Free-living flatworms under the knife: past and present

Traditionally, regeneration research has been closely tied to flatworm research, as flatworms (Plathelminthes) were among the first animals where the phenomenon of regeneration was discovered. Since then, the main focus of flatworm regeneration research was on triclads, for which various phenomena were observed and a number of theories developed. However, free-living flatworms encompass a number of other taxa where regeneration was found to be possible. This review aims to display and to compare regeneration in all major free-living flatworm taxa, with special focus on a new player in the field of regeneration, Macrostomum lignano (Macrostomorpha). Findings on the regeneration capacity of this organism provide clues for links between regeneration and (post-)embryonic development, starvation, and asexual reproduction. The role of the nervous system and especially the brain for regeneration is discussed, and similarities as well as particularities in regeneration among free-living flatworms are pointed out.     

Comparison of two indices for evaluating regeneration ability in rice (Oryza sativa L.) through a diallel analysis

 A full diallel analysis was performed among seven rice cultivars, all of which showed different abilities of regeneration from seed-derived calli. Number of regenerated shoots and regeneration frequency were used as indices of regeneration ability. In both cases, additive effects were significant at the 0.1% level, and dominant genes had a positive effect, that is, they increased regeneration ability. Non-allelic interaction (epistasis) and maternal effects were not detected. Dominance effects were significant at the 1% and the 0.1% level when the number of regenerated shoots and regeneration frequency were used as indices, respectively. Average degree of dominance was 0.531 for the shoot regeneration index and 0.990 for the regeneration frequency index. Since broad-sense heritability was 0.919 for number of regenerated shoots and 0.736 for regeneration frequency, the former was considered to be a better index of regeneration ability than the latter. Received: 10 June 1996 / Accepted: 23 August 1996     

Ethylene influences green plant regeneration from barley callus

The plant hormone ethylene is involved in numerous plant processes including in vitro growth and regeneration. Manipulating ethylene in vitro may be useful for increasing plant regeneration from cultured cells. As part of ongoing efforts to improve plant regeneration from barley (Hordeum vulgare L.), we investigated ethylene emanation using our improved system and investigated methods of manipulating ethylene to increase regeneration. In vitro assays of regeneration from six cultivars, involving 10 weeks of callus initiation and proliferation followed by 8 weeks of plant regeneration, showed a correlation between regeneration and ethylene production: ethylene production was highest from ‘Golden Promise’, the best regenerator, and lowest from ‘Morex’ and ‘DH-20’, the poorest regenerators. Increasing ethylene production by addition of 1-aminocyclopropane 1-carboxylic acid (ACC) during weeks 8–10 increased regeneration from Morex. In contrast, adding ACC to Golden Promise cultures during any of the tissue culture steps reduced regeneration, suggesting that Golden Promise may produce more ethylene than needed for maximum regeneration rates. Blocking ethylene action with silver nitrate during weeks 5–10 almost doubled the regeneration from Morex and increased the Golden Promise regeneration 1.5-fold. Silver nitrate treatment of Golden Promise cultures during weeks 8–14 more than doubled the green plant regeneration. These results indicate that differential ethylene production is related to regeneration in the improved barley tissue culture system. Specific manipulations of ethylene were identified that can be used to increase the green plant regeneration from barley cultivars. The timing of ethylene action appears to be critical for maximum regeneration.     

森林更新与空间异质性

森林更换是一个重要的生态学过程,一直是森林生态系统动态研究中的主要领域之一。森林更新受物理环境、自然干扰、人为干扰、更新树种特性、树种对干扰的反应等因素及其相互作用的影响。这些生物和非生物的因素随空间和时间而不断变化,构成了森林的空间异质性和时间异质性,使森林更新具有空间和时间上的变化特点,表现在异质性的格局和过程中。探索森林更新与空间异质性的内在规律,可揭示空间格局对更新的生态学过程的潜在作用机制。本文主要综述了近年来有关森林更新与空间异质性研究的主要内容和一些观点,分析了更新中空间异质性的来源,着重评述了空间异质性的生境及更新树种的反应、小尺度的空间异质性与更新动态、林分中光有效性的空间异质性与更新格局以及土壤和更新的空间异质性尺度的关联性等方面的研究。     

Ethylene inhibitors and low kanamycin concentrations improve adventitious regeneration from apricot leaves

An improved method for adventitious regeneration from apricot leaves is described. The use of the ethylene inhibitors silver thiosulphate (30-60 micro M) or aminoethoxyvinylglycine (0.5 micro M) increased regeneration percentages in Helena and Canino apricot cultivars and also the consistency of results from different experiments. Use of "Pure Agar" also improved regeneration from Helena leaves as compared with agargel or agarose. Regeneration rates for Canino were dependent on the medium in which the shoots were micropropagated. When different antibiotics were tested for their influence on regeneration, the combination of cefotaxime (0.13 m M) plus vancomycin (0.63 m M), which efficiently controls Agrobacterium growth, also increased regeneration percentages in Helena two-fold but did not affect regeneration in Canino. Kanamycin, an antibiotic widely used for selection of nptII transformed cells, promoted more rapid regeneration and higher regeneration rates from Helena leaves when added at low concentrations (8.6 and 17.1 micro M). With this improved procedure, regeneration from apricot leaves has been increased more than 200% as compared with rates reported previously.     

Regenerative medicine for diseases of the head and neck: principles of in vivo regeneration

The application of endogenous regeneration in regenerative medicine is based on the concept of inducing regeneration of damaged or lost tissues from residual tissues in situ. Therefore, endogenous regeneration is also termed in vivo regeneration as opposed to mechanisms of ex vivo regeneration which are applied, for example, in the field of tissue engineering. The basic science foundation for mechanisms of endogenous regeneration is provided by the field of regenerative biology. The ambitious vision for the application of endogenous regeneration in regenerative medicine is stimulated by investigations in the model organisms of regenerative biology, most notably hydra, planarians and urodeles. These model organisms demonstrate remarkable regenerative capabilities, which appear to be conserved over large phylogenetical stretches with convincing evidence for a homologue origin of an endogenous regenerative capability. Although the elucidation of the molecular and cellular mechanisms of these endogenous regenerative phenomena is still in its beginning, there are indications that these processes have potential to become useful for human benefit. Such indications also exist for particular applications in diseases of the head and neck region. As such epimorphic regeneration without blastema formation may be relevant to regeneration of sensorineural epithelia of the inner ear or the olphactory epithelium. Complex tissue lesions of the head and neck as they occur after trauma or tumor resections may be approached on the basis of relevant mechanisms in epimorphic regeneration with blastema formation.     

Enhancement of in vitro organogenetic capacity of rose by preculture of donor shoots on the medium with thidiazuron

Rose genotypes can be recalcitrant in in vitro regeneration. A lack of competence for regeneration results in unsatisfactory numbers of adventitious shoots, which diminishes achievements in use of genetic engineering in rose modern breeding. Among the factors which can partially overcome this problem is the use of thidiazuron (TDZ) in the induction phase of regeneration. We studied the use of TDZ in concentrations 0.05 and 1.0 mg l⁻¹, in the medium for the culture of donor shoots with the aim of increasing the regeneration competence of five rose cultivars differing in their regeneration ability. All the tested genotypes increased their regeneration potential when the donor shoots were cultured on TDZ media for 15 or 30 days. The strongest effect, doubling to quadrupling the number of shoots, was obtained in the cultivars characterized by the lowest regeneration potential. This enhanced regeneration ability was maintained through two or three subsequent 4-week long regeneration passages.     

Deer antler regeneration: cells, concepts, and controversies

The periodic replacement of antlers is an exceptional regenerative process in mammals, which in general are unable to regenerate complete body appendages. Antler regeneration has traditionally been viewed as an epimorphic process closely resembling limb regeneration in urodele amphibians, and the terminology of the latter process has also been applied to antler regeneration. More recent studies, however, showed that, unlike urodele limb regeneration, antler regeneration does not involve cell dedifferentiation and the formation of a blastema from these dedifferentiated cells. Rather, these studies suggest that antler regeneration is a stem-cell-based process that depends on the periodic activation of, presumably neural-crest-derived, periosteal stem cells of the distal pedicle. The evidence for this hypothesis is reviewed and as a result, a new concept of antler regeneration as a process of stem-cell-based epimorphic regeneration is proposed that does not involve cell dedifferentiation or transdifferentiation. Antler regeneration illustrates that extensive appendage regeneration in a postnatal mammal can be achieved by a developmental process that differs in several fundamental aspects from limb regeneration in urodeles.     

An insight into planarian regeneration

BackgroundPlanarian has attracted increasing attentions in the regeneration field for its usefulness as an important biological model organism attributing to its strong regeneration ability. Both the complexity of multiple regulatory networks and their coordinate functions contribute to the maintenance of normal cellular homeostasis and the process of regeneration in planarian. The polarity, size, location and number of regeneration tissues are regulated by diverse mechanisms. In this review we summarize the recent advances about the importance genetic and molecular mechanisms for regeneration control on various tissues in planarian.MethodsA comprehensive literature search of original articles published in recent years was performed in regards to the molecular mechanism of each cell types during the planarian regeneration, including neoblast, nerve system, eye spot, excretory system and epidermal.ResultsAvailable molecular mechanisms gave us an overview of regeneration process in every tissue. The sense of injuries and initiation of regeneration is regulated by diverse genes like follistatin and ERK signaling. The Neoblasts differentiate into tissue progenitors under the regulation of genes such as egfr‐3. The regeneration polarity is controlled by Wnt pathway, BMP pathway and bioelectric signals. The neoblast within the blastema differentiate into desired cell types and regenerate the missing tissues. Those tissue specific genes regulate the tissue progenitor cells to differentiate into desired cell types to complete the regeneration process.ConclusionAll tissue types in planarian participate in the regeneration process regulated by distinct molecular factors and cellular signaling pathways. The neoblasts play vital roles in tissue regeneration and morphology maintenance. These studies provide new insights into the molecular mechanisms for regulating planarian regeneration.

Genetic and molecular mechanisms for regeneration control on various tissues in planarian.     

Notch signaling inhibits axon regeneration

Many neurons have limited capacity to regenerate their axons after injury. Neurons in the mammalian central nervous system do not regenerate, and even neurons in the peripheral nervous system often fail to regenerate to their former targets. This failure is likely due in part to pathways that actively restrict regeneration; however, only a few factors that limit regeneration are known. Here, using single-neuron analysis of regeneration in?vivo, we show that Notch/lin-12 signaling inhibits the regeneration of mature C.?elegans neurons. Notch signaling suppresses regeneration by acting autonomously in the injured cell to prevent growth cone formation. The metalloprotease and gamma-secretase cleavage events that lead to Notch activation during development are also required for its activity in regeneration. Furthermore, blocking Notch activation immediately after injury improves regeneration. Our results define a postdevelopmental role for the Notch pathway as a repressor of axon regeneration in?vivo.     

Unifying principles of regeneration I: Epimorphosis versus morphallaxis

Because research on regeneration has a long history, some classic definitions and concepts about regeneration which were established in earlier times have been retained without reconsideration for a long time, even though many relevant new findings have accumulated. To clarify the points on which research should be focused on for elucidating the mechanisms of regeneration, we should reconsider such classical definitions and principles of regeneration at the cellular and molecular level. Here, we consider two differing principles of regeneration which have been classically defined as 'epimorphosis' and 'morphallaxis', and propose the abandonment of these classical categories and their replacement by a new unifying principle in order to facilitate regeneration studies.     

The roles of signaling pathways in liver repair and regeneration

The liver has remarkable regeneration potency that restores liver mass and sustains body hemostasis. Liver regeneration through signaling pathways following resection or moderate damages are well studied. Various cell signaling, growth factors, cytokines, receptors, and cell types implicated in liver regeneration undergo controlled hypertrophy and proliferation. Some aspects of liver regeneration have been discovered and many investigations have been carried out to identify its mechanisms. However, for optimizing liver regeneration more should be understood about mechanisms that control the growth of hepatocytes and other liver cell types in adults. The current paper deals with the possible applicability of liver regeneration signaling pathways as a target for therapeutic approaches and preventing various liver damages. Furthermore, the latest findings of spectrum-specific signaling pathway mechanisms that underlie liver regeneration are briefly described.     

The influence of fundamental traits on mechanisms controlling appendage regeneration

One of the most compelling questions in evolutionary biology is why some animals can regenerate injured structures while others cannot. Appendage regeneration appears to be common when viewed across the metazoan phylogeny, yet this ability has been lost in many taxa to varying degrees. Within species, the capacity for regeneration also can vary ontogenetically among individuals. Here we argue that appendage regeneration along the secondary body axis may be constrained by fundamental traits such as body size, aging, life stage, and growth pattern. Studies of the molecular mechanisms affecting regeneration have been conducted primarily with small organisms at early life stages. Such investigations disregard the dramatic shifts in morphology and physiology that organisms undergo as they age, grow, and mature. To help explain interspecific and intraspecific constraints on regeneration, we link particular fundamental traits to specific molecular mechanisms that control regeneration. We present a new synthesis for how these fundamental traits may affect the molecular mechanisms of regeneration at the tissue, cellular, and genomic levels of biological organization. Future studies that explore regeneration in organisms across a broad phylogenetic scale, and within an ontogenetic framework, will help elucidate the proximate mechanisms that modulate regeneration and may reveal new biomedical applications for use in regenerative medicine.     

Protoplast isolation and regeneration in Streptomyces clavuligerus

The regeneration of streptomycete protoplasts is a major step following genetic manipulations such as fusion and DNA-mediated transformation. Reports of studies on the regeneration of protoplasts from Streptomyces clavuligerus are limited and for this reason the experiments described in this paper were carried out. An investigation of protoplast formation and cytology was made to gain further insight into the loss of protoplast viability in osmotically stabilized support media. Protoplasts with the highest regeneration frequency were isolated from mycelium, grown in a two-stage culture system (without glycine), using lysozyme dissolved in a sucrose osmoticum containing 1% bovine serum albumin. The latter promoted improved protoplast viability. A systematic survey was made of the components of regeneration medium R5, previously used for S. clavuligerus, and other potentially advantageous components and conditions, in an attempt to raise the regeneration frequency of the protoplasts. An improved regeneration medium (R6) and protocol which supported higher and more consistent levels of regeneration of S. clavuligerus protoplasts resulted from these experiments. These improved procedures for protoplast isolation and regeneration proved to be suitable for other streptomycete species.     

Sugarbeet (<Emphasis Type="Italic">Beta vulgaris</Emphasis> L.): shoot regeneration from callus and callus protoplasts

The successful application of recombinant DNA technology for crop plants requires efficient regeneration systems. A detailed study on the regeneration potential of callus and callus-derived protoplasts of a recalcitrant species, sugarbeet, was performed. A reproducible and highly efficient method for induction of regenerable friable callus was established from etiolated hypocotyl explants. A reduced sucrose concentration proved beneficial. Successful shoot regeneration could be demonstrated in 10 out of 12 tested lines. Seed germination, followed by callus induction and shoot regeneration required only a single culture medium. Additionally, the regeneration capacity of roots and root-derived callus was demonstrated. Highly efficient plant regeneration was also achieved when using protoplasts isolated from regenerable friable callus induced on etiolated hypocotyls explants. To our knowledge this represents the first report on callus protoplast to plant regeneration in sugarbeet.     

Sugar beet (Beta vulgaris L.) morphogenesis in vitro: effects of phytohormone type and concentration in the culture medium, type of explants, and plant genotype on shoot regeneration frequency

In vitro regeneration techniques have been optimized for seven strains and cultivars of sugar beet (Beta vulgaris L.) bred in Russia. The frequency of shoot regeneration from somatic cells and tissues of sugar beet varies from 10 to 97% depending on the explant type, culture-medium composition, and genotype. The in vitro regeneration potential has been estimated in plants with different genotypes. The effect of medium composition (phytohormones and carbohydrates) on the frequency of the formation of a morphogenic callus competent for plant regeneration has been determined. The effect of the types and concentrations of various cytokines (zeatin, kinetin, and 6-benzylaminopurine) on direct shoot regeneration from cotyledon nodes has been estimated. The culture-medium composition has been optimized for direct shoot regeneration from petioles. The effects of different concentrations of abscisic acid on the frequency of shoot regeneration from a morphogenic callus has been studied. Micropropagation has been used to obtain petiole explants and reproduce the shoots obtained by direct regeneration from cotyledon nodes, petioles, and calluses. Improved shoot-regeneration methods can be used for both agrobacterial and bioballistic genetic transformation of the sugar beet genotypes studied.     

Cell proliferation is necessary for the regeneration of oral structures in the anthozoan cnidarian Nematostella vectensis

The contribution of cell proliferation to regeneration varies greatly between different metazoan models. Planarians rely on pluripotent neoblasts and amphibian limb regeneration depends upon formation of a proliferative blastema, while regeneration in Hydra can occur in the absence of cell proliferation. Recently, the cnidarian Nematostella vectensis has shown potential as a model for studies of regeneration because of the ability to conduct comparative studies of patterning during embryonic development, asexual reproduction, and regeneration. The present study investigates the pattern of cell proliferation during the regeneration of oral structures and the role of cell proliferation in this process. In intact polyps, cell proliferation is observed in both ectodermal and endodermal tissues throughout the entire oral-aboral axis, including in the tentacles and physa. Following bisection, there is initially little change in proliferation at the wound site of the aboral fragment, however, beginning 18 to 24?hours after amputation there is a dramatic increase in cell proliferation at the wound site in the aboral fragment. This elevated level of proliferation is maintained throughout the course or regeneration of oral structures, including the tentacles, the mouth, and the pharynx. Treatments with the cell proliferation inhibitors hydroxyurea and nocodazole demonstrate that cell proliferation is indispensable for the regeneration of oral structures. Although inhibition of regeneration by nocodazole was generally irreversible, secondary amputation reinitiates cell proliferation and regeneration. The study has found that high levels of cell proliferation characterize the regeneration of oral structures in Nematostella, and that this cell proliferation is necessary for the proper progression of regeneration. Thus, while cell proliferation contributes to regeneration of oral structures in both Nematostella and Hydra, Nematostella lacks the ability to undergo the compensatory morphallactic mode of regeneration that characterizes Hydra. Our results are consistent with amputation activating a quiescent population of mitotically competent stem cells in spatial proximity to the wound site, which form the regenerated structures.     

Electrical stimulation accelerates neurite regeneration in axotomized dorsal root ganglion neurons by increasing MMP-2 expression

Electrical stimulation (ES) can be useful for promoting the regeneration of injured axons, but the mechanism underlying its positive effects is largely unknown. The current study aimed to investigate whether ES could enhance the regeneration of injured neurites in dorsal root ganglion explants and regulate the MMP-2 expression level, which is correlated with regeneration. Significantly increased neurite regeneration and MMP-2 expression was observed in the ES group compared with the sham group. However, an MMP inhibitor significantly decreased this ES-induced neurite regeneration. Our data suggest that the positive effect of ES on neurite regeneration could likely be mediated by an increase in MMP-2 expression, thereby promoting the regeneration of injured neurites.     

In vitro regeneration of sesame (Sesamum indicum L.) from seedling cotyledon and hypocotyl explants

The goal of this study was to develop an efficient regeneration protocol to be used for genetic transformation of sesame. Published regeneration methods using benzyladenine (BA) and 1-naphthalene acetic acid (NAA) were unsuccessful for the cultivars used herein. Experiments were carried out using cotyledon and hypocotyl explants from the cultivar Mtwara-2. Later the optimised culture conditions were used to investigate the regeneration response of different genotypes. There was significant interaction between hormone treatments and macronutrients for shoot and root regeneration. Results also showed that shoot regeneration was significantly influenced by explant type. Shoots were only obtained from cotyledons whereas both cotyledons and hypocotyls could produce roots. Modified Murashige and Skoog (MS) medium with N6 macronutrients resulted in twice the shoot regeneration frequency obtained with ½MS macronutrients in the presence of thidiazuron (TDZ). The shoot regeneration frequency was significantly reduced when BA was used in place of TDZ. On shoot regeneration medium containing BA and NAA, only roots were formed. Replacing NAA with indole-3-acetic acid (IAA) greatly improved the regeneration of shoots. The optimum growth regulator combination for shoot regeneration was 20 μM TDZ together with 2.5 μM IAA, which gave a frequency of 63% and 4.4 shoots per regenerating explant for the best cultivar Ex-El. Genotypic differences were significant both for the number of explants regenerating shoots and the number of shoots produced per regenerating explant.