Objective To elucidate the molecular mechanism of sirtuin-3 (SIRT3) in regulating mitochondrial biogenesis in human renal tubular epithelial cells. Methods Cells were stimulated with different concentrations of H₂O₂ and divided into four groups: control (NC), 50 μmol/L H₂O₂, 110 μmol/L H₂O₂ and 150 μmol/L H₂O₂. SIRT3 protein expression was then measured. SIRT3 was knocked down with siRNA, and cells were further assigned to five groups: control (NC), negative-control siRNA (NCsi), SIRT3-siRNA (siSIRT3), NCsi+H₂O₂, and siSIRT3+H₂O₂. After 24 h, cellular adenosine triphosphate (ATP) and mitochondrial superoxide anion (O₂•⁻) levels were determined, together with mitochondrial expression of SIRT3, peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α), nuclear respiratory factor 1 (NRF1), mitochondrial transcription factor A (TFAM), superoxide dismutase 2 (SOD2), acetylated-SOD2 and adenosine monophosphate activated protein kinase α1 (AMPKα1). Results The 110 and 150 μmol/L H₂O₂ decreased SIRT3 protein (both P<0.05). ATP and mitochondrial O₂•⁻ did not differ between NC and NCsi groups (both P>0.05). Compared to the NCsi group, the siSIRT3 group exhibited elevated O₂•⁻ level, decreased SIRT3 protein and increased expression levels of SOD2 and acetylated SOD2 protein (all P<0.05). Compared to the NCsi group, the NCsi+H₂O₂ group exhibited decreased cellular ATP levels, elevated mitochondrial O₂•⁻ levels, and reduced protein expression levels of SIRT3, SOD2, TFAM, AMPKα1, PGC-1α and NRF1 (all P<0.05). Compared with the siSIRT3 group, the siSIRT3+H₂O₂ group showed a decrease in cellular ATP levels, an increase in mitochondrial O₂•⁻ levels, a decrease in SIRT3, SOD2, TFAM, AMPKα1, PGC-1α and NRF1 protein expression levels and a decrease in acetylated SOD2 protein expression levels (all P<0.05). Compared with the NCsi+H₂O₂ group, the siSIRT3+H₂O₂ group showed a decrease in cellular ATP levels, an increase in mitochondrial O₂•⁻ levels, a decrease in SIRT3, AMPKα1, PGC-1α and NRF1, TFAM protein expression levels, and an increase in SOD2 and acetylated SOD2 protein expression levels (all P<0.05). Conclusions SIRT3 promotes mitochondrial biogenesis in tubular epithelial cells via the AMPK/PGC-1α/NRF1/TFAM axis, representing a key mechanism through which SIRT3 ameliorates oxidative stress-induced mitochondrial dysfunction.

Objective To evaluate the value of serum Mac-2 binding protein (M2BP) for assessment of the severity of schisto somiasis-induced liver fibrosis, so as to provide insights into non-invasive diagnosis and disease surveillance of liver fibrosis caused by schistosomiasis. Methods A total of 234 individuals with a history of Schistosoma japonicum infection were sampled from Xinhua Village, Lushan City, Jiangxi Province from 2019 to 2020, and 234 serum samples were collected from all participants. All participants received B-ultrasound examinations of the liver. Serum samples were categorized into four groups (grades 0, Ⅰ, Ⅱ and Ⅲ schistosomiasis-induced liver fibrosis groups) according to B-ultrasound examination results, and then, each group was randomly divided into a receiver operating characteristic (ROC) curve group and an efficacy assessment group at a ratio of 7∶3. Serum M2BP concentration was measured in four groups using the enzyme-linked immunosorbent assay (ELISA), and differences in serum M2BP concentrations were compared with analysis of variance and Spearman correlation analysis. Serum M2BP concentration was subjected to ROC curve analysis among individuals with different grades of schistosomiasis-induced liver fibrosis in the ROC curve group to determine the optimal diagnostic threshold of M2BP concentration at different fibrosis grades, and the area under the ROC curve (AUC) was calculated to evaluate the diagnostic performance. The diagnostic accuracy was verified by comparing the accordance rate and Kappa consistency test in the efficacy assessment group. Results Among 234 serum samples, there were 79 samples with grade 0 schistosomiasis-induced liver fibrosis, 87 samples with Grade Ⅰ, 46 samples with Grade Ⅱ and 22 samples with Grade Ⅲ according to the B-ultrasound examinations. The mean serum M2BP concentrations were (0.40 ± 0.31) [95% confidence interval (CI): (0.33, 0.47)], (0.64 ± 0.48) [95% CI: (0.53, 0.74)], (1.76 ± 0.58) [95% CI: (1.59, 1.93)] μg/mL and (2.56 ± 0.93) [95% CI: (2.14, 2.97)] μg/mL in the four groups, respectively (F = 150.796, P < 0.001), and the severity of schistosomiasis-induced liver fibrosis significantly positively correlated with serum M2BP concentration (rs = 0.715, P < 0. 001). The sample sizes of grades 0, Ⅰ, Ⅱ and Ⅲ schistosomiasis-induced liver fibrosis sera were randomly allocated as follows: 55 versus 24, 61 versus 26, 32 versus 14, and 15 versus 7 in the ROC curve and efficacy assessment groups, respectively, and the serum M2BP concentrations were (0.39 ± 0.29) μg/mL and (0.42 ± 0.36) μg/mL (F = 0.196, P > 0.05), (0.59 ± 0.47) μg/mL and (0.75 ± 0.51) μg/mL (F = 1.967, P > 0.05), (1.73 ± 0.59) μg/mL and (1.85 ± 0.57) μg/mL (F = 0.417, P > 0.05), and (2.46 ± 0.64) μg/mL and (2.76 ± 1.41) μg/mL (F = 0.491, P > 0.05), respectively. ROC curve analysis showed that the optimal diagnostic thresholds of serum M2BP concentration were 0.347 86 μg/mL (AUC = 0.635, P < 0.05), 1.188 83 μg/mL (AUC = 0.938, P < 0.000 1) and 2.021 21 μg/mL (AUC = 0.821, P < 0.000 1) for grade Ⅰ, Ⅱ and Ⅲ schistosomiasis-induced liver fibrosis. In addition, the accordance rates between the optimal diagnostic threshold of serum M2BP and B-ultrasound examinations for predicting grade Ⅰ, Ⅱ and Ⅲ schistosomiasis-induceed liver fibrosis were 69.23%, 85.71% and 71.43% (χ2 = 1.340, P > 0.05), and the overall Kappa consistency test showed moderate consistency [Kappa = 0.608, 95% CI: (0.428, 0.788); Z = 6.609, P < 0.000 1]. Conclusions Serum M2BP may serve as a potential biomarker for assessing moderate to advanced schistosomiasis-induced liver fibrosis; however, its diagnostic value for early-stage schistosomiasis-induced liver fibrosis remains limited.

BACKGROUND: Tissue engineering holds promise for vascular repair and regeneration by mimicking the extracellular matrix of blood vessels. However, achieving a functional and thick vascular wall with aligned fiber architecture by electrospinning remains a significant challenge. METHODS: A novel electrospinning setup was developed that utilizes an auxiliary electrode and a spring. The impact of process parameters on fiber size and morphology was investigated. The structure and functions of the scaffolds were evaluated through material characterization and assessments of cellular biocompatibility. RESULTS: The new setup enabled controlled deposition of fibers in different designed orientations. The fabricated small-diameter vascular scaffolds consisted of an inner layer of longitudinally oriented fibers and an outer layer of circumferentially oriented fibers (L + C vascular scaffold). Key parameters, including rotational speed, the utilization of the auxiliary electrode, and top-to-collector distance (TCD) significantly influenced fiber orientation. Additionally, voltage, TCD, feed rate, needle size, auxiliary electrode and collector-auxiliary electrode distance affected fiber diameter and distribution. Mechanical advantages and improved surface wettability of L + C vascular scaffold were confirmed through tensile testing and water contact angle. Cellular experiments indicated that L + C vascular scaffold facilitated cell adhesion and proliferation, with human umbilical vein endothelial cells and smooth muscle cells attaching and elongating along the fiber direction of the inner and outer layer, respectively. CONCLUSION This study demonstrated the feasibility of fabricating fiber-aligned, thick-walled vascular scaffolds using a modified electrospinning setup. The findings provided insights into how the auxiliary electrode, specific collector influenced fiber deposition, potentially advancing biomimetic vascular scaffold engineering.

Neuropathic pain is a chronic condition caused by damage or dysfunction in the nervous system. While the spleen may influence neuropathic pain, its role has been poorly understood. This study demonstrates that the spleen plays a crucial role in regulating neuropathic pain through the bed nucleus of the stria terminalis (BNST) - paraventricular nucleus of the hypothalamus (PVN) neural circuit in a chronic constriction injury (CCI) mouse model. Splenectomy, splenic denervation, or splenic sympathectomy significantly increased the mechanical withdrawal threshold (MWT) and reduced macrophage infiltration in the dorsal root ganglia (DRG) of CCI mice. Pseudorabies virus injections into the spleen revealed connections to the BNST and PVN in the brain. Chemogenetic inhibition of the BNST-PVN circuit increased macrophage infiltration in the DRG and decreased the MWT; these effects were reversed by splenectomy, splenic denervation, or sympathectomy. These findings underscore the critical role of the spleen, regulated by the BNST-PVN circuit, in neuropathic pain.
Animals ; Neuralgia/pathology* ; Septal Nuclei/physiopathology* ; Male ; Spleen/physiopathology* ; Paraventricular Hypothalamic Nucleus/physiopathology* ; Mice, Inbred C57BL ; Splenectomy ; Mice ; Neural Pathways/physiopathology* ; Disease Models, Animal ; Ganglia, Spinal/physiopathology* ; Sympathectomy ; Macrophages

Radiation-induced thrombocytopenia (RIT) faces a perplexing challenge in the clinical treatment of cancer patients, and current therapeutic approaches are inadequate in the clinical settings. In this research, oxymatrine, a new molecule capable of healing RIT was screened out, and the underlying regulatory mechanism associated with magakaryocyte (MK) differentiation and thrombopoiesis was demonstrated. The capacity of oxymatrine to induce MK differentiation was verified in K-562 and Meg-01 cells in vitro. The ability to induce thrombopoiesis was subsequently demonstrated in Tg (cd41:enhanced green fluorescent protein (eGFP)) zebrafish and RIT model mice. In addition, we carried out network pharmacological prediction, drug affinity responsive target stability assay (DARTS) and cellular thermal shift assay (CETSA) analyses to explore the potential targets of oxymatrine. Moreover, the pathway underlying the effects of oxymatrine was determined by Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses, Western blot (WB), and immunofluorescence. Oxymatrine markedly promoted MK differentiation and maturation in vitro. Moreover, oxymatrine induced thrombopoiesis in Tg (cd41:eGFP) zebrafish and accelerated thrombopoiesis and platelet function recovery in RIT model mice. Mechanistically, oxymatrine directly binds to toll-like receptor 2 (TLR2) and further regulates the downstream pathway stimulator of interferon genes (STING)/nuclear factor-kappaB (NF-κB), which can be blocked by C29 and C-176, which are specific inhibitors of TLR2 and STING, respectively. Taken together, we demonstrated that oxymatrine, a novel TLR2 agonist, plays a critical role in accelerating MK differentiation and thrombopoiesis via the STING/NF-κB axis, suggesting that oxymatrine is a promising candidate for RIT therapy.

This study aimed to characterize and identify the non-volatile components in aqueous and ethanolic extracts of the stems and leaves of Rhododendron tomentosum by using sensitive and efficient ultra-performance liquid chromatography-quadrupole-time of flight mass spectrometry(UHPLC-Q-TOF-MS) combined with a self-built information database. By comparing with reference compounds, analyzing fragment ion information, searching relevant literature, and using a self-built information database, 118 compounds were identified from the aqueous and ethanolic extracts of R. tomentosum, including 35 flavonoid glycosides, 15 phenolic glycosides, 12 flavonoids, 7 phenolic acids, 7 phenylethanol glycosides, 6 tannins, 6 phospholipids, 5 coumarins, 5 monoterpene glycosides, 6 triterpenes, 3 fatty acids, and 11 other types of compounds. Among them, 102 compounds were reported in R. tomentosum for the first time, and 36 compounds were identified by comparing them with reference compounds. The chemical components in the ethanolic and aqueous extracts of R. tomentosum leaves and stems showed slight differences, with 84 common chemical components accounting for 71.2% of the total 118 compounds. This study systematically characterized and identified the non-volatile chemical components in the ethanolic and aqueous extracts of R. tomentosum for the first time. The findings provide a reference for active ingredient research, quality control, and product development of R. tomentosum.
Rhododendron/chemistry* ; Chromatography, High Pressure Liquid/methods* ; Drugs, Chinese Herbal/chemistry* ; Mass Spectrometry/methods* ; Plant Leaves/chemistry*

BACKGROUND: Tissue engineering holds promise for vascular repair and regeneration by mimicking the extracellular matrix of blood vessels. However, achieving a functional and thick vascular wall with aligned fiber architecture by electrospinning remains a significant challenge. METHODS: A novel electrospinning setup was developed that utilizes an auxiliary electrode and a spring. The impact of process parameters on fiber size and morphology was investigated. The structure and functions of the scaffolds were evaluated through material characterization and assessments of cellular biocompatibility. RESULTS: The new setup enabled controlled deposition of fibers in different designed orientations. The fabricated small-diameter vascular scaffolds consisted of an inner layer of longitudinally oriented fibers and an outer layer of circumferentially oriented fibers (L + C vascular scaffold). Key parameters, including rotational speed, the utilization of the auxiliary electrode, and top-to-collector distance (TCD) significantly influenced fiber orientation. Additionally, voltage, TCD, feed rate, needle size, auxiliary electrode and collector-auxiliary electrode distance affected fiber diameter and distribution. Mechanical advantages and improved surface wettability of L + C vascular scaffold were confirmed through tensile testing and water contact angle. Cellular experiments indicated that L + C vascular scaffold facilitated cell adhesion and proliferation, with human umbilical vein endothelial cells and smooth muscle cells attaching and elongating along the fiber direction of the inner and outer layer, respectively. CONCLUSION This study demonstrated the feasibility of fabricating fiber-aligned, thick-walled vascular scaffolds using a modified electrospinning setup. The findings provided insights into how the auxiliary electrode, specific collector influenced fiber deposition, potentially advancing biomimetic vascular scaffold engineering.

Oxidative stress due to aberrant metabolism is considered as a crucial contributor to diabetes and its complications. Hyperglycemia and hyperlipemia boost excessive reactive oxygen species generation by elevated mitochondrial respiration, increased nicotinamide adenine dinucleotide phosphate oxidase activity, and enhanced pro-oxidative processes, including protein kinase C pathways, hexosamine, polyol, and advanced glycation endproducts, which exacerbate oxidative stress. Oxidative stress plays a significant role in the onset of diabetes and its associated complications by impairing insulin production, increasing insulin resistance, maintaining hyperglycemic memory, and inducing systemic inflammation. A more profound comprehension of the molecular processes that link oxidative stress to diabetes is crucial to new preventive and therapeutic strategies. Therefore, this review discusses the mechanisms underlying how oxidative stress contributes to diabetes mellitus and its complications. We also summarize the current approaches for prevention and treatment by targeting the oxidative stress pathways in diabetes.
Oxidative Stress/physiology* ; Humans ; Diabetes Mellitus/physiopathology* ; Diabetes Complications/metabolism* ; Reactive Oxygen Species/metabolism* ; Glycation End Products, Advanced/metabolism* ; Animals

This study explores the mechanism of Guben Jiannao Liquid on Alzheimer's disease(AD) by regulating autophagy based on the liver kinase B1(LKB1)/adenosine monophosphate-activated protein kinase(AMPK)/mammalian target of rapamycin(mTOR) pathway. Male SD rats were randomly divided into the blank group, model group, low-dose and high-dose groups of Guben Jiannao Liquid, and rapamycin group, with 10 rats in each group. Except for the blank group, all other groups of rats were injected bilaterally in the hippocampus with β-amyloid(Aβ)_(1-42) to establish the AD model. The low-dose(6.21 g·kg~(-1)) and high-dose(12.42 g·kg~(-1)) groups of Guben Jiannao Liquid and rapamycin group(1 mg·kg~(-1)) were given the corresponding drugs by gavage, and the blank and model groups were given an equal volume of saline by gavage for four weeks. Morris water maze was used to test the learning and memory ability of rats in each group; hematoxylin-eosin(HE) and Nissl staining were used to observe the morphological and quantitative changes of neurons and Nissl bodies in the CA1 region of rat hippocampus; immunohistochemistry was utilized to detect Aβ-positive cell expression in the CA1 region of rat hippocampus; transmission electron microscopy was employed to observe ultrastructural changes in rat hippocampal tissue, and Western blot was used to examine the protein expression levels of LKB1, p-AMPK/AMPK, p-mTOR/mTOR, Beclin1, p62, and LC3-Ⅱ in the hippocampal tissue of the rats. The results showed that compared with those in the blank group, rats in the model group had elevated evasion latency and decreased number of platform transversal and residence time in the platform quadrant. The number of neurons in the hippocampal area was reduced, and the morphology was impaired. The average integral optical density value of Aβ-positive cells was elevated; the expression levels of LKB1, p-AMPK/AMPK, Beclin1, and LC3-Ⅱ were decreased, and the expression levels of p-mTOR/mTOR and p62 were increased. Compared with those in the model group, rats in the low-dose and high-dose groups of Guben Jiannao Liquid had shorter evasion latency, higher number of platform transversal, longer residence time in the platform quadrant, increased number of neurons, decreased expression of Aβ-positive cells and average integral optical density values, and increased number of autophagic lysosomes in hippocampal tissue. The expression levels of LKB1, Beclin1, and LC3-Ⅱ were elevated in the hippocampus of rats in the low-dose group of Guben Jiannao Liquid. The expression levels of LKB1, p-AMPK/AMPK, Beclin1, and LC3-Ⅱ were elevated in the hippocampal tissue of rats in the high-dose group of Guben Jiannao Liquid, and the expression levels of p-mTOR/mTOR and p62 were decreased. The findings suggest that Guben Jiannao Liquid can improve cognitive impairment in AD rats, and its mechanism of action may be related to the activation of the LKB1/AMPK/mTOR signaling pathway and the up-regulation of autophagy level.
Animals ; Alzheimer Disease/physiopathology* ; Male ; TOR Serine-Threonine Kinases/genetics* ; Autophagy/drug effects* ; Rats, Sprague-Dawley ; Protein Serine-Threonine Kinases/genetics* ; AMP-Activated Protein Kinases/genetics* ; Rats ; Drugs, Chinese Herbal/administration & dosage* ; Signal Transduction/drug effects* ; AMP-Activated Protein Kinase Kinases ; Humans ; Hippocampus/metabolism*

In this study, RAW264.7 cells were employed to investigate the effects of honey-processed Astragalus on their energy metabolism and polarization, and explore the scientific connotation of the enhanced efficacy of honey-processed Astragalus on invigorating spleen-stomach and replenishing Qi. The medicated sera were prepared by intragastric administration of rats with water extracts of crude and honey-processed Astragalus, and the composition changes of medicated sera of crude and honey-processed Astragalus were analyzed by using LC-MS technology. The cell survival rates were detected and the concentrations of medicated sera were screened through CCK-8 assay. The differences of cell phagocytic rates, ATP energy metabolism, and NO secretion between crude and honey-processed Astragalus were evaluated by using neutral red phagocytosis assay, ATP detection kit, and NO detection kit. The effects of crude and honey-processed Astragalus on TNF-α secretion of RAW264.7 cells were detected by employing ELISA kit. The effects of crude and honey-processed Astragalus on polarization of RAW264.7 cells were evaluated by utilizing flow cytometry. The differential metabolites related to glycolysis in cell lysates and culture media were screened by using LC-MS technology. The experiment was approved by the experimental animal ethics committee from Shanxi University of Chinese Medicine (No. AWE202407352). The results showed that the contents of betaine, amino acids, and ononin in the prepared medicated sera of rats treated by intragastric administration with water extracts of honey-processed Astragalus increased compared to those in crude one. The results of CCK-8 experiment showed that there were no cytotoxic effects on RAW264.7 cells in the medicated sera of crude and honey-processd Astragalus at different concentration. The phagocytic index and ATP yield both increased to varying degrees after administration of the medicated sera to cells. The secretion of NO in normal and inflammatory cells increased and decreased respectively, and the effect of honey-processed Astragalus was better than that of crude one. The results of ELISA kit showed that the medicated sera of both crude and honey-processed Astragalus could promote the secretion of TNF-α in a concentration-dependent manner, and the promoting effect of honey-processed Astragalus was stronger than that of crude one. The results of flow cytometry showed that the medicated sera of both crude and honey-processed Astragalus could promote M1-type polarization and inhibit M2-type polarization of RAW264.7 cells, and the effect of honey-processed Astragalus was better than that of crude one. Compared to crude Astragalus, the metabolites related to glycolysis in the cell lysates and culture media of the medicated sera of honey-processed Astragalus were generally on the rise, indicating that the effect on promoting glycolysis of honey-processed Astragalus was better than that of crude one. In summary, honey-processed Astragalus can promote the polarization and energy metabolism of RAW264.7 cells, and participate in positive immune regulation, which is correlated with its enhanced effect of invigorating spleen-stomach and replenishing Qi.