Protective effects and molecular mechanisms of Litopenaeus vannamei treated with l-arginine/l-lysine against myofibrillar proteins oxidation and quality degradation during freeze-thaw cycles.

The storage and processing of Litopenaeus vannamei are often challenged by the freeze-thaw (F-T) cycle phenomenon. This study delved into the influence of pretreatment with l-arginine (Arg) and l-lysine (Lys) on the myofibrillar proteins oxidation and quality of shrimp subjected to F-T cycles. Arg and Lys pretreatment notably improved water-holding capacity (WHC), textural integrity as well as the myofibrillar structure of the shrimps. A lesser reduction in the amounts of immobile and bound water was found in the amino acid-treated groups, and the oxidation of lipids and proteins were both decelerated. Molecular simulation results indicated that Arg and Lys could form hydrogen and salt-bridge bonds with myosin, enhancing the stability of Litopenaeus vannamei. The study concludes that Arg and Lys are effective in alleviating the adverse effects of F-T cycles on the quality of Litopenaeus vannamei, and provides a new solution for the quality maintenance during storage and processing.

Daidzein Inhibits Muscle Atrophy by Suppressing Inflammatory Cytokine- and Muscle Atrophy-Related Gene Expression.

BACKGROUND: Sarcopenic obesity, which is associated with a poorer prognosis than that of sarcopenia alone, may be positively affected by soy isoflavones, known inhibitors of muscle atrophy. Herein, we hypothesize that these compounds may prevent sarcopenic obesity by upregulating the gut metabolites with anti-inflammatory effects. METHODS: To explore the effects of soy isoflavones on sarcopenic obesity and its mechanisms, we employed both in vivo and in vitro experiments. Mice were fed a high-fat, high-sucrose diet with or without soy isoflavone supplementation. Additionally, the mouse C2C12 myotube cells were treated with palmitic acid and daidzein in vitro. RESULTS: The isoflavone considerably reduced muscle atrophy and the expression of the muscle atrophy genes in the treated group compared to the control group (Fbxo32, p = 0.0012; Trim63, p < 0.0001; Foxo1, p < 0.0001; Tnfa, p = 0.1343). Elevated levels of daidzein were found in the muscles and feces of the experimental group compared to the control group (feces, p = 0.0122; muscle, p = 0.0020). The real-time PCR results demonstrated that the daidzein decreased the expression of the palmitate-induced inflammation and muscle atrophy genes in the C2C12 myotube cells (Tnfa, p = 0.0201; Il6, p = 0.0008; Fbxo32, p < 0.0001; Hdac4, p = 0.0002; Trim63, p = 0.0114; Foxo1, p < 0.0001). Additionally, it reduced the palmitate-induced protein expression related to the muscle atrophy in the C2C12 myotube cells (Foxo1, p = 0.0078; MuRF1, p = 0.0119). CONCLUSIONS: The daidzein suppressed inflammatory cytokine- and muscle atrophy-related gene expression in the C2C12 myotubes, thereby inhibiting muscle atrophy.

Filbertone Reduces Senescence in C2C12 Myotubes Treated with Doxorubicin or H2O2 through MuRF1 and Myogenin.

It has been demonstrated that filbertone, the principal flavor compound of hazelnuts, exhibits preventive effects against hypothalamic inflammation, obesity, neurodegenerative diseases, and muscle lipid accumulation. However, its influence on muscle aging has yet to be elucidated. The objective of this study was to investigate the effects of filbertone on muscle aging in C2C12 myotubes subjected to senescence induction by either doxorubicin or hydrogen peroxide. To ascertain the mechanisms by which filbertone exerts its effects, we conducted a series of experiments, including Western blot analysis, reverse transcription quantitative polymerase chain reaction (qRT-PCR), and senescence-associated ß-galactosidase (SA-ß-gal) staining. Filbertone was markedly observed to decrease not only the protein levels of p53 (p < 0.01) in senescence-induced skeletal muscle cells, but also the gene expression levels of p21 (p < 0.05), a direct target of p53. The expression of muscle-related genes, including myogenin and muscle RING-finger protein-1 (MuRF1), was found to be significantly enhanced in senescent muscle cells following treatment with filbertone (p < 0.05). In addition, the number of senescent skeletal muscle cells exhibiting ß-galactosidase activity was found to be markedly reduced in the presence of filbertone (p < 0.01). Collectively, these findings suggest that filbertone plays a pivotal role in the regulation of muscle aging.

ANKRD1 aggravates renal ischaemia‒reperfusion injury via promoting TRIM25-mediated ubiquitination of ACSL3.

BACKGROUND: Renal ischaemia‒reperfusion injury (IRI) is the primary cause of acute kidney injury (AKI). To date, effective therapies for delaying renal IRI and postponing patient survival remain absent. Ankyrin repeat domain 1 (ANKRD1) has been implicated in some pathophysiologic processes, but its role in renal IRI has not been explored. METHODS: The mouse model of IRI-AKI and in vitro model were utilised to investigate the role of ANKRD1. Immunoprecipitation-mass spectrometry was performed to identify potential ANKRD1-interacting proteins. Protein‒protein interactions and protein ubiquitination were examined using immunoprecipitation and proximity ligation assay and immunoblotting, respectively. Cell viability, damage and lipid peroxidation were evaluated using biochemical and cellular techniques. RESULTS: First, we unveiled that ANKRD1 were significantly elevated in renal IRI models. Global knockdown of ANKRD1 in all cell types of mouse kidney by recombinant adeno-associated virus (rAAV9)-mitigated ischaemia/reperfusion-induced renal damage and failure. Silencing ANKRD1 enhanced cell viability and alleviated cell damage in human renal proximal tubule cells exposed to hypoxia reoxygenation or hydrogen peroxide, while ANKRD1 overexpression had the opposite effect. Second, we discovered that ANKRD1's detrimental function during renal IRI involves promoting lipid peroxidation and ferroptosis by directly binding to and decreasing levels of acyl-coenzyme A synthetase long-chain family member 3 (ACSL3), a key protein in lipid metabolism. Furthermore, attenuating ACSL3 in vivo through pharmaceutical approach and in vitro via RNA interference mitigated the anti-ferroptotic effect of ANKRD1 knockdown. Finally, we showed ANKRD1 facilitated post-translational degradation of ACSL3 by modulating E3 ligase tripartite motif containing 25 (TRIM25) to catalyse K63-linked ubiquitination of ACSL3, thereby amplifying lipid peroxidation and ferroptosis, exacerbating renal injury. CONCLUSIONS: Our study revealed a previously unknown function of ANKRD1 in renal IRI. By driving ACSL3 ubiquitination and degradation, ANKRD1 aggravates ferroptosis and ultimately exacerbates IRI-AKI, underlining ANKRD1's potential as a therapeutic target for kidney IRI. KEY POINTS/HIGHLIGHTS: Ankyrin repeat domain 1 (ANKRD1) is rapidly activated in renal ischaemia‒reperfusion injury (IRI) models in vivo and in vitro. ANKRD1 knockdown mitigates kidney damage and preserves renal function. Ferroptosis contributes to the deteriorating function of ANKRD1 in renal IRI. ANKRD1 promotes acyl-coenzyme A synthetase long-chain family member 3 (ACSL3) degradation via the ubiquitin‒proteasome pathway. The E3 ligase tripartite motif containing 25 (TRIM25) is responsible for ANKRD1-mediated ubiquitination of ACSL3.

Buzhong Yiqi decoction attenuates acquired myasthenia by regulating the JAK2/STAT3/AKT signaling pathway, inhibiting inflammation, and improving mitochondrial function.

Acquired myasthenia (AM), a debilitating autoimmune disease, is typically characterized by skeletal muscle fatigue and weakness. Despite advances in myasthenia gravis treatment, current approaches remain unsatisfactory and many result in unexpected side effects. Traditional Chinese medicine has shown great potential in the treatment of myasthenia gravis, including relieving myasthenic symptoms, improving patients' quality of life, and reducing Western medicine side effects. This study investigates the protective effects and mechanism of BZYQD in mice with acquired myasthenia. BZYQD alleviates the reduced grip strength and increased expression of MAFbx and MuRF-1 in mice with acquired myasthenia. It also reduces levels of pro-inflammatory factors IL-1ß, IL-6, and TNF-α in the mouse serum. In addition, BZYQD reduces ROS accumulation and the mitochondrial ROS production rate, while increasing ATP levels and mitochondrial membrane potential in mice with acquired myasthenia. Moreover, BZYQD decreases the expression of p-JAK2, p-STAT3, and p-AKT in the skeletal muscle of mice with acquired myasthenia. In summary, BZYQD reduces inflammation, enhances mitochondrial function, and regulates the JAK2/STAT3/AKT signaling pathway to treat acquired myasthenia.

CCDC78: Unveiling the Function of a Novel Gene Associated with Hereditary Myopathy.

CCDC78 was identified as a novel candidate gene for autosomal dominant centronuclear myopathy-4 (CNM4) approximately ten years ago. However, to date, only one family has been described, and the function of CCDC78 remains unclear. Here, we analyze for the first time a family harboring a CCDC78 nonsense mutation to better understand the role of CCDC78 in muscle. METHODS: We conducted a comprehensive histopathological analysis on muscle biopsies, including immunofluorescent assays to detect multiple sarcoplasmic proteins. We examined CCDC78 transcripts and protein using WB in CCDC78-mutated muscle tissue; these analyses were also performed on muscle, lymphocytes, and fibroblasts from healthy subjects. Subsequently, we conducted RT-qPCR and transcriptome profiling through RNA-seq to evaluate changes in gene expression associated with CCDC78 dysfunction in muscle. Lastly, coimmunoprecipitation (Co-Ip) assays and mass spectrometry (LC-MS/MS) studies were carried out on extracted muscle proteins from both healthy and mutated subjects. RESULTS: The histopathological features in muscle showed novel histological hallmarks, which included areas of dilated and swollen sarcoplasmic reticulum (SR). We provided evidence of nonsense-mediated mRNA decay (NMD), identified the presence of novel CCDC78 transcripts in muscle and lymphocytes, and identified 1035 muscular differentially expressed genes, including several involved in the SR. Through the Co-Ip assays and LC-MS/MS studies, we demonstrated that CCDC78 interacts with two key SR proteins: SERCA1 and CASQ1. We also observed interactions with MYH1, ACTN2, and ACTA1. CONCLUSIONS: Our findings provide insight, for the first time, into the interactors and possible role of CCDC78 in skeletal muscle, locating the protein in the SR. Furthermore, our data expand on the phenotype previously associated with CCDC78 mutations, indicating potential histopathological hallmarks of the disease in human muscle. Based on our data, we can consider CCDC78 as the causative gene for CNM4.

Methods for production and assaying catalysis of isolated recombinant human aspartate/asparagine-ß-hydroxylase.

Aspartate/asparagine-ß-hydroxylase (AspH) is a transmembrane 2-oxoglutarate (2OG)-dependent oxygenase that catalyzes the post-translational hydroxylation of aspartate- and asparagine-residues in epidermal growth factor-like domains (EGFDs) of its substrate proteins. Upregulation of ASPH and translocation of AspH from the endoplasmic reticulum membrane to the surface membrane of cancer cells is associated with enhanced cell motility and worsened clinical prognosis. AspH is thus a potential therapeutic and diagnostic target for cancer. This chapter describes methods for the production and purification of soluble constructs of recombinant human AspH suitable for biochemical and crystallographic studies. The chapter also describes efficient methods for performing turnover and inhibition assays which monitor catalysis of isolated recombinant human AspH in vitro using solid phase extraction coupled to mass spectrometry (SPE-MS). The SPE-MS assays employ synthetic disulfide- or thioether-bridged macrocyclic oligopeptides as substrates; a macrocycle is an apparently essential requirement for productive AspH catalysis and mimics an EGFD disulfide isomer that is not typically observed in crystal and NMR structures. SPE-MS assays can be used to monitor catalysis of 2OG oxygenases other than AspH; the methods described herein are representative for 2OG oxygenase SPE-MS assays useful for performing kinetic and/or inhibition studies.

Insight into the effects of ultrasound-assisted intermittent tumbling on the gelation properties of myofibrillar proteins: Conformational modifications, intermolecular interactions, rheological properties and microstructure.

The aim of the present study was to evaluate the effects of ultrasound-assisted intermittent tumbling (UT) at 300 W, 20 kHz and 40 min on the conformation, intermolecular interactions and aggregation of myofibrillar proteins (MPs) and its induced gelation properties at various tumbling times (4 and 6 h). Raman results showed that all tumbling treatments led the helical structure of MPs to unfold. In comparison to the single intermittent tumbling treatment (ST), UT treatment exerted more pronounced effects on strengthening the intermolecular hydrogen bonds and facilitating the formation of an ordered ß-sheet structure. When the tumbling time was the same, UT treatment caused higher surface hydrophobicity, fluorescence intensity and disulfide bond content in the MPs, inducing the occurrence of hydrophobic interaction and disulfide cross-linking between MPs molecules, thus forming the MPs aggregates. Additionally, results from the solubility, particle size, atomic force microscopy and SDS-PAGE further indicated that, relative to the ST treatment, UT treatment was more potent in promoting the polymerization of myosin heavy chain. The MPs aggregates in the UT group were more uniform than those in the ST group. During the gelation process, the pre-formed MPs aggregates in the UT treatment increased the thermal stability of myosin, rendering it more resistant to heat-induced unfolding of the myosin rod region. Furthermore, they improved the protein tail-tail interaction, resulting in the formation of a well-structured gel network with higher gel strength and cooking yield compared to the ST treatment.

Functional and Mechanistic Dissection of Protein Glutaminase PG3 and Its Rational Engineering for Enhanced Modification of Myofibrillar Proteins.

Protein glutaminases (PG; EC = 3.5.1.44) are enzymes known for enhancing protein functionality. In this study, we cloned and expressed the gene chryb3 encoding protein glutaminase PG3, exhibiting 39.4 U/mg specific activity. Mature-PG3 featured a substrate channel surrounded by aromatic and hydrophobic amino acids at positions 38-45 and 78-84, with Val81 playing a pivotal role in substrate affinity. The dynamic opening and closing motions between Gly65, Thr66, and Cys164 at the catalytic cleft greatly influence substrate binding and product release. Redesigning catalytic pocket and cocatalytic region produced combinatorial mutant MT6 showing a 2.69-fold increase in specific activity and a 2.99-fold increase at t65 °C1/2. Furthermore, MT6 boosted fish myofibrillar protein (MP) solubility without NaCl. Key residues such as Thr3, Asn54, Val81, Tyr82, Asn107, and Ser108 were vital for PG3-myosin interaction, particularly Asn54 and Asn107. This study sheds light on the catalytic mechanism of PG3 and guided its rational engineering and utilization in low-salt fish MP product production.

RBM8A, a new target of TEAD4, promotes breast cancer progression by regulating IGF1R and IRS-2.

BACKGROUND: Breast cancer (BC) is the most common malignant tumor in women worldwide, and further elucidation of the molecular mechanisms involved in BC pathogenesis is essential to improve the prognosis of BC patients. RNA Binding Motif Protein 8 A (RBM8A), with high affinity to a myriad of RNA transcripts, has been shown to play a crucial role in genesis and progression of multiple cancers. We attempted to explore its functional significance and molecular mechanisms in BC. METHODS: Bioinformatics analysis was performed on publicly available BC datasets. qRT-PCR was used to determine the expression of RBM8A in BC tissues. MTT assay, clone formation assay and flow cytometry were employed to examine BC cell proliferation and apoptosis in vitro. RNA immunoprecipitation (RIP) and RIP-seq were used to investigate the binding of RBM8A/EIF4A3 to the mRNA of IGF1R/IRS-2. RBM8A and EIF4A3 interactions were determined by co-immunoprecipitation (Co-IP) and immunofluorescence. Chromatin immunoprecipitation (Ch-IP) and dual-luciferase reporter assay were carried out to investigate the transcriptional regulation of RBM8A by TEAD4. Xenograft model was used to explore the effects of RBM8A and TEAD4 on BC cell growth in vivo. RESULTS: In this study, we showed that RBM8A is abnormally highly expressed in BC and knockdown of RBM8A inhibits BC cell proliferation and induces apoptosis in vitro. EIF4A3, which phenocopy RBM8A in BC, forms a complex with RBM8A in BC. Moreover, EIF4A3 and RBM8A complex regulate the expression of IGF1R and IRS-2 to activate the PI3K/AKT signaling pathway, thereby promoting BC progression. In addition, we identified TEAD4 as a transcriptional activator of RBM8A by Ch-IP, dual luciferase reporter gene and a series of functional rescue assays. Furthermore, we demonstrated the in vivo pro-carcinogenic effects of TEAD4 and RBM8A by xenograft tumor experiments in nude mice. CONCLUSION: Collectively, these findings suggest that TEAD4 novel transcriptional target RBM8A interacts with EIF4A3 to increase IGF1R and IRS-2 expression and activate PI3K/AKT signaling pathway, thereby further promoting the malignant phenotype of BC cells.

Blockade of forkhead box protein O1 signaling alleviates primary sclerosing cholangitis-induced sarcopenia in mice model.

AIMS: Primary sclerosing cholangitis (PSC) is a cholestatic liver disease that affects the hepatic bile ducts, leading to hepatic inflammation and fibrosis. PSC can also impact skeletal muscle through the muscle-liver axis, resulting in sarcopenia, a complication characterized by a generalized loss of muscle mass and strength. The underlying mechanisms and therapy of PSC-induced sarcopenia are not well understood, but one potential regulator is the transcription factor forkhead box protein O1 (FOXO1), which is involved in the ubiquitin proteasome system. Thus, the aim of this study is to assess the pharmacological potential of FOXO1 inhibition for treating PSC-induced sarcopenia. MATERIALS AND METHODS: To establish diet-induced PSC model, we provided mice with a 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) diet for 4 weeks. Mice were intramuscularly injected with AS1842856 (AS), a FOXO1 inhibitor, at a dose of 3.5 mg/kg twice a week for last two weeks. C2C12 myotubes with cholic acid (CA) or deoxycholic acid (DCA) were treated with AS. KEY FINDINGS: We observed a decrease in muscle size and performance in DDC-fed mice with upregulated expression of FOXO1 and E3 ligases such as ATROGIN1 and MuRF1. We found that myotube diameter and MyHC protein level were decreased by CA or DCA in C2C12 myotubes, but treatment of AS reversed these reductions. We observed that intramuscular injection of AS effectively mitigates DDC diet-induced sarcopenia in a rodent PSC model. SIGNIFICANCE: Our study suggests that a FOXO1 inhibitor could be a potential leading therapeutic drug for relieving PSC-induced sarcopenia.

Effects of the FHL2 gene on the development of subcutaneous and intramuscular adipocytes in goats.

BACKGROUND: Adipose tissue affects not only the meat quality of domestic animals, but also human health. Adipocyte differentiation is regulated by a series of regulatory genes and cyclins. Four and half-LIM protein (FHL2) is positively correlated with the hypertrophy of adipocytes and can cause symptoms such as obesity and diabetes. RESULT: In the transcriptome sequencing analysis of intramuscular adipocytes after three days of differentiation, the differentially expressed gene FHL2 was found. To further explore the biological significance of the differentially expressed gene FHL2, which was downregulated in the mature adipocytes. We revealed the function of FHL2 in adipogenesis through the acquisition and loss of function of FHL2. The results showed that the overexpression of FHL2 significantly increased the expression of adipogenic genes (PPARγ, C/EBPß) and the differentiation of intramuscular and subcutaneous adipocytes. However, silencing FHL2 significantly inhibited adipocyte differentiation. The overexpression of FHL2 increased the number of adipocytes stained with crystal violet and increased the mRNA expression of proliferation marker genes such as CCNE, PCNA, CCND and CDK2. In addition, it significantly increased the rate of EdU positive cells. In terms of apoptosis, overexpression of FHL2 significantly inhibited the expression of P53 and BAX in both intramuscular and subcutaneous adipocytes, which are involved in cell apoptosis. However, overexpression of FHL2 promoted the expression of BCL, but was rescued by the silencing of FHL2. CONCLUSIONS: In summary, FHL2 may be a positive regulator of intramuscular and subcutaneous adipocyte differentiation and proliferation, and acts as a negative regulator of intramuscular and subcutaneous adipocyte apoptosis. These findings provide a theoretical basis for the subsequent elucidation of FHL2 in adipocytes.

Circulating Amino Acid Concentration after the Consumption of Pea or Whey Proteins in Young and Older Adults Affects Protein Synthesis in C2C12 Myotubes.

As older adults tend to reduce their intake of animal-source proteins, plant-source proteins may offer valuable resources for better protein intake. The aim of this study was to assess whether the pea proteins can be used to achieve blood amino acid levels that stimulate muscle protein synthesis. We measured variations in plasma amino acid concentrations in young and older adults given pea (NUTRALYS® S85 Plus) or whey proteins either alone or in a standardized meal. The effect of amino acid concentrations on protein synthesis in C2C12 myotubes was determined. In terms of results, plasma amino acid concentrations reflected the difference between the amino acid contents of whey and pea proteins. Blood leucine showed a greater increase of 91 to 130% with whey protein compared to pea protein, while the opposite was observed for arginine (A greater increase of 147 to 210% with pea compared to whey). Culture media prepared with plasmas from the human study induced age-dependent but not protein-type-dependent changes in myotube protein synthesis. In conclusion, pea and whey proteins have the same qualities in terms of their properties to maintain muscle protein synthesis. Pea proteins can be recommended for older people who do not consume enough animal-source proteins.

Binding Behavior of Human Hepatoma-Derived Growth Factor on SMYD1.

The protein-induced fluorescence change technique was employed to investigate the interactions between proteins and their DNA substrates modified with the Cy3 fluorophore. It has been reported that the human hepatoma-derived growth factor (HDGF), containing the chromatin-associated N-terminal proline-tryptophan-tryptophan-proline (PWWP) domain (the N-terminal 100 amino acids of HDGF) capable of binding the SMYD1 promoter, participates in various cellular processes and is involved in human cancer. This project investigated the specific binding behavior of HDGF, the PWWP domain, and the C140 domain (the C-terminal 140 amino acids of HDGF) sequentially using protein-induced fluorescence change. We found that the binding of HDGF and its related proteins on Cy3-labeled 15 bp SMYD1 dsDNA will cause a significant decrease in the recorded Cy3 fluorophore intensity, indicating the occurrence of protein-induced fluorescence quenching. The dissociation equilibrium constant was determined by fitting the bound fraction curve to a binding model. An approximate 10-time weaker SMYD1 binding affinity of the PWWP domain was found in comparison to HDGF. Moreover, the PWWP domain is required for DNA binding, and the C140 domain can enhance the DNA binding affinity. Furthermore, we found that the C140 domain can regulate the sequence-specific binding capability of HDGF on SMYD1.

Sequential window acquisition of all theoretical mass spectra (SWATH-MS) as an emerging proteomics approach for the discovery of dark-cutting beef biomarkers.

Recent advances in "omics" technologies have enabled the identification of new beef quality biomarkers and have also allowed for the early detection of quality defects such as dark-cutting beef, also known as DFD (dark, firm, and dry) beef. However, most of the studies conducted were carried out on a small number of animals and mostly applied gel-based proteomics. The present study proposes for the first time a Sequential Window Acquisition of All Theoretical Mass Spectra (SWATH-MS) proteomics approach to characterize and comprehensively quantify the post-mortem muscle proteome of DFD (pH24 ≥ 6.2) and CONTROL (5.4 ≤ pH24 ≤ 5.6) beef samples within the largest database of DFD/CONTROL beef samples to date (26 pairs of the Longissimus thoracis muscle samples of young bulls from Asturiana de los Valles breed, n = 52). The pairwise comparison yielded 35 proteins that significantly differed in their abundances between the DFD and CONTROL samples. Chemometrics methods using both PLS-DA and OPLS-DA revealed 31 and 36 proteins with VIP > 2.0, respectively. The combination of different statistical methods these being Volcano plot, PLS-DA and OPLS-DA allowed us to propose 16 proteins as good candidate biomarkers of DFD beef. These proteins are associated with interconnected biochemical pathways related to energy metabolism (DHRS7B and CYB5R3), binding and signaling (RABGGTA, MIA3, BPIFA2B, CAP2, APOBEC2, UBE2V1, KIR2DL1), muscle contraction, structure and associated proteins (DMD, PFN2), proteolysis, hydrolases, and activity regulation (AGT, C4A, GLB1, CAND2), and calcium homeostasis (ANXA6). These results evidenced the potential of SWATH-MS and chemometrics to accurately identify novel biomarkers for meat quality defects, providing a deeper understanding of the molecular mechanisms underlying dark-cutting beef condition.

Influence of NaCl and phosphate on gelation properties of chicken breast myofibrillar protein gels and its application to in vitro digestion model.

To investigate the combination effect of sodium chloride and phosphates on chicken breast myofibrillar proteins, MP gels containing various molarity of NaCl (0.15, 0.30 and 0.45 M) and phosphate (0 and 0.05 M) were prepared, their rheological properties were characterized, and applied to an in vitro digestion model. MP mixture containing 0.45 M NaCl and 0.05 M phosphate had the highest viscosity. The gel strength and cooking yield of MP gels was improved by increasing of molarity of NaCl. As NaCl concentration in MP increased, sulfhydryl levels decreased, while disulfide levels increased. As NaCl and phosphate levels increase, MP gels become denser and porosity decreases, which may reduce protein digestibility. In SDS-PAGE, protein bands from MP gels containing low NaCl levels (≤ 0.30 M) degraded more rapidly during in vitro digestion. These results may support the need for the meat industry to develop low-salt meat products with improved digestibility. KEYWORDS: Chicken, Myofibrillar protein, NaCl, Phosphate, Rheological properties, In vitro digestion.

Investigation on synergistic inhibition of protein-bound heterocyclic amines in sarcoplasmic and myofibrillar model systems by amino acid combinations.

The inhibition of amino acids on the formation of protein-bound HAs was assessed in both model systems and roast beef patties, and the synergism between these amino acids was also investigated. The amino acids can promote the formation of protein-bound HAs at low addition amount, and the total content of protein-bound HAs increased from 444.05 ± 4.98 ng/g of the control group to 517.36 ± 16.51 ng/g when 0.05 % cysteine was added. Amino acid combinations exhibited stable inhibitory effects, with the maximum inhibitory rate of 64 % in the treatment with histidine-proline combination (1:4). The synergistic inhibition may be caused by simultaneously scavenging intermediates and competing for the binding sites of muscle proteins, and the reaction with protein-bound HAs to form adduct can serve as supporting factors to co-mitigate the promotion in protein-bound HAs from increased protein solubility. These findings proposed the potential mitigation strategies against protein-bound HAs formation.

Propofol pretreatment inhibits ferroptosis and alleviates myocardial ischemia-reperfusion injury through the SLC16A13-AMPK-GPX4 pathway.

This study investigates the protective effects of propofol on the myocardium by inhibiting the expression of SLC16A13 through in vivo animal experiments, while also exploring its mechanism in ferroptosis to provide new strategies for preventing perioperative myocardial ischemia-reperfusion injury. We randomly divided 30 rats into three groups (n=10 each): sham surgery group, ischemia-reperfusion (I/R) group, and propofol pretreatment group. The results showed that compared with the sham surgery group, the I/R group had a significant decrease in cardiac function and an increase in infarct size. Propofol pretreatment effectively alleviated the damage caused by ischemia-reperfusion (I/R). In the next phase of the study, we administered the PPARα agonist GW7647 to artificially increase the expression of SLC16A13. Fifty rats were randomly divided into five groups (n=10 each), with the GW7647 pretreatment group and propofol+GW7647 pretreatment group added based on the previous three groups. Afterwards, we validated the in vivo results using H9C2 and further explored the mechanism by which propofol inhibits ferroptosis. The study found that L-lactic acid in myocardial tissue of the GW7647 group was further increased compared to the I/R group, and the degree of ferroptosis was aggravated. In addition, upregulation of SLC16A13 significantly inhibited the phosphorylation of AMPK, weakened the protective mechanism of AMPK, and exacerbated cardiac damage. However, propofol pretreatment can effectively inhibit the expression of SLC16A13, maintain normal myocardial cell morphology, and protect cardiac function. These results indicate that propofol inhibits the expression of SLC16A13, alleviates myocardial cell ferroptosis via the AMPK/GPX4 pathway, and reverses damage caused by myocardial ischemia-reperfusion.

FHL1: A novel diagnostic marker for papillary thyroid carcinoma.

Although there are clear morphologic criteria for the diagnosis of papillary thyroid carcinoma (PTC), when the morphology is untypical or overlaps, accurate diagnostic indicators are necessary. Since few studies investigated the role of down-regulated genes in PTC, this article aims to further explore the molecular markers associated with PTC. We conducted bioinformatics analysis of gene microarrays of PTC and normal adjacent tissues. Besides, quantitative real-time quantitative polymerase chain reaction array and immunohistochemical staining were used to investigate the expression of the major down-regulated genes. The results indicated that several important down-regulated genes, including TLE1, BCL2, FHL1, GHR, KIT, and PPARGC1A were involved in the process of PTC. Compared to normal adjacent tissues, the mRNA expression of the major genes was down-regulated in PTC (p＜0.05). Immunohistochemically, FHL1 shows negative or low expression in PTC tissues (p＜0.05). BCL2 did not show a significant difference between PTC and normal thyroid tissues (p > 0.05). TLE1, KIT, PPARGC1A and GHR showed negative expression in both tumor and normal tissues. These results suggested that FHL1 could serve as a novel tumor marker for precise diagnosis of PTC.