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1.
Life Sci ; 248: 117465, 2020 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-32105707

RESUMO

BACKGROUND: Severe peripheral nerve injury leads to skeletal muscle atrophy and impaired limb function that is not sufficiently improved by existing treatments. Fibroblast growth factor 6 (FGF6) is involved in tissue regeneration and is dysregulated in denervated rat muscles. However, the way that FGF6 affects skeletal muscle repair after peripheral nerve injury has not been fully elucidated. METHODS: In this study, we investigated the role of FGF6 in the regeneration of denervated muscles using myoblast cells and an in vivo model of peripheral nerve injury. RESULTS: FGF6 promoted the viability and migration of C2C12 and primary myoblasts in a dose-dependent manner through FGFR1-mediated upregulation of cyclin D1. Low concentrations of FGF6 promoted myoblast differentiation through FGFR4-mediated activation of ERK1/2, which upregulated expression of MyHC, MyoD, and myogenin. FGFR-1, FGFR4, MyoD, and myogenin were not upregulated when FGF6 expression was inhibited in myoblasts by shRNA-mediated knockdown. Injection of FGF6 into denervated rat muscles enhanced the MyHC-IIb muscle fiber phenotype and prevented muscular atrophy. CONCLUSION: These findings indicate that FGF6 reduces skeletal muscle atrophy by relying on the ERK1/2 mechanism and enhances the conversion of slow muscle to fast muscle fibers, thereby promoting functional recovery of regenerated skeletal muscle after innervation.


Assuntos
Fator 6 de Crescimento de Fibroblastos/genética , Proteína Quinase 1 Ativada por Mitógeno/genética , Proteína Quinase 3 Ativada por Mitógeno/genética , Músculo Esquelético/metabolismo , Traumatismos dos Nervos Periféricos/genética , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/genética , Regeneração/genética , Animais , Diferenciação Celular , Linhagem Celular , Movimento Celular , Proliferação de Células , Ciclina D1/genética , Ciclina D1/metabolismo , Fator 6 de Crescimento de Fibroblastos/antagonistas & inibidores , Fator 6 de Crescimento de Fibroblastos/metabolismo , Regulação da Expressão Gênica , Masculino , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Denervação Muscular/métodos , Músculo Esquelético/inervação , Músculo Esquelético/patologia , Proteína MyoD/genética , Proteína MyoD/metabolismo , Mioblastos/metabolismo , Mioblastos/patologia , Miogenina/genética , Miogenina/metabolismo , Cadeias Pesadas de Miosina/genética , Cadeias Pesadas de Miosina/metabolismo , Traumatismos dos Nervos Periféricos/metabolismo , Traumatismos dos Nervos Periféricos/patologia , Cultura Primária de Células , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Ratos , Ratos Sprague-Dawley , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/metabolismo , Receptor Tipo 4 de Fator de Crescimento de Fibroblastos/genética , Receptor Tipo 4 de Fator de Crescimento de Fibroblastos/metabolismo , Nervo Isquiático/lesões
2.
Genes Dev ; 34(3-4): 194-208, 2020 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-31919191

RESUMO

Promoting axon regeneration in the central and peripheral nervous system is of clinical importance in neural injury and neurodegenerative diseases. Both pro- and antiregeneration factors are being identified. We previously reported that the Rtca mediated RNA repair/splicing pathway restricts axon regeneration by inhibiting the nonconventional splicing of Xbp1 mRNA under cellular stress. However, the downstream effectors remain unknown. Here, through transcriptome profiling, we show that the tubulin polymerization-promoting protein (TPPP) ringmaker/ringer is dramatically increased in Rtca-deficient Drosophila sensory neurons, which is dependent on Xbp1. Ringer is expressed in sensory neurons before and after injury, and is cell-autonomously required for axon regeneration. While loss of ringer abolishes the regeneration enhancement in Rtca mutants, its overexpression is sufficient to promote regeneration both in the peripheral and central nervous system. Ringer maintains microtubule stability/dynamics with the microtubule-associated protein futsch/MAP1B, which is also required for axon regeneration. Furthermore, ringer lies downstream from and is negatively regulated by the microtubule-associated deacetylase HDAC6, which functions as a regeneration inhibitor. Taken together, our findings suggest that ringer acts as a hub for microtubule regulators that relays cellular status information, such as cellular stress, to the integrity of microtubules in order to instruct neuroregeneration.


Assuntos
Anilidas/metabolismo , Axônios/fisiologia , Proteínas de Drosophila/metabolismo , Drosophila/fisiologia , Ácidos Hidroxâmicos/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Regeneração/genética , Animais , Proteínas de Drosophila/genética , Regulação da Expressão Gênica no Desenvolvimento/genética , Proteínas Associadas aos Microtúbulos/metabolismo , Microtúbulos/genética , Ligação Proteica , Processamento de RNA/genética , Células Receptoras Sensoriais/fisiologia
3.
J Photochem Photobiol B ; 202: 111714, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31830733

RESUMO

Planarian freshwater flatworms have the unique ability to regenerate due to stem cell activity. The process of regeneration is extremely sensitive to various factors, including light radiation. Here, the effect of low-intensity LED light of different wavelengths on regeneration, stem cell proliferation and gene expression associated with these processes was studied. LED matrices with different wavelengths (red (λmax = 635 nm), green (λmax = 520 nm) and blue (λmax = 463 nm), as well as LED laser diodes (red (λmax = 638.5 nm), green (λmax = 533 nm) and blue (λmax = 420 nm), were used in the experiments. Computer-assisted morphometry, whole-mount immunocytochemical study and RT-PCR were used to analyze the biological effects of LED light exposure on the planarian regeneration in vivo. It was found that a one-time exposure of regenerating planarians with low-intensity red light diodes stimulated head blastema growth in a dose-dependent manner (up to 40%). The green light exposure of planarians resulted in the opposite effect, showing a reduced head blastema growth rate by up to 21%. The blue light exposure did not lead to any changes in the rate of head blastema growth. The maximum effects of light exposure were observed at a dose of 175.2 mJ/cm2. No significant differences were revealed in the dynamics of neoblasts' (planarian stem cells) proliferation under red and green light exposure. However, the RT-PCR gene expression analysis of 46 wound-induced genes revealed their up-regulation upon red LED light exposure, and down-regulation upon green light exposure. Thus, we have demonstrated that the planarian regeneration process is rather sensitive to the effects of low-intensity light radiation of certain wavelengths, the biological activity of red and green light being dictated by the different expression of the genes regulating transcriptional activity.


Assuntos
Luz , Planárias/fisiologia , Regeneração/efeitos da radiação , Animais , Proliferação de Células/efeitos da radiação , Expressão Gênica/efeitos da radiação , Proteínas de Helminto/genética , Proteínas de Helminto/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Regeneração/genética , Células-Tronco/citologia
4.
Invest Ophthalmol Vis Sci ; 60(15): 4991-4999, 2019 12 02.
Artigo em Inglês | MEDLINE | ID: mdl-31794598

RESUMO

Purpose: Genomic reprogramming and cellular dedifferentiation are critical to the success of de novo tissue regeneration in lower vertebrates such as zebrafish and axolotl. In tissue regeneration following injury or disease, differentiated cells must retain lineage while assuming a progenitor-like identity in order to repopulate the damaged tissue. Understanding the epigenetic regulation of programmed cellular dedifferentiation provides unique insights into the biology of stem cells and cancer and may lead to novel approaches for treating human degenerative conditions. Methods: Using a zebrafish in vivo model of adult muscle regeneration, we utilized chromatin immunoprecipitation followed by massively parallel DNA sequencing (ChIP-seq) to characterize early changes in epigenetic signals, focusing on three well-studied histone modifications-histone H3 trimethylated at lysine 4 (H3K4me3), and histone H3 trimethylated or acetylated at lysine 27 (H3K27me3 and H3K27Ac, respectively). Results: We discovered that zebrafish myocytes undergo a global, rapid, and transient program to drive genomic remodeling. The timing of these epigenetic changes suggests that genomic reprogramming itself represents a distinct sequence of events, with predetermined checkpoints, to generate cells capable of de novo regeneration. Importantly, we uncovered subsets of genes that maintain epigenetic marks paradoxical to changes in expression, underscoring the complexity of epigenetic reprogramming. Conclusions: Within our model, histone modifications previously associated with gene expression act for the most part as expected, with exceptions suggesting that zebrafish chromatin maintains an easily editable state with a number of genes paradoxically marked for transcriptional activity despite downregulation.


Assuntos
Reprogramação Celular/genética , DNA/genética , Epigênese Genética , Músculos Oculomotores/fisiologia , Regeneração/genética , Animais , Imunoprecipitação da Cromatina , Histonas/genética , Modelos Animais , Regiões Promotoras Genéticas , Análise de Sequência de DNA , Peixe-Zebra
5.
C R Biol ; 342(9-10): 291-298, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31786144

RESUMO

Cell regeneration is a natural repair of different types of tissue after an injury or a lesion, and is associated with asexual reproduction in some animals such as planarians. Its understanding and improvement could have repercussions for tissue repair and regeneration as far as humans are concerned. In this context, we have proceeded to an essential step, which is the identification of the genes involved in planarian regeneration in the model species. Dugesia sicula Lepori (D. sicula) is distributed around the Mediterranean Sea, and this population is found in most of Tunisian dams. The collection of identified genes is already known in other species. DjFoxG, DjPC2, DjotxA, and Cathepsin-D were identified by the PCR technique and their expression was confirmed by RT-PCR and in situ hybridization. DjFoxG gene, the FoxG1 homolog, is expressed throughout the planarian body, abundantly on stem cells. Consecutively, we choose a central nervous system (CNS) marker; the prohormone convertase 2 (DjPC2) gene. DjotxA was observed in the brain and especially in the region surrounding the eyes (visual cells). The regenerative cells of the gut of D. sicula were scored by the Cathepsin-D gene expression, which belongs to the aspartyl protease family. We found significant results through RT-PCR and In Situ Hybridization (ISH) techniques, confirming the expression of DjFoxG, DjPC2, DjotxA and Cathepsin-D genes in our specimens.


Assuntos
Planárias/genética , Regeneração/genética , Animais , Encéfalo/fisiologia , Olho/metabolismo , Mar Mediterrâneo , Tunísia
6.
Nat Med ; 25(11): 1691-1698, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31700187

RESUMO

Millions of people worldwide with incurable end-stage lung disease die because of inadequate treatment options and limited availability of donor organs for lung transplantation1. Current bioengineering strategies to regenerate the lung have not been able to replicate its extraordinary cellular diversity and complex three-dimensional arrangement, which are indispensable for life-sustaining gas exchange2,3. Here we report the successful generation of functional lungs in mice through a conditional blastocyst complementation (CBC) approach that vacates a specific niche in chimeric hosts and allows for initiation of organogenesis by donor mouse pluripotent stem cells (PSCs). We show that wild-type donor PSCs rescued lung formation in genetically defective recipient mouse embryos unable to specify (due to Ctnnb1cnull mutation) or expand (due to Fgfr2cnull mutation) early respiratory endodermal progenitors. Rescued neonates survived into adulthood and had lungs functionally indistinguishable from those of wild-type littermates. Efficient chimera formation and lung complementation required newly developed culture conditions that maintained the developmental potential of the donor PSCs and were associated with global DNA hypomethylation and increased H4 histone acetylation. These results pave the way for the development of new strategies for generating lungs in large animals to enable modeling of human lung disease as well as cell-based therapeutic interventions4-6.


Assuntos
Pneumopatias/terapia , Pulmão/crescimento & desenvolvimento , Células-Tronco Pluripotentes/metabolismo , Regeneração/genética , Acilação/genética , Animais , Blastocisto/metabolismo , Diferenciação Celular/genética , Metilação de DNA/genética , Modelos Animais de Doenças , Histonas/genética , Humanos , Pulmão/patologia , Pneumopatias/patologia , Camundongos , Organogênese/genética , Células-Tronco Pluripotentes/transplante , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/genética , beta Catenina/genética
7.
Nat Commun ; 10(1): 4898, 2019 10 25.
Artigo em Inglês | MEDLINE | ID: mdl-31653858

RESUMO

Osteoarthritis (OA) is a prevalent degenerative disease, which involves progressive and irreversible destruction of cartilage matrix. Despite efforts to reconstruct cartilage matrix in osteoarthritic joints, it has been a difficult task as adult cartilage exhibits marginal repair capacity. Here we report the identification of tankyrase as a regulator of the cartilage anabolism axis based on systems-level factor analysis of mouse reference populations. Tankyrase inhibition drives the expression of a cartilage-signature matrisome and elicits a transcriptomic pattern that is inversely correlated with OA progression. Furthermore, tankyrase inhibitors ameliorate surgically induced OA in mice, and stem cell transplantation coupled with tankyrase knockdown results in superior regeneration of cartilage lesions. Mechanistically, the pro-regenerative features of tankyrase inhibition are mainly triggered by uncoupling SOX9 from a poly(ADP-ribosyl)ation (PARylation)-dependent protein degradation pathway. Our findings provide insights into the development of future OA therapies aimed at reconstruction of articular cartilage.


Assuntos
Cartilagem Articular/efeitos dos fármacos , Condrócitos/metabolismo , Matriz Extracelular/efeitos dos fármacos , Transplante de Células-Tronco Mesenquimais , Osteoartrite do Joelho/metabolismo , Poli ADP Ribosilação/efeitos dos fármacos , Fatores de Transcrição SOX9/efeitos dos fármacos , Tanquirases/antagonistas & inibidores , Animais , Cartilagem Articular/metabolismo , Cartilagem Articular/fisiologia , Simulação por Computador , Inibidores Enzimáticos , Matriz Extracelular/metabolismo , Técnicas de Silenciamento de Genes , Células HEK293 , Humanos , Camundongos , Osteoartrite/genética , Osteoartrite/metabolismo , Osteoartrite do Joelho/genética , Poli ADP Ribosilação/fisiologia , Ratos , Regeneração/genética , Fatores de Transcrição SOX9/metabolismo , Tanquirases/genética , Tanquirases/metabolismo
8.
Nat Commun ; 10(1): 4659, 2019 10 11.
Artigo em Inglês | MEDLINE | ID: mdl-31604915

RESUMO

MG53 is a muscle-specific TRIM-family protein that presides over the cell membrane repair response. Here, we show that MG53 present in blood circulation acts as a myokine to facilitate tissue injury-repair and regeneration. Transgenic mice with sustained elevation of MG53 in the bloodstream (tPA-MG53) have a healthier and longer life-span when compared with littermate wild type mice. The tPA-MG53 mice show normal glucose handling and insulin signaling in skeletal muscle, and sustained elevation of MG53 in the bloodstream does not have a deleterious impact on db/db mice. More importantly, the tPA-MG53 mice display remarkable dermal wound healing capacity, enhanced muscle performance, and improved injury-repair and regeneration. Recombinant human MG53 protein protects against eccentric contraction-induced acute and chronic muscle injury in mice. Our findings highlight the myokine function of MG53 in tissue protection and present MG53 as an attractive biological reagent for regenerative medicine without interference with glucose handling in the body.


Assuntos
Proteínas de Membrana/fisiologia , Cicatrização , Animais , Cálcio/metabolismo , Glucose/metabolismo , Teste de Tolerância a Glucose , Insulina/metabolismo , Proteínas de Membrana/sangue , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Músculo Esquelético/metabolismo , Regeneração/genética , Biologia de Sistemas
9.
PLoS Genet ; 15(10): e1008408, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31626629

RESUMO

Satellite cells (SCs) are muscle stem cells that remain quiescent during homeostasis and are activated in response to acute muscle damage or in chronic degenerative conditions such as Duchenne Muscular Dystrophy. The activity of SCs is supported by specialized cells which either reside in the muscle or are recruited in regenerating skeletal muscles, such as for instance macrophages (MΦs). By using a dystrophic mouse model of transient MΦ depletion, we describe a shift in identity of muscle stem cells dependent on the crosstalk between MΦs and SCs. Indeed MΦ depletion determines adipogenic conversion of SCs and exhaustion of the SC pool leading to an exacerbated dystrophic phenotype. The reported data could also provide new insights into therapeutic approaches targeting inflammation in dystrophic muscles.


Assuntos
Diferenciação Celular/genética , Macrófagos/metabolismo , Distrofia Muscular de Duchenne/genética , Regeneração/genética , Animais , Linhagem da Célula/genética , Modelos Animais de Doenças , Distrofina/genética , Humanos , Macrófagos/patologia , Camundongos , Camundongos Endogâmicos mdx , Músculo Esquelético/metabolismo , Músculo Esquelético/patologia , Distrofia Muscular de Duchenne/metabolismo , Distrofia Muscular de Duchenne/patologia , Mioblastos/metabolismo , Células Satélites de Músculo Esquelético/metabolismo , Células Satélites de Músculo Esquelético/patologia
10.
PLoS Genet ; 15(10): e1008401, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31626630

RESUMO

Wnt signaling regulates primary body axis formation across the Metazoa, with high Wnt signaling specifying posterior identity. Whether a common Wnt-driven transcriptional program accomplishes this broad role is poorly understood. We identified genes acutely affected after Wnt signaling inhibition in the posterior of two regenerative species, the planarian Schmidtea mediterranea and the acoel Hofstenia miamia, which are separated by >550 million years of evolution. Wnt signaling was found to maintain positional information in muscle and regional gene expression in multiple differentiated cell types. sp5, Hox genes, and Wnt pathway components are down-regulated rapidly after ß-catenin RNAi in both species. Brachyury, a vertebrate Wnt target, also displays Wnt-dependent expression in Hofstenia. sp5 inhibits trunk gene expression in the tail of planarians and acoels, promoting separate tail-trunk body domains. A planarian posterior Hox gene, Post-2d, promotes normal tail regeneration. We propose that common regulation of a small gene set-Hox, sp5, and Brachyury-might underlie the widespread utilization of Wnt signaling in primary axis patterning across the Bilateria.


Assuntos
Padronização Corporal/genética , Genes Homeobox/genética , Planárias/genética , Regeneração/genética , Animais , Diferenciação Celular/genética , Regulação da Expressão Gênica no Desenvolvimento/genética , Desenvolvimento Muscular/genética , Proteínas Nucleares/genética , Planárias/crescimento & desenvolvimento , Proteínas Wnt/genética , Via de Sinalização Wnt/genética
11.
PLoS Genet ; 15(10): e1008354, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31589606

RESUMO

Recent evidence implicates mononuclear diploid cardiomyocytes as a proliferative and regenerative subpopulation of the postnatal heart. The number of these cardiomyocytes is a complex trait showing substantial natural variation among inbred mouse strains based on the combined influences of multiple polymorphic genes. One gene confirmed to influence this parameter is the cardiomyocyte-specific kinase Tnni3k. Here, we have studied Tnni3k alleles across a number of species. Using a newly-generated kinase-dead allele in mice, we show that Tnni3k function is dependent on its kinase activity. In an in vitro kinase assay, we show that several common human TNNI3K kinase domain variants substantially compromise kinase activity, suggesting that TNNI3K may influence human heart regenerative capacity and potentially also other aspects of human heart disease. We show that two kinase domain frameshift mutations in mice cause loss-of-function consequences by nonsense-mediated decay. We further show that the Tnni3k gene in two species of mole-rat has independently devolved into a pseudogene, presumably associated with the transition of these species to a low metabolism and hypoxic subterranean life. This may be explained by the observation that Tnni3k function in mice converges with oxidative stress to regulate mononuclear diploid cardiomyocyte frequency. Unlike other studied rodents, naked mole-rats have a surprisingly high (30%) mononuclear cardiomyocyte level but most of their mononuclear cardiomyocytes are polyploid; their mononuclear diploid cardiomyocyte level (7%) is within the known range (2-10%) of inbred mouse strains. Naked mole-rats provide further insight on a recent proposal that cardiomyocyte polyploidy is associated with evolutionary acquisition of endothermy.


Assuntos
Evolução Molecular , Cardiopatias/genética , Proteínas Serina-Treonina Quinases/genética , Alelos , Animais , Diferenciação Celular/genética , Linhagem da Célula/genética , Proliferação de Células/genética , Cardiopatias/metabolismo , Ventrículos do Coração/crescimento & desenvolvimento , Ventrículos do Coração/metabolismo , Humanos , Mutação com Perda de Função/genética , Camundongos , Ratos-Toupeira/genética , Miocárdio/metabolismo , Miócitos Cardíacos/metabolismo , Estresse Oxidativo/genética , Poliploidia , Regeneração/genética
12.
Nat Commun ; 10(1): 4402, 2019 09 27.
Artigo em Inglês | MEDLINE | ID: mdl-31562306

RESUMO

T lymphocytes must be produced throughout life, yet the thymus, where T lymphocytes are made, exhibits accelerated atrophy with age. Even in advanced atrophy, however, the thymus remains plastic, and can be regenerated by appropriate stimuli. Logically, thymic atrophy is thought to reflect senescent cell death, while regeneration requires proliferation of stem or progenitor cells, although evidence is scarce. Here we use conditional reporters to show that accelerated thymic atrophy reflects contraction of complex cell projections unique to cortical epithelial cells, while regeneration requires their regrowth. Both atrophy and regeneration are independent of changes in epithelial cell number, suggesting that the size of the thymus is regulated primarily by rate-limiting morphological changes in cortical stroma, rather than by their cell death or proliferation. Our data also suggest that cortical epithelial morphology is under the control of medullary stromal signals, revealing a previously unrecognized endocrine-paracrine signaling axis in the thymus.


Assuntos
Células Epiteliais/metabolismo , Regeneração/genética , Células Estromais/metabolismo , Linfócitos T/metabolismo , Timo/metabolismo , Animais , Atrofia/genética , Atrofia/metabolismo , Perfilação da Expressão Gênica/métodos , Ontologia Genética , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Microscopia Confocal , Tamanho do Órgão/genética , Regeneração/fisiologia , Timo/patologia , Timo/fisiopatologia
13.
Int J Mol Sci ; 20(18)2019 Sep 10.
Artigo em Inglês | MEDLINE | ID: mdl-31509934

RESUMO

In addition to its role as an endocrine messenger, growth hormone (GH) also acts as a neurotrophic factor in the central nervous system (CNS), whose effects are involved in neuroprotection, axonal growth, and synaptogenic modulation. An increasing amount of clinical evidence shows a beneficial effect of GH treatment in patients with brain trauma, stroke, spinal cord injury, impaired cognitive function, and neurodegenerative processes. In response to injury, Müller cells transdifferentiate into neural progenitors and proliferate, which constitutes an early regenerative process in the chicken retina. In this work, we studied the long-term protective effect of GH after causing severe excitotoxic damage in the retina. Thus, an acute neural injury was induced via the intravitreal injection of kainic acid (KA, 20 µg), which was followed by chronic administration of GH (10 injections [300 ng] over 21 days). Damage provoked a severe disruption of several retinal layers. However, in KA-damaged retinas treated with GH, we observed a significant restoration of the inner plexiform layer (IPL, 2.4-fold) and inner nuclear layer (INL, 1.5-fold) thickness and a general improvement of the retinal structure. In addition, we also observed an increase in the expression of several genes involved in important regenerative pathways, including: synaptogenic markers (DLG1, NRXN1, GAP43); glutamate receptor subunits (NR1 and GRIK4); pro-survival factors (BDNF, Bcl-2 and TNF-R2); and Notch signaling proteins (Notch1 and Hes5). Interestingly, Müller cell transdifferentiation markers (Sox2 and FGF2) were upregulated by this long-term chronic GH treatment. These results are consistent with a significant increase in the number of BrdU-positive cells observed in the KA-damaged retina, which was induced by GH administration. Our data suggest that GH is able to facilitate the early proliferative response of the injured retina and enhance the regeneration of neurite interconnections.


Assuntos
Hormônio do Crescimento/farmacologia , Ácido Caínico/toxicidade , Regeneração/efeitos dos fármacos , Retina/efeitos dos fármacos , Animais , Animais Recém-Nascidos , Fator Neurotrófico Derivado do Encéfalo/genética , Embrião de Galinha , Galinhas , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Neurogênese/efeitos dos fármacos , Neurogênese/genética , Neurogênese/fisiologia , Fármacos Neuroprotetores/farmacologia , Neurotoxinas/toxicidade , Receptor Notch1/genética , Regeneração/genética , Regeneração/fisiologia , Retina/metabolismo , Retina/fisiopatologia , Fatores de Transcrição SOXB1/genética
14.
Nat Commun ; 10(1): 4365, 2019 09 25.
Artigo em Inglês | MEDLINE | ID: mdl-31554796

RESUMO

Epithelia are exposed to diverse types of stress and damage from pathogens and the environment, and respond by regenerating. Yet, the proximal mechanisms that sense epithelial damage remain poorly understood. Here we report that p38 signaling is activated in adult Drosophila midgut enterocytes in response to diverse stresses including pathogenic bacterial infection and chemical and mechanical insult. Two upstream kinases, Ask1 and Licorne (MKK3), are required for p38 activation following infection, oxidative stress, detergent exposure and wounding. Ask1-p38 signaling in enterocytes is required upon infection to promote full intestinal stem cell (ISC) activation and regeneration, partly through Upd3/Jak-Stat signaling. Furthermore, reactive oxygen species (ROS) produced by the NADPH oxidase Nox in enterocytes, are required for p38 activation in enterocytes following infection or wounding, and for ISC activation upon infection or detergent exposure. We propose that Nox-ROS-Ask1-MKK3-p38 signaling in enterocytes integrates multiple different stresses to induce regeneration.


Assuntos
Proteínas de Drosophila/metabolismo , Intestinos/fisiopatologia , MAP Quinase Quinase 3/metabolismo , MAP Quinase Quinase Quinases/metabolismo , NADPH Oxidases/metabolismo , Regeneração/fisiologia , Transdução de Sinais , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Animais , Animais Geneticamente Modificados , Infecções Bacterianas/microbiologia , Proteínas de Drosophila/genética , Drosophila melanogaster/genética , Drosophila melanogaster/metabolismo , Enterócitos/metabolismo , Enterócitos/microbiologia , Mucosa Intestinal/metabolismo , Mucosa Intestinal/microbiologia , Mucosa Intestinal/fisiopatologia , Intestinos/microbiologia , Intestinos/patologia , MAP Quinase Quinase 3/genética , MAP Quinase Quinase Quinases/genética , NADPH Oxidases/genética , Estresse Oxidativo , Regeneração/genética , Células-Tronco/metabolismo , Células-Tronco/microbiologia , Estresse Mecânico , Proteínas Quinases p38 Ativadas por Mitógeno/genética
15.
Nat Commun ; 10(1): 4368, 2019 09 25.
Artigo em Inglês | MEDLINE | ID: mdl-31554819

RESUMO

The colonic epithelial turnover is driven by crypt-base stem cells that express the R-spondin receptor Lgr5. Signals that regulate epithelial regeneration upon stem cell injury are largely unknown. Here, we explore the dynamics of Wnt signaling in the colon. We identify two populations of cells with active Wnt signaling: highly proliferative Lgr5+/Axin2+ cells, as well as secretory Lgr5-/Axin2+ cells. Upon Lgr5+ cell depletion, these cells are recruited to contribute to crypt regeneration. Chemical injury induced by DSS leads to a loss of both Lgr5+ cells and Axin2+ cells and epithelial regeneration is driven by Axin2- cells, including differentiated Krt20+ surface enterocytes. Regeneration requires stromal Rspo3, which is present at increased levels upon injury and reprograms Lgr5- but Lgr4+ differentiated cells. In contrast, depletion of stromal Rspo3 impairs crypt regeneration, even upon mild injury. We demonstrate that Rspo3 is essential for epithelial repair via induction of Wnt signaling in differentiated cells.


Assuntos
Colo/fisiologia , Mucosa Intestinal/fisiologia , Regeneração/fisiologia , Células-Tronco/metabolismo , Trombospondinas/metabolismo , Animais , Proteína Axina/genética , Proteína Axina/metabolismo , Diferenciação Celular/genética , Colite/genética , Colite/metabolismo , Colo/metabolismo , Enterócitos/metabolismo , Perfilação da Expressão Gênica/métodos , Mucosa Intestinal/metabolismo , Queratina-20/genética , Queratina-20/metabolismo , Camundongos Knockout , Camundongos Transgênicos , Receptores Acoplados a Proteínas-G/genética , Receptores Acoplados a Proteínas-G/metabolismo , Regeneração/genética , Células-Tronco/citologia , Trombospondinas/genética , Via de Sinalização Wnt/genética
16.
Zygote ; 27(5): 329-336, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31412969

RESUMO

Mammalian Pou5f1 encodes the POU family class V (POU-V) transcription factor which is essential for the pluripotency of embryonic cells and germ cells. In vertebrates, various POU-V family genes have been identified and classified into the POU5F1 family or its paralogous POU5F3 family. In this study, we cloned two cDNAs named CpPou5f1 and CpPou5f3, which encode POU-V family proteins of the Japanese red bellied newt Cynops pyrrhogaster. In the predicted amino acid sequence encoded by CpPou5f1, the typical MAGH sequence at the N-terminus and deletion of arginine at the fifth position of POU-homeodomain were recognized, but not in the sequence encoded by CpPou5f3. Phylogenetic analysis using Clustal Omega software indicated that CpPou5f1 and CpPou5f3 are classified into the clade of the POU5F1 and POU5F3 families, respectively. In a real-time polymerase chain reaction (RT-PCR) analysis, the marked gene expression of CpPou5f1 was observed during oogenesis and early development up to the tail-bud stage, whereas weak gene expression of CpPou5f3 was detected only in the early stages of oogenesis and gastrula. In adult organs, CpPou5f1 was expressed only in the ovary, while gene expression of CpPou5f3 was recognized in various organs. A regeneration experiment using larval forelimb revealed that transient gene expression of CpPou5f1 occurred at the time of wound healing, followed by gene activation of CpPou5f3 during the period of blastema formation. These results suggest that CpPou5f1 and CpPou5f3 might play different roles in embryogenesis and limb regeneration.


Assuntos
Oogênese/genética , Fatores do Domínio POU/genética , Regeneração/genética , Salamandridae/genética , Animais , Embrião não Mamífero/fisiologia , Extremidades/fisiologia , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Masculino , Família Multigênica , Filogenia , Salamandridae/embriologia , Salamandridae/fisiologia
17.
Genetics ; 213(2): 555-566, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31444245

RESUMO

In larval zebrafish, melanocyte stem cells (MSCs) are quiescent, but can be recruited to regenerate the larval pigment pattern following melanocyte ablation. Through pharmacological experiments, we found that inhibition of γ-aminobutyric acid (GABA)-A receptor function, specifically the GABA-A ρ subtype, induces excessive melanocyte production in larval zebrafish. Conversely, pharmacological activation of GABA-A inhibited melanocyte regeneration. We used clustered regularly interspaced short palindromic repeats/Cas9 to generate two mutant alleles of gabrr1, a subtype of GABA-A receptors. Both alleles exhibited robust melanocyte overproduction, while conditional overexpression of gabrr1 inhibited larval melanocyte regeneration. Our data suggest that gabrr1 signaling is necessary to maintain MSC quiescence and sufficient to reduce, but not eliminate, melanocyte regeneration in larval zebrafish.


Assuntos
Larva/genética , Melanócitos/metabolismo , Receptores de GABA-A/genética , Peixe-Zebra/genética , Animais , Diferenciação Celular/genética , Divisão Celular/genética , Larva/crescimento & desenvolvimento , Pigmentação/genética , Regeneração/genética , Transdução de Sinais/genética , Células-Tronco/metabolismo , Peixe-Zebra/crescimento & desenvolvimento , Ácido gama-Aminobutírico/genética
18.
BMC Plant Biol ; 19(1): 373, 2019 Aug 24.
Artigo em Inglês | MEDLINE | ID: mdl-31445524

RESUMO

BACKGROUND: The ability of severed rootstocks and shoots to re-establish vascular connections is used to generate grafted plants that combine desirable traits from both scions and rootstocks. Clarifying the mechanisms of graft healing is essential for its further application. We performed RNA sequencing of internodes near the cut position, making a distinction between separated or grafted tissues above and below the cut, in order to obtain a genetic description of graft union formation. RESULTS: Using weighted gene co-expression analysis, variable transcripts were clustered into 10 distinct co-expression networks (modules) based on expression profiles, and genes with the most "hubness" ("hub" genes show the most connections in a network) within each module were predicted. A large proportion of modules were related to Position, and represent asymmetric expression networks from different pathways. Expression of genes involved in auxin and sugar transport and signaling, and brassinosteroid biosynthesis was increased above the cut, while stress response genes were up-regulated below the cut. Some modules were related to graft union formation, among which oxidative detoxification genes were co-expressed along with both wounding response and cell wall organization genes. CONCLUSIONS: The present work provides a comprehensive understanding of graft healing-related gene networks in tomato. Also, the candidate pathways and hub genes identified here will be valuable for future studies of grafting in tomato.


Assuntos
Lycopersicon esculentum/fisiologia , Regeneração/genética , Perfilação da Expressão Gênica , Regulação da Expressão Gênica de Plantas , Redes Reguladoras de Genes , Lycopersicon esculentum/genética , Raízes de Plantas/fisiologia , Transdução de Sinais
19.
Mol Med Rep ; 20(2): 1819-1825, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31257532

RESUMO

Tree shrews are most closely related to the primates and so possess a number of advantages in experimental studies; they have been used as an animal model in bacterial and virus infection, cancer, endocrine system disease, and certain nervous system diseases. Their olfactory ensheathing cells (OECs) are able to release several cytokines to promote neuronal survival, regeneration and remyelination. The present study used western blot analysis to identify antibody specificity in protein extracts from whole tree shrew brains to identify the specificity of p75 nerve growth factor receptor (NGFR) derived from rabbits (75 kDa). OECs were cultured and isolated, then stained and identified using the antibodies for p75NGFR. To investigate the capacity of OECs to express cytokines and growth factors, microarray technology was used, and the analysis revealed that OECs were able to express 9,821 genes. Of these genes, 44 genes were from the neurotrophic factor family, which may indicate their potential in transplantation in vivo. The present study considered the function of OECs as revealed by other studies, and may contribute to future research.


Assuntos
Neurônios/metabolismo , Bulbo Olfatório/metabolismo , Receptor de Fator de Crescimento Neural/genética , Tupaia/genética , Animais , Anticorpos/imunologia , Citocinas/biossíntese , Regulação da Expressão Gênica/genética , Humanos , Neuroglia/metabolismo , Bulbo Olfatório/citologia , Regeneração/genética , Remielinização/genética , Tupaia/crescimento & desenvolvimento , Tupaia/metabolismo
20.
BMC Res Notes ; 12(1): 425, 2019 Jul 16.
Artigo em Inglês | MEDLINE | ID: mdl-31311592

RESUMO

OBJECTIVE: Passion fruit improvement efforts by conventional breeding have had limited success calling for research into alternative approaches such as tissue culture and genetic engineering. An efficient and reproducible regeneration system is a prerequisite for successful genetic engineering. Currently, there is no reliable regeneration system for Uganda's passion fruit varieties owing to the high heterogeneity of the Passiflora genus. Therefore, this study aimed at establishing an efficient and reproducible regeneration system for Uganda's Passiflora edulis f. flavicarpa (yellow passion fruit) and Passiflora edulis f. edulis (purple passion fruit) for routine utilization with an ultimate goal of improving its agronomic value. RESULTS: The study successfully induced shoots by both direct and indirect organogenesis for the yellow passion fruit variety. Highest shoot induction frequency (14.85%) was achieved on 8.9 µM BAP while 7.9 µM BAP did not initiate any shoots. Optimal shoot elongation and rooting was achieved on 0.44 µM BAP and 5.37 µM α-naphthaleneacetic (NAA) respectively. Rooted yellow passion fruit plantlets were successfully weaned with over 65% survival rates. It took approximately 6 months to produce a weaned healthy passion fruit plant. The purple passion fruit variety proved to be recalcitrant to tissue culture with no successful shoot or callus induction.


Assuntos
Engenharia Genética/métodos , Passiflora/fisiologia , Folhas de Planta/fisiologia , Regeneração/fisiologia , Técnicas de Cultura de Tecidos/métodos , Ácidos Naftalenoacéticos/farmacologia , Organogênese/efeitos dos fármacos , Organogênese/genética , Passiflora/classificação , Passiflora/genética , Folhas de Planta/genética , Raízes de Plantas/efeitos dos fármacos , Raízes de Plantas/genética , Raízes de Plantas/crescimento & desenvolvimento , Brotos de Planta/genética , Brotos de Planta/crescimento & desenvolvimento , Regeneração/genética , Especificidade da Espécie , Uganda
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