Revealing the angiopathy of lacrimal gland lesion in type 2 diabetes

Purpose: The goals of this study are to compare type 2 diabetes lacrimal gland transcriptome profiling (RNA-seq) with the counterparts. Methods: Lacrimal gland mRNA profiles of 24 weeks T2DM mice (C57 mice with high fat diet + STZ intraperitoneal injection) and counterparts were enriched by NEBNext® Poly(A) mRNA Magnetic Isolation Module (New England Biolabs), and the produced RNA was used for construction library, via KAPA Stranded RNA-Seq Library Prep Kit (Illumina). The prepared RNA-seq libraries were qualified using Agilent 2100 Bioanalyzer and quantified by qPCR absolute quantification method. The sequencing was then performed using Illumina NovaSeq 6000. After quality control, the fragments were 5’, 3’-adaptor trimmed and filtered <=16 bp reads with cutadapt software. The trimmed reads were aligned to a reference genome with Hisat2 software. The transcript abundance for each sample was estimated with StringTie (v1.2.3), and the FPKM values for gene level were calculated with R package Ballgown (v2.6.0). The differentially expressed genes (DEGs) analysis was also performed with Ballgown. Fold change (cutoff 1.5), p-value (cutoff 0.05) and FPKM (≥0.5 mean in one group) were used for filtering differentially expressed genes and transcripts. Conclusions: Our study represents the first detailed analysis of T2DM lacrimal gland transcriptomes, with biologic replicates, generated by RNA-seq technology. The optimized data analysis workflows reported here should provide a framework for comparative investigations of expression profiles. Our results showed that the vascular related genes were altered in the T2DM. Lacrimal gland mRNA profiles of 24 weeks T2DM mice (C57 mice with high fat diet + STZ intraperitoneal injection) and counterparts.