RESUMO
Numerous studies have confirmed that prenatal or early postnatal exposure to pesticides can lead to functional deficits in the developing brain. This study aimed to investigate whether combined exposure to paraquat (PQ) and maneb (MB) during puberty could cause permanent toxic effects in the neural system of rats. In addition, the neuroprotective function of taurine (T) and its possible mechanism were investigated. Rats were administered PQ + MB intragastrically for 12 continuous weeks, while taurine dissolved in water was fed to the rats for 24 continuous weeks. In the behavioral tests, the rats' trajectories became complex, and the reaction latencies and mistake frequencies increased. Significant changes were found in the hippocampal neurons of the PQ + MB groups but not in the taurine treatment groups. PQ + MB stimulated cAMP to reduce the production of protein kinase A (PKA) and inhibited the activation of other elements, such as brain-derived neurotrophic factor (BDNF), cAMP response element binding protein (CREB), phospho-CREB (p-CREB), immediate-early genes (IEGs)Arc, and c-Fos. Importantly, taurine regulated the level of cAMP and the expression of the abovementioned proteins. Together, our findings implied that adolescent exposure to PQ + MB may impact the behavior and cognitive function of rats via the cAMP-PKA-CREB signaling pathway, while taurine may in turn exert neuroprotection by diminishing these impacts.
Assuntos
Hipocampo/metabolismo , Maneb/efeitos adversos , Transtornos do Neurodesenvolvimento , Neurônios/metabolismo , Paraquat/efeitos adversos , Transdução de Sinais/efeitos dos fármacos , Taurina/farmacologia , Animais , Hipocampo/patologia , Masculino , Maneb/farmacologia , Transtornos do Neurodesenvolvimento/induzido quimicamente , Transtornos do Neurodesenvolvimento/metabolismo , Transtornos do Neurodesenvolvimento/patologia , Transtornos do Neurodesenvolvimento/prevenção & controle , Neurônios/patologia , Paraquat/farmacologia , Ratos , Ratos Sprague-DawleyRESUMO
BACKGROUND: During the process of deep decay, when decay approaches the pulp, an immune response is triggered inside the pulp, which activates the complement cascade. The effect of complement component 5a (C5a) on the differentiation of dental pulp mesenchymal stem cells (DPSCs) is related to dentin reparation. The aim of the present study was to stimulate DPSCs with different concentrations of C5a and evaluate the differentiation of odontoblasts using dentin sialoprotein (DSP). METHODS: DPSCs were divided into the following six groups: (i) Control; (ii) DPSCs treated with 50 ng/ml C5a; (iii) DPSCs treated with 100 ng/ml C5a; (iv) DPSCs treated with 200 ng/ml C5a; (v) DPSCs treated with 300 ng/ml C5a; and (vi) DPSCs treated with 400 ng/ml C5a. Flow cytometry and multilineage differentiation potential were used to identify DPSCs. Mineralization induction, Real-time PCR and Western blot were conducted to evaluate the differentiation of odontoblast in the 6 groups. RESULT: DPSCs can express mesenchymal stem cell markers, including CD105, CD90, CD73 and, a less common marker, mesenchymal stromal cell antigen-1. In addition, DPSCs can differentiate into adipocytes, neurocytes, chondrocytes and odontoblasts. All six groups formed mineralized nodules after 28 days of culture. Reverse transcription-quantitative PCR and western blotting indicated that the high concentration C5a groups expressed higher DSP levels and promoted DPSC differentiation, whereas the low concentration C5a groups displayed an inhibitory effect. CONCLUSION: In this study, the increasing concentration of C5a, which accompanies the immune process in the dental pulp, has demonstrated an enhancing effect on odontoblast differentiation at higher C5a concentrations in vitro.
Assuntos
Células-Tronco Mesenquimais , Diferenciação Celular , Proliferação de Células , Células Cultivadas , Polpa Dentária , Humanos , Células-TroncoRESUMO
Aim: To investigate the effect of hDPSC-Exos in flap I/R injury, a condition in which tissue damage increases after blood flow is restored to the flap after ischemia. Materials & methods: HUVECs were used to investigate the influences and mechanisms of hDPSC-Exos on cell proliferation and migration. A rat model was established to verify the role of hDPSC-Exos in flap I/R injuries in vivo. Results: hDPSC-Exos promoted the proliferation, migration and tube formation of HUVECs in a dose-dependent way by activating PI3K/AKT signaling pathway, and improved the survival and microvessel density of the flap and suppressed epithelial cell apoptosis. Conclusion: hDPSC-Exos can enhance flap repair after I/R injury. This process may be mediated by the activation of PI3K/AKT signaling pathway.
Skin flap transplantation is one of the most important methods of repairing refractory wounds and organ reconstruction. I/R injury and insufficiency of neovascularization significantly affect the survival of flaps. Human dental pulp stem cells (hDPSCs) are a type of mesenchymal stem cells (MSCs) present in dental pulp tissue that have attracted increasing attention. They can play a repair role in a variety of ischemic injuries and neovascularization. Exosomes are important paracrine mediators between MSCs and target cells, containing a variety of proteins, mRNA and miRNA. Recent studies have shown that some exosomes derived from MSCs can improve I/R injury, promote angiogenesis and inhibit apoptosis. This study confirmed that hDPSC-Exos could promote the proliferation, migration and tubule formation of vein endothelial cells in a dose-dependent manner. Inhibition of PI3K/AKT signaling pathway can reduce the above promoting effects, suggesting that these processes may depend on the activation of PI3K/AKT signaling pathway. In the rat model, hDPSC-Exos can significantly improve the survival rate and microvessel density of flaps, and inhibit epithelial cell apoptosis.
Assuntos
Exossomos , Células-Tronco Mesenquimais , Traumatismo por Reperfusão , Humanos , Ratos , Animais , Proteínas Proto-Oncogênicas c-akt/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Fosfatidilinositol 3-Quinases/farmacologia , Polpa Dentária , Traumatismo por Reperfusão/terapia , Traumatismo por Reperfusão/metabolismoRESUMO
Dentinogenesis is a necessary prerequisite for dental tissue engineering. One of the steps for dentinogenesis is to obtain large quantities of highly purified odontoblasts. Therefore, we have undertaken an experiment applying different concentrations of ß-glycerophosphate (ß-GP) to induce the differentiation of dental pulp stem cells (DPSCs) in a long-term 28-day culture. In the meanwhile, we have studied the time- and maturation-dependent expression of matrix extracellular phosphoglycoprotein (MEPE) and that of the odontoblast-like marker-dentin sialoprotein (DSP), in order to investigate an optimized mineralized condition. Western blot results revealed that the expression of DSP became lower when accompanied by the increase of the ß-GP concentration, and there was also an influence on MEPE expression when different concentrations of ß-GP were applied. Meanwhile, the mineralized groups had an inhibitory function on the expression of MEPE as compared with the control group. Above all, all experimental groups successfully generated mineralized nodules by Alizarin Red S and the 5 mM ß-GP group formed more mineralized nodules quantitated using the CPC extraction method. In conclusion, there is a significant modulation of the ß-GP during the differentiation of the DPSCs. The degree of odontoblast differentiation is ß-glycerophosphate concentration dependent. A low concentration of ß-GP (5 mM) has been shown to be the optimal concentration for stimulating the maturation of the DPSCs. Moreover, MEPE accompanied with DSP clearly demonstrates the degree of the differentiation.
Assuntos
Diferenciação Celular , Polpa Dentária/efeitos dos fármacos , Glicerofosfatos/farmacologia , Células-Tronco/efeitos dos fármacos , Western Blotting , Meios de Cultura , Polpa Dentária/citologia , Relação Dose-Resposta a Droga , Imunofluorescência , Humanos , Células-Tronco/citologiaRESUMO
Family with sequence similarity 20member C (FAM20C), a recently characterized Golgi kinase, performs numerous biological functions by phosphorylating more than 100 secreted proteins. However, the role of FAM20C in the salivary glands remains undefined. The present study demonstrated that FAM20C is mainly located in the cytoplasm of duct epithelial cells in the salivary glands. Fam20cf/f; MmtvCre mice were created in which Fam20c was inactivated in the salivary gland cells and observed that the number of ducts and the ductal crosssectional area increased significantly, while the number of acinar cells was reduced. The granular convoluted tubules (GCTs) exhibited an accumulation of aberrant secretory granules, along with a reduced expression and altered distribution patterns of ß nerve growth factor, αamylase and bone morphogenetic protein (BMP) 4. This abnormality suggested that the GCT cells were immature and exhibited defects in developmental and secretory functions. In accordance with the morphological alterations and the reduced number of acinar cells, FAM20C deficiency in the salivary glands significantly decreased the salivary flow rate. The Na+, Cl- and K+ concentrations in the saliva were all significantly increased due to dysfunction of the ducts. Furthermore, Fam20c deficiency significantly increased BMP2 and BMP7 expression, decreased BMP4 expression, and attenuated the canonical and noncanonical BMP signaling pathways in the salivary glands. Collectively, the results of the present study demonstrate that FAM20C is a key regulator of acinar and duct structure and duct maturation and provide a novel avenue for investigating novel therapeutic targets for oral diseases including xerostomia.
Assuntos
Células Acinares/patologia , Proteínas de Ligação ao Cálcio/deficiência , Proteínas da Matriz Extracelular/deficiência , Glândulas Salivares/patologia , Células Acinares/metabolismo , Células Acinares/ultraestrutura , Animais , Proteína Morfogenética Óssea 4/metabolismo , Proteínas de Ligação ao Cálcio/metabolismo , Proteínas da Matriz Extracelular/metabolismo , Camundongos Knockout , Reprodutibilidade dos Testes , Saliva/metabolismo , Glândulas Salivares/metabolismo , Glândulas Salivares/ultraestrutura , Salivação , Transdução de Sinais , Glândula Submandibular/patologiaRESUMO
Human dental pulp stem cells (hDPSCs) possess selfrenewal and osteogenic differentiation properties, and have been used for orofacial bone regeneration and periodontal treatment. Aspirin has been demonstrated to enhance the regeneration of bone marrow mesenchymal stem cells (MSCs); however, the impact of aspirin on the osteogenic differentiation of hDPSCs remains unknown. In the present study, hDPSCs were characterized by flow cytometry, while their clonogenic potential and multipotency were assessed using alizarin red, Oil red O and alcian blue staining. The effect of aspirin on hDPSC viability was assessed using Cell Counting Kit8 assay. Osteogenic capacity was examined by alkaline phosphatase activity, alizarin red staining, reverse transcriptionpolymerase chain reaction and western blotting. Furthermore, in vivo cranial defects were established in SpragueDawley rats to evaluate the effect of aspirin on hDPSCbased bone regeneration. Anorganic bovine bone was used as a bone replacement material and as the carrier for hDPSCs. New bone formation was observed through radiographic and histological analysis. The study demonstrated that hDPSCs expressed MSC markers and possessed multipotency in vitro. Aspirin was nontoxic to hDPSCs at a concentration of ≤100 µg/ml and enhanced the osteogenesis of hDPSCs in vitro. Aspirin significantly increased hDPSCbased bone formation in the rat cranial defect model at 8 or 12 weeks postimplantation (P<0.05). The data suggested that aspirin promotes the osteogenic potential of hDPSCs in vitro and in vivo. Overall, the present study indicated that aspirin improves the bone regeneration capacity of hDPSCs.
Assuntos
Aspirina/farmacologia , Diferenciação Celular/efeitos dos fármacos , Polpa Dentária/citologia , Adolescente , Fosfatase Alcalina/metabolismo , Animais , Western Blotting , Feminino , Citometria de Fluxo , Humanos , Masculino , Microscopia Eletrônica , Osteogênese/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Células-Tronco/citologia , Células-Tronco/efeitos dos fármacosRESUMO
Aspirin has positive effects on bone marrow mesenchymal stem cells (BMSCs) osteogenic differentiation. However, researchers did not give much thought to its effect on BMSCs adipogenic differentiation. Here, we analyzed the effect of aspirin on the BMSCs adipogenic differentiation. To detect whether the effect of aspirin on the adipogenic differentiation of BMSCs is associated with the disturbed epigenetic modification, the expression of histone deacetylases (HDACs), activity of HDACs and HAT, global histone H3 acetylation and H3k9 acetylation alterations were investigated. Moreover, to further explore and understand the binding mode between aspirin and HDACs, an attempt was made to identify the interaction between aspirin and the HDACs with the aid of in silico docking study. The results showed that aspirin could induce inhibition of BMSCs adipogenesis. The level of HDAC activity, global histone H3 acetylation, and H3k9 acetylation were all down regulated during adipogenic differentiation, and aspirin can reverse these decreases. Furthermore, the HDAC isoforms have different expression patterns in those progresses. The expression of HDAC9 was increased in a does-dependent manner when aspirin was introduced during BMSCs adipogenic differentiation. Docking study showed that high affinity of HDAC9 to aspirin was existed, suggesting that HDAC9 may has an important role in the process of aspirin-induced suppression of adipogenesis. Further studies are needed to define the intricate mechanisms of the HDAC isoforms, and all of these enable us to understand aspirin and its efficacy of inhibition of adipogenic differentiation and pave the way to aspirin clinical using for the tissue regenerating.
Assuntos
Adipogenia , Tecido Adiposo/citologia , Aspirina/farmacologia , Células da Medula Óssea/citologia , Diferenciação Celular/efeitos dos fármacos , Epigênese Genética , Células-Tronco Mesenquimais/citologia , Acetilação , Tecido Adiposo/efeitos dos fármacos , Tecido Adiposo/metabolismo , Sequência de Aminoácidos , Animais , Anti-Inflamatórios não Esteroides/farmacologia , Apoptose , Células da Medula Óssea/efeitos dos fármacos , Células da Medula Óssea/metabolismo , Proliferação de Células , Células Cultivadas , Histona Desacetilases/química , Histonas/genética , Histonas/metabolismo , Células-Tronco Mesenquimais/efeitos dos fármacos , Células-Tronco Mesenquimais/metabolismo , Simulação de Acoplamento Molecular , Conformação Proteica , Ratos , Ratos Sprague-Dawley , Homologia de SequênciaRESUMO
It has been established that dental pulp stem cells (DPSCs) serve an important role in the restoration and regeneration of dental tissues. DPSCs are present in blood vessels and also exist in the vessel microenvironment in vivo and have a close association with endothelial cells (ECs). The present study aimed to evaluate the influence of ECs and their secretory product endothelin1 (ET1) on the differentiation of DPSCs. In the present study, cells were divided into four groups: i) a DPSConly control group; ii) a DPSC with ET1 administration group; iii) a DPSC and human umbilical vein endothelial cell (HUVEC) direct coculture group; and iv) a DPSC and HUVEC indirect coculture group using a Transwell system. Reverse transcriptionquantitative polymerase chain reaction was used to detect the expression of the odontoblastic differentiationassociated genes, including dentin sialoprotein (DSP) and dentin matrix acidic phosphoprotein 1 (DMP1) at days 4, 7, 14 and 21. Alizarin Red S staining, immunofluorescence and western blot analyses were also conducted to assess the differentiation of the DPSCs in each group. The highest expression levels of odontoblastic differentiationassociated genes were observed on day 7 and in the two coculture groups were increased compared with the DPSConly and DPSC + ET1 culture groups at all four time points. However, expression levels in the DPSC + ET1 group were not downregulated as notably as in the coculture groups on days 14 and 21. The Transwell group exhibited the greatest ability for odontoblastic differentiation compared with the other groups according to staining with Alizarin Red S, immunofluorescence and western blot analysis results. According to the results of the present study, the culture solution with HUVECs affected the differentiation of DPSCs. In addition, ET1 may promote the odontoblastic differentiation of DPSCs.
Assuntos
Polpa Dentária/metabolismo , Endotelina-1/metabolismo , Células Endoteliais da Veia Umbilical Humana/metabolismo , Odontogênese , Células-Tronco/metabolismo , Adolescente , Adulto , Técnicas de Cocultura , Polpa Dentária/citologia , Proteínas da Matriz Extracelular/biossíntese , Feminino , Células Endoteliais da Veia Umbilical Humana/citologia , Humanos , Masculino , Fosfoproteínas/biossíntese , Células-Tronco/citologiaRESUMO
Recent studies have shown that, in numerous species, systemically administered bone marrow-derived mesenchymal stem cells undergo site-specific differentiation. This suggests that osteoblasts, by means of cytokine secretion, may promote dental pulp stem cells (DPSCs) to undergo osteogenesis. The objective of this study was to assess the potential synergistic interaction effect of osteoblasts on DPSCs for promotion of osteogenesis. Stem cells, derived from dental pulp of healthy human donors, were co-cultured with calvaria osteoblasts using a culture insert system. The proliferation rate, calcium deposition, osteogenic-related gene expression of induced DPSCs, including Runx-2, bone sialoprotein, osteocalcin and collagen-1, were assayed using MTT, Alizarin Red S staining and reverse transcriptase polymerase chain reaction, respectively. Co-cultured DPSCs had the highest rate of proliferation compared with those cultured in absence of osteoblasts. The morphology and ultrastructure of DPSCs in the co-cultures showed improvement, with co-cultured DPSCs becoming more osteoblast-like as compared with DPSCs cultured alone, and the mineralization potential of co-cultured DPSCs was enhanced compared with DPSCs cultured alone. Furthermore, osteogenic-related genes were significantly over-expressed in co-cultured DPSCs after osteogenic induction. The results demonstrate that DPSCs successfully differentiate towards osteoblasts and that the paracrine interaction of osteoblasts is likely to contribute to DPSC differentiation. It is believed that this study demonstrates certain useful applications for DPSCs in bone tissue engineering.