Transplantation of Chemical Compound-Induced Cells from Human Fibroblasts Improves Locomotor Recovery in a Spinal Cord Injury Rat Model.

The development of regenerative medicine using cell therapy is eagerly awaited for diseases such as spinal cord injury (SCI), for which there has been no radical cure. We previously reported the direct conversion of human fibroblasts into neuronal-like cells using only chemical compounds; however, it is unclear whether chemical compound-induced neuronal-like (CiN) cells are clinically functional. In this study, we partially modified the method of inducing CiN cells (termed immature CiN cells) and examined their therapeutic efficacy, in a rat model of SCI, to investigate whether immature CiN cells are promising for clinical applications. Motor function recovery, after SCI, was assessed using the Basso, Beattie, and Bresnahan (BBB) test, as well as the CatWalk analysis. We found that locomotor recovery, after SCI in the immature CiN cell-transplanted group, was partially improved compared to that in the control group. Consistent with these results, magnetic resonance imaging (MRI) and histopathological analyses revealed that nerve recovery or preservation improved in the immature CiN cell-transplanted group. Furthermore, transcriptome analysis revealed that immature CiN cells highly express hepatocyte growth factor (HGF), which has recently been shown to be a promising therapeutic agent against SCI. Our findings suggest that immature CiN cells may provide an alternative strategy for the regenerative therapy of SCI.

Defective biosynthesis of ascorbic acid in Sod1-deficient mice results in lethal damage to lung tissue.

Superoxide dismutase 1 (Sod1) plays pivotal roles in antioxidation via accelerating the conversion of superoxide anion radicals into hydrogen peroxide, thus inhibiting the subsequent radical chain reactions. While Sod1 deficient cells inevitably undergo death in culture conditions, Sod1-knockout (KO) mice show relatively mild phenotypes and live approximately two years. We hypothesized that the presence of abundant levels of ascorbic acid (AsA), which is naturally produced in mice, contributes to the elimination of reactive oxygen species (ROS) in Sod1-KO mice. To verify this hypothesis, we employed mice with a genetic ablation of aldehyde reductase (Akr1a), an enzyme that is involved in the biosynthesis of AsA, and established double knockout (DKO) mice that lack both Sod1 and Akr1a. Supplementation of AsA (1.5 mg/ml in drinking water) was required for the DKO mice to breed, and, upon terminating the AsA supplementation, they died within approximately two weeks regardless of age or gender. We explored the etiology of the death from pathophysiological standpoints in principal organs of the mice. Marked changes were observed in the lungs in the form of macroscopic damage after the AsA withdrawal. Histological and immunological analyses of the lungs indicated oxidative damage of tissue and activated immune responses. Thus, preferential oxidative injury that occurred in pulmonary tissues appeared to be primary cause of the death in the mice. These collective results suggest that the pivotal function of AsA in coping with ROS in vivo, is largely in pulmonary tissues that are exposed to a hyperoxygenic microenvironment.

Testis-specific peroxiredoxin 4 variant is not absolutely required for spermatogenesis and fertility in mice.

PRDX4, a member of peroxiredoxin family, is largely concentrated in the endoplasmic reticulum (ER) and plays a pivotal role in the redox relay during oxidative protein folding as well as in peroxidase reactions. A testis-specific PRDX4 variant transcript (PRDX4t) lacks the conventional exon 1, which encodes the signal peptide that is required for entry into the ER lumen, but instead carries alternative exon 1, which is transcribed from the upstream promoter in a testis-specific manner and results in the PRDX4t protein being localized in the cytosol. However, the potential roles of PRDX4t in male genital action remain unknown. Using a CRISPR/Cas9 system, we first disrupted the testis-specific promoter/exon 1 and generated mice that were specifically deficient in PRDX4t. The resulting PRDX4t knockout (KO) mice underwent normal spermatogenesis and showed no overt abnormalities in the testis. Mating PRDX4t KO male mice with wild-type (WT) female mice produced normal numbers of offspring, indicating that a PRDX4t deficiency alone had no effect on fertility in the male mice. We then generated mice lacking both PRDX4 and PRDX4t by disrupting exon 2, which is communal to these variants. The resulting double knockout (DKO) mice were again fertile, and mature sperm isolated from the epididymis of DKO mice exhibited a normal fertilizing ability in vitro. In the meantime, the protein levels of glutathione peroxidase 4 (GPX4), which plays an essential role in the disulfide bond formation during spermatogenesis, were significantly increased in the testis and caput epididymis of the DKO mice compared with the WT mice. Based on these results, we conclude that the disruption of the function of PRDX4t in the spermatogenic process appears to be compensated by other factors including GPX4.

Double Knockout of Peroxiredoxin 4 (Prdx4) and Superoxide Dismutase 1 (Sod1) in Mice Results in Severe Liver Failure.

Mice that are deficient in superoxide dismutase 1 (Sod1), an antioxidative enzyme, are susceptible to developing liver steatosis. Peroxiredoxin 4 (Prdx4) catalyzes disulfide bond formation in proteins via the action of hydrogen peroxide and hence decreases oxidative stress and supports oxidative protein folding for the secretion of lipoproteins. Because elevated reactive oxygen species induce endoplasmic reticulum stress, this negative chain reaction is likely involved in the development of nonalcoholic fatty liver diseases and more advanced steatohepatitis (NASH). In the current study, we generated Prdx4 and Sod1 double knockout (DKO; Prdx4-/ySod1-/-) mice and examined whether the combined deletion of Prdx4 and Sod1 aggravated liver pathology compared to single knockout and wild-type mice. The secretion of triglyceride-rich lipoprotein was strikingly impaired in the DKO mice, leading to aggravated liver steatosis. Simultaneously, the activation of caspase-3 in the liver was observed. The hyperoxidation of Prdxs, a hallmark of oxidative stress, occurred in different isoforms that are uniquely associated with Sod1-/- and Prdx4-/y mice, and the effect was additive in DKO mouse livers. Because DKO mice spontaneously develop severe liver failure at a relatively young stage, they have the potential for use as a model for hepatic disorders and for testing other potential treatments.

Decreased reproductive performance in xCT-knockout male mice.

Sulphoxidation occurs in protamines that are enriched in cysteine and supplies chromatin for packaging. The extracellular fluid contains higher levels of oxidised cysteine (cystine), and some cells utilise system xc-, a cystine transporter in which xCT is the main protein component, to fulfil the need for cysteine. We hypothesised that system xc- might ensure the supply of cysteine needed for spermatogenesis. The reproductive ability of xCT-/- male mice at 6- to 18-weeks of age appeared to be lower than xCT+/+ male mice. The courtship behaviour of the xCT-/- male mice was undynamic, which appeared to be associated with the low reproductive ability of xCT-/- male mice. xCT was found to be expressed in mouse testes, notably in Sertoli cells, as well as in the epididymis and the levels were increased at the time of sexual maturation. Despite the normal histological appearance of testicular tissues, the cauda epididymis of xCT-/- mice contained round, greater numbers of immature spermatogenic cells than that of xCT+/+ mice. However, there were no significant differences in the numbers of sperm stored in the cauda epididymis or in the concentrations of cysteine or glutathione in the testes. The resulting sperm had normal fertilising ability. Thus, system xc- appears to function as a backup system for supplying cysteine to testes and play a pivotal role in supplying cysteine for normal sexual behaviour by a mechanism that is different from that for the supply of cysteine in spermatogenesis.

Protective role of testis-specific peroxiredoxin 4 against cellular oxidative stress.

Peroxiredoxin (PRDX), a newly discovered antioxidant enzyme, has an important role in hydrogen peroxide reduction. Among six PRDX genes (PRDX1-6) in mammals, PRDX4 gene is alternatively spliced to produce the somatic cell form (PRDX4) and the testis specific form (PRDX4t). In our previous study, PRDX4 knockout mice displayed testicular atrophy with an increase in cell death due to oxidative stress. However, the antioxidant function of PRDX4t is unknown. In this study, we demonstrate that PRDX4t plays a protective role against oxidative stress in the mammalian cell line HEK293T. The PRDX4t-EGFP plasmid was transferred into HEK293T cells; protein expression was confirmed in the cytoplasm. To determine the protective role of PRDX4t in cells, we performed image-based analysis of PRDX4t-EGFP expressed cells exposed to UV irradiation and hydrogen peroxide using fluorescent probe CellROX. Our results suggested that PRDX4t-EGFP expressed cells had reduced levels of oxidative stress compared with cells that express only EGFP. This study highlights that PRDX4t plays an important role in cellular antioxidant defense.

xCT deficiency aggravates acetaminophen-induced hepatotoxicity under inhibition of the transsulfuration pathway.

Cystine, an oxidized form of cysteine (Cys), is imported into cells via the protein xCT, which is also associated with the export of glutamate as the counter amino acid. In the current study, we attempted to rationalize roles of xCT in the livers of male mice. While xCT was not expressed in the livers of ordinary mice, it was induced under conditions of glutathione depletion, caused by the administration of acetaminophen (AAP). To differentiate the role between xCT and the transsulfuration pathway on the supply of Cys, we employed an inhibitor of the enzyme cystathionine γ-lyase, propargylglycine (PPG). This inhibitor caused a marked aggravation in AAP-induced hepatic damage and the mortality of the xCT-/- mice was increased to a greater extent than that for the xCT+/+ mice. While a PPG pretreatment had no effect on liver condition or Cys levels, the administration of AAP to the PPG-pretreated mice reduced the levels of Cys as well as glutathione to very low levels in both the xCT+/+ and xCT-/- mice. These findings indicate that the transsulfuration pathway plays a major role in replenishing Cys when glutathione levels are low. Moreover, an ascorbic acid insufficiency, induced by Akr1a ablation, further aggravated the AAP-induced liver damage in the case of the xCT deficiency, indicating that glutathione and ascorbic acid function cooperatively in protecting the liver. In conclusion, while the transsulfuration pathway plays a primary role in supplying Cys to the redox system in the liver, xCT is induced in cases of emergencies, by compensating for Cys supply systems.

Oxidative stress caused by a SOD1 deficiency ameliorates thioacetamide-triggered cell death via CYP2E1 inhibition but stimulates liver steatosis.

We investigated the responses of mice that are defective in the superoxide-scavenging enzyme SOD1 to thioacetamide (TAA)-induced hepatotoxicity. When a lethal dose of TAA (500 mg/kg) was intraperitoneally injected, the wild-type (WT) mice all died within 36 h, but all of the SOD1-knockout (KO) mice survived. Treatment with an SOD1 inhibitor rendered the WT mice resistant to TAA toxicity. To elucidate the mechanism responsible for this, we examined the acute effects of a sublethal dose of TAA (200 mg/kg) on the livers of WT and KO mice. The extent of TAA-induced liver damage was less in the KO mice, but, instead, lipogenesis was further advanced in the SOD1-KO livers. The levels of proteins modified with acetyllysine, a marker for TAA-mediated injury, were lower in the KO mice than the WT mice upon the TAA treatment. The KO mice, which were under oxidative stress per se, exhibited a lower CYP2E1 activity, and this appeared to result in a decrease in the production of reactive oxygen species (ROS) during TAA metabolism. Both cleaved ATF6, a transcriptional regulator that is activated by endoplasmic reticulum (ER) stress, and CHOP, a death signal mediator, were highly elevated in the WT mice as the result of the TAA treatment and consistent with the liver damage. We conclude that elevated TAA metabolites and reactive oxygen species that are produced by CYP-mediated drug metabolism trigger lipogenesis as well as liver damage via ER stress and determine the fate of the mice.

A high-fat diet temporarily renders Sod1-deficient mice resistant to an oxidative insult.

Patients with nonalcoholic fatty liver disease may subsequently develop nonalcoholic steatohepatitis after suffering from a second insult, such as oxidative stress. Aim of this study was to investigate the pathogenesis of the liver injury caused when lipids accumulate under conditions of intrinsic oxidative stress using mice that are deficient in superoxide dismutase 1 (SOD1) and the leptin receptor (Lepr). We established Sod1-/-::Leprdb/db mice and carried out analyses of four groups of genetically modified mice, namely, wild type, Sod1-/-, Leprdb/db and Sod1-/-::Leprdb/db mice. Mice with defects in the SOD1 or Lepr gene are vulnerable to developing fatty livers, even when fed a normal diet. Feeding a high-fat diet (HFD) caused an increase in the number of lipid droplets in the liver to different extents in each genotypic mouse. an HFD caused the accelerated death of db/db mice, but contradictory to our expectations, the death rates for the Sod1-deficient mice were decreased by feeding HFD. Consistent with the improved probability of survival, liver damage was significantly ameliorated by feeding an HFD compared to a normal diet in the mice with an Sod1-deficient background. Oxidative stress markers, hyperoxidized peroxiredoxin and lipid peroxidation products, were decreased somewhat in Sod1-/- mice by feeding HFD. We conclude that lipids reacted with reactive oxygen species and eliminated them in the livers of the young mice, which resulted in the alleviation of oxidative stress, but in advanced age oxidized products accumulated, leading to the aggravation of the liver injury and an increase in fatality rate.

Ascorbic acid prevents acetaminophen-induced hepatotoxicity in mice by ameliorating glutathione recovery and autophagy.

Aldehyde reductase (AKR1A) plays a role in the biosynthesis of ascorbic acid (AsA), and AKR1A-deficient mice produce about 10-15% of the AsA that is produced by wild-type mice. We found that acetaminophen (AAP) hepatotoxicity was aggravated in AKR1A-deficient mice. The pre-administration of AsA in the drinking water markedly ameliorated the AAP hepatotoxicity in the AKR1A-deficient mice. Treatment of the mice with AAP decreased both glutathione and AsA levels in the liver in the early phase after AAP administration, and an AsA deficiency delayed the recovery of the glutathione content in the healing phase. While in cysteine supply systems; a neutral amino acid transporter ASCT1, a cystine transporter xCT, enzymes for the transsulfuration pathway, and autophagy markers, were all elevated in the liver as the result of the AAP treatment, the AsA deficiency suppressed their induction. Thus, AsA appeared to exert a protective effect against AAP hepatotoxicity by ameliorating the supply of cysteine that is available for glutathione synthesis as a whole. Because some drugs produce reactive oxygen species, resulting in the consumption of glutathione during the metabolic process, the intake of sufficient amounts of AsA would be beneficial for protecting against the hepatic damage caused by such drugs.

Trichloroethylene exposure aggravates behavioral abnormalities in mice that are deficient in superoxide dismutase.

Trichloroethylene (TCE) has been implicated as a causative agent for Parkinson's disease (PD). The administration of TCE to rodents induces neurotoxicity associated with dopaminergic neuron death, and evidence suggests that oxidative stress as a major player in the progression of PD. Here we report on TCE-induced behavioral abnormality in mice that are deficient in superoxide dismutase 1 (SOD1). Wild-type (WT) and SOD1-deficient (Sod1(-/-)) mice were intraperitoneally administered TCE (500 mg/kg) over a period of 4 weeks. Although the TCE-administrated Sod1(-/-) mice showed marked abnormal motor behavior, no significant differences were observed among the experimental groups by biochemical and histopathological analyses. However, treating mouse neuroblastoma-derived NB2a cells with TCE resulted in the down regulation of the SOD1 protein and elevated oxidative stress under conditions where SOD1 production was suppressed. Taken together, these data indicate that SOD1 plays a pivotal role in protecting motor neuron function against TCE toxicity.

The SOD1 transgene expressed in erythroid cells alleviates fatal phenotype in congenic NZB/NZW-F1 mice.

Oxidative stress due to a superoxide dismutase 1 (SOD1) deficiency causes anemia and autoimmune responses, which are phenotypically similar to autoimmune hemolytic anemia (AIHA) and systemic lupus erythematosus (SLE) in C57BL/6 mice and aggravates AIHA pathogenesis in New Zealand black (NZB) mice. We report herein on an evaluation of the role of reactive oxygen species (ROS) in a model mouse with inherited SLE, that is, F1 mice of the NZB × New Zealand white (NZW) strain. The ROS levels within red blood cells (RBCs) of the F1 mice were similar to the NZW mice but lower compared to the NZB mice throughout adult period. Regarding SLE pathogenesis, we examined the effects of an SOD1 deficiency or the overexpression of human SOD1 in erythroid cells by establishing corresponding congenic F1 mice. A SOD1 deficiency caused an elevation in ROS production, methemoglobin content, and hyperoxidation of peroxiredoxin in RBC of the F1 mice, which were all consistent with elevated oxidative stress. However, while the overexpression of human SOD1 in erythroid cells extended the life span of the congenic F1 mice, the SOD1 deficiency had no effect on life span compared to wild-type F1 mice. It is generally recognized that NZW mice possess a larval defect in the immune system and that NZB mice trigger an autoimmune reaction in the F1 mice. Our results suggest that the oxidative insult originated from the NZB mouse background has a functional role in triggering an aberrant immune reaction, leading to fatal responses in F1 mice.

Overexpression of Peroxiredoxin 4 Affects Intestinal Function in a Dietary Mouse Model of Nonalcoholic Fatty Liver Disease.

BACKGROUND: Accumulating evidence has shown that methionine- and choline-deficient high fat (MCD+HF) diet induces the development of nonalcoholic fatty liver disease (NAFLD), in which elevated reactive oxygen species play a crucial role. We have reported that peroxiredoxin 4 (PRDX4), a unique secretory member of the PRDX antioxidant family, protects against NAFLD progression. However, the detailed mechanism and potential effects on the intestinal function still remain unclear. METHODS & RESULTS: Two weeks after feeding mice a MCD+HF diet, the livers of human PRDX4 transgenic (Tg) mice exhibited significant suppression in the development of NAFLD compared with wild-type (WT) mice. The serum thiobarbituric acid reactive substances levels were significantly lower in Tg mice. In contrast, the Tg small intestine with PRDX4 overexpression showed more suppressed shortening of total length and villi height, and more accumulation of lipid in the jejunum, along with lower levels of dihydroethidium binding. The enterocytes exhibited fewer apoptotic but more proliferating cells, and inflammation was reduced in the mucosa. Furthermore, the small intestine of Tg mice had significantly higher expression of cholesterol absorption-regulatory factors, including liver X receptor-α, but lower expression of microsomal triglyceride-transfer protein. CONCLUSION: Our present data provide the first evidence of the beneficial effects of PRDX4 on intestinal function in the reduction of the severity of NAFLD, by ameliorating oxidative stress-induced local and systemic injury. We can suggest that both liver and intestine are spared, to some degree, by the antioxidant properties of PRDX4.

An SOD1 deficiency enhances lipid droplet accumulation in the fasted mouse liver by aborting lipophagy.

Under normal feeding conditions, oxidative stress stimulates lipid droplets accumulation in hepatocytes. We found that, despite the low visceral fat in Sod1-knockout (KO) mouse, lipid droplets accumulate in the liver to a greater extent than for the wild-type mouse upon fasting. Liver damage became evident in the KO mice. While fasting caused substantial endoplasmic reticulum stress in KO mice, the expression of genes involved in fatty acid production was suppressed. LC3-II, which is essential for the dynamic process of autophagosome formation, was activated in the wild-type mouse and enhanced in the KO mouse. However, the p62, an adapter protein with the ubiquitin- and LC3-binding activity, accumulated abnormally in the livers of KO mice, implying an abortive lipophagic process as the cause for the impaired lipid metabolism and the hepatic damage that occurs upon fasting.

SOD1 deficiency decreases proteasomal function, leading to the accumulation of ubiquitinated proteins in erythrocytes.

We previously demonstrated that elevated levels of ROS in red blood cells (RBCs) are responsible for anemia in SOD1-deficient mice, suggesting that the oxidative stress-induced massive destruction of RBCs is an underlying mechanism for autoimmune hemolytic anemia. In the current study, we examined the issue of how elevated ROS are involved in the destruction of RBCs and the onset of anemia from the view point of the proteolytic removal of oxidatively-damaged proteins. We found that poly-ubiquitinated proteins had accumulated and had undergone aggregation in RBCs from SOD1-deficient mice and from phenylhydrazine-induced anemic mice. Although the protein levels of the three catalytic components of the proteasome, ß1, ß2, and ß5, were not significantly altered, their proteolytic activities were decreased in the SOD1-deficient RBCs. These data suggest that oxidative-stress triggers the dysfunction of the proteasomal system, which results in the accumulation of the aggregation of poly-ubiquitinated proteins. We conclude that an oxidative stress-induced malfunction in the scavenging activity of proteasomes accelerates the accumulation of damaged proteins, leading to a shortened lifespan of RBCs and, hence, anemia.

SOD1 deficiency induces the systemic hyperoxidation of peroxiredoxin in the mouse.

A deficiency of superoxide dismutase 1 (SOD1) or peroxiredoxin (Prx) 2 causes anemia in mice due to elevated oxidative stress. In the current study, we investigated whether intrinsic oxidative stress caused by a SOD1 deficiency affected the redox status of Prx2 and other isoforms in red blood cells (RBCs) and several organs of mice. We observed a marked elevation in hyperoxidized Prx2 levels in RBCs from SOD1-deficient mice. Hyperoxidized Prx2 reportedly undergoes a rhythmic change in isolated RBCs under culture conditions. We confirmed such changes in RBCs from wild-type mice but observed no evident changes in SOD1-deficient RBCs. In addition, an elevation in hyperoxidized Prxs, notably Prx2 and Prx3, was observed in several organs from SOD1-deficient mice. However, a SOD1 deficiency had no impact on the wheel-running activity of the mice. Thus, although the redox status of some Prxs is systemically shifted to a more oxidized state as the result of a SOD1 deficiency, which is associated with anemia and some diseases, a redox imbalance appears to have no detectable effect on the circadian activity of mice.

Oxidative stress triggers lipid droplet accumulation in primary cultured hepatocytes by activating fatty acid synthesis.

Despite the impaired intestinal lipid absorption and low level of visceral fat, the Sod1-deficient mouse is susceptible to developing liver steatosis. To gain insights into the mechanism responsible for this abnormal lipid metabolism, we analyzed primary cultured hepatocytes obtained from Sod1-deficient and wild-type mice. Lipid droplets began to accumulate in the cultured hepatocytes and was further increased by a Sod1 deficiency. Levels of enzymes involved in lipogenesis were elevated. It thus appears that lipogenesis is activated by oxidative stress, which is more prominent in the case of Sod1 deficiency, and appears to participate in liver steatosis.

Oxidative stress as a potential causal factor for autoimmune hemolytic anemia and systemic lupus erythematosus.

The kidneys and the blood system mutually exert influence in maintaining homeostasis in the body. Because the kidneys control erythropoiesis by producing erythropoietin and by supporting hematopoiesis, anemia is associated with kidney diseases. Anemia is the most prevalent genetic disorder, and it is caused by a deficiency of glucose 6-phosphate dehydrogenase (G6PD), for which sulfhydryl oxidation due to an insufficient supply of NADPH is a likely direct cause. Elevated reactive oxygen species (ROS) result in the sulfhydryl oxidation and hence are another potential cause for anemia. ROS are elevated in red blood cells (RBCs) under superoxide dismutase (SOD1) deficiency in C57BL/6 mice. SOD1 deficient mice exhibit characteristics similar to autoimmune hemolytic anemia (AIHA) and systemic lupus erythematosus (SLE) at the gerontic stage. An examination of AIHA-prone New Zealand Black (NZB) mice, which have normal SOD1 and G6PD genes, indicated that ROS levels in RBCs are originally high and further elevated during aging. Transgenic overexpression of human SOD1 in erythroid cells effectively suppresses ROS elevation and ameliorates AIHA symptoms such as elevated anti-RBC antibodies and premature death in NZB mice. These results support the hypothesis that names oxidative stress as a risk factor for AIHA and other autoimmune diseases such as SLE. Herein we discuss the association between oxidative stress and SLE pathogenesis based mainly on the genetic and phenotypic characteristics of NZB and New Zealand white mice and provide insight into the mechanism of SLE pathogenesis.

Physiological and pathological views of peroxiredoxin 4.

Peroxiredoxins (PRDXs) form an enzyme family that exhibits peroxidase activity using electrons from thioredoxin and other donor molecules. As the signaling roles of hydrogen peroxide in response to extracellular stimuli have emerged, the involvement of PRDX in the hydrogen peroxide-mediated signaling has become evident. Among six PRDX members in mammalian cells, PRDX4 uniquely possesses a hydrophobic signal peptide at the amino terminus, and, hence, it undergoes either secretion or retention by the endoplasmic reticulum (ER) lumen. The role of PRDX4 as a sulfoxidase in ER is now attracting much attention regarding the oxidative protein folding of nascent proteins. Contrary to this role in the ER, the functional significance of PRDX4 in the extracellular milieu is virtually unknown despite its implications as a biomarker under pathological conditions in some diseases. Other than its systemically expressed form, a variant form of PRDX4 is transcribed from the upstream promoter/exon 1 of the systemic promoter/exon 1 and is uniquely expressed in sexually matured testes. Circumstantial evidence, together with deduced functions from the systemic form, suggests that there are potential roles for testicular PRDX4 in the reproductive processes such as the regulation of hormonal signals and the oxidative packaging of sperm chromatin. Elucidation of these PRDX4 functions under in vivo situations is expected to show the whole picture of how PRDX4 has evolved in multicellular organisms.

Reductive detoxification of acrolein as a potential role for aldehyde reductase (AKR1A) in mammals.

Aldehyde reductase (AKR1A), a member of the aldo-keto reductase superfamily, suppresses diabetic complications via a reduction in metabolic intermediates; it also plays a role in ascorbic acid biosynthesis in mice. Because primates cannot synthesize ascorbic acid, a principle role of AKR1A appears to be the reductive detoxification of aldehydes. In this study, we isolated and immortalized mouse embryonic fibroblasts (MEFs) from wild-type (WT) and human Akr1a-transgenic (Tg) mice and used them to investigate the potential roles of AKR1A under culture conditions. Tg MEFs showed higher methylglyoxal- and acrolein-reducing activities than WT MEFs and also were more resistant to cytotoxicity. Enzymatic analyses of purified rat AKR1A showed that the efficiency of the acrolein reduction was about 20% that of glyceraldehyde. Ascorbic acid levels were quite low in the MEFs, and while the administration of ascorbic acid to the cells increased the intracellular levels of ascorbic acid, it had no affect on the resistance to acrolein. Endoplasmic reticulum stress and protein carbonylation induced by acrolein treatment were less evident in Tg MEFs than in WT MEFs. These data collectively indicate that one of the principle roles of AKR1A in primates is the reductive detoxification of aldehydes, notably acrolein, and protection from its detrimental effects.