| With the rapid development of nanotechnology,the interaction of stem cells and nanoparticles is widely used in tissue engineering,especially for the stem cell therapy.Studies have shown that many properties of nanomaterials have important effects on cellular uptake.But the research on biological effects of nanomaterials are mostly based on free magnetic nanoparticles,the biological effect caused by the nanoparticles with weak interaction could be neglected.In addition,there are many factors related to cellular having the effect to nanoparticles internalization,but it remains unclear whether the passage of these cells affect the research results.On this basis,we put forward that whether magnetic nanoparticles with weak interaction have influence on cell uptake?And whether the cell passage number is essential for cell uptake assessment?We first fabricated nanoparticles striped assemblies under magnetic field,then cultured SD-rat adipose tissue-derived mesenchymal stromal cells on the assemblies and studied the cell uptake of free micro-nanoparticles with different dimensions.Finally,we detected the characteristics of the cells with different passage numbers,including cell cycle?viability?senescence level?morphology and etc.We also detected the nanoparticles internalization by ICP-OES and TEM quantitatively and qualitatively.Specific work is as follows:1.To increase the size of nanoparticles by magnetic separation,and find the nanoparticles after magnetic separation are present the clusters phenomenon.Then,we fabricated nanoparticles striped assemblies under a uniform magnetic field,and these assemblies are used as the substrates for cell experiments.2?SD rat adipose tissue-derived mesenchymal stem cells were seeded on different magnetic field strength assemblies to detect their uptake of nanoparticles.It was found that when the cells were cultured on a magnetically assembled substrate,a large amount of nanoparticles appeared around the cells or adsorbed on the cell membrane.There was no nanoparticle labeling in the cells,indicating that for the magnetic nanoparticles with weak interactions,cells have little or no uptake.3.Cultured SD-rat adipose tissue-derived mesenchymal stromal cells on the assemblies and studied the cell uptake of free micro-nanoparticles with different dimensions.The results show that there is a reduction of cell uptake micro-nanoparticles when cultured on the assemblies compared with the control treatment.4.Detected the characteristics of the cells with different passage numbers,including cell cycle?viability?senescence level?morphology and etc.We find that cell viability decreased with the increase of cell passage numbers,and the dissociative bare ?-Fe2O3 nanoparticles have much higher viability for the different passage numbers cells than the stripe-like assemblies.With the increase of cell passage numbers,cell morphology changed significantly,and the aging level increased gradually.5.Detected the nanoparticles internalization by ICP-OES and TEM,we found a iron content reduction with the increase of passage numbers when co-cultivation with dissociative bare magnetically separated ?-Fe2O3 nanopar-ticles.The 120mT assemblies substrates treatments have lower iron content than the dissociative nanoparticles treatments.6.RT-qPCR was used to detect the expression of clathrin,scavenger receptor protein and magnetic induction protein in different treatment groups.The expression of clathrin and SR-A protein showed a decreasing trend with the increase of passage numbers.The expression level of clathrin and SR-A protein was increased compared with that of free nanoparticles when cultured on 120mT assembly.With the increase of passage numbers,the expression of magnetosensing protein present a downward trend and for the same passage numbers,the expression of magnetosensing protein was the highest when ADSCs cultured on the stripe-like assemblies fabricated by 120 mT field. |
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