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Individual Position Lumbar Blend: A planned out Assessment and Meta Evaluation.
Purpose To present a detailed, reliable long range-PCR and sequencing (LR-PCR-Seq) procedure to identify human opsin gene sequences for variations in the long wavelength-sensitive (OPN1LW), medium wavelength-sensitive (OPN1MW), short wavelength-sensitive (OPN1SW), and rhodopsin (RHO) genes. Methods Color vision was assessed for nine subjects using the Farnsworth-Munsell 100 hue test, Ishihara pseudoisochromatic plates, and the Rabin cone-contrast threshold procedure (ColorDX, Konan Medical). The color vision phenotypes were normal trichromacy (n = 3), potential tetrachromacy (n = 3), dichromacy (n = 2), and unexplained low color vision (n = 1). DNA was isolated from blood or saliva and LR-PCR amplified into individual products OPN1LW (4,045 bp), OPN1MW (4,045 bp), OPN1SW (3,326 bp), and RHO (6,715 bp). selleck Each product was sequenced using specific internal primer sets. Analysis was performed with Mutation Surveyor software. Results The LR-PCR-Seq technique identified known single nucleotide polymorphisms (SNPs) in OPN1LW and OPN1MW gene codons (180, 230, 233, 277, and 285), as well as those for lesser studied codons (174, 178, 236, 274, 279, 298 and 309) in the OPN1LW and OPN1MW genes. Additionally, six SNP variants in the OPN1MW and OPN1LW genes not previously reported in the NCBI dbSNP database were identified. An unreported poly-T region within intron 5(c.36+126) of the rhodopsin gene was also found, and analysis showed it to be highly conserved in mammalian species. Conclusions This LR-PCR-Seq procedure (single PCR reaction per gene followed by sequencing) can identify exonic and intronic SNP variants in OPN1LW, OPN1MW, OPN1SW, and rhodopsin genes. There is no need for restriction enzyme digestion or multiple PCR steps that can introduce errors. Future studies will combine the LR-PCR-Seq with perceptual behavior measures, allowing for accurate correlations between opsin genotypes, retinal photopigment phenotypes, and color perception behaviors. Copyright © 2020 Molecular Vision.Purpose To analyze the expression of 440 human cytokines in aqueous humor of high myopic patients with cataracts. Methods Eighty-five patients with cataracts were recruited in this study. In the screening stage, the RayBio G-Series Human Cytokine Antibody Array 440 was used to assay the aqueous humor samples collected from nine high myopic patients with cataracts and eight non-myopic patients with cataracts right before the surgery. The array was further used for verification of the screened cytokines, with aqueous humor samples obtained from 34 eyes of high myopic patients with cataracts and 34 eyes of non-myopic patients with cataracts. Results Compared with the non-myopic patients with cataracts, the expression levels of decorin, receptor activator of NF-kB (RANK), angiopoietin-1 (ANG-1), C-X-C motif ligand 16 (CXCL16), β-inducible gene-h3 (bIG-H3), insulin-like growth factor-binding protein 2 (IGFBP-2), and interleukin-17B (IL-17B) were statistically significantly higher in high myopic patients with cataracts (all p less then 0.000114). The matrix metalloproteinase-2 (MMP-2) level also increased in the aqueous humor of high myopic patients with cataracts (p = 0.0034). The concentrations of ANG-1 and MMP-2 were also increased in the aqueous humor of the confirmatory stage (all p less then 0.05). Conclusions In this study, numerous cytokines in aqueous humor were detected in high myopic patients with cataracts and non-myopic patients with cataracts, and it was confirmed that the MMP-2 level in the aqueous humor of patients with high myopia was statistically significantly increased. Further verification also revealed the elevation of ANG-1 in the aqueous humor of high myopic patients with cataracts, which suggests that ANG-1 may be related to the pathogenesis of high myopia. Copyright © 2020 Molecular Vision.Purpose To analyze whether activation of endogenous wingless (Wnt)/β-catenin signaling in Müller cells is involved in protection of retinal ganglion cells (RGCs) following excitotoxic damage. Methods Transgenic mice with a tamoxifen-dependent β-catenin deficiency in Müller cells were injected with N-methyl-D-aspartate (NMDA) into the vitreous cavity of one eye to induce excitotoxic damage of the RGCs, while the contralateral eye received PBS only. Retinal damage was quantified by counting the total number of RGC axons in cross sections of optic nerves and measuring the thickness of the retinal layers on meridional sections. Then, terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) assay was performed to identify apoptotic cells in retinas of both genotypes. Western blot analyses to assess the level of retinal β-catenin and real-time RT-PCR to quantify the retinal expression of neuroprotective factors were performed. Results Following NMDA injection of wild-type mice, a statistically significant increase in retinal β-catenin protein levels was observed compared to PBS-injected controls, an effect that was blocked in mice with a Müller cell-specific β-catenin deficiency. Furthermore, in mice with a β-catenin deficiency in Müller cells, NMDA injection led to a statistically significant decrease in RGC axons as well as a substantial increase in TUNEL-positive cells in the RGC layer compared to the NMDA-treated controls. Moreover, in the retinas of the control mice a NMDA-mediated statistically significant induction of leukemia inhibitory factor (Lif) mRNA was detected, an effect that was substantially reduced in mice with a β-catenin deficiency in Müller cells. Conclusions Endogenous Wnt/β-catenin signaling in Müller cells protects RGCs against excitotoxic damage, an effect that is most likely mediated via the induction of neuroprotective factors, such as Lif. Copyright © 2020 Molecular Vision.Purpose To detect the differential expression of long non-coding RNAs (lncRNAs) in the ocular posterior poles of two guinea pig myopia models and explore the pathogenic role of lncRNAs in myopia. Methods Form-deprived myopia (FDM) and lens-induced myopia (LIM) models were induced in guinea pig right eyes by wearing a translucent latex balloon head mask and a -10.00 diopter (D) lens, respectively. Ocular biometric parameters were measured biweekly. At 6 weeks after the induction of myopia, the guinea pig eyeballs were processed for hematoxylin and eosin staining to examine the ocular morphology. The ocular posterior poles from the normal control, FDM, and LIM groups were collected to analyze the differential expression of lncRNAs between the groups with high-throughput RNA sequencing (RNA-seq). Further, the lncRNA-mRNA colocation network was delineated to predict the functions of the differentially expressed lncRNAs. Last, Gene Ontology (GO) functional enrichment analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis were performed on the colocated mRNAs of the differentially expressed lncRNAs.
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