Project description:To identify to identify target tissues and molecules involved with refractive myopic shift and axial length elongation in a murine lens-induced myopia model, we performed comprehensive analysis by microRNA array. Negative 30diptor (-30D) lens was fixed on right eye (-30D) of C57BL/6J mice (3weeks old, N=3) for 3 weeks, the refraction and the axial length were measured using a refractometer and a SD-OCT system in all eyes. Eye balls were enucleated and separated to cornea, iris, lens, retina, choroid and sclera. Total RNA was extracted from individual ocular components. MicroRNA expression analysis was carried out using Agilent Mouse miRNA Microarray (8×60K) miRBase21.0 (Agilent). Expression ratio calculation and miRNA varying expression were extracted by GeneSpring GX 14.5 (Agilent). After 3weeks of lens fixing, a refractive change and an axial length elongation change were observed (Normal vs -30D: 0.95 ± 1.85D vs -18.42 ± 3.98D, 0.155 mm ± 0.015mm vs 0.273 ± 0.009 mm), respectively. MiRNA expression changes that induced only by -30D lens fixing was confirmed in each part of the eyeball. By expression ratio calculation and miRNA varying expression analysis, upregulated miRNA (56 in cornea, 13 in iris, 6 in lens, 0 in retina, 29 in choroid and 30 in sclera) and downregulated miRNA (7 in cornea, 28 in iris, 17 in lens, 9 in retina, 7 in choroid and 40 in sclera) were observed. Overlapping miRNAs were also found while each eye tissues. In this study, miRNA varying expression were observed in each ocular part of the murine lens-induced myopia model. These miRNAs dysregulation may be functionally involved with the refractive myopia shift and the axial length elongation.
Project description:Proteomic analyses of the ocular posterior pole tissues at 6 weeks after induction of form-deprived myopia and lens-induced myopia models in guinea pigs.
Project description:To explore the presence of extrachromosomal circular DNA (eccDNA) in the anterior capsule of the lens in the eyes of patients with cataract and with high myopia. Circle-Seq was performed to identify differences in the eccDNA and gene expression between the anterior capsule of the lens of patients with simple nuclear cataract (C, n = 6) and patients with nuclear cataract along with high myopia (HM, n = 6). The expression of eccDNA was confirmed using routine quantitative polymerase chain reaction. The eccDNA ranked in C and HM ranged in length from 0.017 kb – 9.9 Mb with two distinctive peaks detected at 0.2 kb and 0.5 kb, while eccDNA that were differentially expressed (up- and down-regulated) ranged in size from 0.05 kb – 57.8 kb with two distinctive peaks observed at 0.1 kb and 0.5 kb. Only 2.5 % of the eccDNA in C and 2 % in HM were > 25 kb in size. The gene-rich chromosomes contributed to more number of eccDNA/Mb, while several well-known high myopia candidate genes, including catenin delta 2 (CTNND2), ubiquitin-like with PHD and ring finger domains 1 binding protein 1 like (UHRF1BP1L) , exhibited significantly increased levels of eccDNA in the anterior capsule of the lens in patients with high myopia. This study highlighted the topologic analysis of the anterior capsule of eyes with high myopia, which is an emerging direction for research and clinical applications. These findings suggested that eccDNA was commonly detected in eyes with high myopia and cataracts, and the candidate genes for high myopia identified in previous studies were also observed in the eccDNA.
Project description:Purpose: Transcriptome is the entire repertoire of all transcripts present in a cell at any particular time. We undertook next-generation whole transcriptome sequencing approach to gain insight of the transcriptional landscape of the developing mouse lens. Methods: We ascertained mice lenses at six developmental time points including two embryonic (E15 and E18) and four postnatal stages (P0, P3, P6, and P9). The ocular tissue at each time point was maintained as two distinct pools serving as biological replicates for each developmental stage. The mRNA and small RNA libraries were paired-end sequenced on Illumina HiSeq 2000 and subsequently analyzed using bioinformatics tools. Results: Mapping of mRNA and small RNA libraries generated 187.56 and 154.22 million paired-end reads, respectively. We detected a total of 14,465 genes in the mouse ocular lens. Of these, 46 genes exhibited 40-fold differential expression compared to transcriptional levels at E15. Likewise, small RNA profiling identified 379 microRNAs (miRNAs) expressed in mouse lens. Of these, 49 miRNAs manifested an 8-fold or higher differential expression when compared, as above to the microRNA expression at E15. Conclusion: We report the first comprehensive profile of developing murine lens transcriptome including both mRNA and miRNA through next-generation RNA sequencing. A complete repository of the lens transcriptome of six developmental time points will be monumental in elucidating processes essential for development of the ocular lens and maintenance its transparency. Whole transcrtiome and microRNA profilling of mouse lens using 2 embryonic (E15 and E18) and 4 postnatal stages (P0, P3, P6 and P9) in duplicates through high-throughput sequening using Illumina HiSeq2000.
Project description:This is a combined SWATH database of early myopic guinea pig retina. The retinal tissues were divided into lens induced myopia 4 days group (4-day LIM) and control group; 5 individual animals (10 retinas) were included. The potential biomarkers and underlying biochemical pathways during early myopia could be investigated using SWATH based proteomics approach.
Project description:The lens is comprised of the anterior lens epithelium and posterior lens fibers, which form the bulk of the lens. The RNAseq data enables identification of lens epithelium and fiber differentially expressed genes and temporally differentially expressed genes which were also validated by qRTPCR. The present RNA-seq data serves as a comprehensive reference resource for deciphering molecular principles of normal mammalian lens differentiation, mapping a full spectrum of signaling pathways and DNA-binding transcription factors operating in both lens compartments, and predicting novel pathways required to establish lens transparency.
Project description:The proteome is a term used for the entire catalog of proteins present in a cell at any time. In here, we investigated the proteome of developing mouse lens through mass spectrometry-based protein sequencing. We extracted mouse lenses at six developmental time points, which included two embryonic (E15 and E18) and four postnatal stages (P0, P3, P6, and P9). The lenses from each time point were preserved in three distinct pools to serve as biological replicates for each developmental stage. The total cellular protein was extracted from the lens, digested with trypsin and labeled with 6-plex isobaric tandem mass tags (TMT) for three independent 6-plex TMT experiments. A total of 6,117 proteins were identified in the mouse ocular lens in at least one of the above-mentioned six developmental time points. Of these, 6,117 proteins were present in one TMT set, 4,325 in two, and 2,972 were present in three TMT sets, respectively. Majority of the proteins exhibit steady expression; however, we identified 162 proteins that exhibited an 8-fold differential (higher or lower) expression during the developmental time course compared to their levels at E15. The lens proteome is comprised of diverse proteins that have distinct biological properties and functional characteristics including a total of 404 proteins that have been associated with autophagy. We have established a comprehensive profile of the developing murine lens proteome. This repository will be helpful in identifying critical components of lens development and processes essential for the maintenance of its transparency.
Project description:Purpose: Transcriptome is the entire repertoire of all transcripts present in a cell at any particular time. We undertook next-generation whole transcriptome sequencing approach to gain insight of the transcriptional landscape of the developing mouse lens. Methods: We ascertained mice lenses at six developmental time points including two embryonic (E15 and E18) and four postnatal stages (P0, P3, P6, and P9). The ocular tissue at each time point was maintained as two distinct pools serving as biological replicates for each developmental stage. The mRNA and small RNA libraries were paired-end sequenced on Illumina HiSeq 2000 and subsequently analyzed using bioinformatics tools. Results: Mapping of mRNA and small RNA libraries generated 187.56 and 154.22 million paired-end reads, respectively. We detected a total of 14,465 genes in the mouse ocular lens. Of these, 46 genes exhibited 40-fold differential expression compared to transcriptional levels at E15. Likewise, small RNA profiling identified 379 microRNAs (miRNAs) expressed in mouse lens. Of these, 49 miRNAs manifested an 8-fold or higher differential expression when compared, as above to the microRNA expression at E15. Conclusion: We report the first comprehensive profile of developing murine lens transcriptome including both mRNA and miRNA through next-generation RNA sequencing. A complete repository of the lens transcriptome of six developmental time points will be monumental in elucidating processes essential for development of the ocular lens and maintenance its transparency.
Project description:Foxp1 was strongly expressed in developing lens, and its knockout in lens resulted in failure of appropriate lens development. By microarray analysis, we examined effects of loss-of Foxp1 for gene expression pattern of lens at P0 developmental stage.
Project description:We sequenced mRNA from 34 retina/RPE/choroid samples taken from the right eyes of male chicks across a time-course of normal development or refractive error induction (defocus-induced myopia and hyperopia).