Affymetrix human genome u133 plus 2.0 array

NCBI-GEO (http://www.ncbi.nlm.nih.gov/geo/) is a free database repository comprising microarray/gene profile, next-generation sequencing, hybridization array, and chip data. All data were derived from GEO datasets GSE2052, GSE44723, and GSE24206. The microarray data of GSE2052 were based on GPL1739 Platforms (Amersham Biosciences CodeLink Uniset Human I Bioarray, University of Pittsburgh, PA, USA) and included 15 IPF and 11 control lung tissues (submission date: 09 December 2004) [9 (link), 10 (link)]. The GSE44723 data were based on GPL570 Platforms (Affymetrix Human Genome U133 Plus 2.0 Array, Affymetrix, Santa Clara, CA, USA) and included 10 pulmonary fibrosis and 4 normal lung tissues (submission date: 10 April 2013) [11 (link)]. The GSE24206 data were based on GPL570 Platforms (Affymetrix Human Genome U133 Plus 2.0 Array, Affymetrix, Santa Clara, CA, USA) and included 17 IPF and 6 normal lung tissues (submission date: 01 November 2011) [12 (link)]. The total RNA of the samples was extracted to analyze the genomic profile of the RNA. All data came from expression profiling with microarrays conducted for Homo sapiens.

UC expression profiles with reliable sample sources were downloaded from the GEO (https://www.ncbi.nlm.nih.gov/geo/) database using the GEOquery package^{12 (link)}of the R software (version 3.6.5, http://r-project.org/). The dataset GSE38713^{13 (link)} and GSE94648^{14 (link)} with samples from Homo sapiens and platforms based on GPL570 and GPL19109 [HG-U133_Plus_2] Affymetrix Human Genome U133 Plus 2.0 Array were used. The GSE38713 dataset included 30 UC patient samples and 13 normal samples and GSE94648 dataset included 25 UC patient samples and 22 normal samples, both of which were included in this study (Table 1). The raw data of the GSE38713 and GSE94648 datasets were read using the affy package^{15 (link)}; RMA background correction, and data normalization were performed to obtain the gene expression matrices of the two datasets. HUGO Gene Nomenclature Committee (HGNC)^{16 (link)} is responsible for providing a unique, standardized and widely disseminated symbol for all genes on the human genome including protein-coding genes, non-coding RNA genes, methyl genes and other genes; for each human gene, mRNA expression profiles were obtained using the HGNC mRNA gene annotation file.Table1

Related information of dataset Platform (Affymetrix Human Genome U133 Plus 2.0 Array).

Dataset	Patient	Control
GSE38713	30	13
GSE94648	25	22

Raw files of three registered microarray datasets [GSE46687, GSE58435 (Massingham et al., 2014 (link)), and GSE32527 (Li et al., 2012 (link))] containing TS tissue and normal tissue samples were obtained from the GEO database (Barrett et al., 2007 (link)) through the use of the GEOquery R package (Davis and Meltzer, 2007 (link)). The number of samples in the three datasets is shown in Table 1. Homo sapiens was the selected species, and the microarray expression profile was chosen as the data type. The platform for the GSE46687 dataset was the GPL570 [HG-U133_Plus_2] Affymetrix Human Genome U133 Plus 2.0 Array. The GSE58435 platform was the GPL570 [HG-U133_Plus_2] Affymetrix Human Genome U133 Plus 2.0 Array. The expression profiling of the GSE32527 dataset was based on the GPL6244 [HuGene-1_0-st] Affymetrix Human Gene 1.0 ST Array [transcript (gene) version] platform. The GSE46687 and GSE58435 gene expression profiles were merged. To conduct principal component analysis (PCA), we corrected the batch effect using the ComBat function (SVA package) (Parker et al., 2014 (link)) with the normalized read counts (Raychaudhuri et al., 2000 (link)). A total of 16,343 ear-related genes were downloaded from the CTD (http://ctd.mdibl.org) (Mattingly et al., 2006 (link)).

Gene Expression Omnibus (GEO) database (https://www.ncbi.nlm.nih.gov/geo/) was created by NCBI, containing numerous gene expression data from research institutions worldwide. We downloaded two gene expression datasets (GSE35956 and GSE35958) from the GEO database. The expression profiling arrays of GSE35956 were generated through the application of GPL570 [HG-U133_Plus_2] Affymetrix Human Genome U133 Plus 2.0 Array, including five samples of human mesenchymal stem cells from osteoporosis patients and the other five bone marrow from non-osteoporotic donors after total hip arthroplasty. Besides, the expression profiling arrays of GSE35958 were also generated through the application of GPL570 [HG-U133_Plus_2] Affymetrix Human Genome U133 Plus 2.0 Array, including five osteoporosis samples and four control samples.

When selecting an external validation dataset, we followed the same screening principles to minimize bias. We validated the mRNA expression levels of key crosstalk genes in independent external datasets GSE75436 and GSE16134 samples. GSE75436 included 15 normal superficial temporal artery samples, 15 intracranial aneurysms samples, and mRNA sequencing of the samples was based on GPL570 [HG-U133_Plus_2] Affymetrix Human Genome U133 Plus 2.0 Array. GSE16134 contains 241 PD-affected gingival tissue samples and 69 unaffected gingival tissue samples, mRNA sequencing of the samples is based on GPL570 [HG-U133_Plus_2] Affymetrix Human Genome U133 Plus 2.0 Array.