Some regularities of the regeneration of the musculature of internal organs]

The paper summarizes the data of the author and his co-workers on regulation of the musculature of vessels, digestive tract, uterus, ureters and urinary bladder. The smooth muscle cells have different degrees of differentiation. The muscular tissue of the vessels is most differentiated. The uterus musculature is very plastical. After injury mature myocytes are the first to undergo destruction. The intermedial substance is more stable. Myoblasts, young elements of the fibroblast row and the subendothelial layer cells are the origin of muscular regeneration. Figures of mitosis and amitosis are noted. Mature myocytes and intercellular substance are formed in the process of differentiation of the regeneration. The content of RNP in the regeneration cells is high, but in the process of differentiation of its elements it becomes lower. The DNP level has inconsiderable fluctuations. In early experiments PAS-positive substances are revealed in greater degree than in later ones. The content of acid mucopolysaccharides decreases in the process of fibrillogenesis. In all internal organs under study the muscular tissue regenerated. The degree of differentiation, severity of the lesion and functional peculiarities of the organ determine the completeness of the tissue reparation. The musculature of the intestinal tract and vessels regenerates more completely. Mighty layers of connective tissue with de novo formed blood vessels are disposed among the bundles of the repaired muscular tissue of the uterus and urine bladder wall. Simultaneously a part of regeneration cells are destroyed. These are two sides of a single process of development.     

TCAV-1, a monoclonal antibody specific to epithelial pharyngeal cells in the planarian Dugesia (Girardia) tigrina. Application to pattern formation of the pharynx during regeneration

During regeneration in planarians, anterior (head and prepharyngeal) and posterior (postpharyngeal and tail) fragments rebuild one of the most peculiar structures of planarians: the pharynx and the pharynx cavity. Previous studies (see Brønsted, 1969, for a general review, and Asai, 1990, 1991, for anterior regeneration) have shown that within postpharyngeal pieces both structures appear in the old stump from clusters of undifferentiated cells. However, the lineage and differentiation of their elements (inner and outer epithelial cells, muscle layers, gland cells, nerve rings) and the overall pattern of growth and differentiation is not clear.     

Analysis of early processes in wound-induced vascular regeneration using TED3 and ZeHB3 as molecular markers

Interruption of the vascular bundles of Zinnia internodes induced transdifferentiation of cells into tracheary elements (TEs) or sieve elements (SEs) within 4 d of wounding. The early stage of the regeneration processes was analyzed using two molecular marker genes, TED3 and ZeHB3, which are expressed specifically in TE precursor cells and immature phloem cells, respectively. An increase in the numbers of TED3 and ZeHB3 mRNA-expressing cells always preceded an increase in the numbers of TEs and SEs formed. The earliest sign of vascular differentiation was the appearance 24 h after wounding of a layer(s) of TED3 mRNA-expressing cells in the inter- and intrafascicular cambial-like regions along the severed vascular bundles. In contrast, the number of ZeHB3 mRNA-expressing cells decreased dramatically along the severed bundles 24 h after wounding, and increased again 36 h after wounding. These results clearly indicate that xylem and phloem differentiation are not synchronized during vascular regeneration. Treatment with 10(-3) M colchicine abolished the expression of ZeHB3 mRNA in pith parenchyma, but not TED3 mRNA; this suggests that cell division is a prerequisite for the transdifferentiation of pith parenchymal cells into immature phloem cells expressing ZeHB3. In contrast, transdifferentiation of pith parenchymal cells to TE precursor cells does not require preceding cell division. However, the inhibition of cell division prevented the formation of both radial files of TEs and the cambial-like layer(s) of TED3 mRNA-expressing cells, and, ultimately, vascular regeneration altogether. These results imply that wound-induced cambial-like activity in and between severed vascular bundles is essential for vascular regeneration.     

The role of the hemopoiesis-inducing microenvironment in the mechanisms of the regeneration of hemopoiesis after cytostatic exposure]

On the model of adriamycine-induced hemopoiesis hypoplasia it has been shown that the accelerated differentiation of hemopoietic precursors regeneration is of great importance in regeneration of hemopoiesis. This differentiation is provided by preceded recovery of bone marrow structural-functional organization. The important role in regeneration of hemopoiesis belongs to hemopoietic microenvironmental elements which express its induced influence by the increase of colony-stimulating and erythropoietic activities, interleukin-1 and interleukin-3 production.     

Morphological stages of regeneration in the planarian Dugesia tigrina: A light and electron microscopic study

A precise sequence of four morphological stages of head regeneration in the planarian Dugesia tigrina has been determined by light and electron microscopy. Each stage is identified by a particular morphogenetic process: I, wound healing; II, blastema development; III, growth; IV, differentiation. A wound epidermis consisting of a thin, sheet-like layer of cells, rapidly forms from undamaged epidermal cells at the wound margin. The early blastema is comprised of neoblasts which mature into regeneration cells. The maturational changes include the appearance of a nucleolus, nuclear pores, and perinuclear dense aggregates of granulofibrillar material in these cells. These elements are not evident in the neoblasts of the younger blastema. No mitotic cells are encountered in the blastema or wound epidermis. Cytoplasmic expansion of the regeneration cells is correlated with the formation of numerous microtubules radiating from a juxtanuclear centrosphere. During differentiation of muscle cells, distended, granule-studded cisternae, having moderately fibrillar contents, are regularly disposed adjacent to small patches of myofilaments. Presumptive epidermal cells are recognized by prominent “islands” of finely fibrillar cytoplasm. These cytoplasmic zones persist for a time during definitive differentiation when Golgi bodies, vacuoles, mucous droplets, and rhabdites become evident. The newly formed epidermal cells become inserted among the cells of the wound epidermis. Thus, cells of both the blastema and of the wound epidermis contribute to the reconstituted epidermis.     

Bone morphogenetic proteins in dentin regeneration for potential use in endodontic therapy

The human dentition is indispensable for nutrition and physiology. The teeth have evolved for mastication of food. Caries is a common dental problem in which the dentin matrix is damaged. When the caries is deep and the dental pulp is exposed, the pulp has to be removed in many cases, resulting ultimately in loss of the tooth. Therefore, the regeneration of dentin-pulp complex is the long-term goal of operative dentistry and endodontics. The key elements of dentin regeneration are stem cells, morphogens such as bone morphogenetic proteins (BMPs) and a scaffold of extracellular matrix. The dental pulp has stem/progenitor cells that have the potential to differentiate into dentin-forming odontoblasts in response to BMPs. Pulpal wound healing consists of stem/progenitor cells release from dental pulp niche after noxious stimuli such as caries, migration to the injured site, proliferation and differentiation into odontoblasts. There are two main strategies for pulp therapy to regenerate dentin: (1) in vivo method of enhancing the natural healing potential of pulp tissue by application of BMP proteins or BMP genes, (2) ex vivo method of isolation of stem/progenitor cells, differentiation with BMP proteins or BMP genes and transplantation to the tooth. This review summarizes recent advances in application of BMPs for dentin regeneration and possible use in endodotic therapy.     

Anatomical Studies of Regeneration after Ringing of Eucommia ulmoides

The trunk of Eucommia ulmoides Oliv. following ringing about one meter in length, could regenerate new bark without significantly affecting the tree growth. Such regeneration process differed from that following partial stripping in other trees as has been reported previously. After ringing in Eucommia ulmoides, there was primarily a proliferation of xylary mother cells and partly remained cambium cells forming a cork layer which led to the establishmant of a more or less normal complete periderm. At the same time the xylary ray cells near the surface dilated and proliferated until they extended across in continuity with the neighbor rays. No callus formation by the rays emerged from the surface was observed. Among the ray cells following multiplication, some immature xylary elements were also visible. Simultaneously, some immature xylary elements lying internal to region of the dilated and coalesced ray cells gradually transformed into new cambium. Due to the fact that initially part of the xylary ray cells was not completely differentiated, it seemed that some of the remained xylary rays separated the newly formed cambium zone into many segments. Three to four months after ringing, new cambium activity proceeded periclinally so as to establish a complete ring of cambium zone. Subsequent differentiation of newly formed cambium followed the normal pattern of vascular tissue development in this plant.     

A morphological study of regeneration of the goldfish elasmoid scales

Studies were carried out on the model of regeneration of mechanically removed elasmoid scales. Regenerating scales were morphologically analyzed using light and electron microscopy. It was found that the cells responsible for regeneration of the elasmoid scale plates could be classified as osteogenic elements. Little differentiated preosteoblasts were detected in the connective tissue of dermis on day 3 of regeneration, while partially calcified plates underlaid osteoblasts on day 7. The scale cover was fully restored on day 14 and it took two days for each bone plate to be formed. Osteocytes, fully differentiated osteogenic elements, were found in the deepest regions of newly formed scales.     

A morphological study of regeneration of the goldfish elasmoid scales

Studies were carried out on the model of regeneration of mechanically removed elasmoid scales. Regenerating scales were morphologically analyzed using light and electron microscopy. It was found that the cells responsible for regeneration of the elasmoid scale plates could be classified as osteogenic elements. Little differentiated preosteoblasts were detected in the connective tissue of dermis on day 3 of regeneration, while partially calcified plates underlay osteoblasts on day 7. The scale cover was fully restored on day 14 and it took two days for each bone plate to be formed. Osteocytes, fully differentiated osteogenic elements, were found in the deepest regions of newly formed scales.     

Regeneration of a complex of structures of the ophiuroid <Emphasis Type="Italic">Amphipholis kochii</Emphasis> Lütken, 1872 (Ophiurae) after disk autotomy

Regeneration of structures (genital bars and ligaments) which enables ophiuroids to throw away the aboral part of the disc was studied in this work. The succession of disc separation during autotomy has been fixed. It has been revealed that upon restoration of genital bars and ligaments regeneration occurs due to migration of cells from tissues remaining after autotomy, but not due to differentiation of cell elements. The cell sources of regeneration of the studied structures are fibroblasts and sclerocytes.     

Processes of the differentiation and reproduction of different types of muscle cells

The present review is regarding a vast evidence on reproduction, differentiation, and regenerative capacity of various muscle cells on the basis of A. A. Zavarzin's (Senior) parallelism conception. It is specially emphasized that parallelism in the subcellular organization of contractile structures of somatic and of heart muscle goes well together with a totally different principle of organization of these cellular elements (symplasts, or cells, respectively), and with different mechanisms of their histogenesis and regeneration (proliferation of, respectively, myoblasts, or immature myocytes). According to their ultrastructure and pattern of interrelation between proliferation and differentiation process, muscles of lymphatic hearts are closer to somatic muscles rather than to ordinary myocardium. Special attention is called to paradoxical situations, such as the presence of satellite-like cells in the myocardium of Decapoda, DNA-synthesizing capacity observed in the nuclei of growing somatic muscles of the silkworm, or DNA-synthesizing and chromosome-reproducing capacity of adult primate and human cardiomyocytes.     

Physiologically based microenvironment for in vitro neural differentiation of adipose-derived stem cells

The limited capacity of nervous system to promote a spontaneous regeneration and the high rate of neurodegenerative diseases appearance are keys factors that stimulate researches both for defining the molecular mechanisms of pathophysiology and for evaluating putative strategies to induce neural tissue regeneration. In this latter aspect, the application of stem cells seems to be a promising approach, even if the control of their differentiation and the maintaining of a safe state of proliferation should be troubled. Here, we focus on adipose tissue-derived stem cells and we seek out the recent advances on the promotion of their neural differentiation, performing a critical integration of the basic biology and physiology of adipose tissuederived stem cells with the functional modifications that the biophysical, biomechanical and biochemical microenvironment induces to cell phenotype. The pre-clinical studies showed that the neural differentiation by cell stimulation with growth factors benefits from the integration with biomaterials and biophysical interaction like microgravity. All these elements have been reported as furnisher of microenvironments with desirable biological, physical and mechanical properties. A critical review of current knowledge is here proposed, underscoring that a real advance toward a stable, safe and controllable adipose stem cells clinical application will derive from a synergic multidisciplinary approach that involves material engineer, basic cell biology, cell and tissue physiology.     

Activation of insulin-like growth factor II expression during skeletal muscle regeneration in the rat: correlation with myotube formation.

Insulin-like growth factors (IGFs) are important stimulators of proliferation and differentiation of cultured myoblasts. It has previously been shown that IGF-I is induced during muscle regeneration in rodents, however, little is known about the expression of IGF-II. Therefore, two in vivo models were used to analyze IGF-II mRNA expression during skeletal muscle regeneration in the rat: injection of the snake venom notexin and induction of ischemia. During the regeneration process the levels of both IGF-I and IGF-II mRNA were transiently induced, as analyzed by solution hybridization. Both IGF-I-like immunoreactivity and IGF-II-like immunoreactivity were found to be present during muscle regeneration. In a time course study, induction of IGF-II was preceded by IGF-I, both at the mRNA and protein levels. Using alpha- and beta-actin as markers for different stages of skeletal muscle differentiation, together with the immunohistochemistry data, it is concluded that the expression of IGF-I and IGF-II occurs at different differentiation stages, and that IGF-II appears concomitant to the formation of myotubes. These results suggest that each IGF has a distinct role during the differentiation of muscle cells.     

Role of hematopoietic cells-precursors in the regeneration of hematopoiesis after cytostatic action]

Cyclophosphamide was injected at a dose of 150 mg/kg once to CBA mice and caused expressed hypoplasia of bone marrow hemopoiesis (decreased myelokaryocyte count, CFU-S and CFU-D) from day 1 of the experiment. The rise of CFU-S (8) and CFU-S (12) to a baseline level was noted on day 4 (the time of intensive regeneration processes), whereas CFU-D levels did not change. It is suggested that cytostatic action may trigger "shunt" hemopoiesis mechanisms which enhance differentiation of primitive hemopoietic cells and proliferative activity of mature hemopoietic elements.     

Phloem transdifferentiation from immature xylem cells during bark regeneration after girdling in Eucommia ulmoides Oliv

Eucommia ulmoides Oliv. (Eucommiaceae), a traditional Chinesemedicinal plant, was used to study phloem cell differentiationduring bark regeneration after girdling on a large scale. Hereit is shown that new sieve elements (SEs) appeared in the regeneratedtissues before the formation of wound cambium during bark regenerationafter girdling, and they could originate from the transdifferentiationof immature/differentiating axial xylem cells left on the trunk.Assays of water-cultured twigs revealed that girdling blockedsucrose transport until the formation of new SEs, and the regenerationof the functional SEs was not dependent on the substance providedby the axis system outside the girdled areas, while exogenousindole acetic acid (IAA) applied on the wound surface acceleratedSE differentiation. The experiments suggest that the immaturexylem cells can transdifferentiate into phloem cells under certainconditions, which means xylem and phloem cells might share someidentical features at the beginning of their differentiationpathway. This study also showed that the bark regeneration systemcould provide a novel method for studying xylem and phloem celldifferentiation. Key words: Bark regeneration, Eucommia ulmoides Oliv., immature xylem cells, sieve elements, transdifferentiation Received 19 November 2007; Revised 23 January 2008 Accepted 24 January 2008     

From planarians to mammals - the many faces of regeneration

This report presents the history of the involvement of the Department of Cytology in studies of different aspects of regeneration. It can be divided into two major phases; the first focused on the regeneration of Turbellarians and the second on the regeneration of rat skeletal muscles including the differentiation of satellite cells in vitro. Regeneration of Turbellarians was investigated both at the cellular and molecular levels including the role of the protein kinase C (PKC) in this process. Studies on skeletal muscle regeneration initially focused on factors involved in regulation of signal transduction pathways. Next, we explored the influence of growth factors on the modulation of the regeneration process. Another important aspect of our studies was investigating of the distribution and function of different proteins involved in adhesion and fusion of myoblasts. Finally, we are also conducting research on the role of stem cells from other tissues in the regeneration of skeletal muscle.     

Expression patterns of runx2, sparc, and bgp during scale regeneration in the goldfish Carassius auratus

Teleost fish scale is a dermal skeleton equipped with a strong regenerative ability. Owing to this regenerative ability, teleost fish scale can be used as a model for the regeneration of the dermal skeleton. However, there is insufficient fundamental knowledge of the regeneration, and this limits the usage of fish scale. In this study, as a first step toward understanding the molecular mechanism of the cellular differentiation during scale regeneration, we cloned the cDNAs for osteoblast-related proteins (Runx2, Sparc, and Bgp) in goldfish, and analyzed their expressions during scale regeneration. The expression profiles of these genes during scale regeneration were similar to those during mammalian osteoblastic differentiation. Specifically, runx2 expression was increased at the earliest time point, followed by sparc expression and then bgp expression. In the earlier stages, these genes were expressed in cells that formed cellular condensations and the flat cells surrounding them in the scale pocket. As the regeneration proceeded, the expressions became restricted to the episquamal, hyposquamal, and marginal scleroblasts and the cells around the marginal area of the regenerating scale. These results strongly suggest that (1) the differentiation mechanism of scleroblasts is similar to that of mammalian osteoblasts and odontoblasts, (2) scleroblast differentiation occurs around the cellular condensations at the early regeneration stage and is restricted to the marginal area of the scale at the later stage, and (3) the differentiation mechanisms are similar between the episquamal scleroblasts that produce the external layer and the hyposquamal scleroblasts that produce the basal plate.     

Mesenchymal stem cells showed the highest potential for the regeneration of injured liver tissue compared with other subpopulations of the bone marrow

We have previously reported that bone marrow cells (BMCs) participate in the regeneration after liver injury. However, it is not established that this is the result of differentiation of hematopoietic stem cells (HSCs), mesenchymal stem cells (MSCs) or the combination of both. We investigated the contribution of each cell fraction to the regenerative process. First, we confirmed that transplanted stem cells migrate directly to injured liver tissue without dispersing to other organs. Next, we divided green fluorescent protein (GFP)-expressing BMCs into three populations as mononuclear cells, MSCs and HSCs. We then compared the engraftment capacity after transplantation of each fraction of cells into liver-injured mice. Of these, the MSCs transplanted group showed the highest GFP fluorescence intensities in liver tissue by flow cytometry analysis and confocal microscopic observation. Furthermore, MSCs showed differentiation potential into hepatocytes when co-cultured with injured liver cells, which suggests that MSCs showed highest potential for the regeneration of injured liver tissue compared with those of the other two cell refractions.