Proteomic Analysis Reveals Salt-Tolerant Mechanism in Soybean Applied with Plant-Derived Smoke Solution.

Salt stress of soybean is a serious problem because it reduces plant growth and seed yield. To investigate the salt-tolerant mechanism of soybean, a plant-derived smoke (PDS) solution was used. Three-day-old soybeans were subjected to PDS solution under 100 mM NaCl for 2 days, resulting in PDS solution improving soybean root growth, even under salt stress. Under the same condition, proteins were analyzed using the proteomic technique. Differential abundance proteins were associated with transport/formaldehyde catabolic process/sucrose metabolism/glutathione metabolism/cell wall organization in the biological process and membrane/Golgi in the cellular component with or without PDS solution under salt stress. Immuno-blot analysis confirmed that osmotin, alcohol dehydrogenase, and sucrose synthase increased with salt stress and decreased with additional PDS solution; however, H+ATPase showed opposite effects. Cellulose synthase and xyloglucan endotransglucosylase/hydrolase increased with salt and decreased with additional PDS solution. Furthermore, glycoproteins decreased with salt stress and recovered with additional treatment. As mitochondrion-related events, the contents of ATP and gamma-aminobutyric acid increased with salt stress and recovered with additional treatment. These results suggest that PDS solution improves the soybean growth by alleviating salt stress. Additionally, the regulation of energy metabolism, protein glycosylation, and cell wall construction might be an important factor for the acquisition of salt tolerance in soybean.

Blockade of glucagon increases muscle mass and alters fiber type composition in mice deficient in proglucagon-derived peptides.

AIMS/INTRODUCTION: Glucagon is secreted from pancreatic α-cells and plays an important role in amino acid metabolism in liver. Various animal models deficient in glucagon action show hyper-amino acidemia and α-cell hyperplasia, indicating that glucagon contributes to feedback regulation between the liver and the α-cells. In addition, both insulin and various amino acids, including branched-chain amino acids and alanine, participate in protein synthesis in skeletal muscle. However, the effect of hyperaminoacidemia on skeletal muscle has not been investigated. In the present study, we examined the effect of blockade of glucagon action on skeletal muscle using mice deficient in proglucagon-derived peptides (GCGKO mice). MATERIALS AND METHODS: Muscles isolated from GCGKO and control mice were analyzed for their morphology, gene expression and metabolites. RESULTS: GCGKO mice showed muscle fiber hypertrophy, and a decreased ratio of type IIA and an increased ratio of type IIB fibers in the tibialis anterior. The expression levels of myosin heavy chain (Myh) 7, 2, 1 and myoglobin messenger ribonucleic acid were significantly lower in GCGKO mice than those in control mice in the tibialis anterior. GCGKO mice showed a significantly higher concentration of arginine, asparagine, serine and threonine in the quadriceps femoris muscles, and also alanine, aspartic acid, cysteine, glutamine, glycine and lysine, as well as four amino acids in gastrocnemius muscles. CONCLUSIONS: These results show that hyperaminoacidemia induced by blockade of glucagon action in mice increases skeletal muscle weight and stimulates slow-to-fast transition in type II fibers of skeletal muscle, mimicking the phenotype of a high-protein diet.

Circulating miR-20b-5p and miR-330-3p are novel biomarkers for progression of atrial fibrillation: Intracardiac/extracardiac plasma sample analysis by small RNA sequencing.

BACKGROUND: Circulating microRNAs (miRNAs, miR) have been considered as biomarkers reflecting the underlying pathophysiology in atrial fibrillation (AF). Nevertheless, miRNA expression in the peripheral blood samples might not reflect a cardiac phenomenon since most miRNAs are expressed in numerous organs. This study aimed to identify the cardiac-specific circulating miRNAs as biomarkers for AF. METHODS: Plasma samples were obtained from a luminal coronary sinus catheter (CS, cardiac-specific samples) and femoral venous sheath (FV, peripheral samples) in patients with AF and paroxysmal supraventricular tachycardia (control, CTL) undergoing catheter ablation. The circulating miRNA profiles were analyzed by small RNA sequencing. Differently expressed miRNAs between AF and CTL were identified in each sample of the CS and FV; miRNAs exhibiting similar expression patterns in the CS and FV samples were selected as candidates for cardiac-specific biomarkers. The selected miRNAs were related to the outcome of catheter ablation of AF. RESULTS: Small RNA sequencing detected 849 miRNAs. Among the top 30 most differently expressed miRNAs between AF and CTL, circulating hsa-miR-20b-5p, hsa-miR-330-3p, and hsa-miR-204-5p had a similar pattern in the CS and FV samples. Another set of peripheral blood samples was obtained from AF patients undergoing catheter ablation (n = 141). The expression of the miR-20b-5p and miR-330-3p, but not the miR-204-5p, negatively correlated with the echocardiographic left-atrial dimension and was decreased in patients with AF recurrence as compared to those without AF recurrence during a 1-year follow-up. CONCLUSION: Circulating miR-20b-5p and miR-330-3p can be cardiac-specific biomarkers for atrial remodeling progression and arrhythmia recurrence after catheter ablation in AF patients.

Simultaneous loss of skeletal muscle myosin heavy chain IIx and IIb causes severe skeletal muscle hypoplasia in postnatal mice.

The skeletal muscle myosin heavy chain (MyHC) is a fundamental component of the sarcomere structure and muscle contraction. Two of the three adult fast MyHCs, MyHC-IIx and MyHC-IIb, are encoded by Myh1 and Myh4, respectively. However, skeletal muscle disorders have not yet been linked to these genes in humans. MyHC-IIb is barely detectable in human skeletal muscles. Thus, to characterize the molecular function of skeletal muscle MyHCs in humans, investigation of the effect of simultaneous loss of MyHC-IIb and other MyHCs on skeletal muscle in mice is essential. Here, we generated double knockout (dKO) mice with simultaneous loss of adult fast MyHCs by introducing nonsense frameshift mutations into the Myh1 and Myh4 genes. The dKO mice appeared normal after birth and until 2 weeks of age but showed severe skeletal muscle hypoplasia after 2 weeks. In 3-week-old dKO mice, increased expression of other skeletal muscle MyHCs, such as MyHC-I, MyHC-IIa, MyHC-neo, and MyHC-emb, was observed. However, these expressions were not sufficient to compensate for the loss of MyHC-IIb and MyHC-IIx. Moreover, the aberrant sarcomere structure with altered expression of sarcomere components was observed in dKO mice. Our findings imply that the simultaneous loss of MyHC-IIb and MyHC-IIx is substantially detrimental to postnatal skeletal muscle function and will contribute to elucidating the molecular mechanisms of skeletal muscle wasting disorders caused by the loss of skeletal muscle MyHCs.

Proteomic and Biochemical Approaches Elucidate the Role of Millimeter-Wave Irradiation in Wheat Growth under Flooding Stress.

Flooding impairs wheat growth and considerably affects yield productivity worldwide. On the other hand, irradiation with millimeter waves enhanced the growth of chickpea and soybean under flooding stress. In the current work, millimeter-wave irradiation notably enhanced wheat growth, even under flooding stress. To explore the protective mechanisms of millimeter-wave irradiation on wheat under flooding, quantitative proteomics was performed. According to functional categorization, proteins whose abundances were changed significantly with and without irradiation under flooding stress were correlated to glycolysis, reactive-oxygen species scavenging, cell organization, and hormonal metabolism. Immunoblot analysis confirmed that fructose-bisphosphate aldolase and ß tubulin accumulated in root and leaf under flooding; however, even in such condition, their accumulations were recovered to the control level in irradiated wheat. The abundance of ascorbate peroxidase increased in leaf under flooding and recovered to the control level in irradiated wheat. Because the abundance of auxin-related proteins changed with millimeter-wave irradiation, auxin was applied to wheat under flooding, resulting in the application of auxin improving its growth, even in such condition. These results suggest that millimeter-wave irradiation on wheat seeds improves the recovery of plant growth from flooding via the regulation of glycolysis, reactive-oxygen species scavenging, and cell organization. Additionally, millimeter-wave irradiation could promote tolerance against flooding through the regulation of auxin contents in wheat.

Characterization of the MG828507 lncRNA Located Upstream of the FLT1 Gene as an Etiology for Pre-Eclampsia.

BACKGROUND: FLT1 is one of the significantly overexpressed genes found in a pre-eclamptic placenta and is involved with the etiology of this disease. METHODS: We conducted genome-wide expression profiling by RNA-seq of placentas from women with pre-eclampsia and those with normotensive pregnancy. RESULTS: We identified a lncRNA gene, MG828507, located ~80 kb upstream of the FLT1 gene in a head-to-head orientation, which was overexpressed in the pre-eclamptic placenta. MG828507 and FLT1 are located within the same topologically associated domain in the genome. The MG828507 mRNA level correlated with that of the FLT1 in placentas from pre-eclamptic women as well as in samples from uncomplicated pregnancies. However, neither the overexpression nor knockdown of MG828507 affected the expression of FLT1. Analysis of pre-eclampsia-linking genetic variants at this locus suggested that the placental genotype of one variant was associated with the expression of MG828507. The MG828507 transcript level was not found to be associated with maternal blood pressure, but showed a relationship with birth and placental weights, suggesting that this lncRNA might be one of the pivotal placental factors in pre-eclampsia. CONCLUSION: Further characterization of the MG828507 gene may elucidate the etiological roles of the MG828507 and FLT1 genes in pre-eclampsia in a genomic context.

The Functional Role of Long Non-Coding RNA in Myogenesis and Skeletal Muscle Atrophy.

Skeletal muscle is a pivotal organ in humans that maintains locomotion and homeostasis. Muscle atrophy caused by sarcopenia and cachexia, which results in reduced muscle mass and impaired skeletal muscle function, is a serious health condition that decreases life longevity in humans. Recent studies have revealed the molecular mechanisms by which long non-coding RNAs (lncRNAs) regulate skeletal muscle mass and function through transcriptional regulation, fiber-type switching, and skeletal muscle cell proliferation. In addition, lncRNAs function as natural inhibitors of microRNAs and induce muscle hypertrophy or atrophy. Intriguingly, muscle atrophy modifies the expression of thousands of lncRNAs. Therefore, although their exact functions have not yet been fully elucidated, various novel lncRNAs associated with muscle atrophy have been identified. Here, we comprehensively review recent knowledge on the regulatory roles of lncRNAs in skeletal muscle atrophy. In addition, we discuss the issues and possibilities of targeting lncRNAs as a treatment for skeletal muscle atrophy and muscle wasting disorders in humans.

Morphological and Proteomic Analyses of Soybean Seedling Interaction Mechanism Affected by Fiber Crosslinked with Zinc-Oxide Nanoparticles.

Nanoparticles (NPs) enhance soybean growth; however, their precise mechanism is not clearly understood. To develop a more effective method using NPs for the enhancement of soybean growth, fiber crosslinked with zinc oxide (ZnO) NPs was prepared. The solution of ZnO NPs with 200 nm promoted soybean growth at the concentration of 10 ppm, while fibers crosslinked with ZnO NPs promoted growth at a 1 ppm concentration. Soybeans grown on fiber cross-linked with ZnO NPs had higher Zn content in their roots than those grown in ZnO NPs solution. To study the positive mechanism of fiber crosslinked with ZnO NPs on soybean growth, a proteomic technique was used. Proteins categorized in photosynthesis and secondary metabolism accumulated more in soybeans grown on fiber crosslinked with ZnO NPs than in those grown in ZnO NPs solution. Furthermore, significantly accumulated proteins, which were NADPH oxidoreductase and tubulins, were confirmed using immunoblot analysis. The abundance of NADPH oxidoreductase increased in soybean by ZnO NPs application. These results suggest that fiber crosslinked with ZnO NPs enhances soybean growth through the increase of photosynthesis and secondary metabolism. Additionally, the accumulation of NADPH oxidoreductase might relate to the effect of auxin with fiber crosslinked with ZnO NPs on soybean growth.

Morphological, Biochemical, and Proteomic Analyses to Understand the Promotive Effects of Plant-Derived Smoke Solution on Wheat Growth under Flooding Stress.

Wheat is an important staple food crop for one-third of the global population; however, its growth is reduced by flooding. On the other hand, a plant-derived smoke solution enhances plant growth; however, its mechanism is not fully understood. To reveal the effects of the plant-derived smoke solution on wheat under flooding, morphological, biochemical, and proteomic analyses were conducted. The plant-derived smoke solution improved wheat-leaf growth, even under flooding. According to the functional categorization of proteomic results, oppositely changed proteins were correlated with photosynthesis, glycolysis, biotic stress, and amino-acid metabolism with or without the plant-derived smoke solution under flooding. Immunoblot analysis confirmed that RuBisCO activase and RuBisCO large/small subunits, which decreased under flooding, were recovered by the application of the plant-derived smoke solution. Furthermore, the contents of chlorophylls a and b significantly decreased by flooding stress; however, they were recovered by the application of the plant-derived smoke solution. In glycolysis, fructose-bisphosphate aldolase and glyceraldehyde-3-phosphate dehydrogenase decreased with the application of the plant-derived smoke solution under flooding as compared with flooding alone. Additionally, glutamine, glutamic acid, aspartic acid, and serine decreased under flooding; however, they were recovered by the plant-derived smoke solution. These results suggest that the application of the plant-derived smoke solution improves the recovery of wheat growth through the regulation of photosynthesis and glycolysis even under flooding conditions. Furthermore, the plant-derived smoke solution might promote wheat tolerance against flooding stress through the regulation of amino-acid metabolism.

Proteomic, Biochemical, and Morphological Analyses of the Effect of Silver Nanoparticles Mixed with Organic and Inorganic Chemicals on Wheat Growth.

Wheat is vulnerable to numerous diseases; on the other hand, silver nanoparticles (AgNPs) exhibit a sterilizing action. To understand the combined effects of AgNPs with nicotinate and potassium nitrate (KNO3) for plant growth and sterilization, a gel- and label-free proteomics was performed. Root weight was promoted by the treatment of AgNPs mixed with nicotinate and KNO3. From a total of 5557 detected proteins, 90 proteins were changed by the mixture of AgNPs, nicotinate, and KNO3; among them, 25 and 65 proteins increased and decreased, respectively. The changed proteins were mainly associated with redox and biotic stress in the functional categorization. By immunoblot analysis, the abundance of glutathione reductase/peroxiredoxin and pathogen-related protein three significantly decreased with the mixture. Furthermore, from the changed proteins, the abundance of starch synthase and lipoxygenase significantly increased and decreased, respectively. Through biochemical analysis, the starch contents increased with the mixture. The application of esculetin, which is a lipoxygenase inhibitor, increased the weight and length of the root. These results suggest that the AgNPs mixed with nicotinate and KNO3 cause positive effects on wheat seedlings by regulating pathogen-related protein and reactive-oxygen species scavenging. Furthermore, increasing starch and decreasing lipoxygenase might improve wheat growth.

Development of Myostatin Inhibitory d-Peptides to Enhance the Potency, Increasing Skeletal Muscle Mass in Mice.

Myostatin is a key negative regulator of skeletal muscle growth, and myostatin inhibitors are attractive tools for the treatment of muscular atrophy. Previously, we reported a series of 14-29-mer peptide myostatin inhibitors, including a potent derivative, MIPE-1686, a 16-mer N-terminal-free l-peptide with three unnatural amino acids and a propensity to form ß-sheets. However, the in vivo biological stability of MIPE-1686 is a concern for its development as a drug. In the present study, to develop a more stable myostatin inhibitory d-peptide (MID), we synthesized various retro-inverso versions of a 16-mer peptide. Among these, an arginine-containing derivative, MID-35, shows a potent and equivalent in vitro myostatin inhibitory activity equivalent to that of MIPE-1686 and considerable stability against biodegradation. The in vivo potency of MID-35 to increase the tibialis anterior muscle mass in mice is significantly enhanced over that of MIPE-1686, and MID-35 can serve as a new entity for the prolonged inactivation of myostatin in skeletal muscle.

Proteomic and Biological Analyses Reveal the Effect on Growth under Flooding Stress of Chickpea Irradiated with Millimeter Waves.

Chickpea cultivated on marginal lands in arid and semiarid tropics is one of the food legumes, and its growth is reduced by flooding stress. Millimeter-wave irradiation has influences on organisms, and it improves the growth of plants such as soybean. To reveal the dynamic effects of millimeter-wave irradiation on chickpea under flooding, gel- and label-free proteomic analysis was conducted. Millimeter-wave irradiation improved chickpea growth and its tolerance to flooding stress. According to functional categorization, oppositely changed proteins were correlated with photosynthesis, fermentation, and protein degradation. Immunoblot analysis confirmed that RuBisCO activase and large subunits decreased in leaves under flooding; however, they are recovered in irradiated chickpea even if it was in this condition. The activity and accumulation of alcohol dehydrogenase increased in roots under flooding; however, this followed the same pattern. Cell death was significantly increased and decreased by flooding on unirradiated and irradiated chickpeas, respectively. These findings suggest that irradiation with millimeter waves on chickpea seeds improves the recovery of plant growth through regulation of photosynthesis in leaves and fermentation in roots. Furthermore, millimeter-wave irradiation might promote chickpea tolerance under flooding via the regulation of cell death.

Proteomic and Biochemical Analyses of the Mechanism of Tolerance in Mutant Soybean Responding to Flooding Stress.

To investigate the mechanism of flooding tolerance of soybean, flooding-tolerant mutants derived from gamma-ray irradiated soybean were crossed with parent cultivar Enrei for removal of other factors besides the genes related to flooding tolerance in primary generated mutant soybean. Although the growth of the wild type was significantly suppressed by flooding compared with the non-flooding condition, that of the mutant lines was better than that of the wild type even if it was treated with flooding. A two-day-old mutant line was subjected to flooding for 2 days and proteins were analyzed using a gel-free/label-free proteomic technique. Oppositely changed proteins in abundance between the wild type and mutant line under flooding stress were associated in endoplasmic reticulum according to gene-ontology categorization. Immunoblot analysis confirmed that calnexin accumulation increased in both the wild type and mutant line; however, calreticulin accumulated in only the mutant line under flooding stress. Furthermore, although glycoproteins in the wild type decreased by flooding compared with the non-flooding condition, those in the mutant line increased even if it was under flooding stress. Alcohol dehydrogenase accumulated in the wild type and mutant line; however, this enzyme activity significantly increased and mildly increased in the wild type and mutant line, respectively, under flooding stress compared with the non-flooding condition. Cell death increased and decreased in the wild type and mutant line, respectively, by flooding stress. These results suggest that the regulation of cell death through the fermentation system and glycoprotein folding might be an important factor for the acquisition of flooding tolerance in mutant soybean.

An Analysis of Differentially Expressed Coding and Long Non-Coding RNAs in Multiple Models of Skeletal Muscle Atrophy.

The loss of skeletal muscle mass (muscle atrophy or wasting) caused by aging, diseases, and injury decreases quality of life, survival rates, and healthy life expectancy in humans. Although long non-coding RNAs (lncRNAs) have been implicated in skeletal muscle formation and differentiation, their precise roles in muscle atrophy remain unclear. In this study, we used RNA-sequencing (RNA-Seq) to examine changes in the expression of lncRNAs in four muscle atrophy conditions (denervation, casting, fasting, and cancer cachexia) in mice. We successfully identified 33 annotated lncRNAs and 18 novel lncRNAs with common expression changes in all four muscle atrophy conditions. Furthermore, an analysis of lncRNA-mRNA correlations revealed that several lncRNAs affected small molecule biosynthetic processes during muscle atrophy. These results provide novel insights into the lncRNA-mediated regulatory mechanism underlying muscle atrophy and may be useful for the identification of promising therapeutic targets.

Osmotic stress in banana is relieved by exogenous nitric oxide.

Drought is one of the severe environmental stresses threatening agriculture around the globe. Nitric oxide plays diverse roles in plant growth and defensive responses. Despite a few studies supporting the role of nitric oxide in plants under drought responses, little is known about its pivotal molecular amendment in the regulation of stress signaling. In this study, a label-free nano-liquid chromatography-mass spectrometry approach was used to determine the effects of sodium nitroprusside (SNP) on polyethylene glycol (PEG)-induced osmotic stress in banana roots. Plant treatment with SNP improved plant growth and reduced the percentage of yellow leaves. A total of 30 and 90 proteins were differentially identified in PEG+SNP against PEG and PEG+SNP against the control, respectively. The majority of proteins differing between them were related to carbohydrate and energy metabolisms. Antioxidant enzyme activities, such as superoxide dismutase and ascorbate peroxidase, decreased in SNP-treated banana roots compared to PEG-treated banana. These results suggest that the nitric oxide-induced osmotic stress tolerance could be associated with improved carbohydrate and energy metabolism capability in higher plants.

An Optimized Protein Extraction Method for Gel-Free Proteomic Analysis of Opuntia Ficus-Indica.

Opuntia spp. is an economically important vegetable crop with high stress-tolerance and health benefits. However, proteomic analysis of the plant has been difficult due to the composition of its succulent cladodes; the abundant polysaccharides interfere with protein extraction. To facilitate proteomic analysis of this plant, we present a rapid and simple protein extraction method for Opuntia ficus-indica (L.) Miller. The optimized method produced highly reproducible protein patterns and was compatible with a gel-free quantitative workflow without the need for additional purification. We successfully analyzed the cladode mesocarp and exocarp tissues, resulting in the identification of 319 proteins. In addition, we used this method to examine the relative changes in the Opuntia proteome in response to salt stress to determine whether physiological changes could be captured. Qualified observations were obtained, revealing that salt stress increased phosphoenolpyruvate carboxylase abundance and decreased ribulose-bisphosphate carboxylase in young O. ficus-indica plants. These findings suggest that Crassulacean acid metabolism is promoted under salinity. This study highlights the efficacy of our optimized protein extraction method for elucidating the metabolic adaptations of Opuntia using gel-free proteomic analysis.

Myoparr-Associated and -Independent Multiple Roles of Heterogeneous Nuclear Ribonucleoprotein K during Skeletal Muscle Cell Differentiation.

RNA-binding proteins (RBPs) regulate cell physiology via the formation of ribonucleic-protein complexes with coding and non-coding RNAs. RBPs have multiple functions in the same cells; however, the precise mechanism through which their pleiotropic functions are determined remains unknown. In this study, we revealed the multiple inhibitory functions of heterogeneous nuclear ribonucleoprotein K (hnRNPK) for myogenic differentiation. We first identified hnRNPK as a lncRNA Myoparr binding protein. Gain- and loss-of-function experiments showed that hnRNPK repressed the expression of myogenin at the transcriptional level. The hnRNPK-binding region of Myoparr was required to repress myogenin expression. Moreover, hnRNPK repressed the expression of a set of genes coding for aminoacyl-tRNA synthetases in a Myoparr-independent manner. Mechanistically, hnRNPK regulated the eIF2α/Atf4 pathway, one branch of the intrinsic pathways of the endoplasmic reticulum sensors, in differentiating myoblasts. Thus, our findings demonstrate that hnRNPK plays lncRNA-associated and -independent multiple roles during myogenic differentiation, indicating that the analysis of lncRNA-binding proteins will be useful for elucidating both the physiological functions of lncRNAs and the multiple functions of RBPs.

Mesenchymal Bmp3b expression maintains skeletal muscle integrity and decreases in age-related sarcopenia.

Age-related sarcopenia constitutes an important health problem associated with adverse outcomes. Sarcopenia is closely associated with fat infiltration in muscle, which is attributable to interstitial mesenchymal progenitors. Mesenchymal progenitors are nonmyogenic in nature but are required for homeostatic muscle maintenance. However, the underlying mechanism of mesenchymal progenitor-dependent muscle maintenance is not clear, nor is the precise role of mesenchymal progenitors in sarcopenia. Here, we show that mice genetically engineered to specifically deplete mesenchymal progenitors exhibited phenotypes markedly similar to sarcopenia, including muscle weakness, myofiber atrophy, alterations of fiber types, and denervation at neuromuscular junctions. Through searching for genes responsible for mesenchymal progenitor-dependent muscle maintenance, we found that Bmp3b is specifically expressed in mesenchymal progenitors, whereas its expression level is significantly decreased during aging or adipogenic differentiation. The functional importance of BMP3B in maintaining myofiber mass as well as muscle-nerve interaction was demonstrated using knockout mice and cultured cells treated with BMP3B. Furthermore, the administration of recombinant BMP3B in aged mice reversed their sarcopenic phenotypes. These results reveal previously unrecognized mechanisms by which the mesenchymal progenitors ensure muscle integrity and suggest that age-related changes in mesenchymal progenitors have a considerable impact on the development of sarcopenia.

The Effect of Mung Bean (Vigna radiata (L.)) Coat Extract on Mouse Liver Metabolism During Progesterone Withdrawal.

Mung bean (Vigna radiata) is an immunomodulatory medicinal plant, which is recognized as a component of a traditional postpartum diet. The liver plays a crucial role in fatty acid synthesis under the control of various hormones that are affected by pregnancy. This study was designed to establish whether the mung bean water extract, which contains prostaglandins that can regulate corpus luteum maturation, provided any benefits to liver metabolism after the dynamic hormonal change associated with pregnancy. Female C57BL/6J mice were used, and all mice received daily injections of progesterone (5.0 mg/kg) for 5 days, after which progesterone was withdrawn for 3 days. Gel-free/label-free proteomic analysis revealed that the abundance of several proteins was affected in the liver. Hormone manipulation induced changes in lipid metabolism-related protein abundance; oral administration of mung bean coat extract (MBC) for 3 days mitigated the changes and downregulated the expression of Cpt1α, Akr1ß, and Srebp1 in the liver. Together with immunological leukocyte modulation assessed via proteomic analysis, we suggest that MBC may exert health-promoting effects through the modulation of lipid synthesis during postpartum recovery.

Methods for Accurate Assessment of Myofiber Maturity During Skeletal Muscle Regeneration.

Adult skeletal muscle has a remarkable ability to regenerate. Regeneration of mature muscle fibers is dependent on muscle stem cells called satellite cells. Although they are normally in a quiescent state, satellite cells are rapidly activated after injury, and subsequently proliferate and differentiate to make new muscle fibers. Myogenesis is a highly orchestrated biological process and has been extensively studied, and therefore many parameters that can precisely evaluate regenerating events have been established. However, in some cases, it is necessary to evaluate the completion of regeneration rather than ongoing regeneration. In this study, we establish methods for assessing the myofiber maturation during muscle regeneration. By carefully comparing expression patterns of several muscle regeneration-related genes, we found that expression of Myozenin (Myoz1 and Myoz3), Troponin I (Tnni2), and Dystrophin (Dmd) is gradually increased as muscle regeneration proceeds. In contrast, commonly used regeneration markers such as Myh3 and Myh8 are transiently upregulated after muscle injury but their expression decreases as regeneration progresses. Intriguingly, upregulation of Myoz1, Myoz3 and Tnni2 cannot be achieved in cultured myotubes, indicating that these markers are excellent indicators to assess myofiber maturity. We also show that analyzing re-expression of Myoz1 and dystrophin in individual fiber during regeneration enables accurate assessment of myofiber maturity at the single-myofiber level. Together, our study provides valuable methods that are useful in evaluating muscle regeneration and the efficacy of therapeutic strategies for muscle diseases.