| Objective: The relevant literature information in the field of stem cell organoid niche is bibliometric and visual analysis,and the classical stem cell organoid niche with mature research is screened out,and the culture system of tongue cancer stem cell organoid is constructed.The key genes associated with stem of tongue cancer were identified by bioinformatics methods,and the protein interaction network of key genes and stem cell organoid niche was used to preliminarily verify the application of classical stem cell organoid niche in tongue cancer.Finally,through cytology experiments,the expression of stem cell markers was detected to preliminarily verify the effect of organoid culture system composed of stem cell organoid niche on the enrichment of tongue cancer stem cells.Methods: The experiment is divided into three parts.(1)Application of bibliometric methods to analyze research papers in the niche field of stem cell organoids.Collect data from several aspects such as time,institution,author,keywords and cited literature,and visualize it with software such as citespace,VOSviewer,R package bibliometrix,etc.Through the node literature and node keywords,the classical stem cell organoid niche with mature research in the field of stem cell organoid niche were sought,and the culture system of tongue cancer stem cell organoids was constructed.(2)Tongue cancer RNA-seq data and related clinical information are downloaded from the Cancer Genome Atlas(TCGA).WGCNA is used to modularize differential genes.Based on the stem cell index m RNAsi,the modules and key genes associated with the stem of important tongue cancers were identified.The key genes GO and KEGG functions were enriched,and the protein interaction network of key genes and stem cell organoid niche was used to initially verify the adaptability of stem cell organoid niche in tongue cancer.(3)Taking tongue cancer cells CAL-27 as the object,the expression of stem markers of tongue cancer cells was detected by experiments such as PCR,and the organoid culture system composed of stem cell organoid niche was gradually verified for the enrichment of tongue cancer stem cells.Results:(1)The Web of Science Core Collection Database contains 1053 research papers in stem cell organoid niche areas,and the most productive institution is Harvard med sch The author with the highest number of articles is Clevers,and the authors and institutions around the world are very closely cooperating;classic stem cell organoid culture systems,including EGF,have been extracted from the node literature Rspondin1 and Noggin seeds and serve as the basic framework for organoid culture systems for tongue cancer stem cells(referred to as ERN conditions in this study).Keyword analysis found that β-catenin pathway and FGF are the more mature stem cell organoid niche in this field,and can be used as candidate genemes to strengthen ERN The enrichment effect of conditions.(2)Differential expression analysis in the TCGA database showed that there was a significant difference in m RNAsi between normal tissue and tongue cancer tissue,and tongue cancer tissue was significantly higher than that of normal tissue,P < 0.05.In the survival analysis,patients with high m RNAsi scores were observed to have worse overall survival than patients with low m RNAsi scores,P < 0.05.Based on m RNAsi,22 key genes were identified,and GO and KEGG functional enrichment found that the key genes were concentrated in the cell cycle,The Fanconi anemia pathway,P < 0.05,FDR < 0.25.The protein interaction network of the top 5 key genes and stem cell organoid niche with gene significance was found: In addition to AUNIP,all kinds of stem cell organoid niche and key genes are CDK1 A close connection has been established for the center.This validated the esters EGF,Rspondin1,Noggin,FGF and β-The association of contex proteins with tongue cancer stem,and preliminary verification of their suitability for use in tongue cancer stem cells.(3)Observation under an inverted microscope showed that tongue cancer cells proliferated into balls in organoid culture mode and were tightly arranged,but the cell spheroids under THE ERN condition were larger than those under the 10% serum condition,denser,less common cell debris.Cells under conventional 10% serum 2D culture conditions are polygonally and loosely tiled at the bottom of the bottle.The real-time PCR results showed that the expression of CD44 m RNA,a stem marker of tongue cancer cells,was significantly upregulated compared with the 3D 10%serum group,and the difference was statistically significant,P < 0.05.Compared with the 2D10% serum group,the expression of the stem marker CD44 m RNA showed only an upward trend,and the difference was not statistically significant,P > 0.05.In addition,the expression of the stem marker CD44 for tongue cancer was significantly upregulated in the 3D ERN+C+F group relative to the 3D 10% serum group the difference was statistically significant,P <0.05.Moreover,the relative expression of CD44 in the 3D ERN + C+ F group is higher than that of 3D ERN group and the 3D ERN + F group were significantly increased,and the difference was statistically significant,P < 0.05.However,3D ERN groups,3D ERN + C groups,and 3D ERN+ groups Compared with the 3D 10% serum group,the difference in CD44 expression was not statistically significant in the F group,P > 0.05,but its overall upward trend.The results of the Western Blot experiment suggested that the protein expression level of tongue cancer cells CD44 in the 3D ERN + C+ F group was higher than that in the 3D ERN group or the 3D 10% serum group is high and shows an upward trend.Conclusion:(1)Publications in the niche field of stem cell organoids were hierarchically sorted out and visually analyzed from the perspective of bibliometrics.At the same time,it was found that the β-catenin pathway and FGF are the more maturestem cell organoid niche in this field.The combination of EGF,Rspondin1 and Noggin stem cell organoid niche is a classic stem cell organoid culture system.(2)A high m RNAsi score in patients with tongue cancer may indicate a poor prognosis,and 22 key genes related to stem of tongue cancer were identified.(3)In this study,stem cell organoid niche of tongue cancer were screened and culture systems were constructed,including EGF,Rspondin1,Noggin,FGF、CHIR99021.Under this system,the expression of stem cell-related markers of tongue cancer was significantly upregulated. |
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