Mechanisms of plant regeneration during succession after shifting cultivation in eastern Amazonia

The role of seed bank, seed rain, and regeneration from seedlings and sprouts after swidden agriculture was compared in 5-, 10- and 20-year-old secondary forest and in a primary forest in Bragantina, Pará, Brazil. The seed bank (0–5 cm soil depth) was largest in the 5-year-old forest (1190 ± 284 seeds m⁻²) and decreased nearly ten-fold with age to 137 ± 19 seeds m⁻² in the primary forest. The highest seed rain was in the 5-year-old forest (883 ± 230 seeds m⁻² year⁻¹) and the least in the primary forest (220 ± 80 seeds m⁻² year⁻¹). Large plants (≥5 cm dbh) had more individuals and species that regenerated from sprouts than from seeds and the most abundant tree species in the secondary forest stands of all ages appear to be maintained by sprouting. The smaller individuals (≥1 m tall, <5 cm dbh) in the 5-year-old forest were mainly from sprouts, but those in the older secondary forests originated mainly from seeds. These results show that at the beginning of succession, although many species can be introduced to swidden fallow from seed bank and seed rain, it is the sprout that is the main source of recruits of primary forest species in secondary forests in Bragantina.     

Mechanisms of improved knee flexion after rectus femoris transfer surgery

Rectus femoris transfer is frequently performed to treat stiff-knee gait in subjects with cerebral palsy. In this surgery, the distal tendon is released from the patella and re-attached to one of several sites, such as the sartorius or the iliotibial band. Surgical outcomes vary, and the mechanisms by which the surgery improves knee motion are unclear. The purpose of this study was to clarify the mechanism by which the transferred muscle improves knee flexion by examining three types of transfers. Muscle-actuated dynamic simulations were created of ten children diagnosed with cerebral palsy and stiff-knee gait. These simulations were altered to represent surgical transfers of the rectus femoris to the sartorius and the iliotibial band. Rectus femoris transfers in which the muscle remained attached to the underlying vasti through scar tissue were also simulated by reducing but not eliminating the muscle's knee extension moment. Simulated transfer to the sartorius, which converted the rectus femoris’ knee extension moment to a flexion moment, produced 32±8° improvement in peak knee flexion on average. Simulated transfer to the iliotibial band, which completely eliminated the muscle's knee extension moment, predicted only slightly less improvement in peak knee flexion (28±8°). Scarred transfer simulations, which reduced the muscle's knee extension moment, predicted significantly less (p<0.001) improvement in peak knee flexion (14±5°). Simulations revealed that improved knee flexion following rectus femoris transfer is achieved primarily by reduction of the muscle's knee extension moment. Reduction of scarring of the rectus femoris to underlying muscles has the potential to enhance knee flexion.     

Harvesting and cryopreservation of lymphatic endothelial cells for lymphatic tissue engineering

In order to provide a suitable source of cells for lymphatic tissue engineering, the present study was designed to investigate techniques for harvesting and cryopreservation of human dermal lymphatic endothelial cells (LECs) in vitro. The LECs were isolated from children’s foreskins and then cultured in endothelial growth medium-2 MV (EGM-2-MV) with 5% FBS. The second passage LECs were suspended in cryopreservation solution containing 40% FBS and 10% Me₂SO in EGM-2-MV, cooled to −80 °C at about 1 °C/min and stored in liquid nitrogen. Samples were thawed quickly in a 37 °C water bath, and the cryoprotectant was removed by serial elution. The membrane integrity of thawed LECs was determined by trypan blue staining exclusion, and their proliferation was evaluated using the MTT method. The expanded cells of two groups were identified using immunofluorescence staining and RT-PCR with lymphatic-specific markers such as Podoplanin and VEGFR-3. Uptake of fluorescent DiI-Ac-LDL and microtubular formation in three-dimensional cultures were used to detect the function of LECs. Flow cytometry was applied to identify cells and to measure the apoptosis rate as well. Cryopreservation resulted in a retrieval of 67 ± 4% and an intact cell rate of 80 ± 3%. The early apoptosis rate of thawed LECs (9.15 ± 0.34%) was higher than that of fresh control LECs (5.31 ± 0.23%). The growth curves of thawed LECs were similar to those of fresh LECs. The thawed LECs were propagated for at least 6-7 passages without alterations in phenotype and function. Highly purified LECs can be isolated by immunomagnetic beads from human dermis. The cryopreserved/thawed and recultivated LECs are proven to have high vitality and growth potential in vitro and may be considered suitable seed cells for lymphatic tissue engineering.     

Acute brain vascular regeneration occurs via lymphatic transdifferentiation

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Mechanisms of muscle dedifferentiation during regeneration

For many years people have known that amphibians have an amazing ability to regenerate lost body parts. In contrast humans have limited regeneration capacity and even simple wound healing results in scarring. Despite more than a century of scientific inquiry, this remarkable phenomenon remains poorly understood. Recent research has begun to provide insight into how this unique process that is now fully accepted to occur via the reversal of cell differentiation is executed at the molecular level. As more and more is known about regeneration and dedifferentiation we can begin to address the question: if given the right signals could mammals also regenerate body structures?     

Lung tissue regeneration after induced injury in Runx3 KO mice

Runx3 is essential for normal murine lung development, and Runx3 knockout (KO) mice, which die soon after birth, exhibit alveolar hyperplasia. Wound healing, tissue repair, and regeneration mechanisms are necessary in humans for proper early lung development. Previous studies have reported that various signaling molecules, such as pErk, Tgf-ß1, CCSP, pJnk, Smad3, and HSP70 are closely related to wound healing. In order to confirm the relationship between lung defects caused by the loss of function of Runx3 and wound healing, we have localized various wound-healing markers after laser irradiation in wild-type and in Runx3 KO mouse lungs at post-natal day 1. Our results indicate that pERK, Tgf-β1, CCSP, pJnk, and HSP70 are dramatically down-regulated by loss of Runx3 during lung wound healing. However, Smad3 is up-regulated in the Runx3 KO laser-irradiated lung region. Therefore, the lung wound-healing mechanism is inhibited in the Runx3 KO mouse, which shows abnormal lung architecture, by reduced pErk, Tgf-β1, CCSP, pJnk, and HSP70 and by induced Smad3.     

Molecular features of secondary vascular tissue regeneration after bark girdling in Populus

Regeneration is a common strategy for plants to repair damage to their tissue after attacks from other organisms or physical assaults. However, how differentiating cells acquire regenerative competence and rebuild the pattern of new tissues remains largely unknown. Using anatomical observation and microarray analysis, we investigated the morphological process and molecular features of secondary vascular tissue regeneration after bark girdling in trees. After bark girdling, new phloem and cambium regenerate from differentiating xylem cells and rebuild secondary vascular tissue pattern within 1 month. Differentiating xylem cells acquire regenerative competence through epigenetic regulation and cell cycle re-entry. The xylem developmental program was blocked, whereas the phloem or cambium program was activated, resulting in the secondary vascular tissue pattern re-establishment. Phytohormones play important roles in vascular tissue regeneration. We propose a model describing the molecular features of secondary vascular tissue regeneration after bark girdling in trees. It provides information for understanding mechanisms of tissue regeneration and pattern formation of the secondary vascular tissues in plants.     

Regulation of lymphatic capillary regeneration by interstitial flow in skin

Decreased interstitial flow (IF) in secondary lymphedema is coincident with poor physiological lymphatic regeneration. However, both the existence and direction of causality between IF and lymphangiogenesis remain unclear. This is primarily because the role of IF and its importance relative to the action of the prolymphangiogenic growth factor vascular endothelial growth factor (VEGF)-C (which signals primarily through its receptor VEGFR-3) are poorly understood. To clarify this, we explored the cooperative roles of VEGFR-3 and IF in a mouse model of lymphangiogenesis in regenerating skin. Specifically, a region of lymphangiogenesis was created by substituting a portion of mouse tail skin with a collagen gel within which lymphatic capillaries completely regenerate over a period of 60 days. The relative importance of IF and VEGF-C signaling were evaluated by either inhibiting VEGFR-3 signaling with antagonistic antibodies or by reducing IF. In some cases, VEGF-C signaling was then increased with exogenous protein. To clarify the role of IF, the distribution of endogenous matrix metalloproteinases (MMPs) and VEGF-C within the regenerating region was determined. It was found that inhibition of either VEGFR-3 or IF suppressed endogenous lymphangiogenesis. Reduction of IF was found to decrease lymphatic migration and transport of endogenous MMP and VEGF-C through the regenerating region. Therapeutic VEGF-C administration restored lymphangiogenesis following inhibition of VEGFR-3 but did not increase lymphangiogenesis following inhibition of IF. These results identify IF as an important regulator of the pro-lymphangiogenic action of VEGF-C.     

Epigenetic reprogramming during tissue regeneration

Epigenetic control of gene regulation is fundamental to the maintenance of cellular identities during all stages of metazoan life. Tissue regeneration involves cellular reprogramming processes, like dedifferentiation, re-differentiation, and trans-differentiation. Hence, in these processes epigenetic maintenance of gene expression programs requires a resetting through mechanisms that we are only beginning to understand. Here we summarize the current status of these studies, in particular regarding the role of epigenetic mechanisms of cellular reprogramming during tissue regeneration.     

Gene therapy for tissue regeneration

Tissue repair and regeneration are the normal biological responses of many different tissues in the body to injury. During the healing process, profound changes occur in cell composition and extracellular matrix (ECM) formation. Fibroblasts and equivalent reparative cells migrate to the wounded area and subsequently proliferate. These cells and reparative cells from the surrounding tissue are responsible for the rapid repair which results in tissue regeneration. Growth factors, one of which is transforming growth factor-beta (TGF-beta), stimulate fibroblasts and smooth muscle cells to proliferate and synthesize ECM proteins. This process of early repair provides a rapid way to restore new tissue and mechanical integrity. This early tissue repair process is normally followed by involution, which requires the production and activation of proteases, tissue maturation and remodeling, reorganization and finally regeneration. Alternately, failure to replace the critical components of the ECM, including elastin and basement membrane, results in abnormal regeneration of the epithelial cell layer. Although remodeling should occur during healing, provisional repair may be followed by excessive synthesis and deposition of collagen, which results in irreversible fibrosis and scarring. This excessive fibrosis which occurs in aberrant healing is at least in part mediated by persistent TGF-beta. Because of the central role of collagen in the wound healing process, the pharmacological control of collagen synthesis has been of paramount importance as a possible way to abrogate aberrant healing and prevent irreversible fibrosis. Fibrosis is an abnormal response to tissue injury.     

Cord blood for tissue regeneration

Umbilical cord blood (CB) has become a commonly accepted source of hematopoietic stem cells for transplantation in children and adults. It is readily available and outperforms bone marrow (BM) as well as peripheral blood stem cells in terms of tolerance for HLA‐mismatches between donor and recipient and its decreased graft‐versus‐host disease. Clinical use has been expanded from hematological malignancies to various areas such as treatment of metabolic genetic disorders or to induce angiogenesis. For the last years CB has been under intense experimental investigation in in vitro differentiation models as well as in preclinical animal models. Since CB‐derived stem cells offer multiple advantages over adult stem cells from other sources like BM, CB may provide a future source of stem cells for tissue repair and regeneration. To facilitate the use of CB‐derived stem cells in clinical scenarios, the biology of these cells needs to be further explored in detail particularly with regard to the fact that different non‐hematopoietic stem cell populations occur within CB. Here we explore the most consistent and the most contradictory data referring to the differentiation potential of CB‐derived stem cells and give an outlook on their potential clinical value including and possible reprogramming into IPS cells. J. Cell. Biochem. 108: 762–768, 2009. © 2009 Wiley‐Liss, Inc.