Mesodermal Cells Research Articles

Poly(vinyl alcohol)/Gelatin Scaffolds Allow Regeneration of Nasal Tissues

Need for regeneration and repair of nasal tissues occurs as a consequence of several pathologies affecting the nose, including, but not limited to infective diseases, traumas and tumor resections. A platform for nasal tissue regeneration was set up using poly(vinyl alcohol)/gelatin sponges with 20%–30% (w/w) gelatin content to be used as scaffolds, for their intrinsic hydrophilic, cell adhesive and shape recovery properties. We propose mesodermal progenitor cells (MPCs) isolated from the bone marrow as a unique stem cell source for obtaining different connective tissues of the nose, including vascular tissue. Finally, epithelial cell immune response to these scaffolds was assessed in vitro in an environment containing inflammatory molecules. The results showed that mesenchymal stromal cells (MSCs) deriving from MPCs could be used to differentiate into cartilage and fibrous tissue; whereas, in combination with endothelial cells still deriving from MPCs, into pre-vascularized bone. Finally, the scaffold did not significantly alter the epithelial cell response to inflammatory insults derived from interaction with bacterial molecules.

Peripheral Nerve-Derived Stem Cell Spheroids Induce Functional Recovery and Repair after Spinal Cord Injury in Rodents.

Stem cell therapy is one of the most promising candidate treatments for spinal cord injury. Research has shown optimistic results for this therapy, but clinical limitations remain, including poor viability, engraftment, and differentiation. Here, we isolated novel peripheral nerve-derived stem cells (PNSCs) from adult peripheral nerves with similar characteristics to neural-crest stem cells. These PNSCs expressed neural-crest specific markers and showed multilineage differentiation potential into Schwann cells, neuroglia, neurons, and mesodermal cells. In addition, PNSCs showed therapeutic potential by releasing the neurotrophic factors, including glial cell-line-derived neurotrophic factor, insulin-like growth factor, nerve growth factor, and neurotrophin-3. PNSC abilities were also enhanced by their development into spheroids which secreted neurotrophic factors several times more than non-spheroid PNSCs and expressed several types of extra cellular matrix. These features suggest that the potential for these PNSC spheroids can overcome their limitations. In an animal spinal cord injury (SCI) model, these PNSC spheroids induced functional recovery and neuronal regeneration. These PNSC spheroids also reduced the neuropathic pain which accompanies SCI after remyelination. These PNSC spheroids may represent a new therapeutic approach for patients suffering from SCI.

A simplified approach for derivation of induced pluripotent stem cells from Epstein-Barr virus immortalized B-lymphoblastoid cell lines

Given the limited availability of tissue, especially brain tissue, for neurological diseases and disorders research, the development of alternative biological tools for investigations of underlying molecular and genetic mechanisms is imperative. One important resource for this task is the large repositories that bank immortalized blood cells (i.e. lymphoblastoid cell lines; LCLs) from affected individuals and their unaffected family members. These repositories document demographic, phenotypic, and, in some cases, genotypic information about the donors and thus provide a ready-made sample source for hypothesis testing. Importantly, patient-specific LCLs can be used to generate induced pluripotent stem cells (iPSC) that, in turn, can be used to create specific cell types for use in mechanistic studies. To investigate this concept further, LCLs from two males (proband and sibling) were obtained from one such repository, the Autism Genetics Resource Exchange (AGRE), and iPSCs were generated by transfection with Epi5 Episomal iPSC reprogramming plasmids. Characterization of the resultant cell lines by PCR, RT-PCR, immunocytochemistry, karyotyping, and the Taqman® human pluripotent stem cell Scorecard™ Panel, was used to provide evidence of endogenous pluripotency and then to evaluate the trilineage potential of four representative clones. Results indicated that all four iPSC lines were initially pluripotent and displayed the trilineage potential predictive for successful differentiation to mesoderm, endoderm, or ectoderm-derived cell types. Compared to other published protocols, this study details a somewhat simplified approach, used here specifically for the generation and characterization of induced pluripotent stem cells from well-characterized and banked LCLs.

A mechanical model of early somite segmentation

SummarySomitogenesis is often described using the clock-and-wavefront (CW) model, which does not explain how molecular signaling rearranges the pre-somitic mesoderm (PSM) cells into somites. Our scanning electron microscopy analysis of chicken embryos reveals a caudally-progressing epithelialization front in the dorsal PSM that precedes somite formation. Signs of apical constriction and tissue segmentation appear in this layer 3-4 somite lengths caudal to the last-formed somite. We propose a mechanical instability model in which a steady increase of apical contractility leads to periodic failure of adhesion junctions within the dorsal PSM and positions the future inter-somite boundaries. This model produces spatially periodic segments whose size depends on the speed of the activation front of contraction (F), and the buildup rate of contractility (Λ). The Λ/F ratio determines whether this mechanism produces spatially and temporally regular or irregular segments, and whether segment size increases with the front speed.

A single-cell analysis of the Arabidopsis vegetative shoot apex.

The shoot apical meristem allows for reiterative formation of new aerial structures throughout the life cycle of a plant. We use single-cell RNA sequencing to define the cellular taxonomy of the Arabidopsis vegetative shoot apex at the transcriptome level. We find that the shoot apex is composed of highly heterogeneous cells, which can be partitioned into 7 broad populations with 23 transcriptionally distinct cell clusters. We delineate cell-cycle continuums and developmental trajectories of epidermal cells, vascular tissue, and leaf mesophyll cells and infer transcription factors and gene expression signatures associated with cell fate decisions. Integrative analysis of shoot and root apical cell populations further reveals common and distinct features of epidermal and vascular tissues. Our results, thus, offer a valuable resource for investigating the basic principles underlying cell division and differentiation in plants at single-cell resolution.

Modulation of APLNR Signaling Is Required during the Development and Maintenance of the Hematopoietic System.

SummaryApelin receptor (APLNR/AGTRLl1/APJ) marks a transient cell population during the differentiation of hematopoietic stem and progenitor cells (HSPCs) from pluripotent stem cells, but its function during the production and maintenance of hematopoietic stem cells is not clear. We generated an Aplnr-tdTomato reporter mouse embryonic stem cell (mESC) line and showed that HSPCs are generated exclusively from mesodermal cells that express Aplnr-tdTomato. HSPC production from mESCs was impaired when Aplnr was deleted, implying that this pathway is required for their production. To address the role of APLNR signaling in HSPC maintenance, we added APELIN ligands to ex vivo AGM cultures. Activation of the APLNR pathway in this system impaired the generation of long-term reconstituting HSPCs and appeared to drive myeloid differentiation. Our data suggest that the APLNR signaling is required for the generation of cells that give rise to HSCs, but that its subsequent downregulation is required for their maintenance.

The Drosophila Forkhead/Fox transcription factor Jumeau mediates specific cardiac progenitor cell divisions by regulating expression of the kinesin Nebbish

Forkhead (Fkh/Fox) domain transcription factors (TFs) mediate multiple cardiogenic processes in both mammals and Drosophila. We showed previously that the Drosophila Fox gene jumeau (jumu) controls three categories of cardiac progenitor cell division—asymmetric, symmetric, and cell division at an earlier stage—by regulating Polo kinase activity, and mediates the latter two categories in concert with the TF Myb. Those observations raised the question of whether other jumu-regulated genes also mediate all three categories of cardiac progenitor cell division or a subset thereof. By comparing microarray-based expression profiles of wild-type and jumu loss-of-function mesodermal cells, we identified nebbish (neb), a kinesin-encoding gene activated by jumu. Phenotypic analysis shows that neb is required for only two categories of jumu-regulated cardiac progenitor cell division: symmetric and cell division at an earlier stage. Synergistic genetic interactions between neb, jumu, Myb, and polo and the rescue of jumu mutations by ectopic cardiac mesoderm-specific expression of neb demonstrate that neb is an integral component of a jumu-regulated subnetwork mediating cardiac progenitor cell divisions. Our results emphasize the central role of Fox TFs in cardiogenesis and illustrate how a single TF can utilize different combinations of other regulators and downstream effectors to control distinct developmental processes.

Coxsackievirus B3 Infection of Human iPSC Lines and Derived Primary Germ-Layer Cells Regarding Receptor Expression.

The association of members of the enterovirus family with pregnancy complications up to miscarriages is under discussion. Here, infection of two different human induced pluripotent stem cell (iPSC) lines and iPSC-derived primary germ-layer cells with coxsackievirus B3 (CVB3) was characterized as an in vitro cell culture model for very early human development. Transcriptomic analysis of iPSC lines infected with recombinant CVB3 expressing enhanced green fluorescent protein (EGFP) revealed a reduction in the expression of pluripotency genes besides an enhancement of genes involved in RNA metabolism. The initial distribution of CVB3-EGFP-positive cells within iPSC colonies correlated with the distribution of its receptor coxsackie- and adenovirus receptor (CAR). Application of anti-CAR blocking antibodies supported the requirement of CAR, but not of the co-receptor decay-accelerating factor (DAF) for infection of iPSC lines. Among iPSC-derived germ-layer cells, mesodermal cells were especially vulnerable to CVB3-EGFP infection. Our data implicate further consideration of members of the enterovirus family in the screening program of human pregnancies. Furthermore, iPSCs with their differentiation capacity into cell populations of relevant viral target organs could offer a reliable screening approach for therapeutic intervention and for assessment of organ-specific enterovirus virulence.

Generation of nonviral integration-free human iPS cell line KISCOi001-A from normal human fibroblasts, under defined xeno-free and feeder-free conditions.

KISCOi001-A is a healthy feeder-free and fully characterized human induced pluripotent stem (iPS) cell line cultured under xeno-free and defined conditions. The cell line is generated from normal human foreskin fibroblasts with non-integrating episomal plasmid vectors encoding OCT4, SOX2, KLF4, NANOG, LIN28, nontransforming L-MYC and dominant negative p53. The generated iPS cells are transgene-free and their pluripotency is confirmed by the expression of stem cell markers and capacity to differentiate into the cells of ectoderm, endoderm and mesoderm while their identity and karyotype stability is confirmed with Genomic assays.

4-Octylphenol induces developmental abnormalities and interferes the differentiation of neural crest cells in Xenopus laevis embryos.

Developmental toxicity of 4-octylphenol (OP), an estrogenic endocrine disruptor was verified using frog embryo teratogenesis assay Xenopus. LC50, EC50Malformtion and EC50Melanocyte-dysgenesis of OP were 9.9, 10.5, and 2.4μM, respectively. In tadpoles, despite the low teratogenic index, 2μM OP significantly inhibited head cartilage development and tail malformation. The total length of tadpole was significantly increased at 5μM and decreased at 10μM OP. In OP-treated tadpoles, head cartilages were frequently missed and col2a1 mRNA was decreased at 2μM, indicating a chondrogenic defect in developing head. In the head skin of 1μM OP-treated tadpoles, number of melanocytes and melanogenic pathway genes expression were significantly decreased. In the head-neck junction of stage 22 embryos, OP increased foxd3 and sox10 mRNA and SOX10(+) neural crest cells (NCCs) in somite mesoderm and endoderm, indicating the inhibition of chondrogenic differentiation, ectopic migration to endoderm, and undifferentiation of NCCs by OP. Together, OP-induced head dysplasia and inhibition of melanogenesis may be attributable to deregulation of neural crest cells in embryos. In tadpoles, OP at 1μM significantly increased lipid hydroperoxide and induced spliced xbp1 mRNA, an IRE1 pathway endoplasmic reticulum stress (ERS) marker and p-eIF2α protein, a PERK pathway ERS marker. OP at 10μM induced CHOP mRNA, pro-apoptotic genes expression, DNA fragmentation, and cleaved caspase-3, suggesting that OP differentially induced ERS and apoptosis according to the concentration in embryos. In 5-10μM OP-treated stage 22 embryos and stage 45 tadpole heads, Ki67 was significantly increased, suggesting the apoptosis-induced proliferation of embryonic cells in the OP-treated embryos. Together, OP should be managed as a developmental toxicant altering the behavior of NCCs in vertebrates.

Early patterning of ABCB, ABCC, and ABCG transporters establishes unique territories of small molecule transport in embryonic mesoderm and endoderm

Directed intercellular movement of diverse small molecules, including metabolites, signal molecules and xenobiotics, is a key feature of multicellularity. Networks of small molecule transporters (SMTs), including several ATP Binding Cassette (ABC) transporters, are central to this process. While small molecule transporters are well described in differentiated organs, little is known about their patterns of expression in early embryogenesis. Here we report the pattern of ABC-type SMT expression and activity during the early development of sea urchins. Of the six major ABCs in this embryo (ABCB1, -B4, -C1, -C4, -C5 and -G2), three expression patterns were observed: 1) ABCB1 and ABCC1 are first expressed ubiquitously, and then become enriched in endoderm and ectoderm-derived structures. 2) ABCC4 and ABCC5 are restricted to a ring of mesoderm in the blastula and ABCC4 is later expressed in the coelomic pouches, the embryonic niche of the primordial germ cells. 3) ABCB4 and ABCG2 are expressed exclusively in endoderm-fated cells. Assays with fluorescent substrates and inhibitors of transporters revealed a ring of ABCC4 efflux activity emanating from ABCC4+ mesodermal cells. Similarly, ABCB1 and ABCB4 efflux activity was observed in the developing gut, prior to the onset of feeding. This study reveals the early establishment of unique territories of small molecule transport during embryogenesis. A pattern of ABCC4/C5 expression is consistent with signaling functions during gut invagination and germ line development, while a later pattern of ABCB1/B4 and ABCG2 is consistent with roles in the embryonic gut. This work provides a conceptual framework with which to examine the function and evolution of SMT networks and to define the specific developmental pathways that drive the expression of these genes.

Neuromesodermal Progenitors: A Basis for Robust Axial Patterning in Development and Evolution.

During early development the vertebrate embryo elongates through a combination of tissue shape change, growth and progenitor cell expansion across multiple regions of the body axis. How these events are coordinated across the length of the embryo to generate a well-proportioned body axis is unknown. Understanding the multi-tissue interplay of morphogenesis, growth and cell fate specification is essential for us to gain a complete understanding how diverse body plans have evolved in a robust manner. Within the posterior region of the embryo, a population of bipotent neuromesodermal progenitors generate both spinal cord and paraxial mesoderm derivatives during the elongation of the vertebrate body. Here we summarize recent data comparing neuromesodermal lineage and their underlying gene-regulatory networks between species and through development. We find that the common characteristic underlying this population is a competence to generate posterior neural and paraxial mesoderm cells, with a conserved Wnt/FGF and Sox2/T/Tbx6 regulatory network. We propose the hypothesis that by maintaining a population of multi-germ layer competent progenitors at the posterior aspect of the embryo, a flexible pool of progenitors is maintained whose contribution to the elongating body axis varies as a consequence of the relative growth rates occurring within anterior and posterior regions of the body axis. We discuss how this capacity for variation in the proportions and rates of NM specification might have been important allowing for alterations in the timing of embryo growth during evolution.

Chicken Second Branchial Arch Progenitor Cells Contribute to Heart Musculature in vitro and in vivo

In the past, the heart muscle was thought to originate from a single source of myocardial progenitor cells. More recently, however, an additional source of myocardial progenitors has been revealed to be the second heart field, and chicken embryos were important for establishing this concept. However, there have been few studies in chicken on how this field contributes to heart muscles in vitro. We have developed an ex vivo experimental system from chicken embryos between stages HH17–20 to investigate how mesodermal progenitors in the second branchial arch (BA2) differentiate into cardiac muscles. Using this method, we presented evidence that the progenitor cells within the BA2 arch differentiated into beating cardiomyocytes in vitro. The beating explant cells were positive for cardiac actin, Nkx2.5, and ventricular myosin heavy chain. In addition, we performed a time course for the expression of second heart field markers (Isl1 and Nkx2.5) in the BA2 from stage HH16 to stage HH21 using in situ hybridization. Accordingly, using EGFP-based cell labeling techniques and quail-chicken cell injection, we demonstrated that mesodermal cells from the BA2 contributed to the outflow tract and ventricular myocardium in vivo. Thus, our findings highlight the cardiogenic potential of chicken BA2 mesodermal cells in vitro and in vivo.

Imaging of dynamic actin remodeling reveals distinct behaviors of head and trunk mesoderm in gastrulating Xenopus laevis.

Gastrulation involves coordinated movements of cells, facilitating mesoderm and endoderm internalization and proper patterning of tissues across the germ layers. In Xenopus laevis, head mesoderm migrates collectively along the blastocoel roof fibronectin network towards the animal pole. Meanwhile, the trunk mesodermal cells migrate over each other in convergent thickening and convergent extension movements elongating the body axis. The behaviors of cells in these regions are investigated mainly in tissue explants taken from the respective head or trunk mesodermal regions. How cells behave at the transitional zone between these territories is not described in detail. To learn about cell behaviors around this junction, we imaged cell movements in an explant that encompassed the head and trunk mesoderm. We observed that head mesoderm migration on fibronectin employed lamellipodial protrusions at the leading edge and dynamic actin remodeling in the trailing cells. Trunk mesodermal cells underwent mediolateral cell elongation and intercalation to form the notochord. Lateral edges of the notochord were defined before the anterior edge. Our movie reveals distinct mesodermal cell behaviors occurring simultaneously in different regions of gastrulating embryos. This study highlights the power of applying modern microscopy tools to revisit classical experiments, permitting a greater understanding of the cellular dynamics that shape the embryo.

Autophagy Promoted Neural Differentiation of Human Placenta-derived Mesenchymal Stem Cells.

Human placenta-derived mesenchymal stem cells (hPMSCs) are multipotent and possess neurogenicity. Numerous studies have shown that Notch inhibition and DNA demethylation promote neural differentiation. Here, we investigated the modulation of autophagy during neural differentiation of hPMSCs, induced by DAPT and 5-Azacytidine. hPMSCs were treated with DAPT to induce neural differentiation, and the autophagy regulating molecules were used to assess the impact of autophagy on neural differentiation. The hPMSCs presented with typical mesenchymal stem cell phenotypes, in which the majority of cells expressed CD73, CD90 and CD105. hPMSCs were multipotent, capable of differentiating into mesodermal cells. After treatment with DAPT, hPMSCs upregulated the expression of neuronal genes including SOX2, Nestin, and βIII-tubulin, and the autophagy genes LC3I/II and Beclin. These genes were further increased when 5-Azacytidine was co-supplemented in the culture medium. The inhibition of autophagy by chloroquine impeded the neural differentiation of hPMSCs, marked by the downregulation of βIII-tubulin, while the activation of autophagy by valproic acid (VPA) instigated the emergence of βIII-tubulin-positive cells. During the differentiation process, autophagy was modulated, implying that autophagy could play a significant role during the differentiation of these cells. The blockage and stimulation of autophagy could either hinder or induce the formation of neural-like cells, respectively. Therefore, the refinement of autophagic activity at an appropriate level might improve the efficiency of stem cell differentiation.

Single-Cell Analysis of Embryoids Reveals Lineage Diversification Roadmaps of Early Human Development

Despite its clinical and basic importance, our understanding of early human development remains limited. Stem cell-derived, embryo-like structures (or embryoids) allowing studies of embryonic development without using natural embryos can potentially help fill the knowledge gap of human development. Herein, transcriptome at the single-cell level of a human embryoid model, which recapitulates aspects of lineage diversification and three-dimensional tissue architecture of the human embryo from the implantation to the onset of gastrulation, was profiled at different time points. Molecular maps of lineage diversifications from the pluripotent human epiblast towards the amniotic ectoderm, primitive streak / mesoderm, and primordial germ cells were constructed and compared with in vivo primate data. Similarly, chimpanzee embryoids were generated and profiled to reveal transcriptome dynamics during the early post-implantation chimpanzee development. Our comparative transcriptome analyses reveal a critical role of NODAL signaling in human mesoderm and primordial germ cell specification, which is further functionally validated. Through comparative transcriptome analyses and validations with human blastocysts and in vitro cultured cynomolgus embryos, we further proposed stringent criteria for distinguishing between human trophectoderm and amniotic ectoderm cells. Altogether, this study provides new knowledge of the lineage diversification roadmap of early human development and will serve as a valuable resource for studying human development.

Differentiation of Human Adipose Tissue Derived Mesenchymal Stem Cells to Functional Hepatocytes like Cells: Exploring the Niche and Admscs Conditioned Media

Tissue engineered approaches have been used for hepatic regeneration and growing hepatic cells in-vitro, however it remains challenging due to the lesser proliferative capacity of the hepatic cells as well as cell source availability due to damaged tissues. Stem cells could be an alternative strategy if differentiated to functional hepatic like cells by providing suitable niche or culture conditions. In the present study we tried to explore differentiation of mesodermal origin cells to the endodermal origin hepatic cells under different culture conditions including fibrin niche and adipocytes derived mesenchymal stem cells conditioned medium. Fibrin provides better cell adhesion and proliferation. Mesenchymal stem cells were isolated and characterized. Two step induction was used for differentiating into hepatic like cells. Functionality of the differentiated hepatic like cells was analyzed using Cyto P 450, glycogen storage and urea/albumin assay at different time points. It was observed the fibrin niche supports the hepatic differentiation and differentiated cells shows hepatic functions. This approach may be used to generate alternative cell source for hepatic tissue engineering.

Zebrafish Vascular Development: General and Tissue-Specific Regulation.

Circulation is required for the delivery of oxygen and nutrition to tissues and organs, as well as waste collection. Therefore, the heart and vessels develop first during embryogenesis. The circulatory system consists of the heart, blood vessels, and blood cells, which originate from the mesoderm. The gene expression pattern required for blood vessel development is predetermined by the hierarchical and sequential regulation of genes for the differentiation of mesodermal cells. Herein, we review how blood vessels form distinctly in different tissues or organs of zebrafish and how vessel formation is universally or tissue-specifically regulated by signal transduction pathways and blood flow. In addition, the unsolved issues of mutual contacts and interplay of circulatory organs during embryogenesis are discussed.

The CXCR4/SDF-1 Axis in the Development of Facial Expression and Non-somitic Neck Muscles.

Trunk and head muscles originate from distinct embryonic regions: while the trunk muscles derive from the paraxial mesoderm that becomes segmented into somites, the majority of head muscles develops from the unsegmented cranial paraxial mesoderm. Differences in the molecular control of trunk versus head and neck muscles have been discovered about 25 years ago; interestingly, differences in satellite cell subpopulations were also described more recently. Specifically, the satellite cells of the facial expression muscles share properties with heart muscle. In adult vertebrates, neck muscles span the transition zone between head and trunk. Mastication and facial expression muscles derive from the mesodermal progenitor cells that are located in the first and second branchial arches, respectively. The cucullaris muscle (non-somitic neck muscle) originates from the posterior-most branchial arches. Like other subclasses within the chemokines and chemokine receptors, CXCR4 and SDF-1 play essential roles in the migration of cells within a number of various tissues during development. CXCR4 as receptor together with its ligand SDF-1 have mainly been described to regulate the migration of the trunk muscle progenitor cells. This review first underlines our recent understanding of the development of the facial expression (second arch-derived) muscles, focusing on new insights into the migration event and how this embryonic process is different from the development of mastication (first arch-derived) muscles. Other muscles associated with the head, such as non-somitic neck muscles derived from muscle progenitor cells located in the posterior branchial arches, are also in the focus of this review. Implications on human muscle dystrophies affecting the muscles of face and neck are also discussed.

Pengaruh Pemberian Ekstrak Kulit Buah Manggis (Garcinia mangostana L.) Menghambat Flexi Cranial Embrio Ayam Umur 48 Jam

Embryonic development forms three layers of germinativum that is endoderm, mesoderm and ectoderm. At the beginning of intrauterine fetus exchange of substances through diffusion, but in line with the development of the embryo, nutrition is not obtained through diffusion alone. Endoderm lining mesoderm cells will then be formed angioblasts as an early sign of vascularity. In the development of flexi cranial can be known from the development of the brain and heart which is characterized by bending the head of the embryo and decreased heart from the cranial down. The development of a normal vascular system will have an effect on organogenesis and morphogesis processes. If there are disturbances in the process of growth and development at that stage can cause congenital abnormalities. Xanthones mangosteen peel is rich in benefits, but in research on the effects of xanthones on embryos is still very limited and has not been able to explain their effects on the condition of the fetus. The study was conducted with the aim to determine the effect of mangosteen peel extract inhibiting the flexi cranial of 48 hours chicken embryos using doses of 100 µg/mL, 150 µg/mL and 200 µg/mL. Mangosteen peel extract is injected into chicken eggs less than 7 days after oviposition then incubated for 48 hours. The results showed that mangosteen peel extract inhibited flexi cranial of chicken embryos with Chi-Square test results (p = 0.002). The conclusion of the study was that exposure to mangosteen peel extract inhibited flexi cranial and gave significant results.