肢体再生——来自有尾两栖类的认知

蝾螈等有尾两栖类在其肢体任何节段被截断后,能通过准确的时空模式调节完成具有位置匹配关系的再生修复,该过程由受损肢体残端产生的芽基组织介导完成。芽基细胞的来源目前尚有争议,其产生受局部基质微环境诱导并涉及细胞表观遗传学改变,性状上呈现不完全的细胞再编程特征,增殖分化具有神经依赖性。哺乳类包括人类仅具有极为有限的肢体再生能力,其肢体再生限于指（趾）末端受损离断。深入探讨有尾两栖类等肢体再生过程的细胞分子机制,将为探索新的干细胞损伤修复途径及再生促进策略提供线索。     

Limb regeneration in vertebrates: regulatory factors

Various regulatory factors are required in epimorphic regeneration of an adult newt limb. These factors (namely, amputational injury, the wound and apical epithelium, nerves (mitogenic agents), hormones (the hormonal milieu), bioelectric fields, probably the immune system, and possibly cyclic nucleotides and heretofore unknown regulators) act in concert and contribute to the developing microenvironment of the regenerate in support of normal regrowth and differentiation.     

Retina and lens regeneration in anuran amphibians

Anuran amphibians can regenerate the retina through differentiation of stem cells in the ciliary marginal zone and through transdifferentiation of the retinal pigmented epithelium. By contrast, the regeneration of the lens has been demonstrated only in larvae of species belonging to the Xenopus genus, where the lens regenerates through transdifferentiation of the outer cornea. Retinal pigmented epithelium to neural retina and outer cornea to lens transdifferentiation processes are triggered and sustained by signaling molecules belonging to the family of the fibroblast growth factor. Both during retina and lens regeneration there is a re-activation of many of the genes which are activated during development of the eye, even though the spatial and temporal pattern of gene expression is not a simple repetition of that found in development.     

Limb regeneration: re-entering the cell cycle.

Understanding the cellular plasticity that enables urodeles to regenerate many tissues is important for determining why mammals repair those same tissues with scar. The answer may lie partly in a recently discovered differential responsiveness of urodele cells to factors present in serum at the wound site.     

Limb muscle regeneration in peripheral nerve autografts

In a previous study we demonstrated regenerative growth of extraocular muscle within transplanted peripheral nerve autografts. The present study addresses the feasibility of inducing regeneration of limb muscle within autologous peripheral nerve implants in the gluteus medius of beagles. In six anesthetized animals, a 2-cm segment of the left infraorbital sensory nerve was removed from the nose and implanted between the cut ends of several muscle fascicles in the left gluteus medius. After 4 weeks, the nerve grafts were removed and examined by light and electron microscopy. Muscle fibers were seen surrounded by the epineurium of the implanted nerve along its entire length, growing in parallel with the long axis of the nerve. The regenerating fibers were closely associated with the basal lamina of degenerating myelinated and unmyelinated axons. This study suggests that limb muscle, like extraocular muscle, is capable of organized regenerative growth within peripheral nerve autografts.     

Punctuated cell cycling in the regeneration blastema of urodele amphibians

Abstract. We previously assumed that all cells in the regeneration blastema are randomly distributed throughout the cell cycle and actively cycling towards the next mitotic division. We now show that data from continuous labeling (³H-thymidine) experiments do not support this view and favor instead the hypothesis that the blastema cell cycle is punctuated in the g₁ phase wherein cells can enter what we term a transiently quiescent (TQ) position. We call this hypothesis the punctuated cycling (PC) hypothesis. We further propose that the relative sizes of the quiescent and actively cycling populations explain (1) variations in rates of regeneration in different sizes of urodele amphibians, (2) the rate and success of regeneration in different species, and (3) how various controlling factors, such as injury, nerves, growth factors, wound epidermis, and hormones, influence the initiation and progression of the regeneration process. This PC hypothesis is important for interpreting previous pulse labeling data, is consistent with recently obtained continuous labeling data, and is experimentally testable.     

Early innervation of skeletal muscle during tail regeneration in urodele amphibians.

The innervation pattern of skeletal muscles was studied in the normal and regenerating tail of Notophthalmus viridescens. Silver staining for nerve endings and histochemical localization of acetylcholinesterase (AChE) were used for light microscopy. In In normal musculature, AChE positive reactions were localized at the ends of the muscle fibers where they are anchored on connective tissue septa by myotendinous junctions. At this level, silver staining shows nerve terminals forming endplates. During regeneration, positive reactions for AChE appear de novo as dense plates localized at the ends of the newly formed myotubes. The mechanisms involved in the localization of AChE on this surface seem to operate before previous local contacts by nerve terminals. From the ultrastructural data and immunohistochemical results with anti-laminin antibody, these observations suggest that regenerating muscle fibers determine a region of post-synaptic specialization in close relation with the organization of myotendinous regions and basement membrane formation. Nerve-muscle contacts appear at these levels at stage IV (15-20 days after amputation) in the stump and in the rostral part of the regenerate (transition zone). These nerve terminals are provided by the disorganized peripheral nervous system of the injured segment. In the regenerate a similar pattern of AChE reaction can be seen in every myotube, differentiating according to a rostro-caudal gradient. Innervation at the ends of the muscle fibers is in spatiotemporal relation with the exists of the ventral roots from the regenerating nerve cord as the regenerate continues to grow in length.     

Peripheral and central contribution to the pupillary reflex control in amphibians: pupillographic and theoretical considerations

The amphibian pupillary response is mediated by two mechanisms. One is restricted to the iris itself, the other involves the central nervous system. In the present study we investigated the dynamic of the pupillary response and separated the parts by experiments with lesions of the optic and oculomotor nerve. The pupillary response of urodeles is comparable to that in other vertebrates. Differences are observable in amplitude and time course. Furthermore there are no binocular summation or consensual reactions. The central control of the pupillary response increases amplitude and velocity of the pupillary response in sudden luminance changes, compared to the pure iris related reaction. The separation of central and peripheral pupillary reflex control was shown more quantitatively through the introduction of a theoretical model. The investigation of the dark response gives evidence that there is no dilatator pupillae in urodele amphibians, since comparison to results in anuran species show that this is not a general feature in amphibians.     

Limb regeneration and apolysis in the tick, Ornithodoros tartakovskyi.

Limb regeneration potential and the apolysis process were investigated in the argasid tick, Ornithodoros tartakovskyi. Developmental instars received single or multiple amputations and were subsequently allowed to undergo single or multiple apolyses. Amputated ticks regenerated complete normal limbs but only after four successive apolyses. Following a single apolysis, the majority of regenerated limbs were essentially miniature duplicates of normal legs but commonly lacked normal chaetotaxy and/or tarsal hump(s). The site of amputation distal to the coxa-trochanter joint, number of limbs removed from an individual, and instar amputated did not consistently influence the extent of regeneration. Coagulation and clot formation were observed.The limbs of the tick apolysed within the old leg hulls. Larvae and nymphs amputated relatively early during the period of apolyses regenerated limbs; late amputations precluded regeneration. The process of apolysis was irreversible and not obviously affected by amputations.     

Limb regeneration and survival of prolactin treated hypophysectomized adult newts

Hypophysectomized adult newts exhibited 98% survival and limb regeneration at 23 days post-hypophysectomy when injected intraperitoneally every other day with prolactin (0.015 U/newt) and kept continuously in aquaria with 1 × 10^?7 concentration of thyroxine. Thyroxine alone was no more effective than saline injections. Prolactin (1.2 U/newt every other day) alone increased survival and limb regeneration, but less effectively than did the prolactin-thyroxine combination.     

The role of the myoblasts and connective tissue in the regeneration of the limbs in amphibians

A possibility of tissue metaplasia (transformation of one cell type into another) during limb regeneration in lower vertebrates has been a matter of vivid arguments over the last decades. These discussions are rather irreconcilable in character mainly due to the lack of reliable cell markers which permit to follow all the stages of cell transformation during metaplasia. The final conclusions can be made only if any artifacts of cell labelling are excluded. Latest findings obtained using nuclear and cytoplasmic markers are presented which suggest that many data interpreted previously as a convincing proof of metaplasia may be a consequence of the involvement of nondifferentiated cells in regeneration. Molecular biological approaches are believed to be most promising for the solution of disputable problems of tissue metaplasia. However, recent findings about actin gene hypomethylation are insufficient for any final conclusions about the possibility of metaplasia during limb regeneration. The answer to many unsolved questions of developmental biology can be made only when combined use is made of modern methods of cell and molecular biology.     

Melanogenesis in amphibians

Summary The pigmented epithelium of Rana pipiens tadpole eyes normally develops at least two types of melanosomes: (1) an elongated melanin granule of relatively homogeneous electron density, and (2) a complex melanosome which has an outer electrondense area and one or more less dense cores. Evidence indicates that complex melanosomes are formed by new melanin enclosing preexisting melanosomes. An organized fibrillar premelanosome is demonstrated with the aid of the antimelanogenic compound phenylthiourea (PTU). These premelanosomes are the developing forms of the elongated melanosomes. There is evidence that the premelanosomes originate in the smooth endoplasmic reticulum. Phenylthiourea blocks melanin synthesis in the premelanosomes; however, removal of the PTU allows pigment deposition. This finding of an organized, fibrillar premelanosome in an amphibian marks the lowest phylogenetic group in which these organelles have been described.An Oak Ridge Graduate Fellow from Catholic University of America, Washington, D.C., under appointment from Oak Ridge Associated Universities.The MAN Program is supported by the National Cancer Institute, the National Institute of General Medical Sciences, the National Institute of Allergy and Infectious Diseases, and the U.S. Atomic Energy Commission.Oak Ridge National Laboratory is operated by Union Carbide Corporation Nuclear Division for the U.S. Atomic Energy Commission.     

Cancer resistance in amphibians

While spontaneous tumours may occasionally develop in inbred and isogenic strains of Xenopus laevis, the South African clawed toad, they are extremely rare in natural and laboratory populations. Only two amphibian neoplasms, the renal adenocarcinoma of Rana pipiens and the lymphosarcoma of Xenopus laevis, have been extensively explored. Amphibians are resistant to the development of neoplasia, even following exposure to "direct-acting" chemical carcinogens such as N-methyl-N-nitrosourea, that are highly lymphotoxic, thus diminishing immune reactivity. Regenerative capacity in adults, and a dramatic metamorphosis which remodels much of the larval body to produce the adult form, are unique to amphibian vertebrates, and the control mechanisms involved may protect against cancer. For example, naturally rising corticosteroid titres during metamorphosis will impair some T-cell functions, and the removal of T-regulatory (suppressor) functions inhibits the induction of altered-self tolerance. Altered-self tolerance is not as effectively induced in adult Xenopus laevis as in mammals, so cancer cells with new antigenicity are more likely be rejected in amphibians. Amphibian immunocytes tend to undergo apoptosis readily in vitro, and, unlike mammalian immunocytes, undergo apoptosis without entering the cell cycle. Cells not in the cell cycle that die from nuclear damage (apoptosis), will have no opportunity to express genetic instability leading to cell transformation. We suggest that all these factors, rather than any one of them, may reduce susceptibility to cancer in amphibians.