[Transcriptome data mining combined with clinical and animal experiments for identification of syndrome element marker genes during entire course of steroid-induced osteonecrosis of femoral head].

A research strategy combining transcriptome data mining and experimental verification was adopted to identify the marker genes characterizing the syndrome elements of phlegm, stasis, and deficiency in steroid-induced osteonecrosis of the femoral head(SONFH). Firstly, the common differentially expressed gene sets of SONFH with the syndromes of phlegm-stasis obstructing collaterals, vessel obstruction, and liver-kidney deficiency were obtained from the clinical transcriptomic analysis of a previous study. The differential expression trend analysis and functional gene mining were then employed to predict the candidate marker gene sets representing phlegm, stasis, and deficiency. The whole blood samples from SONFH patients, whole blood samples from SONFH rats, and affected femoral head tissue samples were collected for qPCR, which aimed to determine the expression levels of the candidate marker genes mentioned above. Furthermore, the receiver operating characteristic curve(ROC) was established to objectively evaluate the syndrome differentiation effectiveness of the candidate marker genes mentioned above. The transcriptome data analysis results showed that the candidate marker genes for phlegm was ELOVL fatty acid elongase 6(ELOVL6), and those for stasis were ankyrin 1(ANK1), glycophorin A/B(GYPA/B), and Rh-associated glycoprotein(RHAG). The candidate marker genes for deficiency were solute carrier family 2 member 1(SLC2A1) and stomatin(STOM). The qPCR results showed that compared with that in the non-SONFH group, ELOVL6 had the lowest expression level in the peripheral blood of the SONFH patients with the syndrome of phlegm-stasis obstructing collaterals(P<0.05). Compared with that in the normal control group, ELOVL6 had the lowest expression level in the peripheral blood and affected femoral head tissue of SONFH rats modeled for 4 weeks(P<0.01), and it showed better syndrome differentiation effectiveness of rats modeled for 4 weeks(AUC=0.850, P=0.006) than at other modeling time points(8, 12, 16, and 21 weeks, AUC of 0.689, 0.766, 0.588, and 0.662, respectively). Compared with that in the non-SONFH group, the expression levels of ANK1, GYPA, and RHAG were the lowest in the peripheral blood of SONFH patients with the vessel obstruction syndrome(P<0.05). The expression levels of the three genes were the lowest in the peripheral blood and affected femoral head tissue of SONFH rats modeled for 12 weeks(P<0.05, P<0.01), and their syndrome differentiation effectiveness in the rats modeled for 12 weeks(GYPA: AUC=0.861, P=0.012; ANK1: AUC=0.855, P=0.006; RHAG: AUC=0.854, P=0.009) was superior to that for 4, 8, 16, and 21 weeks(GYPA: AUC=0.646, 0.573, 0.691, and 0.617, respectively; ANK: AUC1=0.630, 0.658, 0.657, and 0.585, respectively; RHAG: AUC=0.592, 0.511, 0.515, and 0.536, respectively). Compared with the non-SONFH group, both SLC2A1 and STOM had the lowest expression levels in the peripheral blood of patients with the syndrome of liver and kidney deficiency(P<0.05). Compared with the normal control group, their expression levels were the lowest in the peripheral blood and affected femoral head tissue of SONFH rats modeled for 21 weeks(P<0.05, except STOM in the peripheral blood of rats). Moreover, the syndrome differentiation effectiveness of SLC2A1 in the rats modeled for 21 weeks(AUC=0.806, P=0.009) was superior to that for 4, 8, 12, and 16 weeks(AUC=0.520, 0.580, 0.741, 0.774, respectively), and STOM was meaningless in syndrome differentiation. In summary, the candidate marker gene for phlegm in SONFH is ELOVL6; the candidate marker genes for stasis are GYPA, RHAG, and ANK1; the candidate marker gene for deficiency is SLC2A1. The results help to reveal the biological connotations of phlegm, stasis, and deficiency in SONFH at the genetic level.

Danggui Sini decoction alleviates oxaliplatin-induced peripheral neuropathy by regulating gut microbiota and potentially relieving neuroinflammation related metabolic disorder.

BACKGROUND: Danggui Sini decoction (DSD), a traditional Chinese medicine formula, has the function of nourishing blood, warming meridians, and unblocking collaterals. Our clinical and animal studies had shown that DSD can effectively protect against oxaliplatin (OXA)-induced peripheral neuropathy (OIPN), but the detailed mechanisms remain uncertain. Multiple studies have confirmed that gut microbiota plays a crucial role in the development of OIPN. In this study, the potential mechanism of protective effect of DSD against OIPN by regulating gut microbiota was investigated. METHODS: The neuroprotective effects of DSD against OIPN were examined on a rat model of OIPN by determining mechanical allodynia, biological features of dorsal root ganglia (DRG) as well as proinflammatory indicators. Gut microbiota dysbiosis was characterized using 16S rDNA gene sequencing and metabolism disorders were evaluated using untargeted and targeted metabolomics. Moreover the gut microbiota mediated mechanisms were validated by antibiotic intervention and fecal microbiota transplantation. RESULTS: DSD treatment significantly alleviated OIPN symptoms by relieving mechanical allodynia, preserving DRG integrity and reducing proinflammatory indicators lipopolysaccharide (LPS), IL-6 and TNF-α. Besides, DSD restored OXA induced intestinal barrier disruption, gut microbiota dysbiosis as well as systemic metabolic disorders. Correlation analysis revealed that DSD increased bacterial genera such as Faecalibaculum, Allobaculum, Dubosiella and Rhodospirillales_unclassified were closely associated with neuroinflammation related metabolites, including positively with short-chain fatty acids (SCFAs) and sphingomyelin (d18:1/16:0), and negatively with pi-methylimidazoleacetic acid, L-glutamine and homovanillic acid. Meanwhile, antibiotic intervention apparently relieved OIPN symptoms. Furthermore, fecal microbiota transplantation further confirmed the mediated effects of gut microbiota. CONCLUSION: DSD alleviates OIPN by regulating gut microbiota and potentially relieving neuroinflammation related metabolic disorder.

Modulation of fried spring roll wrapper quality upon treatment of batter with maltogenic amylase, transglutaminase and bromelain.

BACKGROUND: Fried foods are favored for their unique crispiness, golden color and flavor, but they also face great challenge because of their high oil content, high calories and the existence of compounds such as acrylamide and polycyclic aromatic hydrocarbons. Long-term consumption of fried foods may adversely affect health. Therefore, it is necessary to explore fried foods with lower oil contents and a high quality to meet the demand. RESULTS: A method of enzyme treatment was explored to investigate the effects of maltogenic amylase (MA), transglutaminase (TG) and bromelain (BRO) on the physicochemical properties of the batter and the quality of fried spring roll wrapper (FSRW). The results showed that the MA-, TG- or BRO-treated batters had a significant shear-thinning behavior, especially with an increase in viscosity upon increasing TG contents. FSRW enhanced its fracturability from 419.19 g (Control) to 616.50 g (MA-6 U g-1), 623.49 g (TG-0.75 U g-1) and 644.96 g (BRO-10 U g-1). Meanwhile, in comparison with BRO and MA, TG-0.5 U g-1 endowed batter with the highest density and thermal stability. MA-15 U g-1 and TG-0.5 U g-1 displayed FSRW with uniform and dense pores, and significantly reduced its oil content by 18.05% and 25.02%, respectively. Moreover, compared to MA and TG, BRO-50 U g-1 improved the flavor of FSRW. CONCLUSION: MA, TG or BRO played a key role in affecting the physicochemical properties of the batter and the quality of FSRW. TG-0.5 U g-1 remarkly reduced the oil content of FSRW with a great potential in practical application. © 2024 Society of Chemical Industry.

Chromosome-level genome of Thymus mandschuricus reveals molecular mechanism of aroma compounds biosynthesis.

Background: Thymus mandschuricus is an aromatic and medicinal plant with notable antibacterial and antioxidant properties. However, traditional breeding methods rely on phenotypic selection due to a lack of molecular resources. A high-quality reference genome is crucial for marker-assisted breeding, genome editing, and molecular genetics. Results: We utilized PacBio and Hi-C technologies to generate a high-quality chromosome-level reference genome for T. mandschuricus, with a size of 587.05 Mb and an N50 contig size of 8.41 Mb. The assembled genome contained 29,343 predicted protein-coding genes, and evidence of two distinct whole-genome duplications in T. mandschuricus was discovered. Comparative genomic analysis revealed rapid evolution of genes involved in phenylpropanoid biosynthesis and the CYP450 gene family in T. mandschuricus. Additionally, we reconstructed the gene families of terpenoid biosynthesis structural genes, such as TPS, BAHD, and CYP, and identified regulatory networks controlling the expression of aroma-synthesis genes by integrating transcriptome data from various organs and developmental stages. We discovered that hormones and transcription factors may collaborate in controlling aroma-synthesis gene expression. Conclusion: This study provides the first high-quality genome sequence and gene annotation for T. mandschuricus, an indigenous thyme species unique to China. The genome assembly and the comprehension of the genetic basis of fragrance synthesis acquired from this research could potentially serve as targets for future breeding programs and functional studies.

Diterpenoid tanshinones inhibit gastric cancer angiogenesis through the PI3K/Akt/mTOR signaling pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Salvia miltiorrhiza Bunge is a kind of Chinese herbal medicine known for activating blood circulation and removing blood stasis, with the effect of cooling blood and eliminating carbuncles, and has been proven to have the effect of treating tumors. However, the inhibitory effect of Salvia miltiorrhiza Bunge extracts (Diterpenoid tanshinones) on tumors by inhibiting angiogenesis has not been studied in detail. AIM OF THE STUDY: This study aimed to investigate the anti-gastric cancer effect of diterpenoid tanshinones (DT) on angiogenesis, including the therapeutic effects and pathways. MATERIALS AND METHODS: This experiment utilized network pharmacology was used to identify relevant targets and pathways of Salvia miltiorrhiza Bunge-related components in the treatment of gastric cancer. The effects of DT on the proliferation and migration of human gastric cancer cell line SGC-7901 and human umbilical vein endothelial cell line HUVECs were evaluated, and changes in the expression of angiogenesis-related factors were measured. In vivo, experiments were conducted on nude mice to determine tumor activity, size, immunohistochemistry, and related proteins. RESULTS: The findings showed that DT could inhibit the development of gastric cancer by suppressing the proliferation of gastric cancer cells, inducing apoptosis, and inhibiting invasion and metastasis. In addition, the content of angiogenesis-related factors and proteins was significantly altered in DT-affected cells and animals. CONCLUSIONS: Results suggest that DT has potential as a therapeutic agent for the treatment of gastric cancer, as it can inhibit tumor growth and angiogenesis. It was also found that DT may affect the expression of the angiogenic factor VEGF through the PI3K/Akt/mTOR pathway, leading to the regulation of tumor angiogenesis. This study provides a new approach to the development of anti-tumor agents and has significant theoretical and clinical implications for the treatment of gastric cancer.

Berberine Enhances Intestinal Mucosal Barrier Function by Promoting Vitamin D Receptor Activity.

OBJECTIVE: To evaluate if berberine can act on vitamin D receptors (VDR) and thereby regulate the expression of tight junction proteins (TJPs) in irritable bowel syndrame-diarrhea-predominant (IBS-D) rats. METHODS: The newborn rats were induced into IBS-D rat model via neonatal maternal separation combined with acetic acid chemical stimulation. After modeling, the model was evaluated and rats were divided into the control group and berberine treatment groups (0.85, 1.7 and 3.4 mg/kg, once a day for 2 weeks). The distal colon was obtained and colonic epithelial cells (CECs) were isolated and cultured after IBS-D model evaluation. The vitamin D receptor response element (VDRE) reporter gene was determined in the CECs of IBS-D rats to analyze the effect of berberine on the VDRE promoter. VDR overexpression or silencing technology was used to analyze whether VDR plays a role in promoting intestinal barrier repair, and to determine which region of VDR plays a role in berberine-regulated intestinal TJPs. RESULTS: The IBS-D rat model was successfully constructed and the symptoms were improved by berberine in a dose-dependent manner (P<0.05). The activity of VDRE promoter was also effectively promoted by berberine (P<0.05). Berberine increased the expression of TJPs in IBS-D CECs (P<0.05). VDR expression was significantly increased after transfection of different domains of VDR when compared to normal control and basic plasmid groups (all P<0.05). RT-qPCR and Western blot results showed that compared with the blank group, expressions of occludin and zonula occludens-1 were significantly higher in VDR containing groups (all P<0.05). Berberine plus pCMV-Myc-VDR-N group exerted the highest expression levels of occludin and zonula occludens-1 (P<0.05). CONCLUSION: Berberine enhances intestinal mucosal barrier function of IBS-D rats by promoting VDR activity, and the main site of action is the N-terminal region of VDR.

A novel drug specific mRNA biomarker predictor for selection of patients responding to dovitinib treatment of advanced renal cell carcinoma and other solid tumors.

PURPOSE: Dovitinib is a receptor tyrosine kinase inhibitor of VEGFR1-3, PDGFR, FGFR1/3, c-KIT, FLT3 and topoisomerase 1 and 2. The drug response predictor (DRP) biomarker algorithm or DRP-Dovitinib is being developed as a companion diagnostic to dovitinib and was applied retrospectively. PATIENTS AND METHODS: Archival tumor samples were obtained from consenting patients in a phase 3 trial comparing dovitinib to sorafenib in renal cell carcinoma patients and the DRP-Dovitinib was applied. The biomarker algorithm combines the expression of 58 messenger RNAs relevant to the in vitro sensitivity or resistance to dovitinib, including genes associated with FGFR, PDGF, VEGF, PI3K/Akt/mTOR and topoisomerase pathways as well as ABC drug transport, and provides a likelihood score between 0-100%. RESULTS: The DRP-Dovitinib divided the dovitinib treated RCC patients into two groups, sensitive (n = 49, DRP score >50%) or resistant (n = 86, DRP score ≤ 50%) to dovitinib. The DRP sensitive population was compared to the unselected sorafenib arm (n = 286). Median progression-free survival (PFS) was 3.8 months in the DRP sensitive dovitinib arm and 3.6 months in the sorafenib arm (hazard ratio 0.71, 95% CI 0.51-1.01). Median overall survival (OS) was 15.0 months in the DRP sensitive dovitinib arm and 11.2 months in the sorafenib arm (hazard ratio 0.69, 95% CI 0.48-0.99). The observed clinical benefit increased with increasing DRP score. At a cutoff of 67% the median OS was 20.6 months and the median PFS was 5.7 months in the dovitinib arm. The results were confirmed in five smaller phase II trials of dovitinib which showed a similar trend. CONCLUSION: The DRP-Dovitinib shows promise as a potential biomarker for identifying advanced RCC patients most likely to experience clinical benefit from dovitinib treatment, subject to confirmation in an independent prospective trial of dovitinib in RCC patients.

Promoting Photosynthetic Production of Dammarenediol-II in Chlamydomonas reinhardtii via Gene Loading and Culture Optimization.

Ginsenosides are major bioactive compounds found in Panax ginseng that exhibit various pharmaceutical properties. Dammarenediol-II, the nucleus of dammarane-type ginsenosides, is a promising candidate for pharmacologically active triterpenes. Dammarenediol-II synthase (DDS) cyclizes 2,3-oxidosqualene to produce dammarenediol-II. Based on the native terpenoids synthetic pathway, a dammarane-type ginsenosides synthetic pathway was established in Chlamydomonas reinhardtii by introducing P. ginseng PgDDS, CYP450 enzyme (PgCYP716A47), or/and Arabidopsis thaliana NADPH-cytochrome P450 reductase gene (AtCPR), which is responsible for producing dammarane-type ginsenosides. To enhance productivity, strategies such as "gene loading" and "culture optimizing" were employed. Multiple copies of transgene expression cassettes were introduced into the genome to increase the expression of the key rate-limiting enzyme gene, PgDDS, significantly improving the titer of dammarenediol-II to approximately 0.2 mg/L. Following the culture optimization in an opt2 medium supplemented with 1.5 mM methyl jasmonate under a light:dark regimen, the titer of dammarenediol-II increased more than 13-fold to approximately 2.6 mg/L. The C. reinhardtii strains engineered in this study constitute a good platform for the further production of ginsenosides in microalgae.

Network pharmacology and molecular docking to explore Polygoni Cuspidati Rhizoma et Radix treatment for acute lung injury.

BACKGROUND: Polygoni Cuspidati Rhizoma et Radix (PCRR), a well-known traditional Chinese medicine (TCM), inhibits inflammation associated with various human diseases. However, the anti-inflammatory effects of PCRR in acute lung injury (ALI) and the underlying mechanisms of action remain unclear. AIM: To determine the ingredients related to PCRR for treatment of ALI using multiple databases to obtain potential targets for fishing. METHODS: Recognized and candidate active compounds for PCRR were obtained from Traditional Chinese Medicine Systems Pharmacology, STITCH, and PubMed databases. Target ALI databases were built using the Therapeutic Target, DrugBank, DisGeNET, Online Mendelian Inheritance in Man, and Genetic Association databases. Network pharmacology includes network construction, target prediction, topological feature analysis, and enrichment analysis. Bioinformatics resources from the Database for Annotation, Visualization and Integrated Discovery were utilized for gene ontology biological process and Kyoto Encyclopedia of Genes and Genomes network pathway enrichment analysis, and molecular docking techniques were adopted to verify the combination of major active ingredients and core targets. RESULTS: Thirteen bioactive compounds corresponding to the 433 PCRR targets were identified. In addition, 128 genes were closely associated with ALI, 60 of which overlapped with PCRR targets and were considered therapeutically relevant. Functional enrichment analysis suggested that PCRR exerted its pharmacological effects in ALI by modulating multiple pathways, including the cell cycle, cell apoptosis, drug metabolism, inflammation, and immune modulation. Molecular docking results revealed a strong associative relationship between the active ingredient and core target. CONCLUSION: PCRR alleviates ALI symptoms via molecular mechanisms predicted by network pharmacology. This study proposes a strategy to elucidate the mechanisms of TCM at the network pharmacology level.

Iron-Catalyzed C(Sp3)-H Borylation, Thiolation, and Sulfinylation Enabled by Photoinduced Ligand-to-Metal Charge Transfer.

Catalytic C(sp3)-H functionalization has provided enormous opportunities to construct organic molecules, facilitating the derivatization of complex pharmaceutical compounds. Within this framework, direct hydrogen atom transfer (HAT) photocatalysis becomes an appealing approach to this goal. However, the viable substrates utilized in these protocols are limited, and the site selectivity shows preference to activated and thermodynamically favored C(sp3)-H bonds. Herein, we describe the development of undirected iron-catalyzed C(sp3)-H borylation, thiolation, and sulfinylation reactions enabled by the photoinduced ligand-to-metal charge transfer (LMCT) process. These reactions exhibit remarkably broad substrate scope (>150 examples in total), and most importantly, all of these three reactions show unconventional regioselectivity, with the occurrence of C(sp3)-H borylation, thiolation, and sulfinylation preferentially at the distal methyl position. The procedures are operationally simple and readily scalable and provide access to high-value products from simple hydrocarbons in one step. Mechanistic studies and control experiments indicate that the afforded site selectivity is not only relevant to the HAT species but also largely affected by the use of boron- and sulfone-based radical acceptors.

A thalamic circuit facilitates stress susceptibility via melanocortin 4 receptor-mediated activation of nucleus accumbens shell.

AIMS: Central melanocortin 4 receptor (MC4R) has been reported to induce anhedonia via eliciting dysfunction of excitatory synapses. It is evident that metabolic signals are closely related to chronic stress-induced depression. Here, we investigated that a neural circuit is involved in melanocortin signaling contributing to susceptibility to stress. METHODS: Chronic social defeat stress (CSDS) was used to develop depressive-like behavior. Electrophysiologic and chemogenetic approaches were performed to evaluate the role of paraventricular thalamus (PVT) glutamatergic to nucleus accumbens shell (NAcsh) circuit in stress susceptibility. Pharmacological and genetic manipulations were applied to investigate the molecular mechanisms of melanocortin signaling in the circuit. RESULTS: CSDS increases the excitatory neurotransmission in NAcsh through MC4R signaling. The enhanced excitatory synaptic input in NAcsh is projected from PVT glutamatergic neurons. Moreover, chemogenetic manipulation of PVTGlu -NAcsh projection mediates the susceptibility to stress, which is dependent on MC4R signaling. Overall, these results reveal that the strengthened excitatory neurotransmission in NAcsh originates from PVT glutamatergic neurons, facilitating the susceptibility to stress through melanocortin signaling. CONCLUSIONS: Our results make a strong case for harnessing a thalamic circuit to reorganize excitatory synaptic transmission in relieving stress susceptibility and provide insights gained on metabolic underpinnings of protection against stress-induced depressive-like behavior.

Gut microbiota mediates the pharmacokinetics of Zhi-zi-chi decoction for the personalized treatment of depression.

ETHNOPHARMACOLOGICAL RELEVANCE: Zhi-zi-chi decoction (ZZCD), from "Treatise on Febrile Diseases", is a typical traditional Chinese medicine herb pair, which consists of Gardeniae Fructus (GF) and Semen Sojae Praeparatu (SSP). In clinical research, ZZCD was widely used to fight depression, remove annoyance. Many studies have reported that gut microbiota is critical target for the influence of depress through gut-brain axis, and our previously studies have found that ZZCD exhibiting antidepressant effect was through the gut-brain axis. However, the specific mechanism by which gut microbiota mediates the pharmacokinetics parameters of active compounds from ZZCD during the process of depression treatment has not yet been studied. AIM OF THE STUDY: To explore the differences in pharmacokinetics characters of bioactive iridoids from ZZCD and study the changes of gut microbiota at different stages of depression with the personalized medicine of ZZCD. MATERIALS AND METHODS: A new strategy exploring the relationship among disease phenotypes (D), intestinal microbiota (I), enzymes (E) and traits of metabolism (T) named as "DIET" was established. Firstly, a fast, selective and sensitive ultra-performance liquid chromatography coupled with tandem mass spectrometer (UPLC-MS/MS) was established and validated to quality the main bioactive compounds from ZZCD and compare the pharmacokinetics and bioavailability of different iridoids prototypes and metabolites from ZZCD between normal and chronic unpredictable mild stress rats. Subsequently, the activity of corresponding metabolic enzymes of anti-depressive compounds, ß-glucosidases and sulfotransferases, were analyzed by ρ-nitrophenyl-ß -D-glucopyranoside and sulfotransferases ELISA kits, respectively. Finally, 16S rRNA gene sequencing was adopt to analyze intestinal bacteria composition for the treatment of depression by ZZCD. RESULTS: The antidepressant effect of ZZCD was promoted due to the increased exposures and reduced eliminations of anti-depressive compounds, especially geniposide and genipin 1-gentiobioside, under the depression state. With the ZZCD treatment, the depression was improved, but the exposures of anti-depressive compounds from ZZCD gradually decreased. Meanwhile, there were the corresponding decreased trends on the activity of ß-glucosidases and sulfotransferases. With the consumption of ZZDC and the improvement of depression, the exposures of anti-depressive iridoid glycosides decreased and the activity of metabolism enzymes restored. Meanwhile, the dysbiosis of pathogenic bacteria (Bacteroidota) induced by depression was ameliorated and the probiotics (Firmicutes) at the phylum and genus level raised, the two phyla are closely related to the production of ß-glucosidase and sulfotransferases. CONCLUSIONS: It is the first proposed that ZZCD could personalized to treat depression at different stages targeting gut microbiota and gut microbiome could emerged as a potential diagnostic and therapeutic biomarker in depression.

3D Microfluidic System for Evaluating Inhibitory Effect of Chinese Herbal Medicine Oldenlandia diffusa on Human Malignant Glioma Invasion Combined with Network Pharmacology Analysis.

OBJECTIVE: To investigate the anti-invasion efficacy of the ethanol extract of Oldenlandia diffusa Will. (EEOD) on a three-dimensional (3D) human malignant glioma (MG) cell invasion and perfusion model based on microfluidic chip culture and the possible mechanism of action of Oldenlandia diffusa Will. (OD). METHODS: The comprehensive pharmacodynamic analysis method in this study was based on microfluidic chip 3D cell perfusion culture technology, and the action mechanism of Chinese medicine (CM) on human MG cells was investigated through network pharmacology analysis. First, the components of EEOD were analyzed by ultraperformance liquid chromatography with quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF/MS). Then, cell viability and apoptosis were assessed to determine the optimum concentration of EEOD for invasion experiments, and two-dimensional (2D) migration and invasion abilities of U87 and U251 MG cells were evaluated using scratch wound and Transwell assays. The possible mechanism underlying the effects of EEOD on glioma was analyzed through a network pharmacology approach. RESULTS: Thirty-five compounds of EEOD were detected by UPLC-Q-TOF/MS. EEOD suppressed the viability of MG cells, promoted their apoptosis, and inhibited their migratory and invasive potentials (all P<0.05). Network pharmacology analysis showed that OD inhibited the invasion of MG cells by directly regulating MAPK and Wnt pathways through MAPK, EGFR, MYC, GSK3B, and other targets. The anti-invasion effect of OD was also found to be related to the indirect regulation of microtubule cytoskeleton organization. CONCLUSIONS: ]EEOD could inhibit the invasion of human MG cells, and the anti-invasion mechanism of OD might be regulating MAPK and Wnt signaling pathways and microtubule cytoskeleton organization.

Sexual Dimorphism in Mechanical Hypersensitivity Induced by Subcutaneous Injection of M-CSF / 中山大学学报(医学科学版)

ObjectiveTo investigate whether there exists gender differences in mechanical pain hypersensitivity induced by the subcutaneous injection of macrophage colony-stimulating factor （M-CSF） in normal mice and to explore the preliminary mechanism. MethodsThirty 10-week-old C57BL/6J mice were randomly divided into three groups， (n = 10 mice/group, half male and half female). The albumin control group （BSA， 0.3 μg）， low dose M-CSF group （L M-CSF， 0.075 μg） and high dose M-CSF group （H M-CSF， 0.3 μg） received 50 μL BSA or M-CSF injected subcutaneously into the left medial thigh once daily for 3 consecutive days. Before and after drug administration， von-Frey mechanical sensitivity test was used to detect the mechanical paw withdrawal threshold （PWT） in each group. Immunofluorescence was performed to examine the expression changes of Ionized calcium-binding adaptor molecule 1 (Iba1) in skin， calcitonin gene-related peptide （CGRP） and phosphorylated ERK1/2 （p-ERK） in L5-L6 DRG and lumbar spinal dorsal horn. ResultsIn female mice， only high dose of M-CSF caused mechanical allodynia， whereas in male mice both doses produced marked allodynia. Mechanically， high-dose M-CSF induced massive aggregation of subcutaneous macrophages （marked by Iba1） in male and female mice， but more dramatic dependence in female mice. Similar gender differences were also found in the increase of p-ERK and CGRP expression in dorsal root ganglion （DRGs）. Notably， CGRP expression was especially elevated in the fibers of DRG in male mice. Correspondingly， the expressions of p-ERK and CGRP+ terminals in the superficial spinal dorsal horn of male mice were significantly higher than those of female mice after M-CSF treatment. ConclusionSubcutaneous injection of M-CSF triggers sexual dimorphism in mechanical pain hypersensitivity, which is related with differential changes in peripheral macrophage expansion and sensitization of the nociceptive pathway.

Bushen Huoxue Fang improves recurrent miscarriage in mice by down-regulating the JAK2/STAT3 pathway / 南方医科大学学报

OBJECTIVE@#To investigate the efficacy of Bushen Huoxue Fang (BSHXF, a traditional Chinese medicine formula) for improving recurrent spontaneous abortion (RSA) in mice and the role of tyrosine kinase (JAK2) and transcriptional activator (STAT3) signaling pathway in its therapeutic mechanism.@*METHODS@#Female CBA/J mice were caged with male DBA/2 mice to establish RSA mouse models, which were randomly divided into model group, dydrogesterone group and BSHXF group, with the female mice caged with male BALB/c mice as the control group (n=6). From the first day of pregnancy, the mice were subjected to daily intragastric administration of BSHXF, dydrogesterone, or distilled water (in control and model groups) for 12 days. After the treatments, serum levels of antithrombin III (AT-III), activated protein C (APC), tissue plasminogen activator (t-PA), progesterone, human chorionic gonadotropin (HCG), and estradiol (E2) were detected in each group using ELISA. HE staining was used to observe the morphological changes of the endometrium of the mice. Western blotting was performed to determine the expressions of p-JAK2, p-Stat3 and Bcl-2 in the placenta of the mice.@*RESULTS@#Compared with the control mice, the mouse models of RSA showed a significantly increased embryo loss rate with decreased serum levels of AT-III, T-PA, progesterone, APC and HCG, increased placental expressions of p-JAK2, p-STAT3 and Bax, and decreased expression of Bcl-2 (P < 0.05). Treatments with BSHXF and dydrogesterone both increased serum levels of AT-III, t-PA and HCG in the mouse models; Serum APC level was significantly reduced in BSHXF group and serum progesterone level was significantly increased in dydrogesterone group (P < 0.05).@*CONCLUSION@#BSHXF can improve the prethrombotic state and inhibit cell apoptosis by downregulating the JAK2/STAT3 pathway to increase the pregnancy rate in mouse models of RSA.

Association between the CASC16 rs4784227 polymorphism and breast cancer risk and prognosis in a northeast Chinese Han population.

Background: Breast cancer (BC) poses a serious threat to women worldwide. This research was designed to explore the association between the rs4784227 polymorphism of cancer susceptibility candidate gene 16 (CASC16) and BC susceptibility and prognosis, aiming to provide further information for the early detection of BC and to accelerate comprehensive cancer management. Methods: A total of 1,733 subjects were recruited for this case-control study, of which 828 are BC patients and 905 are healthy individuals. The relevance between SNP rs4784227 and BC risk in diverse genetic models was analyzed by using the SNPStats analysis program and was assessed by odds ratios (ORs) and 95% confidence intervals (CIs) using the binary logistic regression model. Pearson's χ 2 test was used to determine the correlation between the polymorphism and clinical characteristics of BC patients. Additionally, univariate survival analysis was performed by the Kaplan-Meier method and log-rank test, and multivariate survival analysis was performed by Cox regression. Results: SNP rs4784227 was significantly associated with susceptibility to BC in the dominant model (CT/TT versus CC, OR = 1.237, 95% CI = 1.012-1.513, P = 0.038). The minor allele of SNP rs4784227 was significantly linked to an increased risk of BC (OR = 1.197, 95% CI = 1.022-1.401, P = 0.026). In addition, the rs4784227 polymorphism of CASC16 was associated with perineural invasion (P = 0.030), menstrual status (P = 0.016) and histological grade (P = 0.001, P = 0.003, P = 0.025; respectively) of BC patients. There was no significant association between the genotypes of rs4784227 and disease-free survival (DFS) or overall survival (OS) of breast cancer patients (P > 0.05). Conclusions: The rs4784227 polymorphism of CASC16 may affect susceptibility to breast cancer and is associated with perineural invasion, menstrual status and histological grade in BC patients. Additionally, our results could not confirm that this polymorphism was related to breast cancer prognosis.

Prevention of polycystic ovary syndrome and postmenopausal osteoporosis by inhibiting apoptosis with Shenling Baizhu powder compound.

Objective: Shenling Baizhu powder (SBP) has been shown to reverse the abnormal expression of the aromatic hydrocarbon receptor (AHR) mediated by air pollution. Our study aimed to understand the main ingredient of SBP and investigate its action mechanism in preventing polycystic ovary syndrome (POCS) and postmenopausal osteoporosis (PMO). Methods: The active ingredients of SBP with the highest binding affinity to AHR were screened using a Chinese medicine database, and their binding mechanism was simulated using molecular dynamics simulation (MDS). Rutin was utilized to treat ovarian granulosa cell lines and osteoblast cell lines. The cell lines were treated with a gradient of rutin concentration (0.01 mmol/L, 0.05 mmol/L and 0.1 mmol/L) to find the optimal drug dose. PCR was used to detect AHR and apoptosis-related proteins, and WB to detect the expression of AHR, caspase-3 and cleaved-caspase-3. Finally, the CCK-8 cell proliferation assay detected the proliferation of cells. Results: We obtained Rutin through the Chinese medicine database, and dynamics simulation determined its binding sites. Ovarian granulosa cell lines and osteoblast cell lines were treated with Rutin. RT-PCR and western blotting revealed that the expression of apoptosis-associated protein Bcl-2 was elevated, and the expression of AHR, Bax, caspase-3 and PARP were decreased. CCK-8 results showed accelerated proliferation in both cell types. Conclusion: Rutin, the main ingredient of SBP compound, works by binding to AHR, which can improve POCS and PMO by inhibiting cell apoptosis and by promoting cell proliferation.

PD-1 engineered cytomembrane cloaked molybdenum nitride for synergistic photothermal and enhanced immunotherapy of breast cancer.

Incomplete tumor ablation and subsequent tumor metastasis usually occur during photothermal anti-tumor processes. The combination of photothermal and immunotherapy has proven to be a promising method to conquer technical challenges. Inhibiting the programmed death ligand-1 (PD-L1)/programmed cell death protein 1 (PD-1) immune pathway represents one of the most successful immunotherapy strategies. Whereas, the PD-L1 expression level significantly differs, leading to a relatively low response rate to the immune checkpoint blockade (ICB) approaches. Therefore, improving the expression level of PD-L1 becomes one potential method to enhance the response rate. Herein, NIH 3T3 cells were educated to steadily express PD-1 protein. Furthermore, the synthesized molybdenum nitride was then coated with PD-1 protein-modified cytomembrane, which endows it with immune checkpoint blocking capability. Moreover, under the irradiation of near-infrared light, the local mild heat released from the molybdenum nitride causes the apoptosis of tumor cells. More importantly, the elevated temperature simultaneously helps elevate the expression level of PD-L1, further enhancing the response rate of ICB. Finally, the PD-1 cytomembrane coatings interact with the upregulated PD-L1, leading to the activation of the immune system. In summary, we confirmed that the PD-1 protein-coated molybdenum nitride could synergistically ablate tumors and avoid metastasis.

Relaxin-3 Ameliorates Diabetic Cardiomyopathy by Inhibiting Endoplasmic Reticulum Stress.

Background: This study is aimed at investigating whether relaxin-3 exhibits protective effects against cardiomyopathy in diabetic rats by suppressing ERS. Methods: Eighty male SD rats were randomly divided into two groups: controls (n = 20) and diabetes (n = 60). The streptozotocin-treated rats were randomly divided into three groups: diabetic group (DM), low-dose relaxin-3 group (0.2 µg/kg/d), and high-dose relaxin-3 group (2 µg/kg/d). The myocardial tissues and collagen fiber were observed by hematoxylin and eosin (H&E) and Masson staining. Serum brain natriuretic peptide (BNP), troponin (TNI), myoglobin, interleukin (IL-17), interleukin (IL)-1α, and tumor necrosis factor (TNF)-α were determined by ELISA. The protein expression of glucose regulatory protein 78 (GRP78) and C/EBP homologous protein (CHOP) in the heart tissue of each group was detected by Western blot analysis. Results: (1) HE and Masson staining indicated that relaxin-3 could attenuate myocardial lesions and myocardial collagen volume fraction. (2) BNP, TnI, and myoglobin in the DM group at four and eight weeks were significantly higher than in the controls (P < 0.01). The relaxin-3-treated groups showed significantly reduced serum BNP, TnI, and myoglobin levels compared with the DM group (P < 0.05). (3) IL-17, IL-1α, and TNF-α levels in the DM rats at 4 weeks were higher than in the controls (P < 0.05). Low or high dose of relaxin-3-treated groups showed reduced serum IL-17 and TNF-α levels compared with the DM group at four and eight weeks (P < 0.05). (4) CHOP and GRP78 protein expression was increased in the DM group at four and eight weeks compared with the controls (P < 0.01), and small and large doses of relaxin-3 significantly reduced GRP78 and CHOP protein expression. Conclusions: Exogenous relaxin-3 ameliorates diabetic cardiomyopathy by inhibiting ERS in diabetic rats.

Amanitin intoxication: effects of therapies on clinical outcomes - a review of 40 years of reported cases.

BACKGROUND AND AIMS: Amanita phalloides poisoning causes severe liver damage which may be potentially fatal. Several treatments are available, but their effectiveness has not been systematically evaluated. We performed a systematic review to investigate the effect of the most commonly used therapies: N-acetylcysteine (NAC), benzylpenicillin (PEN), and silibinin (SIL) on patient outcomes. In addition, other factors contributing to patient outcomes are identified. METHODS: We searched MEDLINE and Embase for case series and case reports that described patient outcomes after poisoning with amanitin-containing Amanita mushrooms. We extracted clinical characteristics, treatment details, and outcomes. We used the liver item from the Poisoning Severity Score (PSS) to categorize intoxication severity. RESULTS: We included 131 publications describing a total of 877 unique cases. The overall survival rate of all patients was 84%. Patients receiving only supportive care had a survival rate of 59%. The use of SIL or PEN was associated with a 90% (OR 6.40 [3.14-13.04]) and 89% (OR 5.24 [2.87-9.56]) survival rate, respectively. NAC/SIL combination therapy was associated with 85% survival rate (OR 3.85 [2.04, 7.25]). NAC/PEN/SIL treatment group had a survival rate of 76% (OR 2.11 [1.25, 3.57]). Due to the limited number of cases, the use of NAC alone could not be evaluated. Additional analyses in 'proven cases' (amanitin detected), 'probable cases' (mushroom identified by mycologist), and 'possible cases' (neither amanitin detected nor mushroom identified) showed comparable results, but the results did not reach statistical significance. Transplantation-free survivors had significantly lower peak values of aspartate aminotransferase (AST), alanine aminotransferase (ALT), total serum bilirubin (TSB), and international normalized ratio (INR) compared to liver transplantation survivors and patients with fatal outcomes. Higher peak PSS was associated with increased mortality. CONCLUSION: Based on data available, no statistical differences could be observed for the effects of NAC, PEN or SIL in proven poisonings with amanitin-containing mushrooms. However, monotherapy with SIL or PEN and combination therapy with NAC/SIL appear to be associated with higher survival rates compared to supportive care alone. AST, ALT, TSB, and INR values are possible predictors of potentially fatal outcomes.