and CD44 with donor vasoactive drugs (OR, 0.36; 95% CI, 0.13-1) and glutamic oxaloacetic transaminase 1 week increase (OR, 1.01; 95% CI, 1-1.03). DCD immunohistochemical initial liver damage was found to behave similarly to DBD. The increase in complications and cholangiopathy associated with warm ischemia could be related to a different later phenomenon.

Hapln1b, a central organizer of the ECM, modulates kit signaling to control developmental hematopoiesis in zebrafish

AbstractDuring early vertebrate development, hematopoietic stem and progenitor cells (HSPCs) are produced in hemogenic endothelium located in the dorsal aorta, before they migrate to a transient niche where they expand to the fetal liver and the caudal hematopoietic tissue, in mammals and zebrafish, respectively. In zebrafish, previous studies have shown that the extracellular matrix (ECM) around the aorta must be degraded to enable HSPCs to leave the aortic floor and reach blood circulation. However, the role of the ECM components in HSPC specification has never been addressed. In this study, hapln1b, a key component of the ECM, was specifically expressed in hematopoietic sites in the zebrafish embryo. Gain- and loss-of-function experiments all resulted in the absence of HSPCs in the early embryo, showing that hapln1b is necessary, at the correct level, to specify HSPCs in the hemogenic endothelium. Furthermore, the expression of hapln1b was necessary to maintain the integrity of the ECM through its link domain. By combining functional analyses and computer modeling, we showed that kitlgb interacts with the ECM to specify HSPCs. The findings show that the ECM is an integral component of the microenvironment and mediates the cytokine signaling that is necessary for HSPC specification.

STUDYING THE INITIAL STABILITY OF A NEW TYPE OF HIP REPLACEMENT- SPIRON SHORT STEM

Objective: we perform this study with the following objective: Studying the initial stability of a new type of hip replacement-spiron short stem. Subjects and methods: Experiments to determine the deformability, elasticity and force capacity of the femur were used for the short stem and the long stem replacement corresponds to the original femur without the prosthesis. Along with that, we were evaluated short-term results of treatment for 70 patients with 92 Spiron hips replacement in the Institute of Trauma and Orthopedic-108 Military Central Hospital, from 2012 to 2016. Results: The analysis results showed that the Spiron short stem prosthetic had a significantly smaller displacement in the mid-frost bond than the Zweymuller long stem prosthesis. Clinical research: Harris’s score was 45.2 (± 2.4) before surgery. At the final examination time Harris’s score was 91.4 (± 2.5). The rate: very good (75.0%), good (17.4%), average (3.3%), poor (4.3%). There are 02 cases early stem loosening had to stem revision surgery (2.2%). Conclusion: These results show the good initial fixed ability as well as stability when replaced by the Spiron stem.

Electrical stimulation of titanium to promote stem cell orientation, elongation and osteogenesis

Electrical stimulation of cells allows exogenous electric signals as stimuli to manipulate cell growth, preferential orientation and bone remodelling. In this study, commercially pure titanium discs were utilised in combination with a custom-built bioreactor to investigate the cellular responses of human mesenchymal stem cells via in-vitro functional assays. Finite element analysis revealed the homogeneous delivery of electric field in the bioreactor chamber with no detection of current density fluctuation in the proposed model. The custom-built bioreactor with capacitive stimulation delivery system features long-term stimulation with homogeneous electric field, biocompatible, sterilisable, scalable design and cost-effective in the manufacturing process. Using a continuous stimulation regime of 100 and 200 mV/mm on cp Ti discs, viability tests revealed up to an approximately 5-fold increase of cell proliferation rate as compared to non-stimulated controls. The human mesenchymal stem cells showed more elongated and differentiated morphology under this regime, with evidence of nuclear elongation and cytoskeletal orientation perpendicular to the direction of electric field. The continuous stimulation did not cause pH fluctuations and hydrogen peroxide production caused by Faradic reactions, signifying the suitability for long-term toxic free stimulation as opposed to the commonly used direct stimulation regime. An approximate of 4-fold increase in alkaline phosphatase production and approximately 9-fold increase of calcium deposition were observed on 200 mV/mm exposed samples relative to non-stimulated controls. It is worth noting that early stem cell differentiation and matrix production were observed under the said electric field even without the presence of chemical inductive growth factors. Statement of significanceThis manuscript presents a study on combining pure titanium (primarily preferred as medical implant materials) and electrical stimulation in a purpose-built bioreactor with capacitive stimulation delivery system. A continuous capacitive stimulation regime on titanium disc has resulted in enhanced stem cell orientation, nuclei elongation, proliferation and differentiation as compared to non-stimulated controls. We believe that this manuscript creates a paradigm for future studies on the evolution of healthcare treatments in the area of targeted therapy on implantable and wearable medical devices through tailored innovative electrical stimulation approach, thereby influencing therapeutic conductive and electroactive biomaterials research prospects and development.

The Fourier transform infrared spectroscopy from Diplazium esculentum and Rivina humilis analysis to reveals the existence of necessary components in oil palm plantations of Ganoderma boninense control

Abstract. Saragih WS, Purba E, Lisnawita, Basyuni M. 2021. The Fourier transform infrared spectroscopy from Diplazium esculentum and Rivina humilis analysis reveals necessary components in oil palm plantations of Ganoderma boninense control. Biodiversitas 22: 3645-3651. The Fourier transform infrared spectroscopy (FTIR) has been widely utilized for biological samples and biomolecular characterization. We aim to identify Ganoderma boninense through FTIR and obtain a functional group that can facilitate early basal stem rot detection. Here, positive control (KP) was not inoculated with G. boninense and negative control (KN) was inoculated with G. boninense. However, the treatment samples, Diplazium esculentum leaf extract, Rivina humilis leaf extract, and fungicide treatment, were not inoculated with G. boninense. The positive control oil-palm leaf samples exhibited spectral bands similar to those in the D. esculentum extract, R. humilis extract, and fungicide treatment. Strong bonds were observed at wavelengths 3379 cm-1, 2927 cm-1, 1639 cm-1, and 1056 cm-1. Others were moderate to weak, except the negative control samples with strong bonds at 2044 cm-1. This indicates amine N-H functional groups, alkane functional group C-H, functional group alkene C=C, C-O, functional group ester, and functional group isothiocyanate N=C=S (C4H5NS or CH2 = CHCH2N=C=S). The FTIR plot result denotes G. boninense through N=C=S Isothiocyanate functional group presence at 2140-1990 cm-1. This unique structure is only found in infected oil-palm leaf tissues of G. boninense. Our study suggests that FTIR spectroscopy is more beneficial than conventional methods in early detection of G. boninense infection in oil palm.

Early-career STEM faculty members’ perceptions on their teaching praxis

ABSTRACT This study explores the perceptions of early career STEM faculty about their teaching contexts and praxis after pursuing an accredited teaching and learning programme in an Irish university. A social realist approach using Archer’s morphogenetic framework was employed to facilitate an exploration of their perceptions about teaching and the variegated responses that those perceptions produced. The study reveals the different ways in which faculty develops their teaching skills and how their work environment influences their development. The social realist approach provides a lens through which leaders of accredited programmes can understand the challenges and motivations of early career STEM faculty.

Recapitulative haematopoietic development of human pluripotent stem cells in the absence of exogenous haematopoietic cytokines.

To improve the recapitulative quality of human pluripotent stem cell (hPSC) differentiation, we removed exogenous haematopoietic cytokines from the defined differentiation system. Here, we show that endogenous stimuli and VEGF are sufficient to induce robust hPSC‐derived haematopoiesis, intensive generation of haematopoietic progenitors, maturation of blood cells and the emergence of definitive precursor cells including those that phenotypically identical to early human embryonic haematopoietic stem cells (HSCs). Moreover, the cytokine‐free system produces significantly higher numbers of haematopoietic progenitors compared to the published protocols. The removal of cytokines revealed a broad developmental potential of the early blood cells, stabilized the hPSC‐derived definitive precursors and led to spontaneous activation of inflammatory signalling. Our cytokine‐free protocol is simple, efficient, reproducible and applicable for embryonic stem cells (ESCs) and induced PSCs. The spectrum of recapitulative features of the novel protocol makes the cytokine‐free differentiation a preferred model for studying the early human haematopoietic development.

Who I’m Going to Become Is Not Clear! COVID-19’s Impact on Future Professional Identity Clarity

In this study, we investigate the impact of the COVID-19 pandemic, as a natural, strong, and external identity-threatening event, on workers’ future professional identity clarity trajectories. We examine the influence of workers’ current professional identity beliefs (self-efficacy, imposterism) on their pre-COVID-19 and post-COVID-19 future professional identity clarity trajectories. Using a repeated measures design, we test our model with 223 observations collected from a unique sample of early career STEM academics participating in a nine-month collaboration with start-ups. As expected, we find that the COVID-19 pandemic hindered future professional identity clarity growth. Further, we find that both self-efficacy and imposterism encourage future professional identity clarity growth, and that self-efficacy provides a protective benefit for workers’ future professional identity clarity trajectories in the wake of the COVID-19 pandemic. Our study has implications for research on identity, self-efficacy, and imposterism. We provide practical implications of this research.

Molecular Regulation of Paused Pluripotency in Early Mammalian Embryos and Stem Cells.

The energetically costly mammalian investment in gestation and lactation requires plentiful nutritional sources and thus links the environmental conditions to reproductive success. Flexibility in adjusting developmental timing enhances chances of survival in adverse conditions. Over 130 mammalian species can reversibly pause early embryonic development by switching to a near dormant state that can be sustained for months, a phenomenon called embryonic diapause. Lineage-specific cells are retained during diapause, and they proliferate and differentiate upon activation. Studying diapause thus reveals principles of pluripotency and dormancy and is not only relevant for development, but also for regeneration and cancer. In this review, we focus on the molecular regulation of diapause in early mammalian embryos and relate it to maintenance of potency in stem cells in vitro. Diapause is established and maintained by active rewiring of the embryonic metabolome, epigenome, and gene expression in communication with maternal tissues. Herein, we particularly discuss factors required at distinct stages of diapause to induce, maintain, and terminate dormancy.