Scalable in vitro production of defined mouse erythroblasts

Mouse embryonic stem cells (mESCs) can be manipulated in vitro to recapitulate the process of erythropoiesis, during which multipotent cells undergo lineage specification, differentiation and maturation to produce erythroid cells. Although useful for identifying specific progenitors and precursors, this system has not been fully exploited as a source of cells to analyse erythropoiesis. Here, we establish a protocol in which characterised erythroblasts can be isolated in a scalable manner from differentiated embryoid bodies (EBs). Using transcriptional and epigenetic analysis, we demonstrate that this system faithfully recapitulates normal primitive erythropoiesis and fully reproduces the effects of natural and engineered mutations seen in primary cells obtained from mouse models. We anticipate this system to be of great value in reducing the time and costs of generating and maintaining mouse lines in a number of research scenarios.

RAS mutation patterns arise from tissue-specific responses to distinct oncogenic signaling

Despite multiple possible oncogenic mutations in the proto-oncogene KRAS, unique subsets of these mutations are detected in different cancer types. As KRAS mutations occur early, if not being initiating, these mutational biases are ostensibly a product of how normal cells respond to the encoded oncoprotein. Oncogenic mutations can impact not only the level of active oncoprotein, but also engagement with effectors and other proteins. To separate these two effects, we generated four novel inducible Kras alleles encoded by the biochemically distinct mutations G12D versus Q61R encoded by native (nat) rare versus common (com) codons to produce either low or high protein levels. Each allele induced a distinct transcriptional response in normal cells. At one end of the spectrum, the KrasnatG12D allele induced transcriptional hallmarks suggestive of an expansion of multipotent cells, while at the other end, the KrascomQ61R allele exhibited all the hallmarks of oncogenic stress and inflammation. Further, this dramatic difference in the transcriptomes of normal cells appears to be a product of signaling differences due to increased protein expression as well as the specific mutation. To determine the impact of these distinct responses on RAS mutational patterning in vivo, all four alleles were globally activated, revealing that hematolymphopoietic lesions were sensitive to the level of active oncoprotein, squamous tumors were sensitive to the G12D mutant, while carcinomas were sensitive to both these features. Thus, we identify how specific KRAS mutations uniquely signal to promote the conversion of normal hematopoietic, epithelial, or squamous cells towards a tumorigenic state.

Stimulation of the Migration and Expansion of Adult Mouse Neural Stem Cells by the FPR2-Specific Peptide WKYMVm

Neural stem cells (NSCs) are multipotent cells capable of self-renewal and differentiation into different nervous system cells. Mouse NSCs (mNSCs) are useful tools for studying neurogenesis and the therapeutic applications of neurodegenerative diseases in mammals. Formyl peptide receptor 2 (FPR2), expressed in the central nervous system and brain, is involved in the migration and differentiation of murine embryonic-derived NSCs. In this study, we explored the effect of FPR2 activation in adult mNSCs using the synthetic peptide Trp-Lys-Tyr-Met-Val-D-Met-NH2 (WKYMVm), an agonist of FPR2. After isolation of NSCs from the subventricular zone of the adult mouse brain, they were cultured in two culture systems—neurospheres or adherent monolayers—to demonstrate the expression of NSC markers and phenotypes. Under different conditions, mNSCs differentiated into neurons and glial cells such as astrocytes, microglia, and oligodendrocytes. Treatment with WKYMVm stimulated the chemotactic migration of mNSCs. Moreover, WKYMVm-treated mNSCs were found to promote proliferation; this result was confirmed by the expansion of mNSCs in Matrigel and the increase in the number of Ki67-positive cells. Incubation of mNSCs with WKYMVm in a supplement-free medium enhanced the survival rate of the mNSCs. Together, these results suggest that WKYMVm-induced activation of FPR2 stimulates cellular responses in adult NSCs.

Comparative Transcriptomics Identifies Potential Stemness-Related Markers for Mesenchymal Stromal/Stem Cells

Mesenchymal stromal/stem cells (MSCs) are multipotent cells residing in multiple tissues with the capacity for self-renewal and differentiation into various cell types. These properties make them promising candidates for regenerative therapies. MSC identification is critical in yielding pure populations for successful therapeutic applications; however, the criteria for MSC identification proposed by the International Society for Cellular Therapy (ISCT) is inconsistent across different tissue sources. This study aimed to identify potential markers to be used together with the ISCT criteria to provide a more accurate means of MSC identification. Thus, we carried out a comparative analysis of the expression of human and mouse MSCs derived from multiple tissues to identify the common differentially expressed genes. We show that six members of the proteasome degradation system are similarly expressed across MSCs derived from bone marrow, adipose tissue, amnion, and umbilical cord. Also, with the help of predictive models, we found that these genes successfully identified MSCs across all the tissue sources tested. Moreover, using genetic interaction networks, we showed a possible link between these genes and antioxidant enzymes in the MSC antioxidant defense system, thereby pointing to their potential role in prolonging the life span of MSCs. Our results suggest these genes can be used as stemness-related markers.

Mesenchymal Stromal Cells: an Antimicrobial and Host-Directed Therapy for Complex Infectious Diseases

There is an urgent need for new antimicrobial strategies for treating complex infections and emerging pathogens. Human mesenchymal stromal cells (MSCs) are adult multipotent cells with antimicrobial properties, mediated through direct bactericidal activity and modulation of host innate and adaptive immune cells.

Fluctuation of cellular differentiation in limb regeneration is regulated by Pde4b in urodele amphibians

Urodele amphibians, Pleurodeles waltl and Ambystoma mexicanum, have organ level regeneration capability, such as limb regeneration. Multipotent cells are induced by an endogenous mechanism in amphibian limb regeneration. It is well known that dermal fibroblasts receive regenerative signals and turn into multipotent cells, called blastema cells. However, the induction mechanism of the blastema cells from matured dermal cells was unknown. We previously found that BMP2, FGF2, and FGF8 (B2FF) could play sufficient roles in blastema induction in urodele amphibians. Here, we show that B2 FF treatment can induce dermis derived cells that can participate in multiple cell lineage in limb regeneration. We first established a newt dermis derived cell line and confirmed that B2FF treatment on the newt cells provided plasticity in cellular differentiation in limb regeneration. Interspecies comparative analysis clarified that Pde4b upregulation by B2FF specifically took place in the newt cells. Blocking P DE4B signaling by Rolipram suppressed dermis to cartilage transformation and the mosaic knockout animals showed consistent results . Our results are a valuable insight into how dermal fibroblasts acquire multipotency during the early phase of limb regeneration via an endogenous program in amphibian limb regeneration.

SAFETY AND EFFICACY OF STEM CELL THERAPY FOR TREATMENT SEVERE TRAUMATIC BRAIN INJURY

Traumatic brain injury (TBI) is a complicated topic.TBI has expensive treatment, sequelae, high mortality and efficient therapeutic strategies for TBI are urgently needed. Patients with severe TBI account for 10% of all traumatic brain injury patients, but more than 80% of patients who die from traumatic brain injury are in this group of patients. That proves the danger of severe TBI. Mesenchymal stem cells (MSCs) are multipotent cells with high proliferative and self-renewal capacities, as well as immunomodulatory and neuroregenerative effects. MSCshave been shown to treat TBI and significantly reduce inflammation of injured tissue. In the present study, we evaluated the quality and effectiveness of mesenchymal stem cell transplantation with the graft dose of 9x106cells/kg. Clinical indexes (Pulse, temperature, SBP, HATTr, HATTB, GCS) and subclinical (Hepatic index, kidney, cortisol, lactate, glucose, hematological index, respiratory index) were monitored on 60 patients within 6 months. The results showed that the recovery journey is different for every person with the graft dose of 6-9x106 cells/kg, but there were no adverse complications for the patients, the clinical and paraclinical indicators were stable. No significant side effects were observed during the trial as well as after 6 months of treatment. The MSC transfusions significantly increased the life quality patients; reduced brain damage and increased HSC counts. Inflammatory cytokines (IL-6) levels decreased and GCS increased in the MSC transfusion group, indicating that the patient's health improved.

Autophagy, Mesenchymal Stem Cell Differentiation, and Secretion

Mesenchymal stem cells (MSC) are multipotent cells capable to differentiate into adipogenic, osteogenic, and chondrogenic directions, possessing immunomodulatory activity and a capability to stimulate angiogenesis. A scope of these features and capabilities makes MSC a significant factor of tissue homeostasis and repair. Among factors determining the fate of MSC, a prominent place belongs to autophagy, which is activated under different conditions including cell starvation, inflammation, oxidative stress, and some others. In addition to supporting cell homeostasis by elimination of protein aggregates, and non-functional and damaged proteins, autophagy is a necessary factor of change in cell phenotype on the process of cell differentiation. In present review, some mechanisms providing participation of autophagy in cell differentiation are discussed

Roles of MicroRNAs in Osteogenesis or Adipogenesis Differentiation of Bone Marrow Stromal Progenitor Cells

Bone marrow stromal cells (BMSCs) are multipotent cells which can differentiate into chondrocytes, osteoblasts, and fat cells. Under pathological stress, reduced bone formation in favour of fat formation in the bone marrow has been observed through a switch in the differentiation of BMSCs. The bone/fat switch causes bone growth defects and disordered bone metabolism in bone marrow, for which the mechanisms remain unclear, and treatments are lacking. Studies suggest that small non-coding RNAs (microRNAs) could participate in regulating BMSC differentiation by disrupting the post-transcription of target genes, leading to bone/fat formation changes. This review presents an emerging concept of microRNA regulation in the bone/fat formation switch in bone marrow, the evidence for which is assembled mainly from in vivo and in vitro human or animal models. Characterization of changes to microRNAs reveals novel networks that mediate signalling and factors in regulating bone/fat switch and homeostasis. Recent advances in our understanding of microRNAs in their control in BMSC differentiation have provided valuable insights into underlying mechanisms and may have significant potential in development of new therapeutics.

Pericytes for Therapeutic Approaches to Ischemic Stroke

Pericytes are perivascular multipotent cells located on capillaries. Although pericytes are discovered in the nineteenth century, recent studies have found that pericytes play an important role in maintaining the blood—brain barrier (BBB) and regulating the neurovascular system. In the neurovascular unit, pericytes perform their functions by coordinating the crosstalk between endothelial, glial, and neuronal cells. Dysfunction of pericytes can lead to a variety of diseases, including stroke and other neurological disorders. Recent studies have suggested that pericytes can serve as a therapeutic target in ischemic stroke. In this review, we first summarize the biology and functions of pericytes in the central nervous system. Then, we focus on the role of dysfunctional pericytes in the pathogenesis of ischemic stroke. Finally, we discuss new therapies for ischemic stroke based on targeting pericytes.