Notes

Fresh interferon-sensitive genetics introduced through correlation-driven gene variety and also programs the field of biology.
Spermatogonial stem cells (SSCs) are the germ cells at the basis of spermatogenesis in adult mammals. SSCs offer many biotechnological possibilities and are fundamental cells in the study of spermatogenesis (Aponte, World J Stem Cells 7669-680, 2015). This chapter describes detailed procedures for SSC isolation, culture, cryopreservation, and characterization in bovine, murine, and human models.It has been shown that freshly isolated satellite cells from adult muscle constitute a stem cell-like population that exhibits more efficient engraftment and self-renewal activity in regenerating muscle than myoblast. Thus, purification of pure populations of quiescent satellite cells from adult skeletal muscle is highly necessary, not only for understanding the biology of satellite cells and myoblasts but also for improving cell-based therapies for muscle regeneration. This chapter describes a basic protocol used in our laboratory to isolate quiescent muscle satellite cells from adult skeletal muscle by enzymatic dissociation followed by a sequential magnetic-activated cell sorting (MACS). This method is cheap and fast providing and alternative procedure to other purification methods that require fluorescence-activated cell sorting (FACS) machines. Freshly isolated quiescent satellite cells purified by this method can be used in a broad range of experiments including cell transplantation for satellite cell self-renewal experiments or cell therapies.Tissue resident mesenchymal progenitor cells (MPC) are important regulators of tissue repair or regeneration, remodeling, inflammation, and angiogenesis. Here we describe a technology used to define, isolate, and characterize a population of resident lung MPC in both human and mouse explanted tissue. The definition of this population using a defined set of markers facilitates the repeatable isolation of a mesenchymal subpopulation population by flow cytometry and the subsequent translational study of this specific cell type and function.The superior laryngeal nerve (SLN) is known to play an essential role in the laryngeal reflex and swallowing. Damage to the SLN causes difficulty swallowing, that is, dysphagia. We successfully developed a novel rat model of dysphagia by SLN injury, in which we could evaluate the neuroregenerative capacity of stem cell from human exfoliated deciduous teeth (SHED). The dysphagic rats exhibit weight loss and altered drinking patterns. Furthermore, SLN injury induces a delayed onset of the swallowing reflex and accumulation of laryngeal debris in the pharynx. This rat model was used to evaluate the systemic application of SHED-conditioned medium (SHED-CM) as a therapeutic candidate for dysphagia. We found that SHED-CM promoted functional recovery and significant axonal regeneration in SLNs through the polarization shift of macrophages from activated inflammatory macrophages (M1) to anti-inflammatory macrophages (M2) and angiogenesis. This chapter describes the establishment of SLN-injury induced dysphagia rat model and the preparation and application of SHED-CM.Innervation plays a key role in the development, homeostasis, and regeneration of organs and tissues. However, the mechanisms underlying these phenomena are not well understood yet. In particular, the role of innervation in tooth development and regeneration is neglected. Cocultures constitute a valuable method to investigate and manipulate the interactions between nerve fibers and teeth in a controlled and isolated environment. Microfluidic systems for allow cocultures of neurons and different cell types in their appropriate culture media, while permitting the passage of axons from one compartment to the other. Here we describe how to isolate and coculture developing trigeminal ganglia and tooth germs in a microfluidic coculture system. This protocol describes a simple and flexible way to coculture ganglia/nerves and their target tissues and to study the roles of specific molecules on such interactions in a controlled and isolated environment.Among the adult stem cells, multipotent mesenchymal stem cells (MSCs) turned out to be a promising option for cell-based therapies for the treatment of various diseases including autoimmune and cardiovascular disorders. MSCs bear a high proliferation and differentiation capability and exert immunomodulatory functions while being still clinically safe. As tissue-resident stem cells, MSCs can be isolated from various tissue including peripheral or umbilical cord blood, placenta, blood, fetal liver, lung, adipose tissue, and blood vessels, although the most commonly used source for MSCs is the bone marrow. However, the proportion of MSCs in primary isolates from adult tissue biopsies is rather low, and therefore MSCs must be intensively expanded in vitro before the MSCs find particular use in therapies that may require extensive and repetitive cell replacement. Therefore, more easily accessible sources of MSCs are needed. Here, we present a detailed protocol to generate tissue-typical MSCs by direct linage conversion using transcription factors defining target MSC identity from murine induced pluripotent stem cells (iPSCs).Niches for tissue-resident mesenchymal stem cells (MSCs) have been identified in many adult tissues. In particular, MSCs residing in the vascular stem cell niche came into focus the so-called vascular wall-resident MSCs (VW-MSCs) were, based upon their anatomic location, (1) distributed throughout the adult organism, and (2) supposed to be the first line cells which could be addressed in response to a pathologic trigger acting on or in close vicinity to the vascular system. this website Like tissue-resident MSCs in general, VW-MSC contribute to organ integrity and harbor the capacity to suppress inflammation and promote repair during normal vessel homeostasis, although resident MSCs present in the healthy situation of an individual seems not to bear sufficient for protection or repair following injury. In contrast, injury affected MSCs could contribute to disease induction and progression. A detailed understanding of the molecular repertoire as well as of the signaling pathways controlling stem cell fate of VW-MSCs is prerequisite to understand how (1) endogenous VW-MSCs contribute to normal vessel homeostasis as well as diseases that include the vascular system, (2) a potential on-site manipulation of these cells directly within their endogenous niche could be used for therapeutically benefits, and (3) isolated and therapeutically applied VW-MSCs in terms of exogenous MSCs with superior repair capabilities might be logically more efficient to address vascular diseases than MSCs derived from other tissues.
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