Rat neurosphere cells protect axotomized rat retinal ganglion cells and facilitate their regeneration

We investigated the ability of a population of rat neural stem and precursor cells derived from rat embryonic spinal cord to protect injured neurons in the rat central nervous system (CNS). The neonatal rat optic pathway was used as a model of CNS injury, whereby retinal ganglion cells (RGCs) were axotomized by lesion of the lateral geniculate nucleus one day after birth. Neural stem and precursor cells derived from expanded neurospheres (NS) were transplanted into the lesion site at the time of injury. Application of Fast Blue tracer dye to the lesion site demonstrated that significant numbers of RGCs survived at 4 and 8 weeks in animals that received a transplant, with an average of 28% survival, though in some individual cases survival was greater than 50%. No RGCs survived in animals that received a lesion alone. Furthermore, labeled RGCs were also observed when Fast Blue was applied to the superior colliculus (SC) at 4 weeks, suggesting that neurosphere cells also facilitated RGC to regenerate to their normal target. Transplanted cells did not migrate or express neural markers after transplantation, and secreted several neurotrophic factors in vitro. We conclude that NS cells can protect injured CNS neurons and promote their regeneration. These effects are not attributable to cell replacement, and may be mediated via secretion of neurotrophic factors. Thus, neuroprotection by stem cell populations may be a more viable approach for treatment of CNS disorders than cell replacement therapy.

Adult skeletal muscle stem cell migration is mediated by a blebbing/amoeboid mechanism

Adult skeletal muscle possesses a resident stem cell population called satellite cells which are responsible for tissue repair following damage. Satellite cell migration is crucial in promoting rapid tissue regeneration but is a poorly understood process. Furthermore, the mechanisms facilitating satellite cell movement have yet to be elucidated. Here the process of satellite cell migration has been investigated revealing that they undergo two distinct phases of movement; firstly under the basal lamina and then rapidly increasing their velocity when on the myofibre surface. Most significantly we show that satellite cells move using a highly dynamic blebbing based mechanism and not via lamellopodia mediated propulsion. We show that nitric oxide and non-canonical Wnt signalling pathways are necessary for regulating the formation of blebs and the migration of satellite cells. In summary, we propose that the formation of blebs and their necessity for satellite cell migration has significant implications in the future development of therapeutic regimes aimed at promoting skeletal muscle regeneration.

Limitations of the MRL mouse as a model for cardiac regeneration

Objective Myocardial repair following injury in mammals is restricted such that damaged areas are replaced by scar tissue, impairing cardiac function. MRL mice exhibit exceptional regenerative healing in an ear punch wound model. Some myocardial repair with restoration of heart function has also been reported following cryoinjury. Increased cardiomyocyte proliferation and a foetal liver stem cell population were implicated. We investigated molecular mechanisms facilitating myocardial repair in MRL mice to identify potential therapeutic targets in non-regenerative species. Methods Expressions of specific cell-cycle regulators that might account for regeneration (CDKs 1, 2, 4 and 6; cyclins A, E, D1 and B1; p21, p27 and E2F5) were compared by immunoblotting in MRL and control C57BL/6 ventricles during development. Flow cytometry was used to investigate stem cell populations in livers from foetal mice, and infarct sizes were compared in coronary artery-ligated and sham-treated MRL and C57BL/6 adult mice. Key findings No differences in the expressions of cell cycle regulators were observed between the two strains. Expressions of CD34+Sca1+ckit-, CD34+Sca1+ckit+ and CD34+Sca1-ckit+ increased in livers from C57BL/6 vs MRL mice. No differences were observed in infarct sizes, levels of fibrosis, Ki67 staining or cardiac function between MRL and C57BL/6 mice. Conclusions No intrinsic differences were observed in cell cycle control molecules or stem cell populations between MRL and control C57BL mouse hearts. Pathophysiologically relevant ischaemic injury is not repaired more efficiently in MRL myocardium, questioning the use of the MRL mouse as a reliable model for cardiac regeneration in response to pathophysiologically relevant forms of injury.

Age related changes in speed and mechanism of adult skeletal muscle stem cell migration

Skeletal muscle undergoes a progressive age-related loss in mass and function. Preservation of muscle mass depends in part on satellite cells, the resident stem cells of skeletal muscle. Reduced satellite cell function may contribute to the age-associated decrease in muscle mass. Here we focused on characterising the effect of age on satellite cell migration. We report that aged satellite cells migrate at less than half the speed of young cells. In addition, aged cells show abnormal membrane extension and retraction characteristics required for amoeboid based cell migration. Aged satellite cells displayed low levels of integrin expression. By deploying a mathematical model approach to investigate mechanism of migration, we have found that young satellite cells move in a random ‘memoryless’ manner whereas old cells demonstrate superdiffusive tendencies. Most importantly, we show that nitric oxide, a key regulator of cell migration, reversed the loss in migration speed and reinstated the unbiased mechanism of movement in aged satellite cells. Finally we found that although Hepatocyte Growth Factor increased the rate of aged satellite cell movement it did not restore the memoryless migration characteristics displayed in young cells. Our study shows that satellite cell migration, a key component of skeletal muscle regeneration, is compromised during aging. However, we propose clinically approved drugs could be used to overcome these detrimental changes.

Overexpression of coconut AINTEGUMENTA-like gene, CnANT, promotes in vitro regeneration in transgenic Arabidopsis

Knowledge of the molecular biological changes underlying the process of embryogenesis is important for the improvement of somatic embryogenesis of coconut. Among the transcription factors that control the transition from vegetative to embryogenic growth, members of APETALA2/Ethylene-responsive element binding protein domain family play an important role in promoting embryo development. Significant insights into the role of AP2 genes have been obtained by the ectopic expression of AP2 sub family genes in transgenic Arabidopsis. A homolog of the AINTEGUMENTA-like gene that encodes the two AP2 domains and the linker region was identified in the coconut genome. Phylogenetic analysis showed that this gene, CnANT, encodes a protein that branched with BABY BOOM/PLETHORA clade in the AINTEGUMENTA-like major clade and was similar to the oil palm EgAP2-1 protein. According to real time RT-PCR results, higher expression of CnANT was observed in more mature zygotic embryos. Also, high CnANT expression was recorded in embryogenic callus compared to other stages of somatic embryogenesis. We examined the effect of ectopic CnANT expression on the development and regenerative capacity of transgenic Arabidopsis. Overexpression of CnANT in Arabidopsis induced hormone free regeneration of explants. Furthermore, ectopic expression of CnANT enhanced regeneration in vitro and suggested a role for this gene in cell proliferation during in vitro culture.

Generation of Schwann cell-derived multipotent neurospheres isolated from intact sciatic nerve

Schwann cells (SCs) are the supporting cells of the peripheral nervous system and originate from the neural crest. They play a unique role in the regeneration of injured peripheral nerves and have themselves a highly unstable phenotype as demonstrated by their unexpectedly broad differentiation potential. Thus, SCs can be considered as dormant, multipotent neural crest-derived progenitors or stem cells. Upon injury they de-differentiate via cellular reprogramming, re-enter the cell cycle and participate in the regeneration of the nerve. Here we describe a protocol for efficient generation of neurospheres from intact adult rat and murine sciatic nerve without the need of experimental in vivo pre-degeneration of the nerve prior to Schwann cell isolation. After isolation and removal of the connective tissue, the nerves are initially plated on poly-D-lysine coated cell culture plates followed by migration of the cells up to 80% confluence and a subsequent switch to serum-free medium leading to formation of multipotent neurospheres. In this context, migration of SCs from the isolated nerve, followed by serum-free cultivation of isolated SCs as neurospheres mimics the injury and reprograms fully differentiated SCs into a multipotent, neural crest-derived stem cell phenotype. This protocol allows reproducible generation of multipotent Schwann cell-derived neurospheres from sciatic nerve through cellular reprogramming by culture, potentially marking a starting point for future detailed investigations of the de-differentiation process.

The Bone Formation Capabilities of the Anorganic Bone Matrix-Synthetic Cell-Binding Peptide 15 Grafts in an Animal Periodontal Model: A Histologic and Histomorphometric Study in Dogs

Background: The aim of this study is to verify the regenerative potential of particulate anorganic bone matrix synthetic peptide-15 (ABM-P-15) in class III furcation defects associated or not with expanded polytetrafluoroethylene membranes. Methods: Class III furcation defects were produced in the mandibular premolars (P2, P3, and P4) of six dogs and filled with impression material. The membranes and the bone grafts were inserted into P3 and P4, which were randomized to form the test and control groups, respectively; P2 was the negative control group. The animals were sacrificed 3 months post-treatment. Results: Histologically, the complete closure of class III furcation defects was not observed in any of the groups. Partial periodontal regeneration with similar morphologic characteristics among the groups was observed, however, through the formation of new cementum, periodontal ligament, and bone above the notch. Histologic analysis showed granules from the bone graft surrounded by immature bone matrix and encircled by newly formed tissue in the test group. The new bone formation area found in the negative control group was 2.28 +/- 2.49 mm(2) and in the test group it was 6.52 +/- 5.69 mm(2), which showed statistically significant differences for these groups considering this parameter (Friedman test P <0.05). There was no statistically significant difference among the negative control, control, and test groups for the other parameters. Conclusions: The regenerative potential of ABM-P-15 was demonstrated through new bone formation circumscribing and above the graft particles. The new bone also was accompanied by the formation of new cementum and periodontal ligament fibers. J Periodontol 2010;81:594-603.

In Vivo Comparison of Hard Tissue Regeneration with Human Mesenchymal Stem Cells Processed with Either the FICOLL Method or the BMAC Method

Objective: To compare new bone formation in maxillary sinus augmentation procedures using biomaterial associated with mesenchymal stem cells (MSCs) separated by two different isolation methods. Background: In regenerative medicine open cell concentration systems are only allowed for clinical application under good manufacturing practice conditions. Methods: Mononuclear cells, including MSCs, were concentrated with either the synthetic poylsaccharid (FICOLL) method (classic open system-control group, n = 6 sinus) or the bone marrow aspirate concentrate (BMAC) method (closed system-test group, n = 12 sinus) and transplanted in combination with biomaterial. A sample of the cells was characterized by their ability to differentiate. After 4.1 months (SD +/- 1.0) bone biopsies were obtained and analyzed. Results: The new bone formation in the BMAC group was 19.9% (90% confidence interval [CI], 10.9-29), and in the FICOLL group was 15.5% (90% CI, 8.6-22.4). The 4.4% difference was not significant (90% CI, -4.6-13.5; p = 0.39). MSCs could be differentiated into osteogenic, chondrogenic, and adipogenic lineages. Conclusion: MSCs harvested from bone marrow aspirate in combination with bovine bone matrix particles can form lamellar bone and provide a reliable base for dental implants. The closed BMAC system is suited to substitute the open FICOLL system in bone regeneration procedures.

Ridge Preservation With Acellular Dermal Matrix and Anorganic Bone Matrix Cell-Binding Peptide P-15 After Tooth Extraction in Humans

Background: Preventing ridge collapse with the extraction of maxillary anterior teeth is vital to an esthetic restorative result. Several regenerative techniques are available and are used for socket preservation. The aim of this study is to analyze by clinical parameters the use of acellular dermal matrix (ADM) and anorganic bovine bone matrix (ABM) with synthetic cell-binding peptide P-15 to preserve alveolar bone after tooth extraction. Methods: Eighteen patients in need of extraction of maxillary anterior teeth were selected and randomly assigned to the test group (ADM plus ABM/P-15) or the control group (ADM only). Clinical measurements were recorded initially and at 6 months after ridge-preservation procedures. Results: In the clinical measurements (external vertical palatal measurement [EVPM], external vertical buccal measurement [EVBM], and alveolar horizontal measurement [AHM]) the statistical analysis showed no difference between test and control groups initially and at 6 months. The intragroup analysis, after 6 months, showed a statistically significant reduction in the measurements for both groups. In the comparison between the two groups, the differences in the test group were as follows: EVPM = 0.83 +/- 1.53 mm; EVBM = 1.20 +/- 2.02 mm; and AHM = 2.53 +/- 1.81 mm. The differences in the control group were as follows: EVPM = 0.87 +/- 1.13 mm; EVBM = 1.50 +/- 1.15 mm; and AHM = 3.40 +/- 1.39 mm. The differences in EVPM and EVBM were not statistically significant; however, in horizontal measurement (AHM), there was a statistically significant difference (P<0.05). Conclusion: The results of this study show that ADM used as membrane associated with ABM/P-15 can be used to reduce buccal-palatal dimensions compared to ADM alone for preservation of the alveolar ridge after extraction of anterior maxillary teeth. J Periodontol 2011;82:72-79.

A novel regeneration system for a wild passion fruit species (Passiflora cincinnata Mast.) based on somatic embryogenesis from mature zygotic embryos

The objective of the present work was to induce somatic embryogenesis from zygotic embryos of Passiflora cincinnata Masters. Zygotic embryos formed calli on media with different concentrations of 2,4-dichlorophenoxyacetic acid (2,4-D) and 4.5 mu M benzyladenine (BA) after 30 days of in vitro culture. A concentration of 18.1 mu M 2,4-D resulted in the largest number of somatic embryos. Embryogenic calli were yellowish and friable, forming whitish proembryogenic masses. Morphologically, embryogenic cells were small and had large nuclei and dense cytoplasm, whereas non-embryogenic cells were elongated, with small nuclei and less dense cytoplasm. Calli cultured under white light on basal Murashige and Skoog`s medium with activated charcoal produced embryos in all developmental stages. There were differences among the treatments, with some leading to the production of calli with embryos and some only to callus formation. Some abnormalities were associated with somatic embryos, including fused axes, fused cotyledons and polycotyledonary embryos. Production of secondary somatic embryos occurred in the first cycle of primary embryo development. Secondary embryos differentiated from the surface of the protodermal layer of primary embryos with intense cell proliferation, successive mitotic divisions in the initial phase of embryoid development, and a vascular system formed with no connection to the parental tissue. This secondary embryogenic system of P. cincinnata is characterized by intense proliferation and maintenance of embryogenic competence after successive subcultures. This reproducible protocol opens new prospects for massive propagation and is an alternative to the current organogenesis-based transformation protocol.

Early modulation of inflammation by mesenchymal stem cell after acute kidney injury

Therapy with stem cells has showed to be promising for acute kidney injury (AKI), although how it works is still controversial. Modulation of the inflammatory response is one possible mechanism. Most of published data relies on early time and whether the protection is still maintained after that is not known. Here, we analyzed whether immune modulation continues after 24 h of reperfusion. MSC were obtained from male Wistar rats. After 3-5 passages, cells were screened for CD73, CD90, CD44, CD45, CD29 and CD 31. In addition, MSC were submitted to differentiation in adipocyte and in osteocyte. AKI was induced by bilaterally clamping of renal pedicles for 60 min. Six hours after injury, MSC (2 x 105 cells) were administered intravenously. MSC-treated animals presented the lowest serum creatinine compared to non-treated animals (24 h: 1.3 +/- 0.21 vs. 3.23 +/- 0.89 mg/dl, p<0.05). The improvement in renal function was followed by a lower expression of IL-1b, IL-6 and TNF-alpha and higher expression of IL-4 and IL-10. However, 48 h after reperfusion, this cytokine profile has changed. The decrease in Th1 cytokines was less evident and IL-6 was markedly up regulated. PCNA analysis showed that regeneration occurs faster in kidney tissues of MSC-treated animals than in controls at 24 h. And also ratio of Bcl-2/Bad was higher at treated animals after 24 and 48 h. Our data demonstrated that the immunomodulatory effects of MSC occur at very early time point, changing the inflammation profile toward a Th2 profile. (C) 2009 Elsevier B.V. All rights reserved.

Involvement of Eukaryotic Translation Initiation Factor 5A (eIF5A) in Skeletal Muscle Stem Cell Differentiation

The eukaryotic translation initiation factor 5A (eIF5A) contains a special amino acid residue named hypusine that is required for its activity, being produced by a post-translational modification using spermidine as substrate. Stem cells from rat skeletal muscles (satellite cells) were submitted to differentiation and an increase of eIF5A gene expression was observed. Higher content of eIF5A protein was found in satellite cells on differentiation in comparison to non-differentiated satellite cells and skeletal muscle. The treatment with NI-guanyl- 1,7-diaminoheptane (GC7), a hypusination inhibitor, reversibly abolished the differentiation process. In association with the differentiation blockage, an increase of glucose consumption and lactate production and a decrease of glucose and palmitic acid oxidation were observed. A reduction in cell proliferation and protein synthesis was also observed. L-Arginine, a spermidine precursor and partial suppressor of muscle dystrophic phenotype, partially abolished the GC7 inhibitory effect on satellite cell differentiation. These results reveal a new physiological role for eIF5A and contribute to elucidate the molecular mechanisms involved in muscle regeneration.

In vitro morphogenesis and cell suspension culture establishment in Piper solmsianum C. DC. (Piperaceae)

In vitro morphogenesis and cell suspension culture establishment in Piper solmsianum C. DC. (Piperaceae)). Piper solmsianum is a shrub from Southeast Brazil in which many biologically active compounds were identified. The aim of this work was to establish a cell suspension culture system for this species. With this in mind, petiole and leaf explants obtained from in vitro plantlets were cultured in the presence of different plant growth regulator combinations (IAA, NAA, 2,4-D and BA). Root and indirect shoot adventitious formation, detected by histological analysis, was observed. Besides the different combinations of plant growth regulators, light regime and the supplement of activated charcoal (1.5 mg.l(-1)) were tested for callus induction and growth. Cultures maintained in light, on a 0.2 mg.l(-1) 2,4-D and 2 mg.l(-1) BA supplemented medium, and in the absence of activated charcoal, showed the highest calli fresh matter increment. From a callus culture, cell suspension cultures were established and their growth and metabolite accumulation studied. The achieved results may be useful for further characterization of the activated secondary metabolites pathways in in vitro systems of P. solmsianum.

Functionalization of dendrimers for improved gene delivery to mesenchymal stem cell

Disease, injury, and age problems compromise human quality of life and continuously motivate the search for new and more efficacious therapeutic approaches. The field of Tissue Regeneration and Engineering has greatly evolved over the last years, mainly due to the combination of the important advances verified in Biomaterials Science and Engineering with those of Cell and Molecular Biology. In particular, a new and promising area arose – Nanomedicine – that takes advantage of the extremely small size and especial chemical and physical properties of Nanomaterials, offering powerful tools for health improvement. Research on Stem Cells, the self-renewing progenitors of body tissues, is also challenging to the medical and scientific communities, being expectable the appearance of new and exciting stem cell-based therapies in the next years. The control of cell behavior (namely, of cell proliferation and differentiation) is of key importance in devising strategies for Tissue Regeneration and Engineering. Cytokines, growth factors, transcription factors and other signaling molecules, most of them proteins, have been identified and found to regulate and support tissue development and regeneration. However, the application of these molecules in long-term regenerative processes requires their continuous presence at high concentrations as they usually present short half-lives at physiological conditions and may be rapidly cleared from the body. Alternatively, genes encoding such proteins can be introduced inside cells and be expressed using cell’s machinery, allowing an extended and more sustained production of the protein of interest (gene therapy). Genetic engineering of stem cells is particularly attractive because of their self-renewal capability and differentiation potential. For Tissue Regeneration and Engineering purposes, the patient’s own stem cells can be genetically engineered in vitro and, after, introduced in the body (with or without a scaffold) where they will not only modulate the behavior of native cells (stem cell-mediated gene therapy), but also directly participate in tissue repair. Cells can be genetically engineered using viral and non-viral systems. Viruses, as a result of millions of years of evolution, are very effective for the delivery of genes in several types of cells, including cells from primary sources. However, the risks associated with their use (like infection and immunogenic reactions) are driving the search for non-viral systems that will efficiently deliver genetic material into cells. Among them, chemical methods that are promising and being investigated use cationic molecules as carriers for DNA. In this case, gene delivery and gene expression level remain relatively low when primary cells are used. The main goal of this thesis was to develop and assess the in vitro potential of polyamidoamine (PAMAM) dendrimers based carriers to deliver genes to mesenchymal stem cells (MSCs). PAMAM dendrimers are monodispersive, hyperbranched and nanospherical molecules presenting unique characteristics that make them very attractive vehicles for both drug and gene delivery. Although they have been explored for gene delivery in a wide range of cell lines, the interaction and the usefulness of these molecules in the delivery of genes to MSCs remains a field to be explored. Adult MSCs were chosen for the studies due to their potential biomedical applications (they are considered multipotent cells) and because they present several advantages over embryonic stem cells, such as easy accessibility and the inexistence of ethical restrictions to their use. This thesis is divided in 5 interconnected chapters. Chapter I provides an overview of the current literature concerning the various non-viral systems investigated for gene delivery in MSCs. Attention is devoted to physical methods, as well as to chemical methods that make use of polymers (natural and synthetic), liposomes, and inorganic nanoparticles as gene delivery vectors. Also, it summarizes the current applications of genetically engineered mesenchymal stem cells using non-viral systems in regenerative medicine, with special focus on bone tissue regeneration. In Chapter II, the potential of native PAMAM dendrimers with amine termini to transfect MSCs is evaluated. The level of transfection achieved with the dendrimers is, in a first step, studied using a plasmid DNA (pDNA) encoding for the β-galactosidase reporter gene. The effect of dendrimer’s generation, cell passage number, and N:P ratio (where N= number of primary amines in the dendrimer; P= number of phosphate groups in the pDNA backbone) on the level of transfection is evaluated, being the values always very low. In a second step, a pDNA encoding for bone morphogenetic protein-2, a protein that is known for its role in MSCs proliferation and differentiation, is used. The BMP-2 content produced by transfected cells is evaluated by an ELISA assay and its effect on the osteogenic markers is analyzed through several classical assays including alkaline phosphatase activity (an early marker of osteogenesis), osteocalcin production, calcium deposition and mineralized nodules formation (late osteogenesis markers). Results show that a low transfection level is enough to induce in vitro osteogenic differentiation in MSCs. Next, from Chapter III to Chapter V, studies are shown where several strategies are adopted to change the interaction of PAMAM dendrimers with MSCs cell membrane and, as a consequence, to enhance the levels of gene delivery. In Chapter III, generations 5 and 6 of PAMAM dendrimers are surface functionalized with arginine-glycine-aspartic acid (RGD) containing peptides – experiments with dendrimers conjugated to 4, 8 and 16 RGD units were performed. The underlying concept is that by including the RGD integrin-binding motif in the design of the vectors and by forming RGD clusters, the level of transfection will increase as MSCs highly express integrins at their surface. Results show that cellular uptake of functionalized dendrimers and gene expression is enhanced in comparison with the native dendrimers. Furthermore, gene expression is dependent on both the electrostatic interaction established between the dendrimer moiety and the cell surface and the nanocluster RGD density. In Chapter IV, a new family of gene delivery vectors is synthesized consisting of a PAMAM dendrimer (generation 5) core randomly linked at the periphery to alkyl hydrophobic chains that vary in length and number. Herein, the idea is to take advantage of both the cationic nature of the dendrimer and the capacity of lipids to interact with biological membranes. These new vectors show a remarkable capacity for internalizing pDNA, being this effect positively correlated with the –CH2– content present in the hydrophobic corona. Gene expression is also greatly enhanced using the new vectors but, in this case, the higher efficiency is shown by the vectors containing the smallest hydrophobic chains. Finally, chapter V reports the synthesis, characterization and evaluation of novel gene delivery vectors based on PAMAM dendrimers (generation 5) conjugated to peptides with high affinity for MSCs membrane binding - for comparison, experiments are also done with a peptide with low affinity binding properties. These systems present low cytotoxicity and transfection efficiencies superior to those of native dendrimers and partially degraded dendrimers (Superfect®, a commercial product). Furthermore, with this biomimetic approach, the process of gene delivery is shown to be cell surface receptor-mediated. Overall, results show the potential of PAMAM dendrimers to be used, as such or modified, in Tissue Regeneration and Engineering. To our knowledge, this is the first time that PAMAM dendrimers are studied as gene delivery vehicles in this context and using, as target, a cell type with clinical relevancy. It is shown that the cationic nature of PAMAM dendrimers with amine termini can be synergistically combined with surface engineering approaches, which will ultimately result in suitable interactions with the cytoplasmic membrane and enhanced pDNA cellular entry and gene expression. Nevertheless, the quantity of pDNA detected inside cell nucleus is always very small when compared with the bigger amount reaching cytoplasm (accumulation of pDNA is evident in the perinuclear region), suggesting that the main barrier to transfection is the nuclear membrane. Future work can then be envisaged based on the versatility of these systems as biomedical molecular materials, such as the conjugation of PAMAM dendrimers to molecules able to bind nuclear membrane receptors and to promote nuclear translocation.

Studies of gastric mucosa regeneration and safety promoted by Mouriri pusa treatment in acetic acid ulcer model

Ethnopharmacological Relevance: Mouriri pusa Gardn. (Melastomataceae) is a medicinal plant commonly used by people living in the Cerrado to treat gastrointestinal disturbances. This medicinal plant has shown intense gastroprotective action in rodent gastric lesion, but still there are no data about its healing effect on gastric mucosa.Aim of the Study: To evaluate the methanolic extract (MeOH) obtained from Mouriri pusa leaves for its effect on the cicatrisation process of gastric ulcer.Mterials and Methods: The healing effects on gastric ulcers inducted by subserosal injection of acetic acid were evaluated by macroscopic and microscopic measures, imunohistochemistry and cell counting in rats treated with MeOH extract of Mouriri pusa (250 mg/kg, p.o./daily) for 14 or 30 days. The toxicity of Mouriri pusa was also evaluated by body and organ weight measure and clinical biochemical parameters.Results: Mouriri pusa treatments lasting 14 and 30 days showed elevated mucus secretion (PAS) and thicker regenerative gastric mucosa, denoting increased cell proliferation, which was confirmed by PCNA immunohistochemical analysis. Moreover, there was important cell recruitment (neutrophils and mast cells) to the site of the ulcer, which is an important factor in ulcer healing. No toxic effect was observed in all parameters evaluated. Phenolic compounds present in the MeOH extract like tannins, flavonoids and epicatechin are the probable agents involved in the healing effects of this medicinal plant.Conclusions: These findings showed a potential effect of Mouriri pusa in increasing regeneration of damaged gastric mucosa with safety for human use. (C) 2007 Elsevier B.V. All rights reserved.