Methionine Sulfoxide Reductase B3 Research Articles

Exploration of hub genes, lipid metabolism, and the immune microenvironment in stomach carcinoma and cholangiocarcinoma.

BackgroundGastric cancer (GC) is the 5th most common cause of cancer in the world and the 3rd largest cause of cancer-related death. It is usually associated with a variety of cancers, of which cholangiocarcinoma (CCA) combined with GC accounts for about 1.6%. This study sought to examine the hub genes and role of lipid metabolism in the development and diagnosis of GC and CCA.MethodsTo screen potential hub genes, The Cancer Genome Atlas (TCGA) data sets, including the GC (STAD, dataset of GC) and CCA (CHOL, dataset of CCA) data sets, were used to conduct a differentially expressed gene (DEG) analysis and an enrichment analysis of the DEGs. A weighted-gene co-expression network analysis (WGCNA) was conducted to identify the significant gene module and then find the hub genes in the module. To verify the 4 hub genes, we conducted a differentiation analysis of the 4 genes in GC and CCA and found that there were differences. A survival analysis of the hub genes was performed and mutations were mapped. Additionally, tumor immune microenvironment (TIME) and immune analyses were performed to evaluate how lipid metabolism affects the development of GC with CCA.ResultsThe principal component analysis showed that both GC and CCA had distinct up-regulated and down-regulated genes, which are involved in a variety of metabolic processes. Upon WGCNA, the turquoise and blue modules were meaningful, and the hub genes were identified from these 2 modules. Four hub genes were identified: amyloid beta precursor protein binding family B member 1 (APBB1), Homo sapiens armadillo repeat containing X-linked 1 (ARMCX1), DAZ interacting zinc finger protein 1 (DZIP1), and methionine sulfoxide reductase B3 (MSRB3). In survival analysis, increased expression of the 4 hub genes was associated with inferior survival outcomes, with variations in all 4 genes. Additionally, we demonstrated that genes related to lipid metabolism had an effect on immune function.ConclusionsAPBB1, ARMCX1, DZIP1, and MSRB3 affect the development of GC and CCA and can be used as biomarkers. The expression of lipid metabolism genes is related to the TIME of patients with GC and CCA.

Methionine sulfoxide reductase B3 deficiency inhibits the development of diet-induced insulin resistance in mice

Oxidative and endoplasmic reticulum (ER) stress are involved in mediating high-fat diet (HFD)-induced insulin resistance. As the ER-localized methionine sulfoxide reductase B3 (MsrB3) protects cells against oxidative and ER stress, we hypothesized that MsrB3 might be associated with HFD-induced insulin resistance. To test this hypothesis, we examined the effect of MsrB3 deficiency on HFD-induced insulin resistance using MsrB3 knockout (KO) mice. Mice were fed a control diet or HFD for 12 weeks and insulin sensitivity was measured using a hyperinsulinemic-euglycemic clamp. HFD consumption increased the body weight of both wild-type and MsrB3 KO mice, and no significant difference was observed between the genotypes. The HFD increased oxidative stress and induced insulin resistance in the skeletal muscle of wild-type mice, but did not affect either in MsrB3 KO mice. The unfolded protein response (UPR) was increased in MsrB3 KO mice upon consumption of HFD, but not in wild-type mice. Mitochondrial oxidative phosphorylation proteins and the levels of superoxide dismutase 2 and glutathione peroxidase 1 were increased in MsrB3 KO mice upon HFD consumption. The respiratory control ratio was reduced in wild-type mice consuming HFD but not in MsrB3 KO mice. The levels of calcium/calmodulin-dependent protein kinase kinase β, phosphorylated AMP-activated protein kinase, and peroxisome proliferator-activated receptor gamma coactivator 1α were increased in MsrB3 KO mice following HFD consumption. These results suggest that MsrB3 deficiency inhibits HFD-induced insulin resistance, and the increased mitochondrial biogenesis and antioxidant induction might be the mechanisms underlying this phenomenon.

MicroRNA-874-3p Aggravates Doxorubicin-Induced Renal Podocyte Injury via Targeting Methionine Sulfoxide Reductase B3

Clinical application of doxorubicin (Dox) is limited due to its serious side effects including nephrotoxicity, and kidney podocytes play important roles in renal diseases. MicroRNAs (miRNAs) are critical regulators associated with human diseases. The purpose of this study was to explore a novel target in adjusting Dox-induced renal podocyte injury. Through a double luciferase reporter gene experiment, it was found that miR-874-3p directly targeted methionine sulfoxide reductase B3 (MsrB3). During the tests of miR-874-3p inhibitor and MsrB3 siRNA in human podocytes or miR-874-3p antagomir in mice, we found that the expression levels of downstream oxidative stress and apoptosis-related proteins were regulated by miR-874-3p/MsrB3 signal to alleviate or aggravate renal podocyte injury. The data in the present work showed that miR-874-3p aggravated Dox-caused renal podocyte injury by promoting apoptosis and oxidative damage via inhibiting MsrB3. Therefore, miR-874-3p/MsrB3 should be considered as a new therapeutic target in controlling renal podocyte injury induced by Dox.

Identification of Key Gene and Pathways for the Prediction of Peritoneal Metastasis of Gastric Cancer by Co-expression Analysis.

Peritoneal metastasis is the most common pattern in advanced gastric cancer and can predict poor disease prognosis. Early detection of peritoneal tumor dissemination is restricted by small peritoneal deposits. Therefore, it is critical to identify a novel predictive marker and to explore the potential mechanism associated with this process. In the present study, one module that correlated with peritoneal metastasis was identified. Enrichment analysis indicated that the Focal adhesion and the PI3K-Akt signaling pathway were the most significant pathways. Following network and Molecular Complex Detection (MCODE) analysis, the hub-gene cluster that consisted of 19 genes was selected. Methionine sulfoxide reductase B3 (MSRB3) was identified as a seed gene. Survival analysis indicated that high expression levels of MSRB3 were independent predictors of peritoneal disease-free survival (pDFS) as determined by univariate (HR 8.559, 95% CI; 3.339-21.937; P<.001) and multivariate Cox analysis (HR 3.982, 95% CI; 1.509-10.509; P=.005). Furthermore, patients with high levels of MSRB3 exhibited a significantly lower Overall Survival (OS) (log-rank P = 0.007). The external validation was performed by the (The Cancer Genome Atlas (TCGA)) (log-rank P = 0.037) and Kaplan Meier-plotter (KMplotter) (log-rank P = 0.031) data. In vitro experiments confirmed that MSRB3 was a critical protein in regulating gastric cancer cell proliferation and migration. In conclusion, High expression levels of MSRB3 in GC can predict peritoneal metastasis and recurrence as well as poor prognosis. Furthermore, MSRB3 was involved in the regulation of the proliferation and migration of GC cells.

MSRB3 promotes the progression of clear cell renal cell carcinoma via regulating endoplasmic reticulum stress

BackgroundRenal cancer represents about 3 % of all human cancers. Clear cell renal cell carcinoma (ccRCC) is the main type of renal cancer. Methionine sulfoxide reductase B3 (MSRB3) is a protein repair enzyme that specifically catalyzes the reduction of methionine-R-sulfoxide residues and has an antioxidant function. However, MSRB3’s role in ccRCC is still obscure. MethodsImmunohistochemical staining and Real-time PCR were used to compare the expression level of MSRB3 in ccRCC tissues and adjacent tissues. Western blot was used to detect the expression of MSRB3 in cell lines. Chi-square test were applied to evaluate the potential of MSRB3 to function as a cancer biomarker. RNA interference was used to inhibit MSRB3 expression in ccRCC cells, followed by detecting cell proliferation, apoptosis, migration and invasion. The markers of endoplasmic reticulum stress were then detected by western blot. ResultsIn this study, we validated that MSRB3 was significantly up-regulated in ccRCC samples and cell lines. It was also demonstrated that the up-regulation of MSRB3 was associated with several clinicopathologic features. Knockdown of MSRB3 remarkably arrested the proliferation, migration and invasion, while promoted apoptosis, and induced the changes of markers of endoplasmic reticulum stress. ConclusionIn conclusion, we demonstrated that MSRB3 was an oncogene of ccRCC associated with patients’ pathological characteristics and modulated endoplasmic reticulum stress of cancer cells.

A three-gene novel predictor for improving the prognosis of cervical cancer.

Cervical cancer is the most common gynecological malignancy, the third most common malignant tumor in women worldwide, and the most common malignant tumor among Chinese women. However, despite continuous improvement in medical treatment, the number of cervical cancer cases in China is on the increase annually, consistent with the general trend in global cervical cancer incidence. Therefore, it is particularly important to study the pathogenesis of cervical cancer at the genetic level in China. The aim of the present study was to use the TCGA database to identify potential genetic signatures that could predict the prognosis of patients with cervical cancer and provide evidence supporting clinical genetic intervention in cervical cancer. Primarily, an effective three-gene signature was found that predicts prognosis in patients with cervical cancer. This model can provide prima facie evidence for future assessment of patient risk and prognosis, but further testing is required to improve its accuracy. Our results also suggested that centromere protein M, methionine sulfoxide reductase B3 and Zic family member 2 could be promising biomarkers for the prognosis of cervical cancer.

Exploring insertions and deletions (indels) of MSRB3 gene and their association with growth traits in four Chinese indigenous cattle breeds.

Methionine sulfoxide reductase B3 (MSRB3) is instrumental in ossification and fat deposition, which regulate the growth and development of cattle directly. The purpose of this study was aimed to explore insertions and deletions (indels) in MSRB3 gene and investigate their association with growth traits in four indigenous cattle breeds (Luxi cattle, Qinchuan cattle, Nanyang cattle, and Jiaxian Red cattle). Four indels were identified by sequencing with DNA pool. Association analysis showed that three of them were associated with growth traits (). For P1, the DD (deletion and deletion) genotype was significantly associated with body length of Nanyang cattle; for P6, II (insertion and insertion) and/or DD genotypes were significantly associated with enhanced growth traits of Qinchuan cattle; for P7, II genotype was significantly associated with hucklebone width of Luxi cattle. Our results demonstrated that the polymorphisms in bovine MSRB3 gene were significantly associated with growth traits, which could be candidate loci for marker-assisted selection (MAS) in cattle breeding.

Identifying Potential Prognostic Markers for Muscle-Invasive Bladder Urothelial Carcinoma by Weighted Gene Co-Expression Network Analysis.

Muscle-invasive bladder urothelial carcinoma (MIBC) is characterized as a genetic heterogeneous cancer with a high percentage of recurrence and worse prognosis. Identify the prognostic potentials of novel genes for muscle-invasive urothelial bladder cancer could at least provide important information for early detection and clinical treatment. Weighted gene co-expression network analysis (WGCNA) algorithm, a powerful systems biology approach, was utilized to extract co-expressed gene networks from mRNA expression dataset to construct transcriptional modules in MIBC samples, which was associated with demographic and clinical traits of MIBC patients. The potential prognostic markers of MIBC were screened out in the discovery dataset and verified in an independent external validation dataset. A total of 8 co-expression modules were detected through the WGCNA algorithm in the discovery datasets based on 401 MIBC samples. One transcriptional module enriched in cell development was observed to be correlated with the MIBC prognosis in the discovery datasets (HR = 1.48, 95%CI = 1.04-2.11) and independently verified in an external dataset (HR = 3.59, 95%CI = 1.09-11.79). High expression of hub genes including discoidin domain receptor tyrosine kinase 2 (DDR2), PDZ and LIM domain 3 (PDLIM3), zinc finger protein 521 (ZNF521), methionine sulfoxide reductase B3 (MSRB3) were significantly associated with the unfavorable survival of MIBC patients. We identified and validated four novel potential biomarkers associated with prognosis of MIBC patients by constructing genes co-expression networks. The discovery of these genetic markers may provide a new target for the development of MIBC chemotherapeutic drugs.

Molecular cloning of WIF1 and HMGA2 reveals ear-preferential expression while uncovering a missense mutation associated with porcine ear size in WIF1.

Considerable diversity exists in porcine ear size, which is an important morphological feature of pig breeds. Previously, we localized four crucial candidate genes-high mobility group AT-hook 2 (HMGA2), LEM domain-containing 3 (LEMD3), methionine sulfoxide reductase B3 (MSRB3) and Wnt inhibitory factor 1 (WIF1)-on Sus Scrofa chromosome 5 affecting porcine ear size, then cloned LEMD3 and MSBR3. In this study, we performed rapid amplification of cDNA ends to obtain full-length cDNA sequences of 2338-bp WIF1 and 2998-bp HMGA2. Using quantitative real-time PCR, we revealed that WIF1 expression was highest in ear cartilage of 60-day-old pigs and that this is therefore a better candidate gene for ear size than HMGA2. We further screened coding sequence variants in both genes and identified only one missense mutation (WIF1:c.1167C>G) in a conserved epidermal growth factor-like domain from the mammalian WIF1 protein. The protein-altering mutation was significantly associated with ear size across the Large White×Minzhu hybrid and Beijing Black pig populations. When WIF1:c.1167C>G was included as fixed effect in the model to re-run a genome-wide association study in the Large White×Minzhu intercross population the P-value of the peak SNP on SSC5 from re-running the genome-wide association study dropped from 2.45E-12 to 7.33E-05. Taken together, the WIF1:c.1167C>G could be an important mutation associated with ear size. Our findings provide helpful information for further studies of the molecular mechanisms controlling porcine ear size.

Copy number variation in the MSRB3 gene enlarges porcine ear size through a mechanism involving miR-584-5p

BackgroundThe size and type of ears are important conformation characteristics that distinguish pig breeds. A significant quantitative trait locus (QTL) for ear size has been identified on SSC5 (SSC for Sus scrofa chromosome) but the underlying causative gene and mutation remain unknown. Thus, our aim was to identify the gene responsible for enlarged ears in pig.ResultsFirst, we narrowed down the QTL region on SSC5 to a 137.85-kb interval that harbors only the methionine sulfoxide reductase B3 (MSRB3) gene. Then, we identified a 38.7-kb copy number variation (CNV) that affects the last two exons of MSRB3 and could be the candidate causative mutation for this QTL. This CNV showed complete concordance with genotype at the QTL of the founder animals in a white Duroc × Erhualian F2 intercross and was found only in pigs from six Chinese indigenous breeds with large ears and from the Landrace breed with half-floppy ears. Moreover, it accounted for the significant association with ear size on SSC5 across the five pig populations tested. eQTL mapping revealed that this CNV was significantly associated with the expression of the microRNA (miRNA) miR-584-5p, which interacts with MSRB3, one of its target genes. In vivo and in vitro experiments confirmed that miR-584-5p inhibits the translation of MSRB3 mRNA. Taken together, these results led us to conclude that presence of the 38.7-kb CNV in the genome of some pig breeds affects ear size by altering the expression of miR-584-5p, which consequently hinders the expression of one of its target genes (e.g. MSRB3).ConclusionsOur findings shed insight into the underlying mechanism of development of external ears in mammals and contribute to a better understanding of how the presence of CNV can regulate gene expression.

Methionine sulfoxide reductase B3 requires resolving cysteine residues for full activity and can act as a stereospecific methionine oxidase.

The oxidation of methionine residues in proteins occurs during oxidative stress and can lead to an alteration in protein function. The enzyme methionine sulfoxide reductase (Msr) reverses this modification. Here, we characterise the mammalian enzyme Msr B3. There are two splice variants of this enzyme that differ only in their N-terminal signal sequence, which directs the protein to either the endoplasmic reticulum (ER) or mitochondria. We demonstrate here that the enzyme can complement a bacterial strain, which is dependent on methionine sulfoxide reduction for growth, that the purified recombinant protein is enzymatically active showing stereospecificity towards R-methionine sulfoxide, and identify the active site and two resolving cysteine residues. The enzyme is efficiently recycled by thioredoxin only in the presence of both resolving cysteine residues. These results show that for this isoform of Msrs, the reduction cycle most likely proceeds through a three-step process. This involves an initial sulfenylation of the active site thiol followed by the formation of an intrachain disulfide with a resolving thiol group and completed by the reduction of this disulfide by a thioredoxin-like protein to regenerate the active site thiol. Interestingly, the enzyme can also act as an oxidase catalysing the stereospecific formation of R-methionine sulfoxide. This result has important implications for the role of this enzyme in the reversible modification of ER and mitochondrial proteins.

Methionine Sulfoxide Reductase-B3 (MsrB3) Protein Associates with Synaptic Vesicles and its Expression Changes in the Hippocampi of Alzheimer's Disease Patients.

Genome-wide association studies (GWAS) identified susceptibility loci associated with decreased hippocampal volume, and found hippocampal subfield-specific effects at MSRB3 (methionine sulfoxide reductase-B3). The MSRB3 locus was also linked to increased risk for late onset Alzheimer's disease (AD). In this study, we uncovered novel sites of MsrB3 expression in CA pyramidal layer and arteriolar walls by using automated immunohistochemistry on hippocampal sections from 23 individuals accompanied by neuropathology reports and clinical dementia rating scores. Controls, cognitively intact subjects with no hippocampal neurofibrillary tangles, exhibited MsrB3 signal as distinct but rare puncta in CA1 pyramidal neuronal somata. In CA3, however, MsrB3-immunoreactivity was strongest in the neuropil of the pyramidal layer. These patterns were replicated in rodent hippocampi where ultrastructural and immunohistofluorescence analysis revealed MsrB3 signal associated with synaptic vesicles and colocalized with mossy fiber terminals. In AD subjects, the number of CA1 pyramidal neurons with frequent, rather than rare, MsrB3-immunoreactive somatic puncta increased in comparison to controls. This change in CA1 phenotype correlated with the occurrence of AD pathological hallmarks. Moreover, the intensity of MsrB3 signal in the neuropil of CA3 pyramidal layer correlated with the signal pattern in neurons of CA1 pyramidal layer that was characteristic of cognitively intact individuals. Finally, MsrB3 signal in the arteriolar walls in the hippocampal white matter decreased in AD patients. This characterization of GWAS-implicated MSRB3 protein expression in human hippocampus suggests that patterns of neuronal and vascular MsrB3 protein expression reflect or underlie pathology associated with AD.

MsrB3 deficiency induces cancer cell apoptosis through p53-independent and ER stress-dependent pathways

We have previously shown that down-regulation of methionine sulfoxide reductase B3 (MsrB3) induces cancer cell apoptosis through the activation of the intrinsic mitochondrial pathway. However, the mechanism through which MsrB3 deficiency results in cancer cell death is poorly understood. In this study, we investigated whether p53 and endoplasmic reticulum (ER) stress are involved in MsrB3 deficiency-induced cancer cell apoptosis using breast and colon cancer cells. MsrB3 depletion resulted in p53 down-regulation at the post-transcriptional level. MsrB3 deficiency induced cell death to a similar extent in both p53 wild-type (p53+/+) and null (p53−/−) cancer cells, suggesting that MsrB3 deficiency-induced apoptosis occurs independently of p53. MsrB3 deficiency significantly increased ER stress, which resulted in apoptosis. In addition, MsrB3 depletion activated the pro-apoptotic Bim molecule, which is essential for ER stress-induced apoptosis. MsrB3 deficiency increased cytosolic calcium levels, suggesting that MsrB3 down-regulation leads to a disturbance of calcium homeostasis in the ER, which consequently triggers ER stress. MsrB3 overexpression in MsrB3-depleted cells reduced ER stress, and was accompanied by at least partial recovery of cell viability. Taken together, our results suggest that MsrB3 plays a critical role in cancer cell apoptosis through the modulation of ER stress status.

MRNA and protein expression levels of four candidate genes for ear size in Erhualian and Large White pigs

Porcine ear size is an important characteristic for distinguishing among pig breeds. In a previous genome-wide association study of porcine ear size, LEM domain-containing 3 (LEMD3), methionine sulfoxide reductase B3 (MSRB3), high mobility group AT-hook 2 (HMGA2), and Wnt inhibitory factor 1 (WIF1) were implicated as important candidate genes for ear size. This study investigated the expression levels of four candidate genes for ear size in Erhualian and Large White pigs. Ten Erhualian pigs with large ears and eight Large White pigs with small ears at 60 days of age were examined. The mRNA expression levels of the four candidate genes were quantified by real-time polymerase chain reaction. WIF1 mRNA expression was significantly higher in Large White than in Erhualian pigs (P < 0.05), whereas the expression levels of the other three genes were not significantly different between the two breeds. The protein expression levels of the four genes were analyzed using western blot. WIF1 protein expression was significantly higher in Large White than in Erhualian pigs (P < 0.01), whereas MSRB3 protein expression was significantly higher in Erhualian than in Large White pigs (P < 0.05). There were no significant differences between the two breeds in residual protein expression. These results suggest that WIF1 is the main causal gene for ear size in pigs.

Down-regulation of MsrB3 induces cancer cell apoptosis through reactive oxygen species production and intrinsic mitochondrial pathway activation

Methionine sulfoxide reductase B3 (MsrB3) is a protein repair enzyme that specifically catalyzes the reduction of methionine-R-sulfoxide residues and has an antioxidant function. We have previously shown that depletion of MsrB3 suppresses the proliferation of normal mammalian cells by arresting cell cycle. In this study, we report the crucial role of MsrB3 in cancer cell death. Deficiency of MsrB3 induced cancer cell death, while MsrB3 overexpression stimulated cancer cell proliferation. MsrB3 depletion resulted in apoptotic cancer cell death through the activation of the intrinsic mitochondrial pathway. MsrB3 deficiency increased the levels of cellular reactive oxygen species (ROS) and led to redox imbalance, and also increased the Bax to Bcl-2 ratio and cytochrome c release, leading to caspase activation. Treatment of MsrB3-depleted cells with N-acetylcysteine, an ROS scavenger, prevented cell death, suggesting that MsrB3 deficiency-induced cell death is associated with increased ROS production. In addition, MsrB3 depletion activated poly(ADP ribose) polymerase-1 (PARP-1) and led to the translocation of apoptosis-inducing factor (AIF) to the nucleus. Taken together, our results suggest that MsrB3 plays an important role in cancer cell survival through the modulation of the intrinsic apoptosis pathway.

215 - Methionine Sulfoxide Reductase B3 Deficiency Stimulates Heme Oxygenase-1 Expression Via ROS-Dependent and Nrf2 Activation Pathways

Methionine sulfoxide reductase B3 (MsrB3), which is primarily found in the endoplasmic reticulum (ER), is an important protein repair enzyme that stereospecifically reduces methionine- R -sulfoxide residues. We previously found that MsrB3 deficiency arrests the cell cycle at the G 1 /S stage through up-regulation of p21 and p27. In this study, we report a critical role of MsrB3 in gene expression of heme oxygenase-1 (HO-1), which has an anti-proliferative effect associated with p21 up-regulation. Depletion of MsrB3 elevated HO-1 expression in mammalian cells, whereas MsrB3 overexpression had no effect. MsrB3 deficiency increased cellular reactive oxygen species (ROS), particularly in the mitochondria. ER stress, which is associated with up-regulation of HO-1, was also induced by depletion of MsrB3. Treatment with N -acetylcysteine as an ROS scavenger reduced augmented HO-1 levels in MsrB3-depleted cells. MsrB3 deficiency activated Nrf2 transcription factor by enhancing its expression and nuclear import. The activation of Nrf2 induced by MsrB3 depletion was confirmed by increased expression levels of its other target genes, such as γ-glutamylcysteine ligase. Taken together, these data suggest that MsrB3 attenuates HO-1 induction by inhibiting ROS production, ER stress, and Nrf2 activation.

Methionine sulfoxide reductase B3 deficiency stimulates heme oxygenase-1 expression via ROS-dependent and Nrf2 activation pathways

Methionine sulfoxide reductase B3 (MsrB3), which is primarily found in the endoplasmic reticulum (ER), is an important protein repair enzyme that stereospecifically reduces methionine-R-sulfoxide residues. We previously found that MsrB3 deficiency arrests the cell cycle at the G1/S stage through up-regulation of p21 and p27. In this study, we report a critical role of MsrB3 in gene expression of heme oxygenase-1 (HO-1), which has an anti-proliferative effect associated with p21 up-regulation. Depletion of MsrB3 elevated HO-1 expression in mammalian cells, whereas MsrB3 overexpression had no effect. MsrB3 deficiency increased cellular reactive oxygen species (ROS), particularly in the mitochondria. ER stress, which is associated with up-regulation of HO-1, was also induced by depletion of MsrB3. Treatment with N-acetylcysteine as an ROS scavenger reduced augmented HO-1 levels in MsrB3-depleted cells. MsrB3 deficiency activated Nrf2 transcription factor by enhancing its expression and nuclear import. The activation of Nrf2 induced by MsrB3 depletion was confirmed by increased expression levels of its other target genes, such as γ-glutamylcysteine ligase. Taken together, these data suggest that MsrB3 attenuates HO-1 induction by inhibiting ROS production, ER stress, and Nrf2 activation.

Methionine Sulfoxide Reductase B3-Targeted In Utero Gene Therapy Rescues Hearing Function in a Mouse Model of Congenital Sensorineural Hearing Loss.

Methionine sulfoxide reductase B3 (MsrB3), which stereospecifically repairs methionine-R-sulfoxide, is an important Msr protein that is associated with auditory function in mammals. MsrB3 deficiency leads to profound congenital hearing loss due to the degeneration of stereociliary bundles and the apoptotic death of cochlear hair cells. In this study, we investigated a fundamental treatment strategy in an MsrB3 deficiency mouse model and confirmed the biological significance of MsrB3 in the inner ear using MsrB3 knockout (MsrB3(-/-)) mice. We delivered a recombinant adeno-associated virus encoding the MsrB3 gene directly into the otocyst at embryonic day 12.5 using a transuterine approach. We observed hearing recovery in the treated ears of MsrB3(-/-) mice at postnatal day 28, and we confirmed MsrB3 mRNA and protein expression in cochlear extracts. Additionally, we demonstrated that the morphology of the stereociliary bundles in the rescued ears of MsrB3(-/-) mice was similar to those in MsrB3(+/+) mice. To our knowledge, this is the first study to demonstrate functional and morphological rescue of the hair cells of the inner ear in the MsrB3 deficiency mouse model of congenital genetic sensorineural hearing loss using an in utero, virus-mediated gene therapy approach. Our results provide insight into the role of MsrB3 in hearing function and bring us one step closer to hearing restoration as a fundamental therapy.

Porcine methionine sulfoxide reductase B3: molecular cloning, tissue-specific expression profiles, and polymorphisms associated with ear size in Sus scrofa.

BackgroundIn Sus scrofa, methionine sulfoxide reductase B3 (MSRB3) is a crucial candidate gene for ear size, and an important conformational trait of pig breeds. However, challenges in MSRB3 cDNA amplification have prevented further identification of MSRB3 allelic variants influencing pig ear size.ResultsWe cloned a full-length cDNA sequence of porcine MSRB3 by rapid-amplification of cDNA ends. The 3,765-bp gene contained a 5’-untranslated region (UTR) (190 bp), a coding region (552 bp), and a 3’-UTR (3,016 bp) and shared 84 %, 84 %, 87 %, 86 %, and 70 % sequence identities with human, orangutan, mouse, chicken, and zebrafish, respectively. The gene encoded a 183-amino acid protein, which shared 88 %, 91 %, 89 %, 86 %, and 67 % identities with human, orangutan, mouse, chicken, and zebrafish, respectively. Tissue expression analysis using qRT-PCR revealed that MSRB3 was expressed in the heart, liver, lung, kidney, spleen, ear, muscle, fat, lymph, skeletal, and hypothalamic tissues. Three single nucleotide polymorphisms (SNPs) were identified in MSRB3: c.-735C > T in the 5’ flanking region, c.2571 T > C in the 3’-UTR, and a synonymous mutation of c.484 T > C in the coding region. The SNPs c.-735C > T and c.2571 T > C were significantly associated with ear size in a Large White × Minzhu F2 population other than in Beijing Black pigs. Subsequently, at SNP c.-735C > T, the mRNA of MSRB3 was significantly higher expressed in ears of individuals with the TT genotype (Minzhu) than those with CC (Large White).ConclusionsThe porcine MSRB3 owned a 3,765-bp full-length cDNA sequence and was detected to express in ear tissue. Two SNPs of this gene were shown to be significantly associated with ear size in a Large White × Minzhu intercross population instead of Beijing Black pig population. What’s more, the individuals with higher mRNA expression of MSRB3 have larger ear sizes. These results provide useful information for further functional analyses of MSRB3 influencing ear size in pigs.Electronic supplementary materialThe online version of this article (doi:10.1186/s40104-015-0060-x) contains supplementary material, which is available to authorized users.

Protein Expression of Alzheimer's disease‐ and Reduced Hippocampal Volume‐ Risk Loci in Human Hippocampus

We examined the protein expression of novel genome‐wide association studies (GWAS) identified risk alleles implicated in Alzheimer's disease (AD) and low hippocampal volume using postmortem hippocampi with no or mild‐to‐moderate AD‐associated pathological changes. We found BIN1, the most significant late‐onset AD susceptibility locus after APOE, expressed in glia of all the examined cases while the punctate signal in neuropil varied substantially. EPHA1, another confirmed AD risk locus, is an ephrin receptor implicated in neuronal apoptosis. We found EphA1 in the neuronal cytoplasm, with a trend of decreasing signal intensity as AD progressed. Methionine sulfoxide reductase B3 (MSRB3) locus, critical for oxidative stress, was identified in GWAS on hippocampal volume. We found MSRB3 signal mostly as cytoplasmic puncta in pyramidal neurons from all of the examined cases, albeit in various amounts. Finally, LEMD3, a possible causal gene at the MSRB3 locus, was expressed in neurons as well as astrocytes. Neuronal expression of LEMD3 may increase with AD progression. Our data provide first insight into the human hippocampal cell type‐ and cell compartment‐specific protein expression of AD‐ and hippocampal volume reduction‐ risk alleles. We are currently examining the correlation of these proteins' expression with the cognitive status reflected in CDR (Clinical Dementia Rating) scores.