Dental pulp stem cells(DPSCs) secrete neurotrophic factors which may play an important therapeutic role in neural development, maintenance and repair. To test this hypothesis, DPSCs-conditioned medium(DPSCs-CM) was co...Dental pulp stem cells(DPSCs) secrete neurotrophic factors which may play an important therapeutic role in neural development, maintenance and repair. To test this hypothesis, DPSCs-conditioned medium(DPSCs-CM) was collected from 72 hours serum-free DPSCs cultures. The impact of DPSCs-derived factors on PC12 survival, growth, migration and differentiation was investigated. PC12 cells were treated with nerve growth factor(NGF), DPSCs-CM or co-cultured with DPSCs using Transwell inserts for 8 days. The number of surviving cells with neurite outgrowths and the length of neurites were measured by image analysis. Immunocytochemical staining was used to evaluate the expression of neuronal markers NeuN, microtubule associated protein 2(MAP-2) and cytoskeletal marker βIII-tubulin. Gene expression levels of axonal growth-associated protein 43 and synaptic protein Synapsin-I, NeuN, MAP-2 and βIII-tubulin were analysed by quantitative polymerase chain reaction(qRT-PCR). DPSCs-CM was analysed for the neurotrophic factors(NGF, brain-derived neurotrophic factor [BDNF], neurotrophin-3, and glial cell-derived neurotrophic factor [GDNF]) by specific ELISAs. Specific neutralizing antibodies against the detected neurotrophic factors were used to study their exact role on PC12 neuronal survival and neurite outgrowth extension. DPSCs-CM significantly promoted cell survival and induced the neurite outgrowth confirmed by NeuN, MAP-2 and βIII-tubulin immunostaining. Furthermore, DPSCsCM was significantly more effective in stimulating PC12 neurite outgrowths than live DPSCs/PC12 co-cultures over the time studied. The morphology of induced PC12 cells in DPSCs-CM was similar to NGF positive controls;however, DPSCs-CM stimulation of cell survival was significantly higher than what was seen in NGF-treated cultures. The number of surviving PC12 cells treated with DPSCs-CM was markedly reduced by the addition of anti-GDNF, whilst PC12 neurite outgrowth was significantly attenuated by anti-NGF, anti-GDNF and anti-BDNF antibodies. These findings demonstrated that DPSCs were able to promote PC12 survival and differentiation. DPSCs-derived NGF, BDNF and GDNF were involved in the stimulatory action on neurite outgrowth, whereas GDNF also had a significant role in promoting PC12 survival. DPSCs-derived factors may be harnessed as a cell-free therapy for peripheral nerve repair. All experiments were conducted on dead animals that were not sacrificed for the purpose of the study. All the methods were carried out in accordance with Birmingham University guidelines and regulations and the ethical approval is not needed.展开更多
Retinal ganglion cells （RGCs） are responsible for propagat- ing signals derived from visual stimuli in the eye to the brain, along their axons within the optic nerve to the superior colliculus, lateral geniculate nu...Retinal ganglion cells （RGCs） are responsible for propagat- ing signals derived from visual stimuli in the eye to the brain, along their axons within the optic nerve to the superior colliculus, lateral geniculate nucleus and visu- al cortex of the brain. Damage to the optic nerve either through trauma, such as head injury, or degenerative dis- ease, such as glaucoma causes irreversible loss of function through degeneration of non-regenerating RGC axons and death of irreplaceable RGCs, ultimately leading to blindness （Berry et al., 2008）. The degeneration of RGCs and their axons is due to the loss of the necessary source of retrogradely transported neurotrophic factors （NTFs） being hindered by axonal injury. NTFs are survival factors for neurons and play a pivotal part in axon regeneration. Stem cells particularly mesenchymal stem cells （MSCs） have been shown to possess a natural intrinsic capacity for paracrine support, releasing multiple signalling mol- ecules including NTFs. By transplanting MSCs into the vitreous, they are positioned adjacent to the injured reti- na to provide paracrine-mediated therapy for the retinal neuronal cells （Johnson et al., 2010a; Mead et al., 2013）. Additionally, MSCs may be pre-differentiated into sup- portive glial-like cells, such as Schwann cells, which could further increase their potential for paracrine support of injured neurons （Martens et al., 2013）. Thus, MSCs have received considerable attention as a new cellular therapy for both traumatic and degenerative eye disease, acting as an alternative source of NTFs, protecting injured RGCs and promoting regeneration of their axons （Figure 1）.展开更多
The trigeminal nerve and its peripheral branches are susceptible to injury in the dental practice due to surgical removal of impacted third molars and placement of dental implants.Although peripheral trigeminal nerve ...The trigeminal nerve and its peripheral branches are susceptible to injury in the dental practice due to surgical removal of impacted third molars and placement of dental implants.Although peripheral trigeminal nerve injuries can undergo spontaneous regeneration,some injuries may be permanent with varying degrees of continued sensory impairment and neuropathic pain ranging from mild numbness to complete anaesthesia(Tay and Zuniga,2007).Adult neurons require continued neurotrophic support from surrounding cells to sustain neural viability,inhibit death-inducing pathways activating a variety of cell survival pathways(Zheng and Quirion,2004).展开更多
基金funded by Egyptian Cultural and Educational Bureau in London,Egyptian mission sector and ministry of higher education in Egypt(grant No.GAM2649)。
文摘Dental pulp stem cells(DPSCs) secrete neurotrophic factors which may play an important therapeutic role in neural development, maintenance and repair. To test this hypothesis, DPSCs-conditioned medium(DPSCs-CM) was collected from 72 hours serum-free DPSCs cultures. The impact of DPSCs-derived factors on PC12 survival, growth, migration and differentiation was investigated. PC12 cells were treated with nerve growth factor(NGF), DPSCs-CM or co-cultured with DPSCs using Transwell inserts for 8 days. The number of surviving cells with neurite outgrowths and the length of neurites were measured by image analysis. Immunocytochemical staining was used to evaluate the expression of neuronal markers NeuN, microtubule associated protein 2(MAP-2) and cytoskeletal marker βIII-tubulin. Gene expression levels of axonal growth-associated protein 43 and synaptic protein Synapsin-I, NeuN, MAP-2 and βIII-tubulin were analysed by quantitative polymerase chain reaction(qRT-PCR). DPSCs-CM was analysed for the neurotrophic factors(NGF, brain-derived neurotrophic factor [BDNF], neurotrophin-3, and glial cell-derived neurotrophic factor [GDNF]) by specific ELISAs. Specific neutralizing antibodies against the detected neurotrophic factors were used to study their exact role on PC12 neuronal survival and neurite outgrowth extension. DPSCs-CM significantly promoted cell survival and induced the neurite outgrowth confirmed by NeuN, MAP-2 and βIII-tubulin immunostaining. Furthermore, DPSCsCM was significantly more effective in stimulating PC12 neurite outgrowths than live DPSCs/PC12 co-cultures over the time studied. The morphology of induced PC12 cells in DPSCs-CM was similar to NGF positive controls;however, DPSCs-CM stimulation of cell survival was significantly higher than what was seen in NGF-treated cultures. The number of surviving PC12 cells treated with DPSCs-CM was markedly reduced by the addition of anti-GDNF, whilst PC12 neurite outgrowth was significantly attenuated by anti-NGF, anti-GDNF and anti-BDNF antibodies. These findings demonstrated that DPSCs were able to promote PC12 survival and differentiation. DPSCs-derived NGF, BDNF and GDNF were involved in the stimulatory action on neurite outgrowth, whereas GDNF also had a significant role in promoting PC12 survival. DPSCs-derived factors may be harnessed as a cell-free therapy for peripheral nerve repair. All experiments were conducted on dead animals that were not sacrificed for the purpose of the study. All the methods were carried out in accordance with Birmingham University guidelines and regulations and the ethical approval is not needed.
基金funded by the Biotechnology and Biological Sciences Research Council(BBSRC)(No.BB/F017553/1)the Rosetrees Trust
文摘Retinal ganglion cells （RGCs） are responsible for propagat- ing signals derived from visual stimuli in the eye to the brain, along their axons within the optic nerve to the superior colliculus, lateral geniculate nucleus and visu- al cortex of the brain. Damage to the optic nerve either through trauma, such as head injury, or degenerative dis- ease, such as glaucoma causes irreversible loss of function through degeneration of non-regenerating RGC axons and death of irreplaceable RGCs, ultimately leading to blindness （Berry et al., 2008）. The degeneration of RGCs and their axons is due to the loss of the necessary source of retrogradely transported neurotrophic factors （NTFs） being hindered by axonal injury. NTFs are survival factors for neurons and play a pivotal part in axon regeneration. Stem cells particularly mesenchymal stem cells （MSCs） have been shown to possess a natural intrinsic capacity for paracrine support, releasing multiple signalling mol- ecules including NTFs. By transplanting MSCs into the vitreous, they are positioned adjacent to the injured reti- na to provide paracrine-mediated therapy for the retinal neuronal cells （Johnson et al., 2010a; Mead et al., 2013）. Additionally, MSCs may be pre-differentiated into sup- portive glial-like cells, such as Schwann cells, which could further increase their potential for paracrine support of injured neurons （Martens et al., 2013）. Thus, MSCs have received considerable attention as a new cellular therapy for both traumatic and degenerative eye disease, acting as an alternative source of NTFs, protecting injured RGCs and promoting regeneration of their axons （Figure 1）.
文摘The trigeminal nerve and its peripheral branches are susceptible to injury in the dental practice due to surgical removal of impacted third molars and placement of dental implants.Although peripheral trigeminal nerve injuries can undergo spontaneous regeneration,some injuries may be permanent with varying degrees of continued sensory impairment and neuropathic pain ranging from mild numbness to complete anaesthesia(Tay and Zuniga,2007).Adult neurons require continued neurotrophic support from surrounding cells to sustain neural viability,inhibit death-inducing pathways activating a variety of cell survival pathways(Zheng and Quirion,2004).
