**2. Physiological activity of MSC**

Postnatal stem and progenitor cells are responsible for tissue renewal and regeneration throughout the whole human lifetime. Accumulating evidence indicates that stem cells function only within a specific niche. Stem cell niche is considered as a local tissue microenvironment that maintains stem cells and regulates their function by producing factors that act directly on stem cells [1]. This microenvironment provides structural cues and paracrine signals to support stemness. Mesenchymal stromal cells (MSCs) are found in many niches of tissue-specific resident stem cells. The function of MSC in a niche is often critical for its maintenance. MSC plays an important role in coupling information from the environment with stem cell populations. MSCs react to endocrine and nervous system signals: thus, the stimulation of a specific type of MSC by granulocyte colony-stimulating factor (G-CSF) or beta-3-adrenoreceptor agonists decreases the expression of the panel of genes supporting hematopoietic stem cells (HSCs). Furthermore, upon MSC, withdrawal in niche HSC activity and quantity decrease [2]. MSCs also interact with stem cells and other internal components of the niche for effective control over HSC, ensuring they support hematopoiesis without inducing aberrant proliferation. It should be noted that the maintenance of stem cells in the niche is not the only function of MSC. These cells are also involved in the formation and maintenance of a structural component of the niche. MSC isolated from a subendothelial layer of bone marrow stroma can form a microenvironment alike a miniature bone organ, similar to the HSC niche, under heterotopic transplantation. The establishment of subendothelial stromal cells in developing heterotopic BM in vivo occurs via specific, dynamic interactions with developing sinusoids. Subendothelial stromal cells are major producers of angiopoietin-1 (Angpt-1), the principal factor of HSC niche involved in vascular remodeling [3].

MSCs provide their supporting functions through secretory activity in physiological conditions. MSCs secrete a number of factors that are critical for the maintenance of stem cells in their niches [4, 5]. Consistent with other reports, we have shown that the largest functional cluster in MSC secretome is composed of extracellular matrix (ECM) proteins [6]. Such protein profile is in line with the stromal characteristics of adipose MSC. Adequate production of ECM components is necessary for tissue homeostasis and regeneration, because these molecules not only provide a scaffold for cells and soluble molecules but also regulate angiogenesis, neurogenesis, and inflammation. In addition, a large amount of data indicates that apart from soluble factors MSCs secrete regulatory non-coding RNA (e.g., micro RNA) within extracellular vesicles (EVs). The release of these small RNA by MSC can play a role in stem cell niche maintenance by controlling and tuning proliferation, differentiation, and homing. Particularly, microRNA regulates diverse biological processes, including growth and differentiation of stem cells [7, 8].

a pivotal tool in regenerative medicine. It is important to highlight mesenchymal stromal cells (MSCs) as the most popular source for cell therapy and tissue engineering. However, a large amount of experimental data indicate that MSC effects on regeneration are mostly mediated by their ability to produce a wide range of bioactive molecules, and the use of MSCconditioned medium (CM), a complex of the factors secreted to cell culture growth medium, as a distinct biopharmaceutical drug can be a rational alternative to direct MSC therapy. MSC CM could be considered as "сell-free therapeutics" since this product is devoid of MSC themselves in its final formulation, yet possess significant therapeutic potency. It contains

components secreted by MSC and reproduced the effects of MSC-based cell therapy.

quality control of this promising class of biopharmaceuticals.

**2. Physiological activity of MSC**

48 Biopharmaceuticals

"Cell-free therapeutics" have various advantages in overcoming the limitations and risks associated with the cell-based therapy. Despite the outstanding preclinical and clinical efficacy of MSC CM, there is still no generally accepted regulating approaches for CM standardization and quality control. This chapter reviews the current state of art in the development of MSC CM-based medicinal products and describes the crucial issues concerning the production and

Postnatal stem and progenitor cells are responsible for tissue renewal and regeneration throughout the whole human lifetime. Accumulating evidence indicates that stem cells function only within a specific niche. Stem cell niche is considered as a local tissue microenvironment that maintains stem cells and regulates their function by producing factors that act directly on stem cells [1]. This microenvironment provides structural cues and paracrine signals to support stemness. Mesenchymal stromal cells (MSCs) are found in many niches of tissue-specific resident stem cells. The function of MSC in a niche is often critical for its maintenance. MSC plays an important role in coupling information from the environment with stem cell populations. MSCs react to endocrine and nervous system signals: thus, the stimulation of a specific type of MSC by granulocyte colony-stimulating factor (G-CSF) or beta-3-adrenoreceptor agonists decreases the expression of the panel of genes supporting hematopoietic stem cells (HSCs). Furthermore, upon MSC, withdrawal in niche HSC activity and quantity decrease [2]. MSCs also interact with stem cells and other internal components of the niche for effective control over HSC, ensuring they support hematopoiesis without inducing aberrant proliferation. It should be noted that the maintenance of stem cells in the niche is not the only function of MSC. These cells are also involved in the formation and maintenance of a structural component of the niche. MSC isolated from a subendothelial layer of bone marrow stroma can form a microenvironment alike a miniature bone organ, similar to the HSC niche, under heterotopic transplantation. The establishment of subendothelial stromal cells in developing heterotopic BM in vivo occurs via specific, dynamic interactions with developing sinusoids. Subendothelial stromal cells are major producers of angiopoietin-1 (Angpt-1), the principal factor of HSC niche involved in vascular remodeling [3]. MSCs provide their supporting functions through secretory activity in physiological conditions. MSCs secrete a number of factors that are critical for the maintenance of stem cells in their niches [4, 5]. Consistent with other reports, we have shown that the largest functional The function of adult stem cells includes the local or remote replacement of senescent or damaged cells along with maintaining their own pool. Stem cells supported by other niche components can participate in the repair of small lesions of a skin, liver, intestines, kidney, and bone marrow. However, stem cells could not cope with more serious injuries without more substantial support [9]. MSCs are important for maintaining the niche of stem cells; therefore, they can participate in stem cell potentiation to respond to damage, stimulate the survival of stem cells, and, thereby, maintain the structural and functional integrity of the niche. For example, in such serious damage as myocardium ischemia/reperfusion injury, MSC mediated its cardioprotective paracrine effect by secreting exosomes which reduced infarct size in a mouse model [10]. As in majority of animal models and clinical studies, only limited or no engraftment at all was often observed, one should consider paracrine MSC function as principal effector for tissue regeneration after, at least, systemic MSC injection for different injuries [11].
