Regulation Of Stem Cells Research Articles

Photonic control of ligand nanospacing in self-assembly regulates stem cell fate

Extracellular matrix (ECM) undergoes dynamic inflation that dynamically changes ligand nanospacing but has not been explored. Here we utilize ECM-mimicking photocontrolled supramolecular ligand-tunable Azo+ self-assembly composed of azobenzene derivatives (Azo+) stacked via cation-π interactions and stabilized with RGD ligand-bearing poly(acrylic acid). Near-infrared-upconverted-ultraviolet light induces cis-Azo+-mediated inflation that suppresses cation-π interactions, thereby inflating liganded self-assembly. This inflation increases nanospacing of “closely nanospaced” ligands from 1.8 nm to 2.6 nm and the surface area of liganded self-assembly that facilitate stem cell adhesion, mechanosensing, and differentiation both in vitro and in vivo, including the release of loaded molecules by destabilizing water bridges and hydrogen bonds between the Azo+ molecules and loaded molecules. Conversely, visible light induces trans-Azo+ formation that facilitates cation-π interactions, thereby deflating self-assembly with “closely nanospaced” ligands that inhibits stem cell adhesion, mechanosensing, and differentiation. In stark contrast, when ligand nanospacing increases from 8.7 nm to 12.2 nm via the inflation of self-assembly, the surface area of “distantly nanospaced” ligands increases, thereby suppressing stem cell adhesion, mechanosensing, and differentiation. Long-term in vivo stability of self-assembly via real-time tracking and upconversion are verified. This tuning of ligand nanospacing can unravel dynamic ligand-cell interactions for stem cell-regulated tissue regeneration.

ITGA2 as a prognostic factor of glioma promotes GSCs invasion and EMT by activating STAT3 phosphorylation.

Glioma is the most common malignant brain tumor in adults with a high mortality and recurrence rate. Integrin alpha 2 (ITGA2) is involved in cell adhesion, stem cell regulation, angiogenesis and immune cell function. The role of ITGA2 in glioma malignant invasion remains unknown. The function and clinical relevance of ITGA2 were analysed by bioinformatics databases. The expression of ITGA2 in parent cells and GSCs was detected by flow cytometry and immunofluorescence double staining. The role of ITGA2 on the malignant phenotype of GSCs and epithelial-mesenchymal transition (EMT) was identified by stem cell function assays and Western blot. The effect of ITGA2 on glioma progression in vivo was determined by the intracranial orthotopic xenograft model. Immunohistochemistry, Spearman correlation and Kaplan-Meier were used to analyse the relationship of ITGA2 with clinical features and glioma prognosis. Biological analysis showed that ITGA2 might be related to cell invasion and migration. ITGA2, enriched in GSCs and co-expressed with SOX2, promoted the invasion and migration of GSCs by activating STAT3 phosphorylation and enhancing EMT. ITGA2 knockout suppressed the intracranial orthotopic xenograft growth and prolonged the survival of xenograft mice. In addition, the expression level of ITGA2 was significantly correlated to the grade of malignancy, N-cadherin and Ki67. High expression of ITGA2 indicated a worse prognosis of glioma patients. As a biomarker for the prediction of prognosis, ITGA2 promotes the malignant invasion of GSCs by activating STAT3 phosphorylation and enhancing EMT, leading to tumor recurrence and poor prognosis.

Regulation of Stem Cell Proliferation by p38 MAPK Signaling

The p38 Mitogen Activated Protein Kinase (MAPK) is a key member of a highly conserved superfamily of protein kinases. Mostly sensitive to inflammatory cytokines and stress signals, p38 activation leads to a variety of cellular responses, including proinflammatory cytokines secretion, cell survival, apoptosis induction and the regulation of cell proliferation. Here, we carried a focused investigation on the recent literature, across all systems and organisms, concerning the roles of p38 in the regulation of Stem Cell (SC) proliferation. Interestingly, we found that p38 can exert either a pro- or an anti- proliferative function, depending on the specific models and contexts. It is also activated by a variety of upstream signals and regulates SC proliferation through a wide diversity of molecular effectors. Whereas p38 regulates SC proliferation mostly cell-autonomously, an example of a non-cell autonomous p38-dependent regulation of SC proliferation was recently uncovered. Altogether, bringing together these recent findings improve our comprehension of the effects p38 has on SC proliferation, while these mechanisms and effectors appear as being conserved through evolution.

Allopregnanolone pleiotropic action in neurons and astrocytes: calcium signaling as a unifying mechanism.

Allopregnanolone (Allo) is a neurosteroid with pleiotropic action in the brain that includes neurogenesis, oligogenesis, human and rodent neural stem cell regeneration, increased glucose metabolism, mitochondrial respiration and biogenesis, improved cognitive function, and reduction of both inflammation and Alzheimer's disease (AD) pathology. Because the breadth of Allo-induced responses requires activation of multiple systems of biology in the absence of an Allo-specific nuclear receptor, analyses were conducted in both neurons and astrocytes to identify unifying systems and signaling pathways. Mechanisms of Allo action were investigated in embryonic hippocampal neurons and astrocytes cultured in an Aging Model (AM) media. Cellular morphology, mitochondrial function, and transcriptomics were investigated followed by mechanistic pathway analyses. In hippocampal neurons, Allo significantly increased neurite outgrowth and synaptic protein expression, which were paralleled by upregulated synaptogenesis and long-term potentiation gene expression profiles. Mechanistically, Allo induced Ca2+/CREB signaling cascades. In parallel, Allo significantly increased maximal mitochondrial respiration, mitochondrial membrane potential, and Complex IV activity while reducing oxidative stress, which required both the GABAA and L-type Ca2+ channels. In astrocytes, Allo increased ATP generation, mitochondrial function and dynamics while reducing oxidative stress, inflammasome indicators, and apoptotic signaling. Mechanistically, Allo regulation of astrocytic mitochondrial function required both the GABAA and L-type Ca2+ channels. Furthermore, Allo activated NRF1-TFAM signaling and increased the DRP1/OPA1 protein ratio, which led to increased mitochondrial biogenesis and dynamics. Collectively, the cellular, mitochondrial, transcriptional, and pharmacological profiles provide evidence in support of calcium signaling as a unifying mechanism for Allo pleiotropic actions in the brain.

Host-Microbe Interactions Regulate Intestinal Stem Cells and Tissue Turnover in Drosophila.

With the activity of intestinal stem cells and continuous turnover, the gut epithelium is one of the most dynamic tissues in animals. Due to its simple yet conserved tissue structure and enteric cell composition as well as advanced genetic and histologic techniques, Drosophila serves as a valuable model system for investigating the regulation of intestinal stem cells. The Drosophila gut epithelium is in constant contact with indigenous microbiota and encounters externally introduced "non-self" substances, including foodborne pathogens. Therefore, in addition to its role in digestion and nutrient absorption, another essential function of the gut epithelium is to control the expansion of microbes while maintaining its structural integrity, necessitating a tissue turnover process involving intestinal stem cell activity. As a result, the microbiome and pathogens serve as important factors in regulating intestinal tissue turnover. In this manuscript, I discuss crucial discoveries revealing the interaction between gut microbes and the host's innate immune system, closely associated with the regulation of intestinal stem cell proliferation and differentiation, ultimately contributing to epithelial homeostasis.

Role of SIRT3 in bone homeostasis and its application in preventing and treating bone diseases.

Bone homeostasis refers to the balance between osteoblast-mediated bone formation and osteoclast-mediated bone resorption and the maintenance of stable bone mass. SIRT3 is a class of mitochondrial protein deacetylase that influences various mitochondrial functions and is involved in the mechanisms underlying resistance to aging; regulation of bone marrow mesenchymal stem cells, osteoblasts, and osteoclasts; and development of osteoporosis, osteoarthritis, and other bone diseases. Moreover, exercise affects bones through SIRT3. Thus, studies on SIRT3 may provide insights for the treatment of bone diseases. Although SIRT3 can exert multiple effects on bone, the specific mechanism by which it regulates bone homeostasis remains unclear. By evaluating the relevant literature, this review discusses the structure and function of SIRT3, reveals the role and associated mechanisms of SIRT3 in regulating bone homeostasis and mediating bone health during exercise, and highlights the potential pharmacological value of SIRT3 in treating bone diseases.

Regulation of cancer stem cells and immunotherapy of glioblastoma (Review).

Glioblastoma (GB) is one of the most adverse diagnoses in oncology. Complex current treatment results in a median survival of 15 months. Resistance to treatment is associated with the presence of cancer stem cells (CSCs). The present review aimed to analyze the mechanisms of CSC plasticity, showing the particular role of β-catenin in regulating vital functions of CSCs, and to describe the molecular mechanisms of Wnt-independent increase of β-catenin levels, which is influenced by the local microenvironment of CSCs. The present review also analyzed the reasons for the low effectiveness of using medication in the regulation of CSCs, and proposed the development of immunotherapy scenarios with tumor cell vaccines, containing heterogenous cancer cells able of producing a multidirectional antineoplastic immune response. Additionally, the possibility of managing lymphopenia by transplanting hematopoietic stem cells from a healthy sibling and using clofazimine or other repurposed drugs that reduce β-catenin concentration in CSCs was discussed in the present study.

The mechanism of intestinal stem cells differentiation after ischemia-reperfusion injury in a rat model.

Notch and Wnt/β-catenin signaling are responsible for regulation of intestinal stem cells (ISCs) proliferation and differentiation. The purpose of the study was to evaluate Wnt/β-catenin and Notch signaling roles in regulation of ISC differentiation following ischemia-reperfusion (IR) injury in a rat. Rats were assigned into two groups: Sham rats underwent laparotomy without vascular intervention and IR rats underwent occlusion of SMA and portal vein for 20min followed by 48h of reperfusion. Wnt/β-catenin and Notch-related gene expression were determined using Real-Time PCR. Enterocyte proliferation, differentiation and Wnt-related proteins were determined by immunohistochemistry. IR rats demonstrated a significant decrease in β-catenin gene expression, a decrease in cyclin D1 and β-catenin positive cells in jejunum and ileum compared to Sham rats. IR rats demonstrated a significant increase in Notch-related gene expression in jejunum and ileum compared to Sham rats. The number of secretory cells was higher mainly in the jejunum and number of absorptive cells was significantly lower in jejunum and lower in ileum in IR rats compared to Sham rats. Intestinal stem-cell differentiation is toward secretory cells 48h after IR injury; however, Wnt/β-catenin pathway inhibition and Notch-related gene expression stimulation suggest crosstalk between pathways.

Immune privileges as a result of mutual regulation of the immune and stem systems

The immune privileges of cancer stem cells are a well-known and widely studied problem, as the presence of such cells in tumors is associated with refractoriness, recurrence, and metastasis. Accumulating evidence also suggests the presence of immune privileges for non-pathologic stem cells in addition to their other defense mechanisms against damaging factors. This similarity between pathologic and normal stem cells raises the question of why stem cells have such a potentially damaging property. The regulation of vital processes of autoimmunity control and regeneration by interactions between immune cells, stem cells and their microenvironment are reviewed as determinants of stem cell immune privilege formation. A deep mutual integration in the regulation of stem and immune cells is noted. Based on the diversity and complexity of the mutual regulation of stem cells, their microenvironment and the immune system, I propose to use the term “stem system”.

Distinct roles of hematopoietic cytokines in the regulation of leukemia stem cells in murine MLL-AF9 leukemia

Lymphoid-primed multipotent progenitor (LMPP)-like and granulocyte-monocyte progenitor (GMP)-like leukemia stem cells (LSCs) co-exist in the blood of most patients with acute myeloid leukemia (AML). Complete elimination of both types of LSCs is required to cure AML. Using an MLL-AF9-induced murine AML model, we studied the role of hematopoietic cytokines in the survival of LMPP- and GMP-like LSCs. We found that SCF or FLT3L promotes the survival of LMPP-like LSCs by stimulating Stat5-mediated Mcl1 expression, whereas interleukin-3 (IL-3) or IL-6 induces the survival of GMP-like LSCs by stimulating Stat3/nuclear factor κB (NF-κB)-mediated Bcl2 expression. Functional study demonstrated that, compared to AML cells cultured in IL-3 and IL-6 medium, AML cells in SCF- or Flt3L-only culture are highly clonogenic in invitro culture and are highly leukemogenic invivo. Our study suggests that co-inhibition of both STAT5-MCL1 and STAT3/NF-κB-BCL2 signaling might represent an improved treatment strategy against AML, specifically AML cases with a monocytic phenotype and/or FLT3 mutations.

Mind the Viscous Modulus: The Mechanotransductive Response to the Viscous Nature of Isoelastic Matrices Regulates Stem Cell Chondrogenesis.

The design of hydrogels as mimetics of tissues' matrices typically disregards the viscous nature of native tissues and focuses only on their elastic properties. In the case of stem cell chondrogenesis, this has led to contradictory results, likely due to unreported changes in the matrices' viscous modulus. Here, by employing isoelastic matrices with Young's modulus of ≈12kPa, variations in viscous properties alone (i.e., loss tangent between 0.1 and 0.25) are demonstrated to be sufficient to drive efficient growth factor-free chondrogenesis of human mesenchymal stem cells, both in 2D and 3D cultures. The increase of the viscous component of RGD-functionalized polyacrylamide or polyethylene glycol maleimide hydrogels promotes a phenotype with reduced adhesion, alters mechanosensitive signaling, and boosts cell-cell contacts. In turn, this upregulates the chondrogenic transcription factor SOX9 and supports neocartilage formation, demonstrating that the mechanotransductive response to the viscous nature of the matrix can be harnessed to direct cell fate.

PAF1 a Common Self-renewal Regulator of Embryonic and Cancer Stem Cells

PAF1 a Common Self-renewal Regulator of Embryonic and Cancer Stem Cells

Immune Cell Regeneration and Gaining Strength to Attack Multiple Myeloma Cancer

Multiple Myeloma is a rare cancer that primarily affects plasma cells that differentiate into white blood cells (which have important roles in the immune system such as fighting off infections and diseases). Once these plasma cells are transformed into cancerous cells that invade the space of the bone marrow, there is a prevention of the existence of future healthy immune cells that help the human body systems. Due to these viscous effects of the Multiple Myeloma, the blood cell count decreases and patients’ immunity lowers. The weakened immune systems of patients can attack the components of the treatments which leads to the wastage of money, time, and energy of the patients and the medical professionals. With the current research, patients can gain back strength and improve their immune systems. By the usage of the regeneration of stem cells, immunotherapy to increase the resistance of immune cells against the cancer, Chimeric Antigen Receptor T Cell therapy (or CAR-T Cell Therapy), and monoclonal antibody therapies, patients can gain back strength and improve their immune systems in a way that attacks Multiple Myeloma. Yet there is still growth for improvement, since the process of patients receiving treatments must be repeated multiple times due to the intensity and persistence of this cancer. By studying and researching the effects of Multiple Myeloma on immune cells, this paper’s goal is to find ways to improve current treatments and how to regenerate stronger and healthier immune cells which can resist and potentially defeat Multiple Myeloma Cancer.

Therapeutic of Mesenchymal Stem Cells in Diabetes

This study aims to explain and analyze the effective treatment of diabetes mellitus by Mesenchymal Stem Cells with respect to traditional therapy based on oral anti-diabetic drugs. As well as to study its regularly consistent properties. The method used in this research is the case study method. Patient care records were the main source of data for this study including evaluation and analysis. So that the results of the study explain that at the same time the utilized products cultured by "CELLTECH STEM CELL LAB. and BANKING" as an integrated part of the Clinic, also get quality assurance. Effectiveness and efficiency including endogenous beta cell regeneration of pancreatic islets, and other therapeutic issues are novel and belong to cutting-edge treatment. Treatments performed at the Vinski Regenerative Stem Cell Main Clinic have shown positive research results. This has strengthened the level of trust in the clinic's medical services.

Polycom Group Protein Mel18 Inhibits Hematopoietic Stem Cell Self-Renewal through Repressing the Expression of Genes Important for Cellular Senescence and Proliferation

Polycomb group (PcG) proteins are epigenetic gene silencers that have been implicated in stem cell maintenance and cancer development. Genetic and biochemical studies indicate that Polycomb group proteins exist in at least two protein complexes, Polycomb repressive complex 2 (PRC2) and Polycomb repressive complex 1 (PRC1), that act in concert to initiate and maintain stable gene repression. Bmi1 and Mel18 are two major homologs of the PCGF subunit within the canonical PRC1 complex. While Bmi1 is a positive regulator of hematopoietic stem cell (HSC) and leukemia stem cell (LSC) self-renewal, the role of Mel18 in normal and malignant hematopoiesis is not fully understood. To further determine the role of Mel18 in hematopoiesis, we have generated Mel18 conditional knockout mice ( Mel18 f/f-Mx1Cre +). Acute deletion of Mel18 in the hematopoietic compartment did not affect the frequency and survival of hematopoietic stem and progenitor cells (HSPCs). To determine the impact of Mel18 deficiency on HSPC proliferation, we performed serial replating assays and found that loss of Mel18 increases the replating potential of HSPCs in vitro. To determine the role of Mel18 in HSC self-renewal, we first performed serial bone marrow (BM) transplantation assays and found that Mel18 null BM cells show increased engraftment in both primary and secondary transplantation assays. Limiting dilution transplantation showed that there were more functional HSCs in the BM of Mel18 knockout mice. We then performed HSC transplantation assays and found that loss Mel18 enhances HSC self-renewal. Thus, we demonstrated that Mel18 inhibits HSC self-renewal and decreases the number of functional HSCs in vivo. To understand the mechanism by which Mel18 inhibits HSC self-renewal and proliferation, we performed RNA-seq using Mel18 +/+ and Mel18 -/- HSCs. We then performed gene set enrichment analysis (GSEA) to identify potential Mel18 target genes important for HSC behavior. We found that HSC gene signatures are enriched in Mel18 -/- HSCs. We also found that leukemia stem cell gene signatures and CBF-mutant-AML genes are significantly enriched in Mel18 -/- HSCs, suggesting that loss of Mel18 may prime normal HSCs for leukemic transformation. Notably, our RNA-seq assay showed that senescence-related genes, including S100a9 and S100a8, were down-regulated in Mel18 -/- HSPCs. Importantly, loss of Mel18 significantly decreased the number of senescent HSPCs. To understand how Mel18 regulates gene transcription, we performed ATAC-seq assays in Mel18 +/+ and Mel18 -/- HSPCs. Mel18 null HSPCs showed decreased chromatin accessibility near both S100a8 and S100a9 genes. We confirmed that both S100a8 and S100a9 are downregulated in Mel18 -/- HSPCs compared to Mel18 +/+ HSPCs. Given that cell cycle arrest is a common feature of cellular senescence, we also examined the expression of cell cycle related genes in Mel18 -/- HSPCs. The expression of cell cycle regulators, including CDK4 and CCND2, is increased in Mel18 -/- HSPCs. Mel18 can bind to a specific DNA sequence, 5‘-GACTNGACT-3‘, thereby repressing the expression of target genes. Mel18 directly binds to the CCND2 gene locus as revealed by ChIP-seq assays in embryonic stem cells, suggesting that Mel18 may directly repress CCND2 expression in HSPCs. In summary, we demonstrate that Mel18 inhibits HSC self-renewal via repressing the expression of genes that are important for cellular senescence and proliferation.

The LSD1 Inhibitor Ory-1001 (ladademstat) in Combination with Menin Inhibitor SNDX-5613 (revumenib) Has Synergistic in Vitro Activity in KMT2A-Rearranged AML Models

Introduction: Menin is a scaffold protein that binds to gene promoters, enhancers and transcription factors, thereby influencing transcription. Leukemias with rearrangement of KMT2A known as mixed-lineage leukemia (MLL), as well as other abnormalities such as NPM1c, respond to Menin inhibition with promising early clinical results. MLL leukemias affect children and adults and are associated with high rates of resistance to conventional chemotherapy. In addition to these alterations, other leukemia subsets with similar transcriptional dependency could be targeted through Menin inhibition. LSD1 is an essential regulator of murine MLL-AF9 leukemia stem cells (LSCs) and maintains the LSC potential of MLL-AML cells through blocking of differentiation and apoptosis ( Harris et al, 2012). LSD1 acts at genomic loci that are bound by MLL-AF9 to maintain expression of the associated leukemogenic program, thus preventing differentiation and apoptosis. We investigated the pre-clinical efficacy of combined Menin and LSD1 inhibition in AML. PC4 and SF2 interacting protein 1 (PSIP1)/p75, also known as LEDGF, interacts with the N-terminus of MLL and Menin. This complex contributes to the association of MLL and MLL-fusion multiprotein complexes with the chromatin. Several studies have shown that both PSIP1 and Menin are required for efficient MLL-fusion-mediated leukemogenesis and for the expression of MLL-regulated genes. A proposed model of LSD1 interaction with KMT2A-Menin-PSIP1 suprcomplex is depicted in Fig. 1A. Methods: To assess in vitro LSD1 inhibitor (ORY-1001) and Menin inhibitor (SNDX) potency, cell lines were plated and treated with the respective doses of ORY-1001, SNDX, and the combination of these drugs in 96-well plates. We performed synergy assays using increasing concentrations of SNDX and 3µM of ORY-1001. Cell viability and IC50 were measured after 4 days of treatment. The SynergyFinder 2.0 web application was used to calculate synergy scores and generate 2D synergy maps. Western blotting of LSD1, co-immunoprecipitation, and ChIP-qPCR studies were used to assess the depletion of the LSD1 from chromatin. The interaction network of LSD1 interactome proteins was assessed by mass spectrometry. Results: We showed that LSD1 degrades with increasing concentrations of ORY-1001 after 48h of treatment (Fig.1 B). IC 50 curves for ORY-1001, SNDX and synergy effect is shown with increasing concentrations of SNDX and 3µM ORY-1001 in MV4-11 cells (expressing MLL-AF4) (Fig.1 C). ORY-1001 alone shows an IC 50 of ~0.5 µM, and SNDX alone shows an IC 50 of ~10 nM. The combination of LSD1 and SNDX inhibitors synergistically reduced cell viability in MLL-rearranged MOLM-13 and MV4-11 cells (Fig.1 C). We also assayed the LSD1 endogenous protein interactome in MOLM-13 cells (expressing MLL-AF9) by mass spectrometry. Our LSD1 interactome analysis revealed an association of LSD1 with previously identified interactors such as RCOR1, HDAC1, HDAC2, CARM1 and HMG20B. We discovered an unexpected association between LSD1 and PSIP1. Interestingly, PSIP1 interact with MLL1-MEN1 complex and its crucial for assembling MLL-fusion proteins to their gene targets. Conclusion: LSD1 Inhibitor ladademstat in Combination with Menin Inhibitor revumenib has synergistic effect in KMT2A-Rearranged AML models. Surprisingly, we also found that PSIP1, which is essential for inducing MLL-rearranged leukemia, interacts with LSD1. This suggests that PSIP1 may modulate gene expression through its ability to interact with both MLL1 and LSD1. This study opens a new avenue to investigate the role of KMT2A-MEN1 in relation to other epigenetic modifiers, such as LSD1, and guide new potential combination therapies that may further potentiate differentiation and elimination of leukemia LSCs.

Deciphering the Differential Impact of Thrombopoietin/MPL Signaling on Hematopoietic Stem Cell Function in Bone Marrow and Spleen

Introduction Thrombopoietin (TPO) and its receptor MPL are key regulators of hematopoietic stem cell (HSC) function. In humans, ablation of either TPO or MPL leads to bone marrow failure in the early years of life. In contrast, mice lacking TPO or MPL display marked thrombocytopenia and severely reduced HSC numbers but manage to maintain normal survival up to one year of age. In this study, we investigated whether these mice eventually develop marrow failure with longer follow up, with a specific focus on understanding the effects of TPO/MPL signaling on HSC regulation in different hematopoietic niches. Methods Marrow and spleen hematopoiesis in the TPO -/- mice and MPL -/- mice were examined during a 2-yr follow-up. Results At 2 yrs of age, TPO -/- and MPL -/- mice exhibited not only a severely decreased platelet count (TPO -/- vs. MPL -/- vs. control: 38 vs. 107 vs. 1255 x 10 9/L) but also a moderate reduction in neutrophil count compared to age-matched wild-type controls (TPO -/- vs. MPL -/- vs. control: 1.1 vs. 1.7vs.3.3 x 10 3/ul). Femur marrow cell count or spleen cell count was not significantly different between TPO -/- or MPL -/- mice and control mice. There was no significant difference in overall survival between the TPO -/- or MPL -/- mice and wild-type controls. Marrow Lin -cKit +Sca1 +CD150 +CD48 - HSC cell numbers were significantly decreased in young (4-6mo) TPO -/- and MPL -/- mice (~4-fold) compared to age-matched control mice. This HSC loss was further worsened in old (~24mo) TPO -/- (25-fold) and MPL -/- (12-fold) mice compared to control mice. In contrast, there was no significant difference in splenic HSC cell numbers between young TPO -/- or MPL -/- mice and control mice, with only a moderate decrease of HSCs in old TPO -/- and MPL -/- (~4-fold) mice compared to control mice. I n vivo using BrdU labeling revealed an increased proliferation of marrow HSCs in TPO -/- and MPL -/- mice compared to age-matched control mice. In contrast, there was no significant increase in splenic HSC proliferation in TPO -/- or MPL -/- mice. These findings suggest that TPO/MPL signaling has distinct effects on both the number and the function of HSCs in the marrow and spleen. Competitive repopulation assays revealed that HSCs from 2yr old TPO -/- or MPL -/- marrow failed to engraft the recipient mice at 12 weeks post-transplantation. We also conducted competitive repopulation assays in which spleen cells from old TPO -/- mice (CD45.2) were injected together with spleen cells from 2yr old wild-type competitor mice (CD45.1) into lethally irradiated wild-type recipients (CD45.1). At 12 days after transplantation, the TPO -/- spleen donor showed comparable engraftment in the Lin -cKit +Sca1 + stem/progenitor cell compartment, similar to the wild-type competitor spleen donor, suggesting that at least the short-term engraftment capacity of the TPO -/- spleen donor was not impaired. We are currently conducting further investigation to assess the long-term repopulation capacity of TPO -/- spleen HSCs. To investigate how TPO/MPL ablation impacts the hematopoietic niche in the marrow and spleen, we used quantitative flow cytometry and/or fluorescence imaging to examine these niche cells and their expression of niche factors. First, while there was no significant difference in the marrow vascular area (measured by in vivo VE-cadherin labeling) between TPO -/- or MPL -/- mice and control mice, we observed a significant increase in the splenic vascular area in TPO -/- and MPL -/- mice. Second, CXCL12 levels were significantly decreased in marrow endothelial cells (ECs; CD45 -CD31 +) from TPO -/- and MPL -/- mice compared to control mice; in contrast, CXCL12 levels were significantly increased in splenic ECs from TPO -/- and MPL -/- mice. Similar results were also observed with marrow and splenic perivascular stromal cells (CD45 -CD31 -Ter119 -Sca1 -). Therefore, TPO/MPL ablation enhanced both the quantity and quality of the splenic hematopoietic niche, which could contribute to the maintenance of splenic hematopoiesis in TPO -/- and MPL -/- mice. Last, consistent with findings from the TPO/MPL ablated mice, exogenous TPO treatment increased marrow HSC proliferation but did not alter splenic HSC proliferation. In addition, TPO treatment increased CXCL12 expression in marrow ECs, but decreased CXCL12 levels in splenic ECs. Taken together, our study revealed the complexity of TPO/MPL signaling in HSC regulation which differs between the marrow and spleen

Targeting Endothelial PERK-DLL4 Axis to Enhance Hematopoietic Stem Cell and Lymphoid Progenitor Regeneration

Despite our current information on hundreds of genes implicated in hematopoietic stem cells (HSC) biology, major gaps still exist in our understanding of HSC physiology and their regeneration mechanism under disease and stress conditions, hindering the development of strategies to enhance HSC regeneration. HSCs reside in a tightly controlled bone marrow HSC niche that regulates and maintains hematopoietic homeostasis. HSC niche is composed of a heterogeneous population of specialized stroma cells. Among these, the endothelial cells and the perivascular stroma cells provide critical pro-hematopoietic factors to support HSC pool and committed lineage progenitors particularly lymphoid progenitors. In addition, evidence suggests that endothelium-expressing Notch ligand may promote Notch-dependent long-term HSC proliferation. Recent single cell transcriptome profiling revealed that Delta-like 4 (DLL4) is expressed by the vascular endothelial cells at a higher level than other Notch ligands while its expression down-regulates in response to myeloablation. However, the molecular mechanism regulating the dynamic DLL4 expression during hematopoietic regeneration in response to myeloablative stimuli is unknown. Unfolded protein response (UPR) (also known as endoplasmic reticulum stress, ER stress) has been increasingly recognized as crucial for HSC maintenance. During hematopoietic regeneration, UPR is critical for meeting the increased demand for protein synthesis required for rapid cellular proliferation and for adapting to pathological conditions such as increased levels of reactive oxygen species following irradiation or chemotherapy. Unlike HSCs, how HSC niche endothelial cells deal with redox imbalance and higher requirements of vessel regeneration to regulate stress hematopoietic regeneration is not clear. In this study, using genetically engineered mouse models, we examined the dynamic expression and function of PERK, one arm of UPR, and its potential downstream target, Dll4, during the stress hematopoiesis triggered by transplantation. We found that PERK is activated in the mouse bone marrow endothelial cells following irradiation. Ablation of endothelial Perk in mice (referred as Perk iΔEC mice) promoted post-transplantation immediate recovery of blood cells derived from wild type donor mice and enhanced HSC and lymphoid progenitor regeneration with the expansion of B lineage progenitors including pre-proB, pro-B, and pre-B cells. In addition, by whole-mount immunostaining and intravital microscopy, we found that endothelial Perk deletion restrained disorganized angiogenesis and vascular leakage provoked by irradiation. Although irradiation itself had no effect on endothelial Dll4, Perk ablation markedly increased endothelial Dll4 expression. We found that Dll4 is essential for the post-irradiation HSC and lymphoid progenitor rejuvenation as ablation of endothelial Dll4 in mice (referred as Dll4 iΔEC mice) led to impaired short-term and long-term hematopoietic regeneration and markedly decreased wild type donor derived peripheral B cell and lymphoid progenitor recovery. We found that endothelial Dll4 is required for HSC quiescence maintenance, HSC niche retention and HSC self-renewal. Further, ablation of endothelial Dll4 led to disorganized bone marrow angiogenesis and increased marrow vascular leakage. Finally, we showed that deletion of Dll4 in Perk iΔEC mice completely abrogated the increased numbers of HSC and lymphoid progenitors mediated by Perk deletion, suggesting that up-regulation of endothelial Dll4 is likely responsible for Perk ablation-mediated enhanced HSC and lymphoid progenitor regeneration. Taken together, our results revealed a novel regulatory mechanism by which endothelial Dll4 expression is suppressed by PERK of UPR during transplantation while ablating Perk increases Dll4 expression. Endothelial DLL4 in the bone marrow endothelial cells is critical for supporting vascular integrity and myeloablation-induced HSC and lymphoid progenitor regeneration. Our findings suggest that targeting endothelial PERK-DLL4 axis could be a viable option to improve post-irradiation regeneration of HSC and lymphoid progenitors.

The Roles of WNT Signaling Pathways in Skin Development and Mechanical-Stretch-Induced Skin Regeneration.

The WNT signaling pathway plays a critical role in a variety of biological processes, including development, adult tissue homeostasis maintenance, and stem cell regulation. Variations in skin conditions can influence the expression of the WNT signaling pathway. In light of the above, a deeper understanding of the specific mechanisms of the WNT signaling pathway in different physiological and pathological states of the skin holds the potential to significantly advance clinical treatments of skin-related diseases. In this review, we present a comprehensive analysis of the molecular and cellular mechanisms of the WNT signaling pathway in skin development, wound healing, and mechanical stretching. Our review sheds new light on the crucial role of the WNT signaling pathway in the regulation of skin physiology and pathology.

The Epigenetic Regulation of Quiescent in Stem Cells.

This review article discusses the epigenetic regulation of quiescent stem cells. Quiescent stem cells are a rare population of stem cells that remain in a state of cell cycle arrest until activated to proliferate and differentiate. The molecular signature of quiescent stem cells is characterized by unique epigenetic modifications, including histone modifications and deoxyribonucleic acid (DNA) methylation. These modifications play critical roles in regulating stem cell behavior, including maintenance of quiescence, proliferation, and differentiation. The article specifically focuses on the role of histone modifications and DNA methylation in quiescent stem cells, and how these modifications can be dynamically regulated by environmental cues. The future perspectives of quiescent stem cell research are also discussed, including their potential for tissue repair and regeneration, their role in aging and age-related diseases, and their implications for cancer research. Overall, this review provides a comprehensive overview of the epigenetic regulation of quiescent stem cells and highlights the potential of this research for the development of new therapies in regenerative medicine, aging research, and cancer biology.