Cardiotoxicity of (-)-borneol, (+)-borneol, and isoborneol in zebrafish embryos is associated with Na+ /K+ -ATPase and Ca2+ -ATPase inhibition.

Borneol is an example of traditional Chinese medicine widely used in Asia. There are different isomers of chiral borneol in the market, but its toxicity and effects need further study. In this study, we used zebrafish embryos to examine the effects of exposure to three isomers of borneol [(-)-borneol, (+)-borneol, and isoborneol] on heart development and the association with Na+ /K+ -ATPase from 4 h post-fertilization (4 hpf). The results showed that the three isomers of borneol increased mortality and decreased hatching rate when the zebrafish embryo developed to 72 hpf. All three isomers of borneol (0.01-1.0 mM) significantly reduced heart rate from 48 to 120 hpf and reduced the expression of genes related to Ca2+ -ATPase (cacna1ab and cacna1da) and Na+ /K+ -ATPase (atp1b2b, atp1a3b, and atp1a2). At the same time, the three isomers of borneol significantly reduced the activities of Ca2+ -ATPase and Na+ /K+ -ATPase at 0.1 to 1.0 mM. (+)-Borneol caused the most significant reduction (p < 0.05), followed by isoborneol and (-)-borneol. Na+ /K+ -ATPase was mainly expressed in otic vesicles and protonephridium. All three isomers of borneol reduced Na+ /K+ -ATPase mRNA expression, but isoborneol was the most significant (p < 0.01). Our results indicated that (+)-borneol was the least toxic of the three isomers while the isoborneol showed the most substantial toxic effect, closely related to effects on Na+ /K+ -ATPase.

Discussion on Collection of Clinical Questions in International Clinical Practice Guidelines of Acupuncture-Moxibustion / 中国结合医学杂志

The clinical questions of acupuncture-moxibustion (Acup-Mox) guidelines are complicated, including not only the curative effect of Acup-Mox intervention measures, but also the operational elements of Acup-Mox. This paper aimed to put forward the idea and process of collecting clinical questions in developing international acupuncture clinical practice guidelines. The experience was collected and the idea of collecting clinical questions of Acup-Mox was formed through expert consultation and discussion in combination with expert opinions. Based on the characteristics of Acup-Mox discipline. This paper put forward the thinking of collecting elements of clinical questions following the intervention-population-outcome-control (I-P-O-C) inquiry process, according to the discipline of Acup-Mox. It was emphasized that in the process of collecting clinical questions, "treatable population" and "alleviable outcome indicators" for a specific Acup-Mox intervention with certain therapeutic effect should be focused on, so as to highlight the pertinence of clinical questions of Acup-Mox guidelines in terms of population and outcome elements.

Laboratory study of Se speciation in the sediment and oligochaete Lumbriculus variegatus from an aquatic environment.

Sediments are the major sink for selenium (Se) in aquatic environments. Se speciation in sediments is crucial for its bioavailability and toxicity in benthos, but this is relatively understudied. In this study, the background levels of Se in the river sediments, fish flakes, and Lumbriculus variegatus were also detected. Then, the dynamic changes of selenium speciation and concentrations in sediments were investigated after adding selenite (Se(IV)) and seleno-L-methionine (Se-Met) in the sediments for 90 and 7 days, and the accumulation and depuration of Se(IV) and Se-Met for 7 days in the oligochaete L. variegatus were also explored. Without the presence of worms, the levels of Se(IV) in the sediments were relatively stable within 7 days but showed a decreasing trend during the 90 days of aging. In contrast, Se-Met in the sediments showed a sharp decrease within 3 days of aging. The LC50-96 h values of Se(IV) and Se-Met in L. variegatus were 372.6 and 9.4 µg/g, respectively. Interestingly, the dominant Se species in Se(IV)- or Se-Met-treated L. variegatus was Se-Met, whose level was increased with time in 7 days of exposure. Se was barely depurated from L. variegatus during the 8 days of the depuration period. This study has provided indispensable data on the levels of total Se in the abiotic and biotic matrices and the biodynamics of Se in a representative benthos, which could better understand the ecological risk of Se to the freshwater benthic communities.

Inhibitory Effects of Mongolian Medicine Yihe-Tang on Continuous Darkness Induced Liver Steatosis in Zebrafish.

The constant dark induction (DD) causes lipid degeneration and nonalcoholic fatty liver disease (NAFLD) in zebrafish, which might be closely related to the imbalance of gut microbiota and require in-depth study. In this study, a total of 144 zebrafish were divided into four groups, including the control group, Yihe-Tang group, constant dark group, and constant dark + Yihe-Tang group, and were treated with constant darkness (except control and Yihe-Tang groups) for 21 days. The bodyweights of zebrafish were recorded after 8 d, 15 d, and 22 d. The sequencing analysis of gut microbiota, detection of liver histopathological changes, and comparison of lipid metabolism-related gene expression levels were performed on the 22nd day of the experiment. The results showed that the Yihe-Tang could inhibit the constant dark-induced increase in zebrafish weight and liver steatosis. As compared to the control group, the dark treatment could alter the composition of gut microbiota in zebrafish, increase the relative abundance of harmful bacteria, and decrease the Cetobacterium and Bacteroides to Firmicutes ratio in the intestines. The abundance of Proteobacteria in the constant dark + Yihe-Tang group was close to that in the control group and that of Fusobacteria and Cetobacterium increased, especially the Cetobacterium, which increased significantly. The constant dark treatment caused an abnormal expression of liver lipid-related genes, inhibited lipid metabolism, and promoted fat accumulation. However, the Yihe-Tang could restore these changes to the level of the control group. This study indicated that Yihe-Tang could restore the constant dark-induced liver lipid degeneration. We hypothesized that Cetobacterium could significantly inhibit steatosis.

Effects of ferulic acid on muscle development and intestinal microbiota of zebrafish.

Ferulic acid (FA) is one of a common ingredients in Chinese herbal medicine. FA has the interesting property of promoting growth and improving meat quality in livestock, but the mechanism is not understood. This study evaluated both safety and mechanism of efficacy in zebrafish model. At 15 µg/mL or above, FA led to pericardial oedema and delayed growth in zebrafish embryos. Dietary FA promoted growth and feed assimilation in male adult zebrafish. Genes related to myogenic development (myod1, myog and myf5) were significantly upregulated by FA and muscle fibre width in skeletal muscle was increased. At 20 µg/g, FA significantly increased number of goblet cells in zebrafish intestinal tissue, and gut microbiota composition also changed. Based on 16s rRNA gene sequences, 20 µg/g FA decreased Firmicutes and increased Bacteroides. 20 µg/g FA also stimulated the expression of PPAR-α, a gene associated with fat metabolism, and decreased the expression of PPAR-ß and PPAR-γ. These gene expression changes were beneficial to fatty acid synthesis and metabolism and decreased fat deposition. Our overall results indicated that FA can be a safe growth promotor in fish particularly in skeletal muscles.

Analysis and thought on reaching the consensus during the development of acupuncture and moxibustion clinical practice guidelines in China / 中国针灸

Based on the 28 Chinese clinical practice guidelines of acupuncture and moxibustion, this study summarized and analyzed the contents related to reaching consensus during the development process. The results indicated that all the 28 guidelines reported they have used consensus in the "recommendations" section, and provided details on consensus personnel, consensus methods, consensus process and consensus materials. However, it was found that the reporting of consensus was in need of further improvement. The limitations included unclear definition and responsibilities of "expert group", obscure concept between "consensus meeting" and "expert discussion", non-rigorous process of reaching consensus when generating recommendations and lacking of detailed reporting of the consensus reaching process. As such, we suggested that future researchers should conduct researches to further standardized the consensus process when developing acupuncture and moxibustion clinical practice guidelines, so as to improve the quality and clinical applicability of guidelines．.

Interpretation of China national standard Nomenclature and Location of Meridian Points (GB/T 12346-2021) / 中国针灸

To cooperate with the popularization and application of the China national standard Nomenclature and Location of Meridian Points (GB/T 12346 -2021), this study introduced the differences between the 2021 version and the 2006 version, and explained the principles of the revision and the changes in the standard name, terminology, definition and the expression of meridian points' body regions. In addition, the revision of the specific contents, including the adjustment of "bone proportional cun" of several meridian points and the revision basis of location of some meridian points were explained.

Medication Regularity for Chronic Renal Failure and Underlying Mechanism Based on Data Mining， Network Pharmacology， and Molecular Docking / 中国实验方剂学杂志

ObjectiveTo clarify the medication regularity of WU Zhao-dong，a famous chief physician in traditional Chinese medicine （TCM） of Jiangxi province， and investigate the potential mechanism of potential new prescriptions against chronic renal failure （CRF）. MethodThe outpatient prescriptions of WU Zhao-dong from July 2019 to July 2021 were collected. Data mining was carried out by using the Traditional Chinese Medicine Inheritance Auxiliary Platform （V 2.5） to analyze the medication frequency and drug association and obtain potential new prescriptions. The interaction between drug targets in new prescriptions was analyzed by Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform（TCMSP），STRING，and Kyoto Encyclopedia of Genes and Genomes（KEGG）， followed by verification by molecular docking and experiments. ResultA total of 200 prescriptions were screened out， with 217 Chinese medicinal drugs involved， and eight new potential prescriptions were derived. To be specific， Prescription 1： Armeniacae Semen Amarum-Astragali Radix-Platycodonis Radix-Angelicae Sinensis Radix-Smilacis Glabrae Rhizoma-fried Atractylodis Macrocephalae Rhizoma， Prescription 2： Saposhnikoviae Radix-Schizonepetae Herba-Kochiae Fructus-Asteris Radix et Rhizoma-Menthae Haplocalycis Herba，Prescription 3：Armeniacae Semen Amarum-Asteris Radix et Rhizoma-Platycodonis Radix-Eriobotryae Folium-prepared Ephedrae Herba， Prescription 4：Perillae Caulis-Codonopsis Radix-Coptidis Rhizoma-Pseudostellariae Radix， Prescription 5：Ecliptae Herba-Astragali Radix Praeparata Cum Melle-Dryopteridis Crassirhizomatis Rhizoma-Rosae Laevigatae Fructus-Coicis Semen-Ligustri Lucidi Fructus， Prescription 6： Lycopi Herba-Lonice Raejaponicae Caulis-Trachelospermi Caulis et Folium-Alismatis Rhizoma， Prescription 7：Scutellariae Radix-Hirudo-Paeoniae Radix Rubra-Eriobotryae Folium-Glehniae Radix， Prescription 8：Glycyrrhizae Radix et Rhizoma-Scrophulariae Radix-Chrysanthemi Indici Flos-Smilacis Glabrae Rhizoma- Serissae Herba. In Prescription 1，18 main chemical components were screened out. Eighty targets of active components of Prescription 1 were predicted， and 37 potential targets for the treatment of CRF were obtained， including interleukin （IL）-6， vascular endothelial growth factor （VEGF）， cysteinyl aspartate-specific protease-3 （Caspase-3）， nitric oxide synthase 3 （NOS3）， and hypoxia-inducible factor-1 （HIF-1）. The KEGG pathways involved in the targets of Chinese medicinal drugs and disease mainly included the signaling pathways of lipid and atherosclerosis，NF-κB， Toll-like receptors， and HIF-1. Prescription 1 significantly decreased serum creatinine and urea nitrogen， and increased the content of NO and NOS3 in renal tissues of CRF rats. ConclusionPrescription 1 shows the multi-component and multi-target characteristics of action，and its mechanism may be related to its inhibition of renal fibrosis，anti-inflammation，improvement of intestinal microecology，and improvement of renal hypoxia and ischemia.

Inhibitory Effects of Euphorbia ebracteolata Hayata Extract ECB on Melanoma-Induced Hyperplasia of Blood Vessels in Zebrafish Embryos.

Melanoma is a serious malignant form of skin cancer. Euphorbiaceae compound B (ECB, 2,4-dihydroxy-6-methoxy-3-methylacetophenone) is an acetophenone compound that is isolated from Euphorbia ebracteolata Hayata (EEH), an herbaceous perennial, and has antitumor activity. Here, we transplanted human melanoma cells into zebrafish embryos to establish a zebrafish/melanoma model. We showed that this model can be used to evaluate the therapeutic effect of EEH and ECB and discussed its potential mechanism of action. The results showed that ECB was an active ingredient of EEH in inhibiting melanoma-induced hyperplasia of blood vessels in zebrafish embryos, similar to the angiogenic inhibitor vatalanib. ECB inhibited the number and length of subintestinal veins (p < 0.05), as well as the distribution of melanoma in zebrafish embryos (p < 0.05). More importantly, unlike vatalanib, ECB only inhibited melanoma-induced abnormal and excessive growth of blood vessels in xenografts. In addition, ECB inhibited the mRNA expression of vegfr2 and vegfr3 in zebrafish. Both vegfr2 and vegfr3 are essential genes that regulate blood vessel formation and upregulate the expression of p53 and casp3a genes in zebrafish. Together, the above-mentioned results indicate that ECB has a potential antimelanoma effect in vivo, which may be mediated by inhibiting vascular endothelial growth factor receptors.

Aconitine disrupts serotonin neurotransmission via 5-hydroxytryptamine receptor in zebrafish embryo.

Medicinal plants of the genus Aconitum are one of the most commonly used herbs in traditional medicine in East Asia to treat conditions related to the heart, pain, or inflammation. However, these herbs are also dangerous as accidental poisoning due to misuse is a recurring issue. These plants contain a number of diester-diterpenoid alkaloid compounds and aconitine is the most abundant and active one. This study investigated neurotoxicity of aconitine to zebrafish embryos in early development in relation to serotonin regulation. Experimental results showed that aconitine exposure (1, 10, and 100 µM) increased frequency of coiling behavior in zebrafish embryos in a dose-dependent manner and this effect can be triggered by either exposure to 5-hydroxytryptamine 1A (5-HT1A) receptor agonist (±)-8-hydroxy-2-(dipropylamino)tetralin (8-OH-DPAT) or overexpression of serotonin receptor 5-htr1ab. At the same time, coiling behavior caused by aconitine exposure could be rescued by co-exposure to 5-HT1A receptor antagonist WAY-100635 Maleate (WAY100635) and knockdown of 5-htr1ab using morpholino. Exposure to aconitine also significantly increased serotonin receptor 5-htr1ab and 5-htr1bd gene expression at 24 h post fertilization (hpf), but decreased their expression and protein expression of the serotonin receptor at 96 hpf with the high dose. These results suggest that neurotoxicity caused by aconitine is mediated through the 5-HT receptor.