Oxidative stress is one of the causative factors in progression and etiology of agerelated cataract. Peroxiredoxin 6 (Prdx6), a savior for cells from internal or external environmental stresses, plays a role in cellular signaling by detoxifying reactive oxygen species (ROS) and thereby controlling gene regulation. Using targeted inactivation of the Prdx6 gene, we show that Prdx6-deficient lens epithelial cells (LECs) are more vulnerable to UV-triggered cell death, a major cause of skin disorders including cataractogenesis, and these cells display abnormal protein profiles. PRDX6-depleted LECs showed phenotypic changes and formed lentoid body, a characteristic of terminal cell differentiation and epithelial-mesenchymal transition. Prdx6 Ϫ/Ϫ LECs exposed to UV-B showed higher ROS expression and were prone to apoptosis compared with wild-type LECs, underscoring a protective role for Prdx6. Comparative proteomic analysis using fluorescence-based difference gel electrophoresis along with mass spectrometry and database searching revealed a total of 13 proteins that were differentially expressed in Prdx6 Ϫ/Ϫ cells. Six proteins were upregulated, whereas expression of seven proteins was decreased compared with Prdx6LECs. Among the cytoskeleton-associated proteins that were highly expressed in Prdx6-deficient LECs was tropomyosin (Tm)2. Protein blot and real-time PCR validated dramatic increase of Tm2 and Tm1␣ expression in these cells. Importantly, Prdx6 ϩ/ϩ LECs showed a similar pattern of Tm2 protein expression after transforming growth factor (TGF)- or H2O2 treatment. An extrinsic supply of PRDX6 could restore Tm2 expression, demonstrating that PRDX6 may attenuate adverse signaling in cells and thereby maintain cellular homeostasis. Exploring redox-proteomics (Prdx6 Ϫ/Ϫ ) and characterization and identification of abnormally expressed proteins and their attenuation by PRDX6 delivery should provide a basis for development of novel therapeutic interventions to postpone ROS-mediated abnormal signaling deleterious to cells or tissues.peroxiredoxin; oxidative stress; proteomics; tropomyosin OXIDATIVE STRESS has been identified as one of the major causes of age-related diseases, including cataracts (1, 45, 62, 74 -76, 82). Oxidative stress-induced etiology and progression of diseases may result either from diminished natural antioxidants such as catalase, glutathione peroxidase, and peroxiredoxins (PRDXs) due to aging or from increased generation of reactive oxygen species (ROS) by exposure to environmental factors, X-rays, chemicals, toxins, and ultraviolet (UV) radiation. Environmental stressors such as these have been heavily implicated in the etiology and progression of several diseases by inducing ROS-mediated oxidative stress.UV radiation exposure induces several complex sets of acute or chronic responses that can lead to initiation of diseases through overproduction of ROS. ROS modify cellular signaling. Modulation of signal transduction pathways includes changes in gene expression (18). However, investigation...
Injury to lens epithelial cells (LECs) leads to epithelial–mesenchymal transition (EMT) with resultant fibrosis. The tropomyosin (Tpm) family of cytoskeleton proteins is involved in regulating and stabilizing actin microfilaments. Aberrant expression of Tpms leads to abnormal morphological changes with disintegration of epithelial integrity. The EMT of LECs has been proposed as a major cause of posterior capsule opacification (PCO) after cataract surgery. Using in vivo rodent PCO and human cataractous LECs, we demonstrated that the aberrant expression of rat Tpm and human Tpm1α/2β suggested their association in remodelling of the actin cytoskeleton during EMT of LECs. Expression analysis from abnormally growing LECs after lens extraction revealed elevated expression of α-smooth muscle actin (α-SMA), a marker for EMT. Importantly, these cells displayed increased expression of Tpm1α/2β following EMT/PCO formation. Expression of Tpm1α/2β was up-regulated in LECs isolated from cataractous lenses of Shumiya Cataract Rats (SCRs), compared with non-cataractous lenses. Also, LECs from human patients with nuclear cataract and anterior subcapsular fibrosis (ASF) displayed significantly increased expression of Tpm2β mRNA, suggesting that similar signalling invokes the expression of these molecules in LECs of cataractous SCR and human lenses. EMT was observed in LECs overexpressed with Tpm1α/2β, as evidenced by increased expression of α-SMA. These conditions were correlated with remodelling of actin filaments, possibly leading to EMT/PCO and ASF. The present findings may help clarify the condition of the actin cytoskeleton during morphogenetic EMT, and may contribute to development of Tpm-based inhibitors for postponing PCO and cataractogenesis.
Peripheral neuropathy develops in human subjects with prediabetes and metabolic syndrome, prior to overt hyperglycemia. The contributions of impaired glucose tolerance and insulin signaling, hypertriglyceridemia and/or increased NEFA, and hypercholesterolemia to this condition remain unknown. Niacin and its derivatives alleviate dyslipidemia with a minor effect on glucose homeostasis. This study evaluated the roles of impaired glucose tolerance versus dyslipidemia in prediabetic neuropathy using Zucker fatty (fa/fa) rats and the niacin derivative acipimox, as well as the interplay of hypertriglyceridemia, increased NEFA, and oxidative-nitrosative stress. 16 wk-old Zucker fatty rats with impaired glucose tolerance, obesity, hyperinsulinemia, hypertriglyceridemia, hypercholesterolemia, and increased NEFA, displayed sensory nerve conduction velocity deficit, thermal and mechanical hypoalgesia, and tactile allodynia. Acipimox (100 mgkg−1d−1, 4 weeks) reduced serum insulin, NEFA, and triglyceride concentrations without affecting glucose tolerance and hypercholesterolemia. It alleviated sensory nerve conduction velocity deficit, changes in behavioral measures of sensory function, and corrected oxidative-nitrosative stress, but not impaired insulin signaling, in peripheral nerve. Elevated NEFA increased total and mitochondrial superoxide production and NAD(P)H oxidase activity in cultured human Schwann cells. In conclusion, hypertriglyceridemia and/or increased NEFA concentrations cause prediabetic neuropathy through oxidative-nitrosative stress. Lipid-lowering agents and antioxidants may find use in management of this condition.
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