Integrating Epigenetics, Proteomics, and Metabolomics to Reveal the Involvement of Wnt/ß-Catenin Signaling Pathway in Oridonin-Induced Reproductive Toxicity.

Oridonin is the primary active component in the traditional Chinese medicine Rabdosia rubescens, displaying anti-inflammatory, anti-tumor, and antibacterial effects. It is widely employed in clinical therapy for acute and chronic pharyngitis, tonsillitis, as well as bronchitis. Nevertheless, the clinical application of oridonin is significantly restricted due to its reproductive toxicity, with the exact mechanism remaining unclear. The aim of this study was to investigate the mechanism of oridonin-induced damage to HTR-8/SVneo cells. Through the integration of epigenetics, proteomics, and metabolomics methodologies, the mechanisms of oridonin-induced reproductive toxicity were discovered and confirmed through fluorescence imaging, RT-qPCR, and Western blotting. Experimental findings indicated that oridonin altered m6A levels, gene and protein expression levels, along with metabolite levels within the cells. Additionally, oridonin triggered oxidative stress and mitochondrial damage, leading to a notable decrease in WNT6, ß-catenin, CLDN1, CCND1, and ZO-1 protein levels. This implied that the inhibition of the Wnt/ß-catenin signaling pathway and disruption of tight junction might be attributed to the cytotoxicity induced by oridonin and mitochondrial dysfunction, ultimately resulting in damage to HTR-8/SVneo cells.

Euphorbia factor L1 inhibited transport channel and energy metabolism in human colon adenocarcinoma cell line Caco-2.

Euphorbia factor L1 (EFL1) is a kind of lathyrane-type diterpenoid and is isolated from the medical herb Euphorbia lathyris L. (Euphorbiaceae); it has been reported with the toxicity that causes intestinal irritation, but the underlying mechanisms are still obscure. The objective of this study was to assess the EFL1-induced intestinal cytotoxicity in human colon adenocarcinoma Caco-2 cells. The Caco-2 cells were treated with EFL1, and the intracellular calcium ion concentration, mitochondrial membrane potential (MMP), mitochondrial permeability transition pore (mPTP), adenosine 5'-triphosphate (ATP) content, ATPase activities, TGF-ß1 concentration, and transepithelial electrical resistance (TEER) were detected. The interaction between EFL1 and the tight junction proteins Occludin, Claudin-4, Tricellulin, ZO-1, JAM-1, and E-cadherin was simulated by molecular docking. The expression of proteins involved in the energy metabolism, the ion transporters and aquaporins, the tight junction, and the F-actin cytoskeleton were detected by Western blotting and cell immunofluorescence. As a result, EFL1 decreased the intracellular Ca2+, MMP, mPTP, ATP content, and ATPase activities in the Caco-2 cells. The AMPK/SIRT1/PGC-1α signaling pathway, which regulates the energy metabolism, was inhibited. The ion transporters NEH and CFTR, as well as the aquaporins in the Caco-2 cells, were decreased. The tight junction proteins were down-regulated, and the integrity of the intestinal barrier was injured; TGF-ß1 was compensatively increased; so, the intestinal permeability was increased and was characterized by decreased TEER. The morphology of the F-actin cytoskeleton was destroyed. These findings indicated that EFL1 caused cytotoxicity in the human intestinal Caco-2 cells through mitochondrial damage, inhibition of the energy metabolism, and suppression of the ion and water molecule transporters, as well as the down-regulation tight junction and cytoskeleton protiens.

The Application of a Physiologically Based Toxicokinetic Model in Health Risk Assessment.

Toxicokinetics plays a crucial role in the health risk assessments of xenobiotics. Classical compartmental models are limited in their ability to determine chemical concentrations in specific organs or tissues, particularly target organs or tissues, and their limited interspecific and exposure route extrapolation hinders satisfactory health risk assessment. In contrast, physiologically based toxicokinetic (PBTK) models quantitatively describe the absorption, distribution, metabolism, and excretion of chemicals across various exposure routes and doses in organisms, establishing correlations with toxic effects. Consequently, PBTK models serve as potent tools for extrapolation and provide a theoretical foundation for health risk assessment and management. This review outlines the construction and application of PBTK models in health risk assessment while analyzing their limitations and future perspectives.

Cyclodextrin-Modified Amphiphilic Microgel with Bifunctional Domains for Infected Wound Healing via Photothermal Antibacterial Therapy and Nitric Oxide Release.

Bacterial infections are one of the major contributing factors to human mortality, which can cause secondary damage to the injured area, such as leading to inflammation, tissue death, and even personal death. Herein, we developed a novel cyclodextrin (CD)-modified amphiphilic microgel with a 3D network nanostructure that encapsulates hydrophilic indocyanine green (ICG) as a trigger for photothermal therapy (PTT) and hydrophobic N,N'-disubstituted-butyl-N,N'-dinitro-1,4-benzenediamine (BNN6) as a heat-sensitive nitric oxide (NO) donor (CD@I-B) to cope with bacteria-infected wound therapy. This biocompatible microgel showed excellent broad-spectrum antibacterial capability under near-infrared (NIR) laser irradiation, while the photothermal conversion process promotes the deswelling of the microgel and release of NO, which synergistically accelerates wound healing. The therapy strategy by synergizing NO delivery with PTT promoted the formation of neovascularization and collagen fiber as well as the elimination of inflammation cells, thus facilitating wound healing. Our study further demonstrates the fantastic opportunities of applying high-performance microgels to provide all-in-one sites for treating wound sterilization and healing.

Nuciferine Effectively Protects Mice against Acetaminophen-Induced Liver Injury.

Acetaminophen (APAP) overdose still poses a major clinical challenge and is a leading cause of acute liver injury (ALI). N-acetylcysteine (NAC) is the only approved antidote to treat APAP toxicity while NAC therapy can trigger side effects including severe vomiting and even shock. Thus, new insights in developing novel therapeutic drugs may pave the way for better treatment of APAP poisoning. Previous research has reported that nuciferine (Nuci) possesses anti-inflammatory and antioxidant properties. Therefore, the objective of this study was proposed to investigate the hepatoprotective effects of Nuci and explore its underlying mechanisms. Mice were intraperitoneally (i.p.) administered with APAP (300 mg/kg) and subsequently injected with Nuci (25, 50, and 100 mg/kg, i.p.) at 30 min after APAP overdose. Then, all mice were sacrificed at 12 h after APAP challenge for further analysis. Nuci-treated mice did not show any side effects and our results revealed that treating Nuci significantly attenuated APAP-induced ALI, as confirmed by histopathological examinations, biochemical analysis, and diminished hepatic oxidative stress and inflammation. The in silico prediction and mRNA-sequencing analysis were performed to explore the underlying mechanisms of Nuci. GO and KEGG enrichment of the predicted target proteins of Nuci includes reactive oxygen species, drug metabolism of cytochrome P450 (CYP450) enzymes, and autophagy. Furthermore, the mRNA-sequencing analyses indicated that Nuci can regulate glutathione metabolic processes and anti-inflammatory responses. Consistently, we found that Nuci increased the hepatic glutathione restoration but decreased APAP protein adducts in damaged livers. Western blot analysis further confirmed that Nuci effectively promoted hepatic autophagy in APAP-treated mice. However, Nuci could not affect the expression levels of the main CYP450 enzymes (CYP1A2, CYP2E1, and CYP3A11). These results demonstrated that Nuci may be a potential therapeutic drug for APAP-induced ALI via amelioration of the inflammatory response and oxidative stress, regulation of APAP metabolism, and activation of autophagy.

Genomics of clonal evolution in a rare essential thrombocythemia with coexisting Type 2 CALR and MPL S204P mutations.

Essential thrombocythemia (ET) with double driver mutations is a rare disease. ET patients with both MPL and Type 1 CALR mutations have been reported. Here, we report the first case of an ET patient with both MPL S204P and Type 2 CALR mutations and a summary of our literature review findings. In the patient whose case is reported here, the disease progressed to an accelerated phase 3.5 months after diagnosis. CALR mutation disappeared and new mutations emerged as the disease progressed, such as ASXL1, CBL, ETV6, and PTPN11 mutations. This case highlights that screening for additional mutations using NGS should be considered in patients with ET to assess the prognosis, especially as the disease progresses.

Integrated Analysis of Transcriptome and microRNA Profile Reveals the Toxicity of Euphorbia Factors toward Human Colon Adenocarcinoma Cell Line Caco-2.

Euphorbia factors, lathyrane-type diterpenoids isolated from the medical herb Euphorbia lathyris L. (Euphorbiaceae), have been associated with intestinal irritation toxicity, but the mechanisms underlying this phenomenon are still unknown. The objective of this study was to evaluate the transcriptome and miRNA profiles of human colon adenocarcinoma Caco-2 cells in response to Euphorbia factors L1 (EFL1) and EFL2. Whole transcriptomes of mRNA and microRNA (miRNA) were obtained using second generation high-throughput sequencing technology in response to 200 µM EFL treatment for 72 h, and the differentially expressed genes and metabolism pathway were enriched. Gene structure changes were analyzed by comparing them with reference genome sequences. After 72 h of treatment, 16 miRNAs and 154 mRNAs were differently expressed between the EFL1 group and the control group, and 47 miRNAs and 1101 mRNAs were differentially expressed between the EFL2 group and the control. Using clusters of orthologous protein enrichment, the sequenced mRNAs were shown to be mainly involved in transcription, post-translational modification, protein turnover, chaperones, signal transduction mechanisms, intracellular trafficking, secretion, vesicular transport, and the cytoskeleton. The differentially expressed mRNA functions and pathways were enriched in transmembrane transport, T cell extravasation, the IL-17 signaling pathway, apoptosis, and the cell cycle. The differentially expressed miRNA EFLs caused changes in the structure of the gene, including alternative splicing, insertion and deletion, and single nucleotide polymorphisms. This study reveals the underlying mechanism responsible for the toxicity of EFLs in intestinal cells based on transcriptome and miRNA profiles of gene expression and structure.

Effect of Humantenine on mRNA m6A Modification and Expression in Human Colon Cancer Cell Line HCT116.

Humantenine, an alkaloid isolated from the medicinal herb Gelsemium elegans (Gardner & Chapm.) Benth., has been reported to induce intestinal irritation, but the underlying toxicological mechanisms remain unclear. The object of the present study was to investigate the RNA N6-methyladenosine (m6A) modification and distinct mRNA transcriptome profiles in humantenine-treated HCT116 human colon cancer cells. High-throughput MeRIP-seq and mRNA-seq were performed, and bioinformatic analysis was performed to reveal the role of abnormal RNA m6A modification and mRNA expression in humantenine-induced intestinal cell toxicity. After humantenine treatment of HCT116 cells, 1401 genes were in the overlap of differentially m6A-modified mRNA and differentially expressed mRNA. The Kyoto Encyclopedia of Genes and Genomes and Gene Ontology annotation terms for actin cytoskeleton, tight junctions, and adherens junctions were enriched. A total of 11 kinds of RNA m6A methylation regulators were differentially expressed. The m6A methylation levels of target genes were disordered in the humantenine group. In conclusion, this study suggested that the HCT116 cell injury induced by humantenine was associated with the abnormal mRNA expression of m6A regulators, as well as disordered m6A methylation levels of target genes.

Oxidation and Antioxidation of Natural Products in the Model Organism Caenorhabditis elegans.

Natural products are small molecules naturally produced by multiple sources such as plants, animals, fungi, bacteria and archaea. They exert both beneficial and detrimental effects by modulating biological targets and pathways involved in oxidative stress and antioxidant response. Natural products' oxidative or antioxidative properties are usually investigated in preclinical experimental models, including virtual computing simulations, cell and tissue cultures, rodent and nonhuman primate animal models, and human studies. Due to the renewal of the concept of experimental animals, especially the popularization of alternative 3R methods for reduction, replacement and refinement, many assessment experiments have been carried out in new alternative models. The model organism Caenorhabditis elegans has been used for medical research since Sydney Brenner revealed its genetics in 1974 and has been introduced into pharmacology and toxicology in the past two decades. The data from C. elegans have been satisfactorily correlated with traditional experimental models. In this review, we summarize the advantages of C. elegans in assessing oxidative and antioxidative properties of natural products and introduce methods to construct an oxidative damage model in C. elegans. The biomarkers and signaling pathways involved in the oxidative stress of C. elegans are summarized, as well as the oxidation and antioxidation in target organs of the muscle, nervous, digestive and reproductive systems. This review provides an overview of the oxidative and antioxidative properties of natural products based on the model organism C. elegans.

Integrated Profiles of Transcriptome and mRNA m6A Modification Reveal the Intestinal Cytotoxicity of Aflatoxin B1 on HCT116 Cells.

Aflatoxin B1 (AFB1) is widely prevalent in foods and animal feeds and is one of the most toxic and carcinogenic aflatoxin subtypes. Existing studies have proved that the intestine is targeted by AFB1, and adverse organic effects have been observed. This study aimed to investigate the relationship between AFB1-induced intestinal toxicity and N6-methyladenosine (m6A) RNA methylation, which involves the post-transcriptional regulation of mRNA expression. The transcriptome-wide m6A methylome and transcriptome profiles in human intestinal cells treated with AFB1 are presented. Methylated RNA immunoprecipitation sequencing and mRNA sequencing were carried out to determine the distinctions in m6A methylation and different genes expressed in AFB1-induced intestinal toxicity. The results showed that there were 2289 overlapping genes of the differentially expressed mRNAs and differentially m6A-methylation-modified mRNAs. After enrichment of the signaling pathways and biological processes, these genes participated in the terms of the cell cycle, endoplasmic reticulum, tight junction, and mitophagy. In conclusion, the study demonstrated that AFB1-induced HCT116 injury was related to the disruptions to the levels of m6A methylation modifications of target genes and the abnormal expression of m6A regulators.

DKK1 suppresses WWP2 to enhance bortezomib resistance in multiple myeloma via regulating GLI2 ubiquitination.

Bortezomib-based chemotherapy represents the most prevalent regimens for multiple myeloma (MM), whereas acquired drug resistance remains a major obstacle. Myeloma cells often produce excessive amount of dickkopf-1 (DKK1), giving rise to myeloma bone disease. However, it remains obscure about the effects and mechanisms of DKK1 in the progression and bortezomib responsiveness of MM cells. In the current study, we found WWP2, an E3 ubiquitin-protein ligase, was downregulated in the bortezomib-resistant cells along with high expression of DKK1. Further investigation revealed that WWP2 was a direct target of Wnt/ß-catenin signaling pathway, and DKK1 suppressed the expression of WWP2 via canonical Wnt signaling. We further identified that WWP2 mediated the ubiquitination and degradation of GLI2, a main transcriptional factor of the Hedgehog (Hh) pathway. Therefore, DKK1-induced WWP2 downregulation improved GLI2 stability and activation of Hh signaling pathway, contributing to the resistance to bortezomib of MM cells. Clinical data also validated that WWP2 expression was associated with the treatment response and clinic outcomes of MM patients. WWP2 overexpression restricted MM progression and enhanced cell sensitivity to bortezomib treatment in vitro and in vivo. Taken together, our findings demonstrate that DKK1 facilitates the generation of bortezomib resistance in MM via downregulating WWP2 and activating Hh pathway. Thus, the manipulation of DKK1-WWP2-GLI2 axis might sensitize myeloma cells to proteasome inhibitors.

A fixed nitrous oxide/oxygen mixture as an analgesic for patients with postherpetic neuralgia: study protocol for a randomized controlled trial.

BACKGROUND: The pain management of postherpetic neuralgia (PHN) remains a major challenge, with no immediate relief. Nitrous oxide/oxygen mixture has the advantages of quick analgesic effect and well-tolerated. The purpose of this study is to investigate the analgesic effect and safety of nitrous oxide/oxygen mixture in patients with PHN. METHODS/DESIGN: This study is a single-center, two-group (1:1), randomized, placebo-controlled, double-blind clinical trial. A total of 42 patients with postherpetic neuralgia will be recruited and randomly divided into the intervention group and the control group. The control group will receive routine treatment plus oxygen, and the intervention group will receive routine treatment plus nitrous oxide/oxygen mixture. Data collectors, patients, and clinicians are all blind to the therapy. The outcomes of each group will be monitored at baseline (T0), 5 min (T1), and 15 min (T2) after the start of the therapy and at 5 min after the end of the therapy (T3). The primary outcome measure will be the pain intensity. Secondary outcomes included physiological parameters, adverse effects, patients' acceptance of analgesia, and satisfaction from patients. DISCUSSION: Previous studies have shown that nitrous oxide/oxygen mixture can effectively relieve cancer patients with breakthrough pain. This study will explore the analgesic effect of oxide/oxygen mixture on PHN. If beneficial to patients with PHN, it will contribute to the pain management of PHN. TRIAL REGISTRATION: Chinese Clinical Trial Register ChiCTR1900023730 . Registered on 9 June 2019.

Structure elucidation and NMR assignments of a new sesquiterpene of volatile oil from Artemisia frigida Willd.

A new sesquiterpene, artefrigin (5), together with four known sesquiterpenes were isolated from the volatile oil of Artemisia frigida Willd. The structure of five was elucidated by spectroscopic methods, including UV, IR, HR-ESI-MS and extensive 1D and 2D NMR techniques.

Two new compounds from supercritical fluid extract of Artemisia integrifolia L.

Two new compounds, namely integrin A (1) and integrin B (2), were isolated from the supercritical fluid extract (SFE) of Artemisia integrifolia L. Their structures were elucidated by spectroscopic methods, including UV, IR, HR-ESI-MS and extensive 1D and 2D NMR techniques.

Synthesis and antitumor activity of sacroflavonoside.

Sacroflavonoside, a new derivative of diphenylethene, was isolated from Artemisia sacrorum, which have been found to possess the inhibitory effect on the proliferation of gastric carcinoma cells (MKN-45) in vitro in our previous studies. With anisaldehyde (SM-A) as starting material, the sacroflavonoside was synthesized by nucleophilic addition, electrophilic substitution and dehydration cyclization. The structure of sacroflavonoside was established by 1 D (1H NMR and 13C NMR) and 2 D-NMR (HSQC and HMBC) spectral analysis. The antitumor activity and potential mechanism against MKN-45 cells of sacroflavonoside were evaluated in vitro. The results showed that sacroflavonoside could significantly induce MKN-45 cells apoptosis and autophagy by increasing the expression of Bax, Caspase-3, Beclin1 and LC3-II proteins and decreasing the expression of Bcl-2 protein at low micromole level. This investigation provided a valuable lead structure for the development of antitumor drugs.

Structure elucidation and NMR assignments of a new alkaloid from Panzerina lanata (L.) Soják.

A new alkaloid, named as panalasin (1), was isolated from the ethyl acetate extract from Panzerina lanata (L.) Soják. The structure of 1 was elucidated by spectroscopic methods, including UV, IR, HR-ESI-MS and extensive 1D and 2D NMR techniques.

Pre-mixed nitrous oxide/oxygen mixture treatment of pain induced by postoperative dressing change for perianal abscess: Study protocol for a randomized, controlled trial.

AIM: This study aims to evaluate the safety and analgesic efficacy of pre-mixed nitrous oxide/oxygen mixture treatment of pain induced by dressing change for perianal abscess. DESIGN: This protocol is a randomized, double-blind, placebo-controlled trial. METHODS: This study will be implemented in the Hospital of Traditional Chinese Medicine. Subjects enrolled in this study are hospitalized patients who suffered from moderate to severe pain due to dressing change after incision and drainage. Two hundred patients will be selected and randomly assigned to either an intervention or a control group. The intervention group will get routine pain treatment plus pre-mixed nitrous oxide/oxygen mixture treatment and the control group will be treated with routine pain management plus medical air treatment. All these patients, medical staff and investigators are blind to the nature of the gas in each cylinder, which is randomized. Data will be collected at baseline (T0), 5 min (T1) after the starting of intervention and 5 min post intervention (T2) for each group. The primary outcome is the level of pain relief at T1 and T2. The secondary outcomes cover physiological parameters, adverse events, satisfaction of patients and health professionals and the acceptance from patients. DISCUSSION: Results of this study will be discussed and the safety and effect of nitrous oxide/oxygen treatment of pain induced by dressing change will be proven. IMPACT: When the finding of this study has an active effect on the treatment of pain caused by dressing change, it may provide more options for nursing staff to choose nurse-led analgesia techniques and then improving the level and quality of pain care as well as patients' overall satisfaction with the Anorectal Department in China.

Structure elucidation and NMR assignments of a new dihydrochalcone from Empetrum nigrum subsp. asiaticum (Nakai ex H.Ito) Kuvaev.

A new dihydrochalcone, namely 2',5'-dimethyl-3'-methoxy-4',6'-dihydroxyl-dihydrochalcone (1) together with five known compounds were isolated from the CHCl3 extract from Empetrum nigrum L. var. japonicum K. Koch (E. nigrum). The structures of 1 was elucidated by spectroscopic methods, including UV, IR, HR-ESI-MS and extensive 1D and 2D NMR techniques.

Isolation and structure elucidation of two new compounds from artemisia Ordosica Krasch.

Two new compounds, namely arteordoyn A (1) and arteordoyn B (2), together with four known compounds, were isolated from the petroleum ether extract of Artemisia ordosica Krasch. The structures elucidation of 1 and 2 were carried out by 1D-NMR (1H and 13C NMR), 2D-NMR (COSY, HSQC, HMBC and NOESY) and HR-ESI-MS spectral analysis.

Two new compounds from Artemisia ordosica Krasch.

Two new compounds, named ordosacid A (5) and ordosacid B (6), along with four known compounds: 3,4-dihydroxybenzaldehyde (1), p-hydroxybenzoic acid (2), p-hydroxycinnamic acid (3) and o-hydroxycinnamic acid (4), were isolated from the ethyl acetate extract of Artemisia ordosica Krasch. The structures of new compounds were elucidated on the basis of spectroscopic methods including UV, IR, ESI-MS, 1D NMR, 2D NMR, HR-ESI-MS and modified Mosher's method.