Early Stem Research Articles

Ubiquitin-Specific Protease 3 Deubiquitinates and Stabilizes Oct4 Protein in Human Embryonic Stem Cells.

Oct4 is an important mammalian POU family transcription factor expressed by early human embryonic stem cells (hESCs). The precise level of Oct4 governs the pluripotency and fate determination of hESCs. Several post-translational modifications (PTMs) of Oct4 including phosphorylation, ubiquitination, and SUMOylation have been reported to regulate its critical functions in hESCs. Ubiquitination and deubiquitination of Oct4 should be well balanced to maintain the pluripotency of hESCs. The protein turnover of Oct4 is regulated by several E3 ligases through ubiquitin-mediated degradation. However, reversal of ubiquitination by deubiquitinating enzymes (DUBs) has not been reported for Oct4. In this study, we generated a ubiquitin-specific protease 3 (USP3) gene knockout using the CRISPR/Cas9 system and demonstrated that USP3 acts as a protein stabilizer of Oct4 by deubiquitinating Oct4. USP3 interacts with endogenous Oct4 and co-localizes in the nucleus of hESCs. The depletion of USP3 leads to a decrease in Oct4 protein level and loss of pluripotent morphology in hESCs. Thus, our results show that USP3 plays an important role in controlling optimum protein level of Oct4 to retain pluripotency of hESCs.

Two Enhancers Regulate HoxB Genes Expression During Retinoic Acid-Induced Early Embryonic Stem Cells Differentiation Through Long-Range Chromatin Interactions.

Homeobox B cluster (HoxB) genes play important roles in retinoic acid (RA)-induced early embryonic stem cells (ESCs) differentiation. Knowledge of regulation network of HoxB is important to further unveil the mechanism of ESCs differentiation. In this study, we identified two enhancers that were activated by RA treatment and 4C data showed long-range interactions between HoxB genes and the two enhancers. CRISPR/Cas9-mediated individual or compound deletion of the two enhancers significantly inhibits HoxB gene expression, and transcriptome analysis revealed that RA-induced early ESCs differentiation was blocked in the enhancer KO cells. We propose new mechanism by which two enhancers regulate HoxB gene expression by different regulation modes during RA-induced early ESCs differentiation through long-range chromatin interactions.

T cell receptor beta locus sequencing early post-allogeneic stem cell transplant identifies patients at risk of initial and recurrent cytomegalovirus infection.

Identification of patients at risk of initial & recurrent cytomegalovirus (CMV) reactivation following allogeneic stem cell transplant (alloSCT) may help guide prophylactic strategies. T-cell receptor beta (TRB) deep sequencing was used to identify and enumerate the T-cell repertoire harbouring TRB sequences with annotated specificity to CMV (pubCMVrep), as well as the overall T-cell receptor (TCR) repertoire diversity at day +30 & day +60 post-alloSCT for 65 patients. T-cells harbouring TRB sequences with annotated specificity for CMV were identifiable in all patients. 56% of patients required CMV treatment and 23% of the cohort developed recurrent CMV. PubCMVrep size at day +30 was not associated with reactivation, however amongst patients with antecedent CMV viremia a low day +60 pubCMVrep was associated with a greater incidence of recurrent CMV (75% vs. 21%, HR 6.16, 95% CI 1.29-29.40, P = 0.0008). Moreover, patients with high pubCMVrep only developed recurrent CMV in the setting of GVHD. Low TCR diversity at day +30 was associated with a greater incidence of initial CMV reactivation (71% vs. 22%, HR 5.39, 95% CI 1.70-17.09, p = 0.0002). pubCMVrep and TCR diversity are promising biomarkers to identify patients at risk of initial & recurrent CMV who may benefit from novel prophylactic strategies.

Biomarkers for Early Complications of Endothelial Origin After Allogeneic Hematopoietic Stem Cell Transplantation: Do They Have a Potential Clinical Role?

Endothelial cell (EC) dysfunction causes a number of early and life-threatening post hematopoietic stem cell transplant (HCT) complications that result in a rapid clinical decline. The main early complications are graft-vs.-host disease (GVHD), transplant associated thrombotic microangiopathy (TA-TMA), and sinusoidal obstruction syndrome (SOS). Post-HCT endothelial dysfunction occurs as a result of chemotherapy, infections, and allogeneic reactivity. Despite major advances in transplant immunology and improvements in supportive care medicine, these complications represent a major obstacle for successful HCT. In recent years, different biomarkers have been investigated for early detection of post-transplant endothelial cell dysfunction, but few have been validated. In this review we will define GVHD, TA-TMA and SOS, summarize the current data available in HCT biomarker research and identify promising biomarkers for detection and diagnosis of early HCT complications.

Update on imaging modalities for ocular surface pathologies.

To discuss recent studies of imaging modalities for ocular surface pathologies. Novel micro-ocular coherence tomography technology can produce high-resolution images of corneal cellular and nervous structures. Ocular coherence tomography angiography can aid in detecting early stage limbal stem cell deficiency. Several studies used in vivo confocal microscopy to evaluate corneal nerve metrics and morphology. The applications of anterior segment optical coherence tomography, anterior segment optical coherence tomography angiography, ultrasound biomicroscopy, and in vivo confocal microscopy are useful technologies for imaging the ocular surface. Several studies have used artificial intelligence in combination with imaging technologies to create reliable and effective systems to detect and visualize ocular surface pathologies. AS OCT continues to be a key imaging tool and future development of μOCT technology may further enhance its utility.

Root-based characterization of intergeneric hybrids with Triticum and Aegilops species in early vegetative and stem elongation growth stages

Cultivated and wild species of wheat are valuable breeding resources used for the development of new cultivars with superior root traits suited to drought and non-stressed conditions. The present study aimed to determine genotypes with superior root traits and phenotypic variability among intergeneric hybrids with Triticum and Aegilops species in the early vegetative (Z11) and stem elongation (Z31) growth stages. Results indicated that phenotypic variability in rooting depth was 3.2- and 3.4-fold among the genotypes in Z11 and Z31, and it was as great as 20- and 23.8-fold for root biomass, respectively. Hierarchical clustering among 35 genotypes for root traits in both growth stages identified four major clusters, grouping the six deep-rooted genotypes in cluster 2 and three genotypes with high root biomass in cluster 1. In both growth stages, significant associations were found among the root traits. Also, the relationship was stronger between the root and shoot biomass in Z11 (r2=0.83) than in Z31 (r2=0.44). As an overall assessment, the suggested genotypes with superior root characteristics such as deep roots and/or high root biomass sustained in both growth stages might be used for the development of new cultivars.

Cryptic introgressions contribute to transgressive segregation for early blight resistance in tomato.

We identified cryptic early blight resistance introgressions in tomato breeding lines and demonstrated efficient genotypic selection for resistance in the context of a tomato breeding program. Early blight is a widespread and problematic disease affecting tomatoes (Solanum lycopersicum). Caused by the fungal pathogen Alternaria linariae (syn. A. tomatophila), symptoms include lesions on tomato stems, fruit, and foliage, often resulting in yield losses. Breeding tomatoes with genetic resistance would enhance production sustainability. Using cross-market breeding populations, we identified several quantitative trait loci (QTL) associated with early blight resistance. Early blight resistance putatively derived from 'Campbell 1943' was confirmed in modern fresh market tomato breeding lines. This resistance offered substantial protection against early blight stem lesions (collar rot) and moderate protection from defoliation. A distinctive and potentially novel form of early blight foliar resistance was discovered in a processing tomato breeding line and is probably derived from S. pimpinellifolium via 'Hawaii 7998'. Additional field trials validated the three most promising large-effect QTL, EB-1.2, EB-5, and EB-9. Resistance effects for EB-5 and EB-9 were consistent across breeding populations and environments, while EB-1.2's effect was population specific. Using genome-wide marker-assisted backcrossing, we developed fresh market tomato lines that were near-isogenic for early blight QTL. Resistance in these lines was largely mediated by just two QTL, EB-5 and EB-9, that together captured 49.0 and 68.7% of the defoliation and stem lesion variance, respectively. Our work showcases the value of mining cryptic introgressions in tomato lines, and across market classes, for use as additional sources of disease resistance.

The future belongs to those who prepare for it today

Hippocampal neural stem cells (NSCs) and neurogenesis decline sharply with age, though a small population remains. Two articles in this issue of Cell Stem Cell by Ibrayeva etal. (2021) and Harris etal. (2021) indicate the presence of subpopulations of NSCs whose dynamics of activation and self-renewal change over time and may be key to NSC preservation.

Grade Expectations: The Role of First-Year Grades in Predicting the Pursuit of STEM Majors for First- and Continuing-Generation Students

ABSTRACT Earning poor grades in early science, technology, engineering, and mathematics (STEM) college courses decreases the likelihood that a student will major in STEM. However, grades in first-year STEM courses do not impact students evenly, making it hard to know if early “gatekeeping” courses selectively push some students out of STEM. This descriptive study examines the relationship between grades and STEM persistence for first- and continuing-generation students. Using transcript and survey data from three moderately-selective postsecondary institutions, I find that among students with high STEM GPAs, first-generation students are less likely than their continuing-generation peers to persist in STEM, net background preparation and characteristics. Moreover, first-year STEM grades alone account for a substantial portion of the differences in the likelihoods of studying STEM. While first-generation students are slightly less responsive to their early STEM grades than continuing-generation students, their comparatively lower early STEM grades are a significant driver of persistence differences.

Early stem cell aging in the mature brain.

Stem cell dysfunction drives many age-related disorders. Identifying mechanisms that initially compromise stem cell behavior represent early targets to promote tissue function later in life. Here, we pinpoint multiple factors that disrupt neural stem cell (NSC) behavior in the adult hippocampus. Clonal tracing showed that NSCs exhibit asynchronous depletion by identifying short-term NSCs (ST-NSCs) and long-term NSCs (LT-NSCs). ST-NSCs divide rapidly to generate neurons and deplete in the young brain. Meanwhile, multipotent LT-NSCs are maintained for months but are pushed out of homeostasis by lengthening quiescence. Single-cell transcriptome analysis of deep NSC quiescence revealed several hallmarks of molecular aging in the mature brain and identified tyrosine-protein kinase Abl1 as an NSC aging factor. Treatment with the Abl inhibitor imatinib increased NSC activation without impairing NSC maintenance in the middle-aged brain. Our study indicates that hippocampal NSCs are particularly vulnerable and adaptable to cellular aging.

Persistence of Positive RT-PCR for SARS-CoV-2 in an Early Allogenic Stem Cells Transplantation Recipient: A Case Study

The new coronavirus SARS-CoV-2 has rapidly spread over the world causing the disease by WHO called COVID-19, is the defining global health crisis of our time and the greatest threat we have faced during this century [1]. According to the World Health Organizations (WHO) data, in February 2021, almost one hundred fifteen million of the population has been infected with more than two million deaths [2]. To date, in the absence of any specific treatment, our knowledge about this disease remains very limited and is subject to rapid change. This virus represents a serious danger for patients with hematologic malignancies due to myeloablative conditioning and immunosuppressive treatments. People receiving chemotherapy with compromised immune systems and complications after stem cell transplant have an increased risk for infection [3]. COVID-19 infection may complicate clinical symptoms with higher risk of respiratory distress [4]. Actually, the experience in the management of COVID-19 infection in the stem cell transplant recipients is limited.

Analysis of Clinical Characteristics and Prognosis in Children with Acute Megakaryoblastic Leukemia without Down Syndrome

To analyze the clinical characteristics and treatment effects of children with acute megakaryoblastic leukemia without down syndrome (non-DS-AMKL). The clinical data of 19 children with non-DS-AMKL treated in the Pediatric Hematology Ward in Sun Yat-sen Memorial Hospital of Sun Yat-sen University from May 2008 to April 2018 were analyzed retrospectively. The clinical characteristics, laboratory test and treatment methods of the children were concluded. All patients were followed up to evaluate the effect of treatment. The 19 cases of children included nine male and ten female, the median age of onset was 2 years old. The clinical manifestations showed nonspecific. The median white blood cell of peripheral blood was 15.88×109/L, the median hemoglobin was 67 g/L and median platelet was 16×109/L. An increase of primitive and naive megakaryocytes was found in the bone marrow sample. The immunophenotypes of bone marrow detected by flow cytometry in 19 children were all positive expressed for CD41, CD61. Genetic tests showed that five cases carrying EVI1, including one complicating with MLL/AF10, one patient carrying HOX11, one carrying GATA1, IKZF1 and DDX11 mutations, one patient carrying missense mutation of NRAS, one with missense mutation of KRAS and two cases with high expression of WT1. Karyotype analysis were performed in 11 cases of children, including four with normal karyotype, four with complex karyotypes, one with trisomy 8, one with 7/13 trisomy 21 without Down syndrome manifestations (considered as abnormal somatic karyotype) and one Robertsonian translocation carrier involving chromosomes 14 and 21. Ten children received treatment, including three cases with allogeneic hematopoietic stem cell transplantation (allo-HSCT) after complete remission (CR), two cases with complete donor implantation, and one without implanted but hematopoietic recovered, these three children were followed up for 26, 15 and 12 months respectively and the minimal residual disease (MRD) were all less than 10-4. Another three cases achieving CR after chemotherapy but relapsed in 5, 10 and 12 months after the onset respectively, and all the three children were died eventually. One children died of pulmonary hemorrhage during chemotherapy and three children died from discontinuation of treatment after non remission. The remaining nine children died because of without chemotherapy treatment. Non-DS-AMKL was rare in children and difficult to be diagnosed. Determination of MICM classification as early as possible was helpful for diagnosis, and genetic testing played an important role for diagnosis and prognosis evaluation. Early hematopoietic stem cell transplantation in patients with CR after chemotherapy might be an effective way to cure AMKL.

DNA methylation and histone modifications are essential for regulation of stem cell formation and differentiation in zebrafish development.

The complex processes necessary for embryogenesis require a gene regulatory network that is complex and systematic. Gene expression regulates development and organogenesis, but this process is altered and fine-tuned by epigenetic regulators that facilitate changes in the chromatin landscape. Epigenetic regulation of embryogenesis adjusts the chromatin structure by modifying both DNA through methylation and nucleosomes through posttranslational modifications of histone tails. The zebrafish is a well-characterized model organism that is a quintessential tool for studying developmental biology. With external fertilization, low cost and high fecundity, the zebrafish are an efficient tool for studying early developmental stages. Genetic manipulation can be performed in vivo resulting in quick identification of gene function. Large-scale genome analyses including RNA sequencing, chromatin immunoprecipitation and chromatin structure all are feasible in the zebrafish. In this review, we highlight the key events in zebrafish development where epigenetic regulation plays a critical role from the early stem cell stages through differentiation and organogenesis.

Shaping the human brain.

Lancaster and colleagues describe a morphological change in early primate brain stem cells, the timing of which could underlie the differences in brain size between human and apes.

اثر سطوح مختلف رطوبتی و سولفات آهن بر رشد و محتوی برخی از عناصر در گندم (.Triticum aestivum L) رقم روشن

بسیاری از اراضی کشاورزی در مناطق خشک و نیمه‌خشک، با کمبود آب و عناصر غذائی مخصوصا عناصر غذائی کم ­نیاز مواجه هستند. در این راستا، این تحقیق با هدف بررسی اثرات کاربرد خاکی سولفات آهن در شرایط رطوبتی مختلف بر روی خصوصیات رشدی گندم رقم روشن، به‌صورت آزمایش فاکتوریل دوعاملی در قالب طرح بلوک­های کامل تصادفی با 4 تکرار در گلخانه تحقیقاتی دانشکده کشاورزی دانشگاه بیرجند در سال 1391 انجام شد. عامل اول شامل چهار سطح سولفات آهن (صفر، 50، 100 و 150 میلی­ گرم بر کیلوگرم خاک) و عامل دوم نیز شامل دو سطح مختلف رطوبتی (50 و 100 درصد رطوبت ظرفیت زراعی خاک) بود که از طریق توزین روزانه گلدان­ها کنترل می­شد. برداشت اندام هوائی گیاه بعد از گذشت 8 هفته صورت گرفت و صفاتی چون ارتفاع بوته، تعداد برگ، سطح برگ، شاخص سبزینگی و محتوای نسبی آب برگ، درصد نیتروژن، درصد پروتئین، درصد پتاسیم و محتوای آهن برگ اندازه ­گیری شد. نتایج نشان داد که مصرف سولفات آهن روی درصد نیتروژن برگ و محتوای آهن برگ تأثیر معنی­ داری داشت. کاهش سطح تأمین رطوبت منجر به کاهش 31 درصدی ماده خشک تولیدی، 5.8 درصدی ارتفاع بوته، 37 درصدی تعداد برگ، 22 درصدی تعداد ساقه در بوته، 33 درصدی سطح برگ، 16.8 درصدی محتوی نسبی آب برگ، 5.45 درصدی شاخص سبزینگی و همچنین افزایش 18.2 درصدی نیتروژن برگ، 17.9 درصدی پتاسیم برگ و 23.7 درصدی پروتئین برگ شد. افزایش کاربرد 100 میلی­گرم سولفات آهن بر کیلوگرم خاک نیز محتوای آهن برگ را در حدود 100 درصد افزایش داد. به‌طورکلی کاربرد کود سولفات آهن منجر به بهبود رشد گندم تحت تنش خشکی شد.

Canonical and early lineage-specific stem cell types identified in planarian SirNeoblasts

BackgroundThe pluripotent stem cells in planarians, a model for tissue and cellular regeneration, remain further identification. We recently developed a method to enrich piwi-1+ cells in Schmidtea mediterranea, by staining cells with SiR-DNA and Cell Tracker Green, named SirNeoblasts that permits their propagation and subsequent functional study in vivo. Since traditional enrichment for planarian neoblasts by Hoechst 33342 staining generates X1 cells, blocking the cell cycle and inducing cytotoxicity, this method by SiR-DNA and Cell Tracker Green represents a complementary technological advance for functional investigation of cell fate and regeneration. However, the similarities in heterogeneity of cell subtypes between SirNeoblasts and X1 remain unknown.ResultsIn this work, we performed single cell RNA sequencing of SirNeoblasts for comparison with differential expression patterns in a publicly available X1 single cell RNA sequencing data. We found first that all of the lineage-specific progenitor cells in X1 were present in comparable proportions in SirNeoblasts. In addition, SirNeoblasts contain an early muscle progenitor that is unreported in X1. Analysis of new markers for putative pluripotent stem cells identified here, with subsequent sub-clustering analysis, revealed earlier lineages of epidermal, muscular, intestinal, and pharyngeal progenitors than have been observed in X1. Using the gcm as a marker, we also identified a cell subpopulation resided in previously identified tgs-1+ neoblasts. Knockdown of gcm impaired the neoblast repopulation, suggesting a function of gcm in neoblasts.ConclusionsIn summary, the use of SirNeoblasts will enable broad experimental advances in regeneration and cell fate specification, given the possibility for propagation and transplantation of recombinant and mutagenized pluripotent stem cells that are not previously afforded to this rapid and versatile model system.

Nurturing Kindergarteners’ Early STEM Academic Identity Through Makerspace Pedagogy

Children’s attitudes about STEM are formed early. As such, the ecology of early childhood classrooms can either afford or deny access to relevant experiences that help children nurture early self-understandings and ways of positioning themselves in relation to STEM. This early positioning and the funds of identity they afford are critical to early STEM academic identity development. While Fasso and Knight (2020) offer Makerspace Pedagogy for fostering identity development, Marsh et al. (2017) note a lack of wide-spread application and related research on deliberate employment of Makerspace Pedagogy in early childhood classrooms. To address this gap and to explore the adoption of Makerspace pedagogy in an early public-school setting, a university research team joined with a kindergarten teacher to pilot an emergent curriculum focused on tinkering and making. Initial findings suggest that design-based activities may support kindergartners taking on the role of STEM practitioners, that Makerspace activities seem to help produce a syncretism of early STEM and multimodal literacy practices and that Makerspace Pedagogy may offer a unique opportunity to explore the intersection of various aspects of children’s early identity formation.

Osteology and digital reconstruction of the skull of the early tetrapod Whatcheeria deltae

ABSTRACT The Early Carboniferous stem tetrapod Whatcheeria deltae is among the earliest-branching limbed tetrapods represented by multiple near-complete specimens, making it an important taxon in understanding the vertebrate water-to-land transition. However, all preserved skulls of Whatcheeria suffer from post-mortem crushing and lateral compression, which has made cranial reconstruction problematic. In this study, computed tomography data of three Whatcheeria specimens were segmented using visualization software to digitally separate each individual skull bone from matrix. Digital methods were used to repair and retrodeform the bones and produce the first complete three-dimensional skull reconstruction of Whatcheeria. We provide a revised description of the cranial and lower jaw anatomy of Whatcheeria based on CT data, focusing on sutural morphology and previously unknown anatomical details. Our findings suggest that Whatcheeria had one of the narrowest skulls of any known early tetrapod, a gap between the nasals, and significant overlap of the lacrimal onto the nasal and prefrontal. Sutural morphology is used to infer loading regime in the skull during feeding and suggests the skull of Whatcheeria was well adapted to resist stresses induced by biting large prey with its enlarged anterior fangs.

Femoral Stem Subsidence and its Associated Factors after Cementless Bipolar Hemiarthroplasty in Geriatric Patients

Introduction:Early femoral stem subsidence has been a concern as a predictor of the beginning of implant loosening, especially on cementless hip arthroplasty implants. This study aimed to determine the factors that affect femoral stem subsidence and outcome following hemiarthroplasty in the geriatric population.Materials and Methods:This is a retrospective study of 179 patients who underwent cementless bipolar hemiarthroplasty during the 2011-2019 period at an orthopaedic and traumatology hospital. Data on the patient's demography, pre-operative American Society Anaesthesiologist (ASA) score, body mass index (BMI), canal flare index (CFI), Dorr classification, and stem alignment were obtained. The primary outcomes were post-operative femoral stem subsidence, post-operative pain, and functional outcome using Harris Hip Score (HHS). Statistical analysis was conducted to identify risk factors associated with the primary outcome.Results:The mean femoral stem subsidence was 2.16 ±3.4 mm. The mean post-operative Visual Analog Score (VAS) on follow-up was 1.38 ± 1. Mean HHS on follow-up was 85.28±10.3. American Society Anaesthesiologist score 3 (p = 0.011, OR = 2.77) and varus alignment (p=0.039, OR = 6.963) were related to worse stem subsidence. Otherwise, neutral alignment (p = 0.045 and OR = 0.405) gave protection against femoral stem subsidence. The female gender (p = 0.014, OR 2.53) was associated with postoperative pain onset. Neutral alignment had significant relationship with functional outcomes (p = 0.01; OR 0.33).Conclusion:A higher ASA score and varus stem alignment were related to a higher risk of femoral stem subsidence. Meanwhile, neutral stem alignment had a protective effect on the femoral stem subsidence and outcome.

Transcriptional networks identify synaptotagmin-like 3 as a regulator of cortical neuronal migration during early neurodevelopment

Human brain development is a complex process involving neural proliferation, differentiation, and migration that are directed by many essential cellular factors and drivers. Here, using the NetBID2 algorithm and developing human brain RNA sequencing dataset, we identify synaptotagmin-like 3 (SYTL3) as one of the top drivers of early human brain development. Interestingly, SYTL3 exhibits high activity but low expression in both early developmental human cortex and human embryonic stem cell (hESC)-derived neurons. Knockout of SYTL3 (SYTL3-KO) in human neurons or knockdown of Sytl3 in embryonic mouse cortex markedly promotes neuronal migration. SYTL3-KO causes an abnormal distribution of deep-layer neurons in brain organoids and reduces presynaptic neurotransmitter release in hESC-derived neurons. We further demonstrate that SYTL3-KO-accelerated neuronal migration is modulated by high expression of matrix metalloproteinases. Together, based on bioinformatics and biological experiments, we identify SYTL3 as a regulator of cortical neuronal migration in human and mouse developing brains.