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Abstract Recently, the 3D spheroid cell culture application has been extensively used in the treatment of bone defects. A wide variety of methodologies have been used, which has made the comparison of results complex. Therefore, this systematic review has two aims: (i) to perform an analysis focused on the role of 3D spheroid cell culture in bone regeneration strategies; and (ii) address the main challenges in clinical application. A search of the following keywords "3D cell culture", "spheroid", and "bone regeneration" was carried out in the PubMed, Scopus, and ScienceDirect databases and limited to the years 2010-2020. Studies were included if their primary objective was the behavior of cell aggregates to formed spheroids structures by different 3D cell culture techniques focused on the regeneration of bone tissue. To address the risk of bias for in vitro studies, the United States national toxicology program tool was applied, and descriptive statistics of the data were performed, with the SPSS V.22 program. A total of 16 studies were included, which met the established criteria corresponding to in vitro and in vitro/in vivo studies; most of these studies used stem cells for the 3D cell spheroids. The most often methods used for the 3D formation were low adherence surface and rotational methods, moreover, mesenchymal stem cells were the cell line most frequently used because of their regenerative potential in the field of bone tissue engineering. Although the advances in research on the potential use of 3D spheroids in bone regeneration have made great strides, the constant innovation in cell spheroid formation methodologies means that clinical application remains in the future as strategy for 3D tissue bioprinting.
Resumen Recientemente, la aplicación del cultivo 3D de esferoides se ha utilizado ampliamente en el tratamiento de defectos óseos. La variedad de metodologías para lograr los cultivos 3D de esferoides ha hecho compleja la comparación de resultados. Por tanto, esta revisión sistemática tiene dos objetivos: (i) realizar un análisis centrado en el papel de los cultivos 3D de esferoides en las estrategias de regeneración ósea; y (ii) abordar los principales desafíos en la aplicación clínica. Se realizó una búsqueda de las siguientes palabras clave "cultivo celular 3D", "esferoide" y "regeneración ósea" en las bases de datos PubMed, Scopus y ScienceDirect y se limitó a los años 2010-2020. Se incluyeron los estudios si su principal objetivo era el comportamiento de agregados celulares para generar las estructuras esferoidales desarrollados por diferentes técnicas de cultivo celular 3D enfocadas a la regeneración del tejido óseo. Para abordar el riesgo de sesgo de los estudios in vitro, se aplicó la herramienta del programa nacional de toxicología de Estados Unidos y se realizaron estadísticas descriptivas de los datos, con el programa SPSS V.22. Se incluyeron un total de 16 estudios, que cumplieron con los criterios establecidos correspondientes a estudios in vitro e in vitro/in vivo; la mayoría de estos estudios utilizaron células troncales para generar los esferoides celulares 3D. Los métodos más utilizados para la formación de los esferoides 3D fueron la superficie de baja adherencia y los métodos de rotación, asimismo, la línea celular de células troncales mesenquimales fueron las más utilizadas debido a su gran potencial regenerativo en el campo de la ingeniería de tejidos óseos. Aunque los avances en la investigación sobre el uso potencial de los cultivos celulares de esferoides 3D en la regeneración ósea han logrado grandes avances, la constante innovación en las metodologías de la generación de esferoides 3D deja claro que la aplicación clínica de estos permanecerá en el futuro como estrategia en la bioimpresión tisular.
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Regeneración Ósea , Ingeniería de Tejidos , Esferoides CelularesRESUMEN
Objective:To study the soil physical and chemical properties， microorganisms， and metabolites in different culture environments of <italic>Gastrodia elata</italic>， so as to provide scientific basis for subsequent cultivation of <italic>G. elata</italic> in multiple environments. Method:The tubersphere soil of <italic>G. elata</italic> cultured in different environments was collected for analyzing the soil nutrients， microbial numbers， and metabolite differences using the agrochemical method， plate-count method， and gas chromatography-time-of-flight mass spectrometry （GC-TOF-MS）-based non-targeted metabonomic approach. Result:The analysis of soil physical and chemical properties revealed the highest soil moisture， pH， available potassium， and available phosphorus in the spinney and the highest electrical conductivity， total nitrogen， alkali-hydrolyzable nitrogen， and organic matter in the pinewood. As demonstrated by the quantitative analysis of soil microorganisms， the cultivable microorganisms were bacteria， actinomycetes， and fungi， with the bacterial population and total microbial biomass in the spinney and the number of fungi and actinomycetes in the barren slope detected to be the largest. The ratio of bacteria to fungi （B/F value） in the pinewood was the highest， while that in the barren slope was the lowest. The results of metabonomic research demonstrated that the compositions and quantities of soil metabolites in the spinney （Z group）， pinewood （S group）， and barren slope （HD group） varied. Through comparisons between S and Z groups， between HD and Z groups， as well as between HD and S groups by orthogonal partial least squares discriminant analysis （OPLS-DA）， 18， 35， and 24 differential metabolites were separately screened out， and the Kyoto Encyclopedia of Genes and Genomes （KEGG） pathway enrichment analysis yielded 5， 9， and 13 metabolic pathways. There existed a significant causal relationship of the soil physical and chemical properties and microbial numbers with the metabolites. Conclusion:The soil physical and chemical properties， microbial numbers， and metabolite changes differed significantly in different culture environments of <italic>G. elata</italic>， which were sorted by the suitability in a descending order as follows： spinney > pinewood >barren slope. The soil physical and chemical properties and microbial numbers are the crucial factors driving changes in soil metabolites， suggesting that regulating the soil physical and chemical characteristics and microbial characteristics in the culture environment is an important mechanism for maintaining the <italic>G. elata</italic>-soil-microbial symbiotic system.
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Objective To investigate the species of Cheyletoidea mites breeding in the culture environment of Eupolyphaga sinensis and to observe the morphology of Eucheyletia reticulata Cunliffe. Methods The soil samples from an E. sinensis farm in northern Anhui were collected. The mites in the soil samples were separated directly under a microscope and the glass speci?mens were made to observe the morphological feature of the mites under a light microscope,then the mites species were identi?fied and classified based on the morphological characteristics. Results In the culture soil of E. sinensis,totally 7 kinds of Chey?letoidea mites were isolated,namely Eucheyletia reticulata Cunliffe,Cheyletus eruditus Schrank,Cheyletus malaccensis Oudemans,Cheyletus troussarti Oudemans,Cheyletus aveisor Rohdendorz,Acaropsis sollers Rohdendorz and Cheletomor?pha lepidopterorum Shaw. They belonged to genera Eucheyletia,Cheletomorpha,Acaropsis and Cheyletus of Cheyletidae Leach family. The Eucheyletia reticulata Cunliffe was firstly found in the culture environment of E. sinensis,and its gnathosoma was large,the pedipalpal femurs were expanding and there were two strips of comb hair and two smooth bristles on the pedipal?pal tarsus,and the back of the body was covered with two pieces of tergum,which were decorated with reticular pattern. The body and foot setae were fan?shape. Conclusions There are various of Cheyletoidea mites found in the breeding environment of E. sinensis. These mites are important species for pest control in the culture environments of E. sinensis. Related measures should be taken to prevent the excessive growth of Cheyletoidea mites,so as to avoid the adverse effects on the quality and quan?tity of E. sinensis.
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Objective: To investigate the changes of gene expression in CD34+ hematopoietic stem and progenitor cells (HSPCs) under different growth environments. Methods: Umbilical cord blood mononuclear cells (UCB MNCs) were cultured in static and stirred systems. After 7 days of culture, CD34+ cells were isolated and total RNA was extracted. Gene expression patterns of CD34+ cells from fresh, static and stirred cultures were compared using differential display (DD). Results: 30 gene fragments displayed differential expression levels based on the conditions of DD. One of differentially expressed genes was identified as RAN, which is a member of oncogene RAS family. This gene may be associated with proliferation of hematopoietic cells. Conclusion: Different growth environments induced differential gene expression patterns of CD34+ HSPCs. These differentially expressed genes would give new insights into optimizing in vitro environments for expanding hematopoietic cells.