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OBJECTIVE: Velvet antler polypeptide (VAP) has been shown to play important roles in the immune and nervous systems. The purpose of this study was to investigate the protective effects of VAP on cerebral ischemic injury with the involvement of NF-κB signaling pathway in vitro. MATERIALS AND METHODS: PC-12 cells stimulated by oxygen-glucose deprivation/reperfusion (OGD/R) was used to mimic cerebral ischemic injury in vitro. The levels of ROS, SOD, and intracellular concentrations of Ca2+ were measured by the relevant kits. Meanwhile, the expressions of inflammatory cytokines (IL-6, IL-1ß, and TNF-α) were determined by ELISA kit assay. In addition, MTT, EdU, and flow cytometry assays were used to measure the cell proliferation and apoptosis. Besides which, the related proteins of NF-κB signaling pathway were measured by western blotting assay. RESULTS: VAP alleviated cerebral ischemic injury by reducing OGD/R-induced oxidative stress, inflammation, and apoptosis in PC-12 cells in a time dependent manner. Mechanistically, VAP inhibited the levels of p-p65 and p-IkB-α in a time dependent manner, which was induced by OGD/R operation. Moreover, NF-κB agonist diprovocim overturned the suppression effects of VAP on OGD/R-induced oxidative stress, inflammation, and apoptosis in PC-12 cells. CONCLUSIONS: The results demonstrate that VAP may alleviate cerebral ischemic injury by suppressing the activation of NF-κB signaling pathway.
Subject(s)
Antlers , Reperfusion Injury , Humans , Animals , NF-kappa B/metabolism , Antlers/metabolism , Signal Transduction , Oxygen/metabolism , Cytokines/metabolism , Inflammation/metabolism , Reperfusion Injury/prevention & control , Reperfusion Injury/metabolism , Apoptosis , GlucoseABSTRACT
Objective To study the effect of velvet antler polypeptides (VAP) on Rho/ROCK pathway in APP/ PSl double transgenic mice. Methods APP/PSl double transgenic mice were randomly divided into model group and velvet antler polypeptide group, 20 mice in each group, and control group consisting of 20 mice of the same litter and the same gender negative. The mice in VAP group were given velvet antler polypeptide 100 mg/kg by intragastric administration once a day for 28 days. After treatment, the water maze experiment was detected and recorded the escape latency and the number of crossing platforms of the mice; the ultrastructures of the synapse were observed by transmission electron microscopy; the expression of Rhs homolog gene family member A(RhoA) and Rho associated coiled-coil forming protein kinase II(ROCKII) in the hippocampal CAI area were observed by immunofluorescence. The expression levels of RhoA and ROCKII protein in the hippocampus were detected by Western blotting. The contents of hippocampus amyloid (3-protein(A(3),
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Three-dimensional (3D) culture of neural progenitor cells (NPCs) in hydrogels represents a powerful means for recapitulating neurodevelopment, disease modelling and drug discovery. However, the differentiation of NPCs to oligodendrocytes in 3D scaffolds remains a great challenge. In this study, polyvinyl alcohol (PVA) - sodium alginate (SA) composite hydrogels intended for NPC culture in 3D were fabricated by ionic crosslinking between SA and calcium ions. It was demonstrated that adding PVA to the composite hydrogels resulted in increases in pore size and swelling rate and decreases in elastic moduli as the PVA proportion was enhanced. In addition, the composite hydrogels were biocompatible with mouse NPCs and improved the proliferation of the encapsulated NPCs compared with SA hydrogels. Moreover, when velvet antler polypeptides (VAPs), which were capable of facilitating the differentiation of NPCs to oligodendrocyte fate in 2D, were loaded into PVA-SA hydrogels, NPCs differentiated into neurons, astrocytes and oligodendrocytes, with the presence of VAPs promoting oligodendrogenesis in a dose-dependant manner. The present composite hydrogels provide a suitable scaffold for the construction of neural tissue engineering and neurological disease modelling.
Subject(s)
Antlers , Neural Stem Cells , Alginates , Animals , Cell Differentiation , Hydrogels , Mice , Oligodendroglia , Peptides , Polyvinyl AlcoholABSTRACT
Objective: To discuss the protective effect of velvet antler polypeptides (VAP) combined with Schwann cells (SCs) modified by glial cell line derived neurotrophic factor (GDNF) gene on the apoptosis of spinal cord neurons induced by beta-amyloid 25-35 (Aβ25-35). Methods: The spinal cord cells of fetal mice were prepared and the spinal cord neurons in logarithmic growth phase were taken; the apoptosis of spinal cord neurons was induced by Aβ25-35. The spinal cord neurons were divided into normal cell group (the normal spinal cord neurons), induced apoptosis group (the apoptotic spinal cord neurons induced by Aβ25-35, SCs group (the apoptotic spinal cord neurons induced by Aβ25-35 + SCs), GDNF group (the apoptotic spinal cord neurons induced by Aβ25-35 + GDNF), SCs + GDNF group (the apoptotic spinal cord neurons induced by Aβ25-35 + GDNF-transfected SCs) and VAP combination group (the apoptotic spinal cord neurons induced by Aβ25-35 + VAP combined with GDNF-transfected SCs). Flow cytometry was used to detect the apoptotic rates of spinal cord neurons in various groups; immunohistochemical staining was used to detect the number of caspase-3 positive cells in the spinal cord neurons in various groups. Results; After suspension inoculation of fetal spinal cord neurons, most of them were round at the initial stage. The flow cytometry results showed that compared with induced apoptosis group, the apoptotic rates of spinal cord neurons in SCs, GDNF, SCs + GDNF, and VAP combination groups were decreased (P<0.05). Compared with SCs + GDNF group, the apoptotic rate of spinal cord neurons in VAP combination group was decreased (P<0. 05). The immunohistochemistry results showed that the expression of caspase-3 in spinal cord neurons in various groups could be found. There were no significant differences in the number of caspase-3 positive cells and cell staining between SCs group, GDNF group and SCs + GDNF group; but the number of caspase-3 positive cells in SCs group, GDNF group and SCs + GDNF group were significantly higher than that in induced apoptosis group (P<0. 05). Conclusion: VAP combined with GDNF-transfected SCs has the protective effect on the apoptosis of spinal cord cells by reducing the expression of caspase-3 in spinal cord neurons.
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OBJECTIVE To investigate the protective effect of velvet antler polypeptides(VAP)on hydrogen peroxide( H2 O2 )-induced injury in vascular endothelial cells and the possible mechanism.METHODS The EVC-304 cells cultured invitrowere incubated with H2 O2 for another 12 h after pretreat-ment with VAP 20,40 and 80 mg·L-1 for 24 h. Cell viability was determined by MTT assay, Hoechst333258 staining was used to observe cell morphology,the activity of superoxide dismutase (SOD)and the level of malondialdehyde( MDA)were detected with kits and the expression of heat shock protein(HSP70)and caspase 3 was detected by Western blotting. RESULTS Compared with the normal control group,the cell survival rate was decreased significantly in H2 O2 injury group( P ﹤0.01),cell shrinkage,chromatin condensation,and nuclear fragmentation were seen,the intracellular SOD activity decreased while MDA content increased(P﹤0.01),and caspase 3 and HSP70 expression increased(P﹤0.01). Compared with H2 O2 group,the cell survival rate in VAP 20,40 and 80 mg·L-1 pre-treatment groups increased significantly(P﹤0.01),the apoptosis ratio declined from(25.3±1.0)% to (15.2±1.2)%,(10.3±0.9)% and(7.9±1.4)%(P﹤0.01),the SOD activity increased to 19.2±0.5,22.3± 1.7 and(24.9±0.6)kU·g-1 protein(P﹤0.01),MDA concentration decreased to 1.51±0.2,1.48±0.3 and (1.02±0.1)μmol·g-1 protein(P﹤0.01),and the expression of caspase 3 and HSP70 declined significant-ly(P﹤0.01). CONCLUSION VAP has exert protective effect on H2 O2-induced injury in vascular endothe-lial cells. The possible mechanism might be related to improvement of intracellular oxidative stress level.
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[Objective] To study effects of the velvet antler polypeptides on spinal nerve cell apoptosis induced by ?-amyloid peptide. [Method]Spinal nerve cells were performed serial subcultivation,and entered into experiment during the exponential phase of growth.The viability of spinal nervecells 24h after induction by ?-amyloid peptide with different concentrations were detected by MTT assay,percentages of the cell apoptosis and expression of casepase-3 were detected after induction by 25?mol/L ?-amyloid peptide.[Result]It was revealed that 24 h after treatment with ?-amyloid peptide of different concentrations,the viability of spinal nerve cells was significantly decreased in a dose-dependent manner(P
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Objective To study the rehabilitative effect of velvet antler polypeptides-PLGA compound membrane on peripheral nerve injury.Methods Two doses(3 and 15 mg?g~(-1)) of velvet antler polypeptides-PLGA compound membrane(thickness 50 ?m) were used to wrap the sutures of severed sciatic nerves,which were compared with nerve sutures only.2,4 and 6 weeks post-operation,general morphological,electrophysiological,histological and electron microscopic observations and examinations were investigated.Results There were no ulcers in the nars and toes in the treatment group,light conglutination appeared in nerve anastomosis and around tissues.At 6th week post-operation,conglutination was alleviated obviously than 2 weeks post-operation;findings from latency inducing potentials of calf triceps dominated by sciatic nerves showed that recover ratios(%) in treatment group((2 weeks:)9.07?1.44,8.02?1.41;6 weeks: 49.87?9.69,50.11?6.11) were better than those in control group((2 weeks:)2.52?1.83;6 weeks:30.31?6.32) obviously in each time(P