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Specific DNA identification of Pheretima in the Naoxintong capsule.

ABSTRACT: Background:Pheretima is a minister drug in Naoxintong capsule (NXTC), a well-known traditional Chinese medicine (TCM) formula for the treatment of cardiovascular and cerebrovascular diseases. Owing to the loss of morphological and microscopic characteristics and the lack of recognized chemical marker, it is difficult to identify Pheretima in NXTC. This study aims to evaluate the feasibility of using DNA techniques to authenticate Pheretima, especially when it is processed into NXTC. Methods:DNA was extracted from crude drugs of the genuine and adulterant species, as well as nine batches of NXTCs. Based on mitochondrial cytochrome c oxidase subunit I (COI) gene, specific primers were designed for two genera of genuine species, Metaphire and Amynthas, respectively. PCR amplification was performed with the designed primers on crude drugs of Pheretima and NXTCs. The purified PCR products were sequenced and the obtained sequences were identified to species level with top hit of similarity with BLAST against GenBank nucleotide database. Results:Primers MF2R2 and AF3R1 could amplify specific DNA fragments with sizes around 230-250 bp, both in crude drugs and NXTC. With sequencing and the BLAST search, identities of the tested samples were found. Conclusion:This study indicated that the molecular approach is effective for identifying Pheretima in NXTC. Therefore, DNA identification may contribute to the quality control and assurance of NXTC.

SUBMITTER: Zhu X

PROVIDER: S-EPMC6767644 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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Specific DNA identification of Pheretima in the Naoxintong capsule.

Zhu Xiaoxiao X Wu Hoi-Yan HY Shaw Pang-Chui PC Peng Wei W Su Weiwei W

Chinese medicine 20190930

<h4>Background</h4>Pheretima is a minister drug in Naoxintong capsule (NXTC), a well-known traditional Chinese medicine (TCM) formula for the treatment of cardiovascular and cerebrovascular diseases. Owing to the loss of morphological and microscopic characteristics and the lack of recognized chemical marker, it is difficult to identify Pheretima in NXTC. This study aims to evaluate the feasibility of using DNA techniques to authenticate Pheretima, especially when it is processed into NXTC.<h4>M ...[more]

PMID: 31583012

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Project description:Naoxintong capsule (NXT), a Chinese medicine, has performed excellent effects on the prevention and treatment against cardiovascular diseases. NXT is a fine powder mixture without any herb extraction, and there must be lots of ingredients hard to be absorbed. However, little is known about the correlation between the NXT's cardioprotective effects and gut microbiota. Herein, we report the effect of NXT on the development of cardiovascular diseases and clarify the correlation between NXT's cardioprotective effects and gut microbiota. In the current study, minipigs were selected and fed with high-fat diet and NXT daily for successive 8 months. During the process, up to 18 biomedical parameters were monthly determined to observe the dynamic changes after NXT treatment. At the end of experimental process, pathological examinations of heart, coronary artery, carotid artery, thoracic aorta, and abdominal aorta were conducted by HE staining and 16SrDNA sequencing, and analyzing of gut microbiota were conducted. Our results showed that NXT's effects against cardiovascular diseases were through regulating blood lipid profiles, inhibiting vascular inflammation, enhancing antioxidant capacity, and alleviating myocardial injury, without damages on liver and kidney particularly. Concurrently, we also found that long-term administration of NXT increased the diversity of gut microbiota, influenced the microbiome structure and composition stably, and revered the increase of the ratio of the Firmicutes to Bacteroidetes (F/B ratio) in relative abundance. Specifically, our results revealed some key bacterium of Caproiciproducens (enhanced), Sutterella (enhanced), Erysipelotrichaceae (enhanced), and Romboutsia (decreased) that were closely involved in NXT's effects. Taken together, our study demonstrates that NXT can inhibit the development of cardiovascular diseases by ameliorating high-fat diet-induced metabolic disorders and partly through improving gut microbiota.

Project description:AimsTo systematically evaluate the comprehensive effect of combining Naoxintong capsule (NXT) with Western medicine (WM) on coronary heart disease post-percutaneous coronary intervention (PCI).MethodsRandomized controlled trials (RCTs) of NXT for patients with CHD after PCI were systematically searched across multiple databases, including the Cochrane Library, PubMed, Embase, Chinese National Knowledge Infrastructure (CNKI), Chinese Science and Technology Journal Database (VIP), and Wan Fang, from inception until 31 January 2023. Study selection, data extraction, and quality assessment were performed by two independent reviewers. The quality of the included studies was evaluated using version 2 of the Cochrane risk-of-bias tool (RoB 2), and data analysis was performed using R4.2.2.ResultsFifteen RCTs conducted between 2011 and 2022 and involving 1,551 patients were identified, with 774 and 777 patients in the experimental and control groups respectively. It was found that the NXT and WM combination was superior to the WM therapy alone in terms of the effective clinical rate (odds ratio [OR] = 4.69, 95% confidence interval [CI] = 2.13-10.30), effective rate in electrocardiogram (OR = 6.92, 95% CI = 3.44-13.92), effective rate in angina (OR = 5.90, 95% CI = 3.04-11.46), left ventricular ejection fraction (mean difference [MD] = 4.94, 95% CI = 2.89-6.99), brain natriuretic peptide (MD = -294.00, 95% CI = -584.60 to -3.39), creatine kinase-MB (MD = -7.82, 95% CI = -13.26 to -2.37), major adverse cardiovascular events (OR = 0.24, 95% CI = 0.14-0.43), maximum platelet aggregation rate (MD = -8.33, 95% CI = -11.64 to -5.01), and Chinese medicine evidence score (OR = 9.79, 95% CI = 3.57-26.85). However, there was no significant difference in cardiac troponin I level reduction (MD = -0.13, 95% CI = 0.35-0.09) or the occurrence of adverse medicine events (OR = 0.92, 95% CI = 0.41-2.05). Meta-regression and subgroup analyses indicated that NXT capsule dosage, treatment duration, and patient baseline characteristics contributed to the heterogeneity.ConclusionA combination of NXT and WM can improve clinical outcomes in patients undergoing PCI. However, further studies are needed to confirm the reliability and safety of this combined treatment approach.Systematic review registrationPROSPERO, https://www.crd.york.ac.uk/PROSPERO/display_record.php?RecordID=369174, Identifier CRD42022369174.

Project description:Thrombosis is a disease that seriously endangers human health, with a high rate of mortality and disability. However, current treatments with thrombolytic drugs (such as recombinant tissue-plasminogen activator) and the oral anticoagulants (such as dabigatran and rivaroxaban) are reported to have a tendency of major or life-threatening bleeding, such as intracranial hemorrhage or massive gastrointestinal bleed with non-specific antidotes. In contrast, lumbrokinase is very specific to fibrin as a substrate and does not cause excessive bleeding. It can dissolve the fibrin by itself or convert plasminogen to plasmin by inducing endogenous t-PA activity to dissolve fibrin clots. Therefore, searching for potentially new therapeutic molecules from earthworms is significant. In this study, we first collected a strong fibrinolytic extract (PvQ) from the total protein of the Pheretima vulgaris with AKTA pure protein purification systems; its fibrinolytic bioactivity was verified by the fibrin plate assay and zebrafish thrombotic model of vascular damage. Furthermore, according to the cell culture model of human umbilical vein endothelial cells (HUVECs), the PvQ was proven to exhibit the ability to promote the secretion of tissue-type plasminogen activator (t-PA), which further illustrated that it has an indirect thrombolytic effect. Subsequently, extensive chromatographic techniques were applied to reveal the material basis of the extract. Fortunately, six novel earthworm fibrinolytic enzymes were obtained from the PvQ, and the primary sequences of those functional proteins were determined by LC-MS/MStranscriptome cross-identification and the Edman degradation assay. The secondary structures of these six fibrinolytic enzymes were determined by circular dichroism spectroscopy and the three-dimensional structures of these proteases were predicted by MODELLER 9.23 based on multi-template modelling. In addition, those six genes encoding blood clot-dissolving proteins were cloned from P. vulgaris by RT-PCR amplification, which further determined the accuracy of proteins primary sequences identifications and laid the foundation for subsequent heterologous expression.

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Specific DNA identification of Pheretima in the Naoxintong capsule.

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Specific DNA identification of Pheretima in the Naoxintong capsule.

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