Quercetin inhibits necroptosis in cardiomyocytes after ischemia-reperfusion via DNA-PKcs-SIRT5-orchestrated mitochondrial quality control.

We investigated the mechanism by which quercetin preserves mitochondrial quality control (MQC) in cardiomyocytes subjected to ischemia-reperfusion stress. An enzyme-linked immunosorbent assay was employed in the in vivo experiments to assess myocardial injury markers, measure the transcript levels of SIRT5/DNAPK-cs/MLKL during various time intervals of ischemia-reperfusion, and observe structural changes in cardiomyocytes using transmission electron microscopy. In in vitro investigations, adenovirus transfection was employed to establish a gene-modified model of DNA-PKcs, and primary cardiomyocytes were obtained from a mouse model with modified SIRT5 gene. Reverse transcription polymerase chain reaction, laser confocal microscopy, immunofluorescence localization, JC-1 fluorescence assay, Seahorse energy analysis, and various other assays were applied to corroborate the regulatory influence of quercetin on the MQC network in cardiomyocytes after ischemia-reperfusion. In vitro experiments demonstrated that ischemia-reperfusion injury caused changes in the structure of the myocardium. It was seen that quercetin had a beneficial effect on the myocardial tissue, providing protection. As the ischemia-reperfusion process continued, the levels of DNA-PKcs/SIRT5/MLKL transcripts were also found to change. In vitro investigations revealed that quercetin mitigated cardiomyocyte injury caused by mitochondrial oxidative stress through DNA-PKcs, and regulated mitophagy and mitochondrial kinetics to sustain optimal mitochondrial energy metabolism levels. Quercetin, through SIRT5 desuccinylation, modulated the stability of DNA-PKcs, and together they regulated the "mitophagy-unfolded protein response." This preserved the integrity of mitochondrial membrane and genome, mitochondrial dynamics, and mitochondrial energy metabolism. Quercetin may operate synergistically to oversee the regulation of mitophagy and the unfolded protein response through DNA-PKcs-SIRT5 interaction.

Methyl rosmarinate is an allosteric inhibitor of SARS-CoV-2 3 CL protease as a potential candidate against SARS-cov-2 infection.

The coronavirus disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been ongoing for more than three years and urgently needs to be addressed. Traditional Chinese medicine (TCM) prescriptions have played an important role in the clinical treatment of patients with COVID-19 in China. However, it is difficult to uncover the potential molecular mechanisms of the active ingredients in these TCM prescriptions. In this paper, we developed a new approach by integrating the experimental assay, virtual screening, and the experimental verification, exploring the rapid discovery of active ingredients from TCM prescriptions. To achieve this goal, 4 TCM prescriptions in clinical use for different indications were selected to find the antiviral active ingredients in TCMs. The 3-chymotrypsin-like protease (3CLpro), an important target for fighting COVID-19, was utilized to determine the inhibitory activity of the TCM prescriptions and single herb. It was found that 10 single herbs had better inhibitory activity than other herbs by using a fluorescence resonance energy transfer (FRET) - based enzymatic assay of SARS-CoV-2 3CLpro. The ingredients contained in 10 herbs were thus virtually screened and the predicted active ingredients were experimentally validated. Thus, such a research strategy firstly removed many single herbs with no inhibitory activity against SARS-CoV-2 3CLpro at the very beginning by FRET-based assay, making our subsequent virtual screening more effective. Finally, 4 active components were found to have stronger inhibitory effects on SARS-CoV-2 3CLpro, and their inhibitory mechanism was subsequently investigated. Among of them, methyl rosmarinate as an allosteric inhibitor of SARS-CoV-2 3CLpro was confirmed and its ability to inhibit viral replication was demonstrated by the SARS-CoV-2 replicon system. To validate the binding mode via docking, the mutation experiment, circular dichroism (CD), enzymatic inhibition and surface plasmon resonance (SPR) assay were performed, demonstrating that methyl rosmarinate bound to the allosteric site of SARS-CoV-2 3CLpro. In conclusion, this paper provides the new ideas for the rapid discovery of active ingredients in TCM prescriptions based on a specific target, and methyl rosmarinate has the potential to be developed as an antiviral therapeutic candidate against SARS-CoV-2 infection.

Screening and verification of hemostatic effective components group of Panax Notoginseng based on spectrum-effect relationships.

ETHNOPHARMACOLOGICAL RELEVANCE: Panax Notoginseng (PN) can disperse blood stasis, hemostasis, and detumescence analgesic, which can be used for hemoptysis, hematemesis and another traumatic bleeding, and it is known as "A miracle hemostatic medicine". Studies show that the chemical composition of PN is relatively comprehensive, however, its hemostatic active ingredients have not been fully clarified. AIM OF STUDY: This study aimed to clarify the hemostatic effective components group (HECG) of PN, provide a foundation for the assessment of PN's quality and its comprehensive development, and for further studies on the pharmacodynamic material basis of other Traditional Chinese Medicines (TCMs). MATERIALS AND METHODS: UPLC-MS was used to establish the fingerprint and identify the common peaks in 44 batches of PN extracts (PNE). In addition, the plasma recalcification time and in vitro coagulation time were measured. For spectrum-effect analysis, bivariate correlation analysis (BCA) and partial least squares regression analysis (PLSR) were used to screen the hemostasis candidate active monomers of PN. The monomers were prepared by combining several preparative chromatography techniques. The efficacy was verified by plasma recalcification time, in vitro coagulation time, and a rat model of gastric hemorrhage. RESULTS: A total of 30 common peaks and hemostatic efficacy indexes of 44 batches of PNE were obtained. A total of 18 components were positively correlated with the comprehensive coagulation index by two statistical methods. Six and eleven monomers were obtained respectively by chromatographic preparation and procurement, and one monomer was eliminated due to preparation difficulty and other reasons. Seven active monomers with direct hemostatic effect and one active monomer with synergistic hemostatic effect were screened through plasma recalcification time, and their combinations were used as candidate HECG for hemostatic effect verification. The results of in vitro experiments showed that plasma recalcification time and in vitro coagulation time were significantly reduced (P < 0.05) in the HECG group, compared to the PNE group. The results of in vivo experiment also indicated that the hemostatic effect of HECG was comparable to that of PNE and PN powder. CONCLUSION: The composition and efficacy of the HECG of PN were screened and verified using the spectral correlation method and in vivo and in vitro efficacy verification; the HECG included Dencichine, Ginsenoside Rg1, Ginsenoside Rd, Ginsenoside Rh1, Ginsenoside F1, Notoginsenoside R1, Notoginsenoside Ft1 and Notoginsenoside Fe. These results laid a foundation for the quality evaluation of PN and provided a reference for the basic research of pharmacodynamic material basis of other TCMs.

Nardostachys jatamansi and levodopa combination alleviates Parkinson's disease symptoms in rats through activation of Nrf2 and inhibition of NLRP3 signaling pathways.

CONTEXT: Levodopa combined with traditional Chinese medicine has a synergistic effect on Parkinson's disease (PD). Recently, we demonstrated that Nardostachys jatamansi (D. Don) DC. [syn. Patrinia jatamansi D.Don, N. grandiflora DC.] (Valerianaceae) (NJ) can alleviate PD. OBJECTIVE: To explore the synergistic effect of NJ combined with levodopa against PD. MATERIALS AND METHODS: The PD model was established by injecting rotenone. Eighty-four Sprague-Dawley rats were randomly divided into seven groups: sham, model, different doses of NJ (0.31, 0.62, or 1.24 g/kg) combined with levodopa (25 mg/kg), and levodopa alone (25 and 50 mg/kg) groups. The synergistic effect of the combination was investigated by pharmacodynamic investigation and detection of expression of nuclear factor erythro2-related factor 2 (Nrf2) and NLR family proteins containing Pyrin-related domain 3 (NLRP3) pathways. RESULTS: Compared with the model group, NJ + levodopa (1.24 g/kg + 25 mg/kg) increased the moving distance of PD rats in the open field (2395.34 ± 668.73 vs. 1501.41 ± 870.23, p < 0.01), enhanced the stay time on the rotating rod (84.86 ± 18.15 vs. 71.36 ± 17.53, p < 0.01) and the combination was superior to other treatments. The synergistic effects were related to NJ + levodopa (1.24 g/kg + 25 mg/kg) increasing the neurotransmitter levels by 38.80%-88.67% in PD rats, and inhibiting oxidative stress and NLRP3 pathway by activating Nrf2 pathway. DISCUSSION AND CONCLUSIONS: NJ combined with levodopa is a promising therapeutic candidate for PD, which provides a scientific basis for the subsequent clinical combination therapy of levodopa to enhance the anti-PD effect.

Medicinal plant-derived mtDNA via nanovesicles induces the cGAS-STING pathway to remold tumor-associated macrophages for tumor regression.

Plant-derived nanovesicles (PDNVs) have been proposed as a major mechanism for the inter-kingdom interaction and communication, but the effector components enclosed in the vesicles and the mechanisms involved are largely unknown. The plant Artemisia annua is known as an anti-malaria agent that also exhibits a wide range of biological activities including the immunoregulatory and anti-tumor properties with the mechanisms to be further addressed. Here, we isolated and purified the exosome-like particles from A. annua, which were characterized by nano-scaled and membrane-bound shape and hence termed artemisia-derived nanovesicles (ADNVs). Remarkably, the vesicles demonstrated to inhibit tumor growth and boost anti-tumor immunity in a mouse model of lung cancer, primarily through remolding the tumor microenvironment and reprogramming tumor-associated macrophages (TAMs). We identified plant-derived mitochondrial DNA (mtDNA), upon internalized into TAMs via the vesicles, as a major effector molecule to induce the cGAS-STING pathway driving the shift of pro-tumor macrophages to anti-tumor phenotype. Furthermore, our data showed that administration of ADNVs greatly improved the efficacy of PD-L1 inhibitor, a prototypic immune checkpoint inhibitor, in tumor-bearing mice. Together, the present study, for the first time, to our knowledge, unravels an inter-kingdom interaction wherein the medical plant-derived mtDNA, via the nanovesicles, induces the immunostimulatory signaling in mammalian immune cells for resetting anti-tumor immunity and promoting tumor eradication.

Mitochondrial disorder and treatment of ischemic cardiomyopathy: Potential and advantages of Chinese herbal medicine.

Mitochondrial dysfunction is the main cause of damage to the pathological mechanism of ischemic cardiomyopathy. In addition, mitochondrial dysfunction can also affect the homeostasis of cardiomyocytes or endothelial cell dysfunction, leading to a vicious cycle of mitochondrial oxidative stress. And mitochondrial dysfunction is also an important pathological basis for ischemic cardiomyopathy and reperfusion injury after myocardial infarction or end-stage coronary heart disease. Therefore, mitochondria can be used as therapeutic targets against myocardial ischemia injury, and the regulation of mitochondrial morphology, function and structure is a key and important way of targeting mitochondrial quality control therapeutic mechanisms. Mitochondrial quality control includes mechanisms such as mitophagy, mitochondrial dynamics (mitochondrial fusion/fission), mitochondrial biosynthesis, and mitochondrial unfolded protein responses. Among them, the increase of mitochondrial fragmentation caused by mitochondrial pathological fission is the initial factor. The protective mitochondrial fusion can strengthen the interaction and synthesis of paired mitochondria and promote mitochondrial biosynthesis. In ischemia or hypoxia, pathological mitochondrial fission can promote the formation of mitochondrial fragments, fragmented mitochondria can lead to damaged mitochondrial DNA production, which can lead to mitochondrial biosynthesis dysfunction, insufficient mitochondrial ATP production, and mitochondrial ROS. Burst growth or loss of mitochondrial membrane potential. This eventually leads to the accumulation of damaged mitochondria. Then, under the leadership of mitophagy, damaged mitochondria can complete the mitochondrial degradation process through mitophagy, and transport the morphologically and structurally damaged mitochondria to lysosomes for degradation. But once the pathological mitochondrial fission increases, the damaged mitochondria increases, which may activate the pathway of cardiomyocyte death. Although laboratory studies have found that a variety of mitochondrial-targeted drugs can reduce myocardial ischemia and protect cardiomyocytes, there are still few drugs that have successfully passed clinical trials. In this review, we describe the role of MQS in ischemia/hypoxia-induced cardiomyocyte physiopathology and elucidate the relevant mechanisms of mitochondrial dysfunction in ischemic cardiomyopathy. In addition, we also further explained the advantages of natural products in improving mitochondrial dysfunction and protecting myocardial cells from the perspective of pharmacological mechanism, and explained its related mechanisms. Potential targeted therapies that can be used to improve MQS under ischemia/hypoxia are discussed, aiming to accelerate the development of cardioprotective drugs targeting mitochondrial dysfunction.

ß-tubulin contributes to Tongyang Huoxue decoction-induced protection against hypoxia/reoxygenation-induced injury of sinoatrial node cells through SIRT1-mediated regulation of mitochondrial quality surveillance.

BACKGROUND: TYHX-Tongyang Huoxue decoction has been used clinically for nearly 40 years. The ingredients of TYHX are Radix Astragali (Huangqi), Red Ginseng (Hongshen), Rehmannia Glutinosa (Dihuang), Common Yam Rhizome (Shanyao) and Cassia-bark-tree Bark (Rougui). Our previous experiments confirmed that TYHX can protect sinoatrial node cells. However, its mechanism of action is not completely understood yet. PURPOSE: The present study aimed to determine the protective effects of TYHX against Sinus node cell injury under hypoxic stress and elucidate the underlying mechanisms of protection. METHODS: Through RNA sequencing analysis and network pharmacology analysis, we found significant differences in mitochondrial-related genes before and after hypoxia-mimicking SNC, resolved the main regulatory mechanism of TYHX. Through the intervention of TYHX on SNC, a series of detection methods such as laser confocal, fluorescence co-localization, mitochondrial membrane potential and RT-PCR. The regulatory effect of TYHX on ß-tubulin in sinoatrial node cells was verified by in vitro experiments. The mechanism of action of TYHX and its active ingredient quercetin to maintain mitochondrial homeostasis and protect sinoatrial node cells through mitophagy, mitochondrial fusion/fission and mitochondrial biosynthesis was confirmed. RESULTS: Through RNA sequencing analysis, we found that there were significant differences in mitochondrial related genes before and after SNC was modeled by hypoxia. Through pharmacological experiments, we showed that TYHX could inhibit the migration of Drp1 to mitochondria, inhibit excessive mitochondrial fission, activate mitophagy and increase the mitochondrial membrane potential. These protective effects were mainly mediated by ß-tubulin. Furthermore, the active component quercetin in TYHX could inhibit excessive mitochondrial fission through SIRT1, maintain mitochondrial energy metabolism and protect SNCs. Our results showed that protection of mitochondrial function through the maintenance of ß-tubulin and activation of SIRT1 is the main mechanism by which TYHX alleviates hypoxic stress injury in SNCs. The regulatory effects of TYHX and quercetin on mitochondrial quality surveillance are also necessary. Our findings provide empirical evidence supporting the use of TYHX as a targeted treatment for sick sinus syndrome. CONCLUSION: Our data indicate that TYHX exerts protective effects against sinus node cell injury under hypoxic stress, which may be associated with the regulation of mitochondrial quality surveillance (MQS) and inhibition of mitochondrial homeostasis-mediated apoptosis.

Ginsenoside Rb1 reduces oxidative/carbonyl stress damage and ameliorates inflammation in the lung of streptozotocin-induced diabetic rats.

CONTEXT: Ginsenoside Rb1 (Rb1) is a biologically active component of ginseng [Panax ginseng C.A. Meyer (Araliaceae)]. OBJECTIVE: This study determined the underlying mechanisms of Rb1 treatment that acted on diabetes-injured lungs in diabetic rats. MATERIALS AND METHODS: Streptozotocin (STZ)-induced diabetic rat model was used. Male Sprague-Dawley (SD) rats were divided into four groups (n = 10): control, Rb1 (20 mg/kg), insulin (15 U/kg to attain the euglycaemic state) and diabetic (untreated). After treatment for six weeks, oxidative stress assay; histological and ultrastructure analyses; TNF-α, TGF-ß, IL-1 and IL-6 protein expression analyses; and the detection of apoptosis were performed. RESULTS: There was decreased activity of SOD (3.53-fold), CAT (2.55-fold) and GSH (1.63-fold) and increased levels of NO (4.47-fold) and MDA (3.86-fold) in the diabetic group from control. Rb1 treatment increased SOD (2.4-fold), CAT (1.9-fold) and GSH (1.29-fold) and decreased the levels of NO (1.76-fold) and MDA (1.51-fold) as compared with diabetic rats. The expression of IL-6 (5.13-fold), IL-1α (2.35-fold), TNF-α (2.35-fold) and TGF-ß (2.39-fold) was increased in diabetic rats from control. IL-6 (2.43-fold), IL-1α (2.27-fold), TNF-α (1.68-fold) and TGF-ß (2.3-fold) were decreased in the Rb1 treatment group. Diabetes increased the apoptosis rate (2.23-fold vs. control), and Rb1 treatment decreased the apoptosis rate (1.73-fold vs. the diabetic rats). Rb1 and insulin ameliorated lung tissue injury. DISCUSSION AND CONCLUSIONS: These findings indicate that Rb1 could be useful for mitigating oxidative damage and inflammatory infiltration in the diabetic lung.

Protective Effect of Natural Medicinal Plants on Cardiomyocyte Injury in Heart Failure: Targeting the Dysregulation of Mitochondrial Homeostasis and Mitophagy.

Heart failure occurs because of various cardiovascular pathologies, such as coronary artery disease or cardiorenal syndrome, eventually reaching end-stage disease. Various factors contribute to cardiac structural or functional changes that result in systolic or diastolic dysfunction. Several studies have confirmed that the key factor in heart failure progression is myocardial cell death, and mitophagy is the major mechanism regulating myocardial cell death in heart failure. The clinical mechanisms of heart failure are well understood in practice. However, the essential role of mitophagic regulation in heart failure has only recently received widespread attention. Receptor-mediated mitophagy is involved in various mitochondrial processes like oxidative stress injury, energy metabolism disorders, and calcium homeostasis, which are also the main causes of heart failure. Understanding of the diverse regulatory mechanisms in mitophagy and the complexity of its pathophysiology in heart failure remains incomplete. Related studies have found that various natural medicinal plants and active ingredients, such as flavonoids and saponins, can regulate mitophagy to a certain extent, improve myocardial function, and protect myocardial cells. This review comprehensively covers the relevant mechanisms of different types of mitophagy in regulating heart failure pathology and controlling mitochondrial adaptability to stress injury. Further, it explores the relationship between mitophagy and cardiac ejection dysfunction. Natural medicinal plant-targeted regulation strategies and scientific evidence on mitophagy were provided to elucidate current and potential strategies to apply mitophagy-targeted therapy for heart failure.

Clinical Value of Cytokine Assay in Diagnosis and Severity Assessment of Lung Cancer.

Purpose: To investigate the clinical value of interleukin 2 (IL-2), interleukin 4 (IL-4), interleukin 6 (IL-6), interleukin 10 (IL-10), tumor necrosis factor α (TNF-α), and interferon-γ (IFN-γ) in diagnosis and severity assessment of lung cancer. Methods: In this observational study, 50 physical examination healthy subjects were included in the control group and 100 lung cancer patients were included in the study group. In the study group, 53 cases with pleural effusion were subgrouped to the pleural effusion group (n = 53), while 47 patients were assigned to the nonpleural effusion group (n = 47). Plasma cytokines IL-2, IL-4, IL-6, IL-10, TNF-α, IFN-γ, and Acute Physiology and Chronic Health Evaluation II (APACHE II) scores of all eligible subjects were collected and compared. Results: The study group showed significantly higher levels of plasma cytokines IL-2, IL-4, IL-6, IL-10, TNF-α, and IFN-γ versus healthy subjects (P < 0.05). Deterioration of lung cancer was associated with increased plasma cytokine levels and APACHE II scores. The combination assay of the above plasma cytokines showed significantly better diagnostic efficacy for lung cancer versus the single assay of the cytokines. Dead patients had higher plasma cytokine levels versus survived patients. The accuracy of plasma IL-2, IL-4, IL-6, IL-10, TNF-α, and IFN-γ levels in the severity assessment of lung cancer was comparable with that of the APACHE II scale. Conclusion: The plasma cytokines IL-2, IL-4, IL-6, IL-10, TNF-α, and IFN-γ are effective markers for the diagnosis of lung cancer. The combined assay contributes to the early diagnosis of lung cancer patients, and the persistent elevation of cytokines suggests an increased risk of death in lung cancer patients, so the detection of cytokine levels facilitates the severity assessment of lung cancer.

Pyridoxine-responsive KCNQ2 epileptic encephalopathy: Additional cases and literature review.

BACKGROUND: Typical patients with KCNQ2 (OMIM# 602235) epileptic encephalopathy present early neonatal-onset intractable seizures with a burst suppression EEG pattern and severe developmental delay or regression, and those patients always fail first-line treatment with sodium channel blockers. Vitamin B6, either pyridoxine or pyridoxal 50-phosphate, has been demonstrated to improve seizure control in intractable epilepsy. METHODS: Here, we collected and summarized the clinical data for four independent cases diagnosed with pyridoxine-responsive epileptic encephalopathy, and their exome sequencing data. Moreover, we reviewed all published cases and summarized the clinical features, genetic variants, and treatment of pyridoxine-responsive KCNQ2 epileptic encephalopathy. RESULTS: All four cases showed refractory seizures during the neonatal period or infancy, accompanied by global development delay. Four pathogenetic variants of KCNQ2 were uncovered and confirmed by Sanger sequencing: KCNQ2 [NM_172107.4: c.2312C > T (p.Thr771Ile), c.873G > C (p.Arg291Ser), c.652 T > A (p.Trp218Arg) and c.913-915del (p. Phe305del)]. Sodium channel blockers and other anti-seizure medications failed to control their seizures. The frequency of seizures gradually decreased after treatment with high-dose pyridoxine. In case 1, case 2, and case 4, clinical seizures relapsed when pyridoxine was withdrawn, and seizures were controlled again when pyridoxine treatment was resumed. CONCLUSION: Our study suggests that pyridoxine may be a promising adjunctive treatment option for patients with KCNQ2 epileptic encephalopathy.

[Effects of Nardostachys jatamansi on gut microbiota of rats with Parkinson's disease].

Under the guidance of the traditional Chinese medicine(TCM) theory of "Zangfu-organs of spleen and stomach" and the modern theory of "microbiota-gut-brain axis", this study explored the effects of Nardostachys jatamansi on the gut microbiota of rats with Parkinson's disease(PD). The 40 SD rats were randomly divided into the control group, PD model group, levodopa group, and Nardostachys jatamansi ethanol extract group. The PD model was established by subcutaneous injection of rotenone in the neck and back area. After 14 days of intragastric administration, the PD rats' behaviors were analyzed through open field test, inclined plane test, and pole test. After the behavioral tests, the striatum, colon, and colon contents of rats in each group were collected. Western blot was employed to detect the protein expression of tyrosine hydroxylase(TH) and α-synuclein(α-syn) in striatum and that of α-syn in colon. Enzyme linked immunosorbent assay(ELISA) was used to detect the levels of tumor necrosis factor-α(TNF-α), interleukin-1ß(IL-1ß), and nuclear factor-kappa B(NF-κB) in striatum and colon. High-throughput sequencing of 16 S rRNA gene was conducted to detect the differences in microbial diversity, abundance, differential phyla, and dominant bacteria of rats between groups. The results indicated that Nar. ethanol extract could relieve dyskinesia, reverse the increased levels of α-syn, TNF-α, IL-1ß, and NF-κB in striatum, and improve the protein expression of TH in striatum of PD rats. The α diversity analysis indicated a significant decrease in diversity and abundance of gut microbiota in the PD model. The results of linear discriminant analysis effect size(LEfSe) of dominant bacteria indicated that Nardostachys jatamansi ethanol extract increased the relative abundance of Clotridiaceae, Lachnospiraceae, and Anaerostipes, and reversed the increased relative abundance of Proteobacteria, Gammaproteobacteria, Enterobacteriaceae, and Escherichia-Shigella in PD model group to exhibit the neuroprotective effect. In summary, the results indicated that Nar. ethanol extract exert the therapeutic effect on PD rats. Specifically, the extract may regulate gut microbiota, decrease the levels of proinflammatory cytokines, and reduce the protein aggregation of α-syn in the colon and striatum to alleviate intestinal inflammation and neuroinflammation. This study provides a basis for combining the theory of "Zangfu-organs of spleen and stomach" with the theory of "microbiota-gut-brain axis" to treat PD.

Lycium barbarum extract promotes M2 polarization and reduces oligomeric amyloid-ß-induced inflammatory reactions in microglial cells.

Lycium barbarum (LB) is a traditional Chinese medicine that has been demonstrated to exhibit a wide variety of biological functions, such as antioxidation, neuroprotection, and immune modulation. One of the main mechanisms of Alzheimer's disease is that microglia activated by amyloid beta (Aß) transform from the resting state to an M1 state and release pro-inflammatory cytokines to the surrounding environment. In the present study, immortalized microglial cells were pretreated with L. barbarum extract for 1 hour and then treated with oligomeric Aß for 23 hours. The results showed that LB extract significantly increased the survival of oligomeric Aß-induced microglial cells, downregulated the expression of M1 pro-inflammatory markers (inducible nitric oxide synthase, tumor necrosis factor α, interleukin-6, and interleukin-1ß), and upregulated the expression of M2 anti-inflammatory markers (arginase-1, chitinase-like protein 3, and interleukin-4). LB extract also inhibited the oligomeric Aß-induced secretion of tumor necrosis factor α, interleukin-6, and interleukin-1ß in microglial cells. The results of in vitro cytological experiments suggest that, in microglial cells, LB extract can inhibit oligomeric Aß-induced M1 polarization and concomitant inflammatory reactions, and promote M2 polarization.

Hyperthermia Selectively Destabilizes Oncogenic Fusion Proteins.

The PML/RARα fusion protein is the oncogenic driver in acute promyelocytic leukemia (APL). Although most APL cases are cured by PML/RARα-targeting therapy, relapse and resistance can occur due to drug-resistant mutations. Here we report that thermal stress destabilizes the PML/RARα protein, including clinically identified drug-resistant mutants. AML1/ETO and TEL/AML1 oncofusions show similar heat shock susceptibility. Mechanistically, mild hyperthermia stimulates aggregation of PML/RARα in complex with nuclear receptor corepressors leading to ubiquitin-mediated degradation via the SIAH2 E3 ligase. Hyperthermia and arsenic therapy destabilize PML/RARα via distinct mechanisms and are synergistic in primary patient samples and in vivo, including three refractory APL cases. Collectively, our results suggest that by taking advantage of a biophysical vulnerability of PML/RARα, thermal therapy may improve prognosis in drug-resistant or otherwise refractory APL. These findings serve as a paradigm for therapeutic targeting of fusion oncoprotein-associated cancers by hyperthermia. SIGNIFICANCE: Hyperthermia destabilizes oncofusion proteins including PML/RARα and acts synergistically with standard arsenic therapy in relapsed and refractory APL. The results open up the possibility that heat shock sensitivity may be an easily targetable vulnerability of oncofusion-driven cancers.See related commentary by Wu et al., p. 300.

[Spectrum-effect relationship of hemostatic effects of Notoginseng Radix et Rhizoma with different commodity specifications].

This article aims to establish the fingerprints, determine the hemostatic pharmacodynamic indicators, and explore the spectrum-effect relationship of Notoginseng Radix et Rhizoma in 12 different specifications. Firstly, HPLC and liquid chromatography-mass spectrometry(LC-MS) were employed to establish the fingerprints of Notoginseng Radix et Rhizoma. The rat plasma recalcification experiment and the rat gastric bleeding experiment were conducted to determine the pharmacodynamic indicators, including plasma recalcification time(PRT), thrombin time(TT), prothrombin time(PT), and activated partial thromboplastin time(APTT). Afterwards, the partial least squares method was employed to explore the spectrum-effect relationship of Notoginseng Radix et Rhizoma in different specifications. Twenty-six common peaks were detected in the HPLC fingerprints of different specifications of Notoginseng Radix et Rhizoma, and 11 out of the 26 common peaks represented saponins. The content of dencichine was determined by LC-MS. The rat experiments showed that the pharmacodynamic indicators were significantly different among different specifications of Notoginseng Radix et Rhizoma. The spectrum-effect relationship was explored between 27 common components and pharmacodynamic indicators. Among them, 16 components had positive effects on the pharmacodynamic indicators of Notoginseng Radix et Rhizoma, and 11 exerted negative effects. This study provides a basis for the precision medication and quality control of Notoginseng Radix et Rhizoma.

Tongyang Huoxue Decoction (TYHX) Ameliorating Hypoxia/Reoxygenation-Induced Disequilibrium of Calcium Homeostasis and Redox Imbalance via Regulating Mitochondrial Quality Control in Sinoatrial Node Cells.

Sick sinus syndrome (SSS) is a disease with bradycardia or arrhythmia. The pathological mechanism of SSS is mainly due to the abnormal conduction function of the sinoatrial node (SAN) caused by interstitial lesions or fibrosis of the SAN or surrounding tissues, SAN pacing dysfunction, and SAN impulse conduction accompanied by SAN fibrosis. Tongyang Huoxue Decoction (TYHX) is widely used in SSS treatment and amelioration of SAN fibrosis. It has a variety of active ingredients to regulate the redox balance and mitochondrial quality control. This study mainly discusses the mechanism of TYHX in ameliorating calcium homeostasis disorder and redox imbalance of sinoatrial node cells (SANCs) and clarifies the protective mechanism of TYHX on the activity of SANCs. The activity of SANCs was determined by CCK-8 and the TUNEL method. The levels of apoptosis, ROS, and calcium release were analyzed by flow cytometry and immunofluorescence. The mRNA and protein levels of calcium channel regulatory molecules and mitochondrial quality control-related molecules were detected by real-time quantitative PCR and Western Blot. The level of calcium release was detected by laser confocal. It was found that after H/R treatment, the viability of SANCs decreased significantly, the levels of apoptosis and ROS increased, and the cells showed calcium overload, redox imbalance, and mitochondrial dysfunction. After treatment with TYHX, the cell survival level was improved, calcium overload and oxidative stress were inhibited, and mitochondrial energy metabolism and mitochondrial function were restored. However, after the SANCs were treated with siRNA (si-ß-tubulin), the regulation of TYHX on calcium homeostasis and redox balance was counteracted. These results suggest that ß-tubulin interacts with the regulation of mitochondrial function and calcium release. TYHX may regulate mitochondrial quality control, maintain calcium homeostasis and redox balance, and protect SANCs through ß-tubulin. The regulation mechanism of TYHX on mitochondrial quality control may also become a new target for SSS treatment.

Yiqi Jiemin decoction alleviates allergic rhinitis in a guinea pig model by suppressing inflammation, restoring Th1/Th2 balance, and improving cellular metabolism.

We investigated the mechanisms underlying the therapeutic effects of Yiqi Jiemin decoction (YJD), a traditional Chinese medicine (TCM), in the ovalbumin (OVA)-induced allergic rhinitis (AR) model in guinea pigs. YJD significantly decreased infiltration of mast cells and eosinophils into the nasal mucosa of AR model guinea pigs. YJD also increased expression of TGF-ß in the nasal mucosa, restored the balance of Th1/Th2 immune cell responses, and decreased serum levels of various pro-inflammatory mediators, including histamine (HA), neuropeptide Y (NPY), acetylcholine (ACH), norepinephrine and immunoglobulin E (IgE). Metabolic analyses using liquid chromatography coupled with high-resolution mass spectrometry revealed that YJD improved cellular metabolism in AR model guinea pigs and increased serum levels of glycocholic acid while decreasing levels 1-palmitoyl lysophosphatidic acid. RNA-sequencing analysis identified BPIFB2 as a potential diagnostic biomarker and therapeutic target for AR. Functional enrichment analyses showed that YJD significantly inhibited cytokine secretion pathways in AR model guinea pigs. These findings demonstrate that YJD protects against OVA-induced AR in guinea pigs by suppressing inflammation in the nasal mucosa, restoring Th1/Th2 balance, and improving cellular metabolism.

Expert consensus of Chinese Association for the Study of Pain on the application of ozone therapy in pain medicine.

This consensus was compiled by first-line clinical experts in the field of pain medicine and was organized by the Chinese Association for the Study of Pain. To reach this consensus, we consulted a wide range of opinions and conducted in-depth discussions on the mechanism, indications, contraindications, operational specifications and adverse reactions of ozone iatrotechnique in the treatment of pain disorders. We also referred to related previous preclinical and clinical studies published in recent years worldwide. The purpose of this consensus is to standardize the rational application of ozone iatrotechnique in pain treatment, to improve its efficacy and safety and to reduce and prevent adverse reactions and complications in this process.

Network pharmacology-based and clinically relevant prediction of active ingredients and potential targets of Chinese herbs on stage IV lung adenocarcinoma patients.

AIM: This study is designed to ascertain the relative molecular targets of effective Chinese herbs in treating stage IV lung adenocarcinoma based on clinical data and network pharmacology. In addition, we showed that Chinese Herbal Medicine (CHM) treatment was associated with survival benefit for patients with stage IV lung adenocarcinoma and identified 18 herbs beneficial to survival through correlation analysis. BACKGROUND: Increasing evidence has shown that CHM has efficient therapeutic effects for advanced lung adenocarcinoma, while active ingredients and potential targets remain unclear. METHODS: Kaplan-Meier method and Cox regression analysis were used to evaluate the survival benefit of CHM treatment, and correlation analysis was applied to identify the most effective components in the formulas. A network pharmacological approach was used to decipher the potential therapeutic mechanisms of CHM. RESULTS: CHM treatment was an independent protective factor. The hazard ratio (HR) was 0.487 (95% CI 0.293-0.807; P = 0.005). Patients in the CHM group had a longer median survival time (31 months) compared with the non-CHM group (19 months; P < 0.001). 18 out of the total 241 herbs were significantly correlated with favorable survival outcomes (P < 0.05), likely representing the most effective components in these formulas. Bioinformatics analysis suggested that the 18 herbs realize anti-lung-adenocarcinoma activity mainly through (1) inhibiting the activity of some growth factors' receptors, such as HGFR, EGFR, and IGFR. (2) Suppressing angiogenesis not only through VEGFR and PDGFR, but also through the function of neurotransmitters such as dopamine and serotonin. (3) Inhibiting the Ras signaling pathway directly through Ras as well as through ALK and FNTA/FNTB. CONCLUSIONS: We performed a network pharmacological method to decipher the underlying mechanisms, which provides a good foundation for herbal research based on clinical data.

Preservation of Retinal Function Through Synaptic Stabilization in Alzheimer's Disease Model Mouse Retina by Lycium Barbarum Extracts.

In Alzheimer's disease (AD), amyloid ß deposition-induced hippocampal synaptic dysfunction generally begins prior to neuronal degeneration and memory impairment. Lycium barbarum extracts (LBE) have been demonstrated to be neuroprotective in various animal models of neurodegeneration. In this study, we aimed to investigate the effects of LBE on the synapse loss in AD through the avenue of the retina in a triple transgenic mouse model of AD (3xTg-AD). We fed 3xTg-AD mice with low (200 mg/kg) or high (2 g/kg) dose hydrophilic LBE daily for 2 months from the starting age of 4- or 6-month-old. For those started at 6 month age, at 1 month (though not 2 months) after starting treatment, mice given high dose LBE showed a significant increase of a wave and b wave in scotopic ERG. After 2 months of treatment with high dose LBE, calpain-2, calpain-5, and the oxidative RNA marker 8-OHG were downregulated, and presynaptic densities in the inner plexiform layer but not the outer plexiform layer of the retina were significantly increased, suggesting the presynaptic structure of retina was preserved. Our results indicate that LBE feeding may preserve synapse stability in the retina of 3xTg-AD mice, probably by decreasing both oxidative stress and intracellular calcium influx. Thus, LBE might have potential as a neuroprotectant for AD through synapse preservation.