RESUMO
Colorectal cancer (CRC) is one of the most commonly diagnosed malignant tumors of the digestive tract. H2-calponin (CNN2), an actin cytoskeleton-binding protein, is an isoform of the calponin protein family whose role in CRC is still unknown. Research based on clinical samples showed the up-regulation of CNN2 in CRC and its association with tumor development, metastasis, and poor prognosis of patients. Both in vitro loss-of-function and gain-of-function experiments showed that CNN2 participates in CRC development through influencing malignant cell phenotypes. In vivo, xenografts formed by CNN2 knockdown cells also showed a slower growth rate and smaller final tumors. Furthermore, EGR1 was identified as a downstream of CNN2, forming a complex with CNN2 and YAP1 and playing an essential role in the CNN2-induced regulation of CRC development. Mechanistically, CNN2 knockdown down-regulated EGR1 expression through enhancing its ubiquitination, thus decreasing its protein stability in a YAP1-dependent manner. In summary, CNN2 plays an EGR1-dependent promotion role in the development and progression of CRC, which may be a promising therapeutic target for CRC treatment.
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Neoplasias Colorretais , Proteínas dos Microfilamentos , Humanos , Proteínas dos Microfilamentos/genética , Proteínas dos Microfilamentos/metabolismo , Proteínas de Ligação ao Cálcio/metabolismo , Neoplasias Colorretais/metabolismo , Ubiquitinação , Proteína 1 de Resposta de Crescimento Precoce/genética , Proteína 1 de Resposta de Crescimento Precoce/metabolismo , CalponinasRESUMO
AIMS: Engineered conduction tissues (ECTs) fabricated from cardiac progenitor cells (CPCs) and collagen sponges were precisely targeted for the treatment of atrioventricular conduction block in our previous studies. However, obvious shrinkage and deformation of ECTs was observed during in vitro culture. According to the literature, it can be speculated that basic fibroblast growth factor (bFGF) may downregulate alpha-smooth muscle actin (α-SMA) produced by CPCs to prevent the shrinkage of CPC-engineered conduction tissues. MAIN METHODS: In this study, culture media with or without bFGF were used for both cell culture and 3D tissue construction. The expression of α-SMA and the size change of engineered tissue were analyzed to evaluate the feasibility of adding bFGF to regulate α-SMA expression and shrinkage of constructs. In addition, cardiac-specific examinations were performed to evaluate the effect of bFGF on cardiac tissue formation. KEY FINDINGS: Supplementation with bFGF efficiently relieved shrinkage of engineered tissue by downregulating the expression of α-SMA at both the cellular and 3D tissue levels. Moreover, bFGF had a positive influence on cardiac tissue formation in terms of cell viability, tissue organization and electrical conduction velocity. SIGNIFICANCE: This study provides a guide for both shape control and quality improvement of CPC-engineered cardiac tissues.
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Actinas/genética , Meios de Cultura/farmacologia , Fator 2 de Crescimento de Fibroblastos/farmacologia , Miocárdio/citologia , Células-Tronco/citologia , Engenharia Tecidual/métodos , Animais , Tamanho Celular/efeitos dos fármacos , Células Cultivadas , Regulação para Baixo/efeitos dos fármacos , Feminino , Ratos Sprague-Dawley , Células-Tronco/efeitos dos fármacos , Células-Tronco/metabolismo , Alicerces Teciduais/químicaRESUMO
Motor neuron disease (MND) is a kind of common clinical nervous system disease with typical characteristic of progressive motor neurons degeneration or death. Motor neuron derived from stem cells or motor neuron progenitor cells will be a good choice to be used for treatment of the disease. In this study, we used the combination of 5 small molecular including CHIR99021 (CHIR), SB431542 (SB), DMH1 (DMH), retinoic acid (RA) and Purmorphamine (Pur) to induce hair follicles neural crest stem cells (hfNCSCs) to motor neurons progenitors (MNPs). Valproic acid (VPA) was used to make MNPs proliferation. RA and Pur were used to try to induce MNPs toward motor neurons (MNs) and CpdE was tried for MNs maturation. Nestin, ß-tubulin Ш (Tuj1), microtubule associated protein 2 (MAP2), Olig2, choline acetyltransferase (ChAT)and TUBB3 were examined at protein and mRNA levels by immunofluoresence cytochemistry, western blot and real time PCR at 6, 16 and 22 days. Our data showed cells changed into bipolar or multipolar shape forming the cell clusters like scattered rosettes. Nestin expression decreased significantly at 22 days. Compared to 6 days, percentage of Olig2 + MNPs was higher, (88.53 ± 6.67)%, and Olig2 expression at protein and gene level was lower at 22 days. Percentage of MAP2 positive cells increased to (90.62 ± 2.31) % and ChAT positive cells increased to (83.29 ± 6.62) % at 22 days. But no expression of ChAT was examined by western blot and real time PCR. It indicates that these 5 molecular can differentiate hfNCSCs into Olig2 positive cells with a unipotent differentiation toward motor neurons.
Assuntos
Folículo Piloso/metabolismo , Crista Neural/metabolismo , Células-Tronco Neurais/metabolismo , Fator de Transcrição 2 de Oligodendrócitos/metabolismo , Animais , Diferenciação Celular/fisiologia , Colina O-Acetiltransferase/metabolismo , Folículo Piloso/citologia , Masculino , Crista Neural/citologia , Células-Tronco Neurais/citologia , Ratos , Ratos Sprague-DawleyRESUMO
OBJECTIVE: Colorectal cancer (CRC) remains a major cause of cancer-related death worldwide. Proteasome 26S subunit ATPase 2 (PSMC2) plays vital roles in regulating cell cycle and transcription and has been confirmed to be a gene potentially associated with some human tumors. However, the expression correlation and molecular mechanism of PSMC2 in CRC are still unclear. This study aimed to investigate the role of PSMC2 in malignant behaviors in CRC. METHODS: The high protein levels of PSMC2 in CRC samples were identified by tissue microarray analysis. Lentivirus was used to silence PSMC2 in HCT116 and RKO cells; MTT and colony formation assay were performed to determine cell proliferation. Wound healing and Transwell assay were used to detect cell migration and invasion. Flow cytometry assay was applied to detect cell cycle and apoptosis. RESULT: The results showed that, among the 96 CRC patients, the expression of PSMC2 was a positive correlation with the clinicopathological features of the patients with CRC. Furthermore, the low PSMC2 expression group showed a higher survival rate than the high PSMC2 expression group. The expression levels of PSMC2 in cancer tissue were dramatically upregulated compared with adjacent normal tissues. In vitro, shPSMC2 was designed to inhibit the expression of PSMC2 in CRC cells. Compared with shCtrl, silencing of PSMC2 significantly suppressed cell proliferation, decreased single cell colony formation, enhanced apoptosis, and accelerated G2 phase and/or S phase arrest. CONCLUSION: Survival analysis indicated that high expression of PSMC2 in the CRC samples was associated with poorer survival rate than low expression of PSMC2, while the anti-tumor effect of PSMC2 silencing was also confirmed at the cellular level in vitro. Our results suggested that PSMC2 potentially worked as a regulator for CRC, and the silencing of PSMC2 may be a therapeutic strategy for CRC.
Assuntos
ATPases Associadas a Diversas Atividades Celulares/metabolismo , Apoptose , Proliferação de Células , Neoplasias Colorretais/patologia , Complexo de Endopeptidases do Proteassoma/metabolismo , Interferência de RNA , ATPases Associadas a Diversas Atividades Celulares/antagonistas & inibidores , ATPases Associadas a Diversas Atividades Celulares/genética , Idoso , Linhagem Celular Tumoral , Movimento Celular , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/mortalidade , Feminino , Humanos , Estimativa de Kaplan-Meier , Masculino , Prognóstico , Complexo de Endopeptidases do Proteassoma/genética , RNA Interferente Pequeno/metabolismo , Pontos de Checagem da Fase S do Ciclo CelularRESUMO
The present study aimed to investigate the efficacy of transplantation of bone marrow neural tissue-committed stem cell-derived sensory neuron-like cells for the repair of peripheral nerve sensory impairments in rats. Bone marrow was isolated and cultured to obtain the neural tissue-committed stem cells (NTCSCs), and the differentiation of these cells into sensory neuron-like cells was induced. Bone marrow mesenchymal stem cells (BMSCs), bone marrow NTCSCs, and bone marrow NTCSC-derived sensory neurons (NTCSC-SNs) were transplanted by microinjection into the L4 and L5 dorsal root ganglions (DRGs) in an animal model of sensory defect. On the 2nd, 4th, 8th, and 12th week after the transplantation, the effects of the three types of stem cells on the repair of the sensory functional defect were analyzed via behavioral observation, sensory function evaluation, electrophysiological examination of the sciatic nerve, and morphological observation of the DRGs. The results revealed that the transplanted BMSCs, NTCSCs, and NTCSC-SNs were all able to repair the sensory nerves. In addition, the effect of the NTCSC-SNs was significantly better than that of the other two types of stem cells. The general posture and gait of the animals in the sensory defect model exhibited evident improvement over time. Plantar temperature sensitivity and pain sensitivity gradually recovered, and the sensation latency was reduced, with faster sensory nerve conduction velocity. Transplantation of NTCSC-SNs can improve the repair of peripheral nerve sensory defects in rats.
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Transplante de Células-Tronco Mesenquimais , Células-Tronco Mesenquimais/citologia , Regeneração Nervosa , Tecido Nervoso/citologia , Traumatismos dos Nervos Periféricos/fisiopatologia , Traumatismos dos Nervos Periféricos/terapia , Células Receptoras Sensoriais/transplante , Potenciais de Ação , Animais , Comportamento Animal , Separação Celular , Forma Celular , Sobrevivência Celular , Modelos Animais de Doenças , Masculino , Proteínas do Tecido Nervoso/metabolismo , Condução Nervosa , Neurônios/citologia , Traumatismos dos Nervos Periféricos/patologia , Ratos Sprague-Dawley , Células Receptoras Sensoriais/citologia , Esferoides Celulares/citologiaRESUMO
AIMS: Transplantation of a tissue engineered cardiac pacemaker (TECP) may represent a novel therapy for cardiac sinus node dysfunction. We previously reported that cardiac progenitor cells (CPCs) derived from embryonic heart tubes could differentiate into cardiac pacemaking cells after endothelin-1 treatment. We aimed to examine the feasibility of TECP fabricated from CPCs-derived pacemaking cells and vascularization of TECP fabricated from CPCs-derived pacemaking cells and endothelial progenitor cells (EPCs) in vitro and in vivo implantation. MAIN METHODS: TECP created using CPCs-derived pacemaking cells and vTECP created by mixing CPCs and EPCs in vitro were implanted into rat hearts. Sinus node damaged was induced by formaldehyde insult. KEY RESULTS: Spontaneous beating tissues, namely TECP, were obtained after seeding CPCs-derived pacemaking cells into Matrigel. ECG and epicardial multielectrode array (MEA) measurements confirmed implanted TECP have electrical activity. TECP implantation promoted individual survival in sinus node damage models (15/22 animals lived versus 0/17 control). vTECP fabricated by mixing the both EPCs and CPCs-derived pacemaking cells with Matrigel in equal proportions optimally formed pre-vascularization in vitro. The implantation of vTECP enhanced electrical activity in vivo, which may correlate with increased vascularization. PI3K-Akt-VEGF/VEGFR signaling was involved with vascular ingrowth in vTECP. SIGNIFICANCE: Our data supports the therapeutic potential of TECP fabricated with the CPCs-derived pacemaking cells for sinus node dysfunction. Vascularization by the addition of EPCs is an important factor to sustain viability of the TECP in vivo.
Assuntos
Colágeno , Células Progenitoras Endoteliais/citologia , Laminina , Proteoglicanas , Síndrome do Nó Sinusal/terapia , Células-Tronco/citologia , Engenharia Tecidual/métodos , Animais , Diferenciação Celular , Modelos Animais de Doenças , Combinação de Medicamentos , Endotelina-1/química , Masculino , Ratos , Ratos Sprague-Dawley , Transdução de SinaisRESUMO
Objective. The aim of this research is to evaluate the protective effects of methane-rich saline (MS) on lipopolysaccharide- (LPS-) induced acute lung injury (ALI) and investigate its potential antioxidative, anti-inflammatory, and antiapoptotic activities. Methods. LPS-induced (20 mg/kg) ALI rats were injected with MS (2 ml/kg and 20 ml/kg) before the initiation of LPS induction. Survival rate was determined until 96 h after LPS was induced. Lung injury was assayed by oxygenation index, lung permeability index (LPI), wet-to-dry weight (W/D), and histology. The cells in the bronchoalveolar lavage fluid (BALF) were counted. Oxidative stress was examined by the level of malondialdehyde (MDA) and superoxide dismutase (SOD). Inflammatory factors including tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß), and interleukin-6 (IL-6) in BALF were determined by ELISA. Lung tissue apoptosis was detected by TUNEL staining and western blotting of caspase-3. Results. It was found that methane significantly prolonged the rat survival, decreased the lung W/D ratio and the content of the inflammatory factors, and reduced the amount of caspase-3 and apoptotic index. In addition, MS increased the level of SOD and decreased the level of MDA significantly. Conclusions. MS protects the LPS-challenged ALI via antioxidative, anti-inflammatory, and antiapoptotic effect, which may prove to be a novel therapy for the clinical management of ALI.
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Lesão Pulmonar Aguda/induzido quimicamente , Lesão Pulmonar Aguda/tratamento farmacológico , Metano/uso terapêutico , Substâncias Protetoras/uso terapêutico , Cloreto de Sódio/uso terapêutico , Lesão Pulmonar Aguda/patologia , Lesão Pulmonar Aguda/fisiopatologia , Animais , Antioxidantes/farmacologia , Apoptose/efeitos dos fármacos , Líquido da Lavagem Broncoalveolar , Citocinas/metabolismo , Lipopolissacarídeos , Pulmão/efeitos dos fármacos , Pulmão/patologia , Pulmão/fisiopatologia , Pulmão/ultraestrutura , Masculino , Metano/farmacologia , Neutrófilos/efeitos dos fármacos , Neutrófilos/metabolismo , Tamanho do Órgão , Permeabilidade , Substâncias Protetoras/farmacologia , Ratos Sprague-Dawley , Cloreto de Sódio/farmacologia , Regulação para Cima/efeitos dos fármacosRESUMO
A cell-sourced biological pacemaker is a promising therapeutic approach for sick sinus syndrome (SSS) or severe atrial ventricular block (AVB). Adipose tissue-derived stem cells (ATSCs), which are optimal candidate cells for possible use in regenerative therapy for acute or chronic myocardial injury, have the potential to differentiate into spontaneous beating cardiomyocytes. However, the pacemaker characteristics of the beating cells need to be confirmed, and little is known about the underlying differential mechanism. In this study, we found that brown adipose tissue-derived stem cells (BATSCs) in mice could differentiate into spontaneous beating cells in 15% FBS Dulbecco's modified Eagle's medium (DMEM) without additional treatment. Subsequently, we provide additional evidence, including data regarding ultrastructure, protein expression, electrophysiology, and pharmacology, to support the differentiation of BATSCs into a cardiac pacemaker phenotype during the course of early cultivation. Furthermore, we found that silencing Tbx18, a key transcription factor in the development of pacemaker cells, terminated the differentiation of BATSCs into a pacemaker phenotype, suggesting that Tbx18 is required to direct BATSCs toward a cardiac pacemaker fate. The expression of Tbx3 and shox2, the other two important transcription factors in the development of pacemaker cells, was decreased by silencing Tbx18, which suggests that Tbx18 mediates the differentiation of BATSCs into a pacemaker phenotype via these two downstream transcription factors.
Assuntos
Tecido Adiposo Marrom/metabolismo , Diferenciação Celular , Sistema de Condução Cardíaco/metabolismo , Células-Tronco/metabolismo , Proteínas com Domínio T/metabolismo , Tecido Adiposo Marrom/citologia , Animais , Sistema de Condução Cardíaco/citologia , Camundongos , Células-Tronco/citologia , Proteínas com Domínio T/genéticaRESUMO
There is no clinical report on the use of natural orifice transluminal endoscopic surgery (NOTES) for the management of patients with large liver cysts.This study aims to evaluate the feasibility and safety of NOTES for liver cyst fenestration in humans using a currently available technique.From February 2009 to June 2010, 4 cases of transgastric endoscopic liver cyst fenestration were performed; in which 3 cases received NOTES only, while 1 case received additional laparoscopic assistance.Mean time to endoscopically locate the liver cyst was 16âminutes (5-22âminutes). Cysts that were present in the left lobe or on the liver surface were easier to locate endoscopically. Transgastric endoscopic liver cyst fenestration was successful in all patients. The use of an occlusion balloon helped in the endoscopic clipping of the gastrotomy incision. Mean operative time was 101.3âminutes (range, 90-112âminutes), and there were no intra- or postoperative complications including infections. All patients recovered well after the surgery, with only minor postoperative throat pain. There was no recurrence at a mean follow-up of 12 months (range, 6-48 months).Small sample size.It may be technically feasible and safe to perform transgastric endoscopic liver cyst fenestration in humans with no recurrence at follow up.
Assuntos
Cistos/cirurgia , Hepatopatias/cirurgia , Cirurgia Endoscópica por Orifício Natural/métodos , Adulto , Idoso , Idoso de 80 Anos ou mais , Estudos de Viabilidade , Feminino , Humanos , Fígado/cirurgia , Masculino , Pessoa de Meia-Idade , Cirurgia Endoscópica por Orifício Natural/efeitos adversos , Duração da CirurgiaRESUMO
Tissue-engineered nerve conduits are widely used for the study of peripheral nerve injury repair. With regard to repairing long nerve defects, stem-cell-derived neurons are recommended as seed cells. As hair-follicle neural crest stem cells (hfNCSCs) are easily to be harvested from patients and have the potential to differentiate into neuronal cells, hfNCSCs-derived neurons are an ideal candidate choice. Acellular nerve grafts, a type of biological material scaffold, with intact collagen structure, with biocompatibility and less toxicity are obtained through removing live cells with 1 % lysolecithin, are also an ideal choice. In the present report, we describe a tissue-engineered nerve conduit seeded with rat hfNCSCs-derived neurons into the beagle acellular sciatic nerve scaffold. Our goal is to provide a novel engineered therapeutic for repairing peripheral nerve injury with long distance defects.
Assuntos
Folículo Piloso/citologia , Regeneração Nervosa , Crista Neural/citologia , Células-Tronco Neurais/citologia , Neurônios/citologia , Engenharia Tecidual , Animais , Células Cultivadas , Células-Tronco Neurais/metabolismo , Traumatismos dos Nervos Periféricos/terapia , Ratos , Alicerces TeciduaisRESUMO
Background. Mesenchymal stem cells are the most commonly used seed cells in biomedical research and tissue engineering. Their secretory proteins have also been proven to play an important role in tissue healing. Methods. We isolated adipose stem cells and placental stem cells and performed analysis examining characteristics. The secretory proteins were extracted from conditioned medium and analyzed by MALDI-TOF/TOF. The antiaging effect of conditioned mediums was evaluated by the results of facial skin application. Results. Adipose stem cells and placental stem cells were found to be very similar in their surface markers and multipotency. The specific proteins secreted from adipose stem cells were more adept at cell adhesion, migration, wound healing, and tissue remodeling, while the proteins secreted by placental stem cells were more adept at angiogenesis, cell proliferation, differentiation, cell survival, immunomodulation, and collagen degradation. While these two types of conditioned medium could improve the facial index, the improvement of Melanin index after injection of the adipose stem cell conditioned medium was much more significant. Conclusion. The results suggest that the secreted proteins are ideal cell-free substances for regeneration medicine, especially in the antiaging field.
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Clinical treatment of chronic deep venous insufficiency remains difficult despite the availability of various therapies. Previous experimental efforts have demonstrated that the tissue-engineered valvedvenous conduit (TEVV) is a promising option to replace the damaged venous valve. The aim of the present study was to evaluate the TEVV by reseeding bone marrow-derived endothelial progenitor cells and multipotent adult progenitor cells into acellular matrix according to International Standard ISO10993, and to clarify their interactions with blood, the local effect after implantation both in vitro and vivo, and immunogenicity. The results showed that the 2-cm long TEVV did not cause haemolysis in vitro and remained patent without thrombosis formation in vivo. However, the luminal surface of TEVV was partially covered by multilayer cells. Compared with the native ovine femoral vein segment, the TEVV beneath the mouse skin produced significant mononuclear cell infiltration, with serum interleukin-6 and tumour necrosis factor-α similar to normal. The TEVV maintained its structural integrity, while the native ovine femoral vein segments fell apart at postoperative week nine. The TEVV implantation did not change serum immunoglobulin G. In addition, the seeds and extracts of the scaffold did not affect the proliferation of mouse lymphocytes. These findings suggest that the histocompatibility, haemocompatibility and immunogenicity of this TEVV are acceptable owing to complete removal of the cellular components of autologous seeds and residues of chemical regents, thus providing an experimental basis for further clinical translation. Copyright © 2014 John Wiley & Sons, Ltd.
Assuntos
Prótese Vascular , Células da Medula Óssea/metabolismo , Células Progenitoras Endoteliais/metabolismo , Matriz Extracelular/química , Veia Femoral , Animais , Autoenxertos , Células da Medula Óssea/citologia , Células Progenitoras Endoteliais/citologia , Feminino , Camundongos , Camundongos Endogâmicos BALB C , Coelhos , OvinosRESUMO
Tissue engineering has been considered a promising approach for creating grafts to replace autologous venous valves. Here, ovine bone marrow-derived endothelial progenitor cells (EPCs) and multipotent adult progenitor cells (MAPCs) were harvested and then loaded into decellularized venous matrix to create tissue-engineered (TE) valved vein. Subsequently, the ovine femoral veins containing the valve were removed and replaced by TE grafts or acellular matrix only. The morphology and function were analysed for up to 1 year by ultrasonography, angiography, H&E staining and scanning electron microscopy (SEM). The differentiation of seeded cells was traced immunofluorochemically. The results showed that decellularized venous matrix could initially and feebly attract endogenous cells, but failed afterwards and were insufficient to restore valve function. On the contrary, the seeded cells differentiated into endothelial cells (ECs) in vivo and formed a monolayer endothelium, and smooth muscle cells within the scaffold therefore produced TE grafts comparable to the native vein valve. This TE graft remained patent and sufficient after implantation into the venous circuit of the ovine lower extremity for at least 6 months. Unfortunately, cells seeded on the luminal surface and both sides of the leaflets lost their biological functions at 12 months, resulting in thrombosis formation and leading to complete occlusion of the TE grafts and impotent venous valves. These findings suggest that this TE valved venous conduit can function physiologically in vivo in the medium term. Before translating this TE venous valve into clinical practice, the durability should be improved and thrombogenicity should be suppressed. Copyright © 2016 John Wiley & Sons, Ltd.
Assuntos
Células da Medula Óssea/metabolismo , Diferenciação Celular , Células Progenitoras Endoteliais/metabolismo , Matriz Extracelular/química , Animais , Células da Medula Óssea/citologia , Células Progenitoras Endoteliais/citologia , Veia Femoral/citologia , Veia Femoral/metabolismo , Músculo Liso Vascular/citologia , Músculo Liso Vascular/metabolismo , Miócitos de Músculo Liso/citologia , Miócitos de Músculo Liso/metabolismo , OvinosRESUMO
Stem cellbased cell therapy has provided a promising method for the treatment of pulmonary diseases, including idiopathic pulmonary fibrosis (IPF). Furthermore, adiposederived stem cells (ADSCs) have been reported to be effective in lung repair and regeneration. In the current study, IPF was induced in mice by intratracheal instillation of bleomycin (BLM), and ADSCs were delivered systemically into the mice via the tail vein to evaluate the effects of ADSC transplantation. The ADSC engraftment rate and morphometric changes in lung tissue samples in vivo were investigated by histochemistry and immunohistochemistry, as well as by western blotting. The results indicated that ADSCs may relieve IPF and provide a significant contribution to lung repair when administered at an early stage.
Assuntos
Tecido Adiposo/citologia , Fibrose Pulmonar Idiopática/patologia , Transplante de Células-Tronco , Células-Tronco/citologia , Células Epiteliais Alveolares/metabolismo , Células Epiteliais Alveolares/patologia , Animais , Apoptose , Biomarcadores , Líquido da Lavagem Broncoalveolar , Citocinas/metabolismo , Modelos Animais de Doenças , Hidroxiprolina/metabolismo , Fibrose Pulmonar Idiopática/metabolismo , Fibrose Pulmonar Idiopática/terapia , Imuno-Histoquímica , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Camundongos , Transplante de Células-Tronco/métodosRESUMO
Effective repair of peripheral nerve defects is difficult because of the slow growth of new axonal growth. We propose that "neural-like cells" may be useful for the protection of peripheral nerve destructions. Such cells should prolong the time for the disintegration of spinal nerves, reduce lesions, and improve recovery. But the mechanism of neural-like cells in the peripheral nerve is still unclear. In this study, bone marrow-derived neural-like cells were used as seed cells. The cells were injected into the distal end of severed rabbit peripheral nerves that were no longer integrated with the central nervous system. Electromyography (EMG), immunohistochemistry, and transmission electron microscopy (TEM) were employed to analyze the development of the cells in the peripheral nerve environment. The CMAP amplitude appeared during the 5th week following surgery, at which time morphological characteristics of myelinated nerve fiber formation were observed. Bone marrow-derived neural-like cells could protect the disintegration and destruction of the injured peripheral nerve.
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Spinal cord injury (SCI) refers to the damage of spinal cord's structure and function due to a variety of causes. At present, many scholars have confirmed that apoptosis is the main method of secondary injury in spinal cord injury. In view of understanding the function of PI3K/Akt pathway on spinal cord injury, this study observed the temporal variation of key molecules (PI3K, Akt, p-Akt) in the PI3K/Akt pathway after spinal cord injury by immunohistochemistry and Western-blot. The results showed that the expression of PI3K, Akt and p-Akt display a sharp increase one day after the spinal cord injury, and then it decreased gradually with the time passing by, but the absolute expression was certainly higher than the normal group. These results indicate that the PI3K/Akt signaling pathway is involved in the spinal cord injury and the mechanism may be related to apoptosis.
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Studies have showed that transplanted stem cells in the inner ear won't regenerate to replace the damaged sensory hair cells. They can spontaneously differentiate into mesenchymal cells and fibrocytes in the damaged inner ear. Only mature sensory cells of MSCs-derived possess the great potency for cell transplantation in the treatment of sensorineural hearing loss. So, we try to establish an efficient generation of the glutamatergic sensory neural phenotype for the cell transplantation of the hearing loss. We isolated MSCs from femoral and tibial bones according to their adherence to culture dishes. After purification, proliferation, and passaged, cells became homogeneous in appearance, showing more uniformity and grew in a monolayer with a typical spindle-shape morphology. The cell surface markers were assessed using FACS to characterize the isolated cells. For neural induction to harvest the glutamatergic sensory neurons, passage 3 MSCs were incubated with preinduced medium for 24 hr, and neural-induced medium for an additional 14 days. The cells exhibit a typical neural shape. RT-PCR analysis indicated that the mRNA levels of the neural cell marker nestin, Tau, MAP-2, ß-tubulin III, GluR-3, and GluR-4 were higher compared with primary MSCs. Immunohistochemistry and western-blotting proofed that nestin, MAP-2, ß-tubulin III, and GluR-4 proteins indeed exhibit their expression difference in the induced cells compared to the MSCs. We show an efficient protocol by the combined applications of Sonic Hedgehog (Shh) and Retinoic Acid (RA) to induce MSCs to differentiate into the glutamatergic sensory neuron which were identified from the morphological, biochemical, and molecular characteristics.
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Proteínas Hedgehog/imunologia , Células-Tronco Mesenquimais/efeitos dos fármacos , Células-Tronco Neurais/efeitos dos fármacos , Tretinoína/farmacologia , Animais , Diferenciação Celular/efeitos dos fármacos , Diferenciação Celular/imunologia , Masculino , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/imunologia , Células-Tronco Neurais/citologia , Células-Tronco Neurais/imunologia , Ratos , Ratos Sprague-Dawley , Relação Estrutura-AtividadeRESUMO
PURPOSE: The objective of this study was to provide the morphological details on small branches of the portal vein in transverse groove of hepatic hilum. METHODS: According to the surgery significance, the small branches of portal vein in transverse groove of hepatic hilum were named as "Short hepatic portal veins (SHPVs)". SHPVs were minutely dissected in 30 adult cadaveric livers. The number, diameter, length, origin points, and entering liver sites of SHPVs were explored and measured. RESULTS: There were 181 SHPVs in 30 liver specimens, including 46% (83/181) from the left portal vein, 31% (56/181) from the bifurcation, and 23% (42/181) from the right portal vein. At the entering liver sites of SHPVs, 22% (40/181) supplied for segment IV, 9% (17/181) for segment V, 4% (7/181) for segment VI, 23% (41/181) for segment VII, and 42% (76/181) for segment I (caudate lobe). There were 6.0 ± 2.4 branches per liver specimen with range 3-12. The mean diameter of SHPVs was 2.25 ± 0.89 mm. The average length of SHPVs was 4.86 ± 2.12 mm. CONCLUSIONS: SHPVs widely existed in each liver specimen. The detailed anatomical study of SHPVs could be useful to avoid damaging the short portal branches during hepatic operations, such as isolated or combined caudate lobectomy.
Assuntos
Veia Porta/anatomia & histologia , Adulto , Ductos Biliares/anatomia & histologia , Pesos e Medidas Corporais/métodos , Cadáver , Feminino , Humanos , Fígado/anatomia & histologia , Fígado/irrigação sanguínea , Masculino , Pessoa de Meia-Idade , Adulto JovemRESUMO
Our previous study showed that after being treated with 5-azacytidine, Nkx2.5(+) human cardiac progenitor cells (CPCs) derived from embryonic heart tubes could differentiate into cardiomyocytes. Although 5-azacytidine is a classical agent that induces myogenic differentiation in various types of cells, the drug is toxic and unspecific for myogenic differentiation. To investigate the possibility of inducing CPCs to differentiate into cardiomyocytes by a specific and non-toxic method, CPCs of passage 15 and mesenchymal stem cells (MSCs) were treated with cardiac ventricular fibroblast-conditioned medium (CVF-conditioned medium). Following this treatment, the Nkx2.5(+) CPCs underwent cardiomyogenic differentiation. Phase-contrast microscopy showed that the morphology of the treated CPCs gradually changed. Ultrastructural observation confirmed that the cells contained typical sarcomeres. The expression of cardiomyocyte-associated genes, such as alpha-cardiac actin, cardiac troponin T, and beta-myosin heavy chain (MHC), was increased in the CPCs that had undergone cardiomyogenic differentiation compared with untreated cells. In contrast, the MSCs did not exhibit changes in morphology or molecular expression after being treated with CVF-conditioned medium. The results indicated that Nkx2.5(+) CPCs treated with CVF-conditioned medium were capable of differentiating into a cardiac phenotype, whereas treated MSCs did not appear to undergo cardiomyogenic differentiation. Subsequently, following the addition of Dkk1 and the blocking of Wnt signaling pathway, CVF-conditioned medium-induced morphological changes and expression of cardiomyocyte-associated genes of Nkx2.5(+) CPCs were inhibited, which indicates that CVF-conditioned medium-induced cardiomyogenic differentiation of Nkx2.5(+) CPCs is associated with Wnt signaling pathway. In addition, we also found that the activation of Wnt signaling pathway was accompanied by higher expression of GATA-4 and the blocking of the pathway inhibited the expression of GATA-4 in CVF-conditioned medium-incubated Nkx2.5(+) CPCs. This finding suggests that Wnt signaling pathway may alter GATA-4 expression and activate the cardiogenic program in the regulation of differentiation. In conclusion, Nkx2.5(+) CPCs have enormous potential for cardiomyogenic differentiation and the CVF-conditioned medium specifically induces CPCs to differentiate into a cardiac phenotype. Wnt signaling pathway is involved in CVF-conditioned medium-induced cardiomyogenic differentiation of Nkx2.5(+) CPCs.
Assuntos
Diferenciação Celular , Meios de Cultivo Condicionados , Fibroblastos/fisiologia , Miócitos Cardíacos/fisiologia , Células-Tronco/fisiologia , Actinas/análise , Animais , Microscopia Eletrônica , Microscopia de Fluorescência , Microscopia de Contraste de Fase , Miócitos Cardíacos/citologia , Cadeias Pesadas de Miosina/análise , Organelas/ultraestrutura , Ratos , Ratos Sprague-Dawley , Transdução de Sinais , Células-Tronco/citologia , Troponina T/análiseRESUMO
We have previously shown that nestin-expressing hair follicle stem cells from the mouse and human are multipotent and can differentiate into many cell types, including neurons and glial cells. The nestin-expressing hair follicle stem cells can effect nerve and spinal cord repair upon transplantation in mouse models. In the present study, nestin-expressing hair follicle stem cells expressing red fluorescent protein (RFP) were induced by retinoic acid and fetal bovine serum to differentiate and then transplanted together with Matrigel into the transected distal sciatic or tibial nerve stump of transgenic nude mice ubiquitously expressing green fluorescent protein (GFP). Control mice were transplanted with Matrigel only. The transplanted cells appeared neuron like, with large round nuclei and long extensions. Immunofluorescence staining showed that some of the transplanted cells in the distal nerve stump expressed the neuron marker Tuj1 as well as motor neuron markers Isl 1/2 and EN1. These transplanted cells contacted each other as well as host nerve fibers. Two weeks post-transplantation, nerve fibers in the distal sciatic nerve stump of the transplanted mice had greater expression of motor neuron markers and neurotrophic factor-3 than those in the Matrigel-only transplanted mice. Muscle fiber areas in the nestin-expressing stem cell plus Matrigel-transplanted animals were much bigger than that in the Matrigel-only transplanted animals after 4 weeks. The present results suggest that transplanted nestin-expressing hair follicle stem cells can differentiate into motor neurons and reduce muscle atrophy after sciatic nerve transection. This study demonstrates a new and accessible neuron source to reduce muscle atrophy after nerve injury.