[Proteomic analysis of lacrimal gland adenoid cystic carcinoma with high-grade transformation]

Abstract

Objective: To analyze the protein expression differences of lacrimal gland adenoid cystic carcinoma (LACC) with high-grade transformation (HGT). Methods: Experimental study. A total of 8 paraffin tissue samples were collected in Tianjin Medical University Eye Hospital from December 2012 to January 2019. According to pathological examination, the samples were divided into the LACC group and the LACC-HGT group, with 4 cases in each group. The LACC group included 2 male samples and 2 female samples, with an average age of 53 years. The LACC-HGT group included 2 male samples and 2 female samples, with an average age of 44 years. Primary cells were cultured from fresh tumor tissue. Isobaric tags for relative and absolute quantification techniques were used to screen the differentially expressed proteins between the two groups, and bioinformatics analysis was conducted for the differentially expressed proteins. Microarray was used to screen differentially expressed mRNAs between LACC and LACC-HGT primary cells. The mass spectrum data were intersected with mRNA microarray data, and quantitative real-time (qRT) PCR was performed to verify the results. Proteomics and microarray data were compared using the independent sample t test. The qRT-PCR data were compared pairwise by one-way analysis of variance. Results: A total of 105 HGT-related differential proteins were detected in this study, including 50 up-regulated proteins and 55 down-regulated proteins. The significantly up-regulated proteins included hemoglobin subunit beta, hemoglobin subunit alpha 1, and collagen type Ⅵ alpha 2 chain; the significantly down-regulated proteins included Cereblon, adenosylhomocysteinase like 2, and ribosomal protein L39 pseudogene 5. Gene ontology analysis results showed that the LACC-HGT differential proteins were mainly located in the cytoplasm, vesicle cavity, and extracellular matrix, had organic acid binding and molecular carrier activity, and participated in the regulation of extracellular matrix composition, immunity, inflammation, apoptosis, and other biological processes. Pathway analysis showed that the LACC-HGT differential proteins were mainly involved in signal pathways such as mitogen-activated protein kinase signal pathway and extracellular matrix proteoglycans and glycan metabolism signal pathway. Protein complex prediction analysis screened out 4 up-regulated protein complexes and 1 down-regulated protein complex. There were 15 LACC-HGT differential proteins that overlapped with mRNA chip differential genes, of which 6 were tumor-related proteins including collagen type XIV alpha 1 chain (COL14A1), EMAP like 4 (EML4), inter-alpha-trypsin inhibitor heavy chain 4 (ITIH4), NDRG family member 2 (NDRG2), osteoglycin (OGN) an Ras homolog family member C (RhoC). The main function was the movement and migration of tumor cells. The qRT-PCR results showed that the relative expression levels of COL14A1, EML4, ITIH4, NDRG2, OGN, and RhoC in primary LACC-1, LACC-2, LACC-HGT-1, and LACC-HGT-2 cells were significantly different (F=1 675.98, 38.53, 27.37, 16.47, 13.38, 25.22, all P<0.01). For example, the relative expression of COL14A1 in primary LACC-HGT-1 (16.09±0.51) and LACC-HGT-2 (9.96±0.34) cells was significantly higher than that in primary LACC-1 (1.00±0.13) and LACC-2 (0.67±0.08) cells (all P<0.05). Conclusion: There are differentially expressed proteins between LACC-HGT and LACC, among which COL14A1, EML4, ITIH4, NDRG2, OGN, and RhoC may play an important role in LACC-HGT and can be used as potential targets of LACC-HGT in further study. (Chin J Ophthalmol, 2021, 57: 531-539).