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DNA walking machines have achieved significant breakthroughs in areas such as biosensing, bioimaging, and early cancer diagnosis, facilitated by the self-assembly of DNA or its combination with other materials, such as magnetic beads and metal nanoparticles. However, current DNA walking machine strategies are constantly challenged by inadequate analytical sensitivity, while sophisticated signal amplification procedures are often indispensable. Single-particle inductively coupled plasma mass spectrometry (SP-ICPMS) provides superior sensitivity and can effectively discriminate between background noise and detected signals due to the large number of metal atoms in a nanoparticle and the concentrating effect of single nanoparticle detection. In this study, we present a novel approach utilizing single nanoparticle counting and duplex-specific nuclease (DSN)-assisted signal amplification to construct a 3D DNA walking machine for detecting the aggressive prostate cancer (PCa) biomarker miRNA-200c. The proposed strategy showed an improvement in sensitivity with a detection limit (LOD) of 0.93 pM (28 amol) and was successfully applied in human serum samples. To the best of our knowledge, this is the first report of the DNA walking machine with single nanoparticle counting study.
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ADN , MicroARNs , Humanos , MicroARNs/análisis , MicroARNs/sangre , ADN/química , Límite de Detección , Neoplasias de la Próstata/diagnóstico , Masculino , Nanopartículas del Metal/química , Espectrometría de MasasRESUMEN
A tapered stent with inclined proximal end is designed for fitting the iliac anatomically. The aim of the present study was to evaluate the safety and performance of the new stent in ovine left iliac veins. The experiment was performed in 30 adult sheep, and one nitinol-based VENA-BT® iliac venous stent (KYD stent) was implanted into each animal's left common iliac vein. Follow-up in all sheep consisted of angiographic, macroscopic, and microscopic examinations at Day 0 (< 24 h), Day 30, Day 90, Day 180 and Day 360 post-stenting (six animals per each time-point). 30 healthy ~ 50 kg sheep were included in this study and randomly divided into five groups according to the follow-up timepoint. All stents were implanted successfully into the left ovine common iliac vein. No significant migration occurred at follow-up. There is no statistically significant difference between the groups (p > 0.05), indicating no serious lumen loss occurred during the follow-up period. Common iliac venous pressure was further measured and the results further indicated the lumen patency at follow-up. Histological examinations indicated that no vessel injury and wall rupture, stent damage, and luminal thrombus occurred. There was moderate inflammatory cell infiltration around the stent in Day-0 and Day-30 groups with the average inflammation score of 2.278 and 2.167, respectively. The inflammatory reaction was significantly reduced in Day-90, Day-180 and Day-360 groups and the average inflammation scores were 0.9444 (p < 0.001, Day-90 vs Day-0), 1.167 (p < 0.001, Day-180 vs Day-0) and 0.667 (p < 0.001, Day-90 vs Day-0), respectively. The microscopic examinations found that the stents were well covered by endothelial cells in all follow-up time points. The results suggested that the KYD stent is feasible and safe in animal model. Future clinical studies may be required to further evaluate its safety and efficacy.
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Aleaciones , Células Endoteliales , Vena Ilíaca , Animales , Vena Ilíaca/diagnóstico por imagen , Vena Ilíaca/cirugía , Inflamación , Estudios Retrospectivos , Ovinos , Stents/efectos adversos , Resultado del Tratamiento , Grado de Desobstrucción VascularRESUMEN
CRISPR (clustered regularly interspaced short palindromic repeats) and CRISPR-associated protein (Cas) have gained extensive applications in bioassays. However, CRISPR-based detection platforms are often hampered by limited analytical sensitivity, while nucleic acid-based amplification strategies are usually indispensable for additional signal enhancement with potential risks of amplification leakages. To address these challenges, an amplification-free CRISPR-based bioassay of aflatoxin B1 (AFB1) was proposed by applying single nanoparticle counting. Single-particle mode inductively coupled plasma mass spectrometry (Sp-ICPMS) has been regarded as a sensitive tool for nanoparticle counting since one nanoparticle can generate considerable signals above backgrounds. With AFB1, activator strands were introduced to initiate the trans-cleavage of CRISPR/Cas12a for cutting the nanoparticles-tagged-magnetic beads, which were transduced to nanoparticle count signals after separation. Finally, a pico-mole level limit-of-detections (LODs) with moderate selectivity was achieved. Certified reference materials (CRMs) analysis and recovery tests were conducted with promising results. To our best knowledge, this is the first report of the single particle counting-based CRISPR/Cas12a biosensing study.
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Técnicas Biosensibles , Nanopartículas , Aflatoxina B1 , Sistemas CRISPR-Cas , Bioensayo , CertificaciónRESUMEN
Tuning the surface chemical property and the local environment of nanocrystals is crucial for realizing a high catalytic performance in various reactions. Herein, we aim to elucidate the structure sensitivity of Pd facets on the surface catalytic hydrogenation reaction and to identify what role the nanoconfinement effect plays in the catalytic properties of Pd nanocrystal catalysts. By controlling the coating structures of mesoporous silica (mSiO2) on Pd nanocrystals with different exposed facets that include {100}, {111}, and {hk0}, we present a series of Pd@mSiO2 nanoreactors in core-shell and yolk-shell structures and the discovery of a partial-coated structure, which can provide different types of nanoconfinement, and we propose a seed size-dominated growth mechanism. We demonstrate that a superior activity was exhibited in Pd nanocrystals enclosed by the {hk0} facet as compared to the Pd{100} and Pd{111} facets, and substantially enhanced efficiency and stability were achieved in Pd@mSiO2 particles with yolk-shell structures, indicating a crucial superiority of optimizing the configuration of crystal facets and nanoconfinement. Our study provides an efficient strategy to rationally design and optimize nanocatalysts for promoting catalytic performance.
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OBJECTIVES: We used single-center data to evaluate the long-term outcome of percutaneous transluminal angioplasty (PTA) for pediatric renovascular hypertension (RVH) and to analyze the factors that influence effectiveness. METHODS: We retrospectively evaluated 33 pediatric RVH patients (18 boys; mean age: 9.1 ± 4 years, range: 2-16) who underwent PTA from January 2007 to December 2019. 15 patients had Takayasu arteritis (TA) and 18 were non-TA. The median follow-up from the initial PTA was 69 months (range: 12-157; IQR: 25.5-89). RESULTS: The technical success rate of 52 PTA procedures was 90.4% in 33 children. Renal artery stents were implanted in two patients, external guidewires were used in two patients, and a drug-coated balloon was used in only one patient. The overall effective rate of PTA was 63.6%, including cured 39.4% and improved 24.2%, at the end of follow-up. Overall clinical outcomes were not statistically different between the TA and non-TA groups (p = 0.316), nor were cure rates (p = 0.072). 15 patients received reintervention due to restenosis after the first successful PTA; the interval was 2-56 months (median: 12 months). Four patients received reintervention due to a failed PTA. A total of four patients received open surgery. Binary logistics regression analysis showed that stenosis length and residual stenosis rate were strongly correlated with effective PTA (p = 0.045, p = 0.044). CONCLUSIONS: As a primary treatment for pediatric RVH, PTA can achieve satisfactory results, which are influenced by lesion length and residual stenosis rate.
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Angioplastia de Balón , Hipertensión Renovascular , Obstrucción de la Arteria Renal , Arteritis de Takayasu , Masculino , Humanos , Niño , Preescolar , Adolescente , Hipertensión Renovascular/diagnóstico por imagen , Hipertensión Renovascular/etiología , Angioplastia de Balón/efectos adversos , Angioplastia de Balón/métodos , Estudios Retrospectivos , Constricción Patológica , Resultado del Tratamiento , Angioplastia/efectos adversos , Obstrucción de la Arteria Renal/diagnóstico por imagen , Obstrucción de la Arteria Renal/terapiaRESUMEN
Precision medicine demands the best application of multiple unambiguous biomarkers to bring uniform decisions in disease prognosis. The remarkable development of heterogeneous immunoassay greatly promotes precision medicine when combined with the biomarker combination strategy. Nevertheless, the cumbersome washing steps in heterogeneous immunoassay have inevitably compromised the accuracy because of the sample losses and nature change of the matrix, challenging the further exploration of a more facile and lower limit-of-detection analysis. The new methodologies with high throughputs and specificity are never out of date to provide simultaneous evaluations and uniform decisions on multiple analytes through a simple process. Herein, we propose a new wash-free immunoassay, named differential assay, for multiplexed biomarker monitoring. The method is based on counting the number difference of unbound nanoparticle tags before and after immunoreactions from a solid support (i.e., magnetic microsphere) by single-particle inductively coupled plasma mass spectrometry (sp-ICP-MS), discarding the tedious washing steps. We primarily explore the proof-of-concept proposal within two types (sandwich and competitive assay), demonstrating the good feasibility for further facile clinical practice. To provide efficient multiplexed evaluations, we synthesized PtNPs with four diameters and screened the most suitable size for efficient differential immunoassay. The wash-free strategy was successfully utilized in simultaneous serological biomarker (CA724, CA199, and CEA) evaluation, with results in good accordance with those measured by the clinical routine method. Potentially, the proposed differential bioassay can be regarded as a more facile and valuable tool in malignancy prognosis and cancer recurrence monitoring.
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Nanopartículas , Neoplasias Gástricas , Biomarcadores de Tumor , Humanos , Inmunoensayo/métodos , Magnetismo , Neoplasias Gástricas/diagnósticoRESUMEN
Taxus trees are major natural sources for the extraction of taxol, an anti-cancer agent used worldwide. Taxus media is a dioecious woody tree with high taxol yield. However, the sexually dimorphic accumulation of taxoids in T. media is largely unknown. Our study revealed high accumulation of taxoids in female T. media trees using a UPLC-MS/MS method. Thereafter, many differential metabolites and genes between female and male T. media trees were identified using metabolomic and transcriptomic analyses, respectively. Most of the taxol-related genes were predominantly expressed in female trees. A female-specific R2R3-MYB transcription factor gene, TmMYB39, was identified. Furthermore, bimolecular fluorescence complementation and yeast two-hybrid assays suggested the potential interaction between TmMYB39 and TmbHLH13. Several taxol biosynthesis-related promoter sequences were isolated and used for the screening of MYB recognition elements. The electrophoretic mobility shift assay indicated that TmMYB39 could bind to the promoters of the GGPPS, T10OH, T13OH, and TBT genes. Interaction between TmMYB39 and TmbHLH13 transactivated the expression of the GGPPS and T10OH genes. TmMYB39 might function in the transcriptional regulation of taxol biosynthesis through an MYB-bHLH module. Our results give a potential explanation for the sexually dimorphic biosynthesis of taxol in T. media.
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Substitution or bypass is the most effective treatment for vascular occlusive diseases.The demand for artificial blood vessels has seen an unprecedented rise due to the limited supply of autologous blood vessels. Tissue engineering is the best approach to provide artificial blood vessels. In this study, a new type of small-diameter artificial blood vessel with good mechanical and biological properties was designed by using electrospinning coaxial fibers. Four groups of coaxial fibers vascular membranes having polyurethane/gelatin core-shell structure were cross-linked by the EDC-NHS system and characterized. The core-shell structure of the coaxial vascular fibers was observed by transmission electron microscope. After the crosslinking, the stress and elastic modulus increased and the elongation decreased, burst pressure of 0.11 group reached the maximum (2844.55 ± 272.65 mmHg) after cross-linking, which acted as the experimental group. Masson staining identified blue-stained ring or elliptical gelatin ingredients in the vascular wall. The cell number in the vascular wall of the coaxial group was found in muscle embedding experiment significantly higher than that of the non-coaxial group at all time points(p < 0.001). Our results showed that the coaxial vascular graft with the ratio of 0.2:0.11 had better mechanical properties (burst pressure reached 2844.55 ± 272.65 mmHg); Meanwhile its biological properties were also outstanding, which was beneficial to cell entry and offered good vascular remodeling performance.Polyurethane (PU); Gelatin (Gel); Polycaprolactone (PCL); polylactic acid (PLA);1-(3-Dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC); N-Hydroxy succinimide (NHS); 4-Morpholine-ethane-sulfonic (MES); phosphate buffered saline (PBS); fetal calf serum (FCS); Minimum Essential Medium (MEM); Dimethyl sulfoxide (DMSO); hematoxylin-eosin (HE).
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Prótesis Vascular , Gelatina/química , Poliuretanos/química , Remodelación Vascular/fisiología , Animales , Línea Celular , Técnicas Electroquímicas , Ensayo de Materiales , Fenómenos Mecánicos , Ratones , Ratas , Ratas WistarRESUMEN
Background: Currently, there is no uniform standard for selecting the left double lumen tubes (LDLT). Advantages, such as safety and convenience of the ultrasonic technology, and measurement accuracy, make it more widely applied in the clinical anesthesia, and computed tomography (CT) multi-planar reconstruction (MPR) technology will certainly provide a more accurate measurement. For better application for thoracic surgery choice LDLT, relieving the injury to patients, and reducing the complications, this study will compare the two approaches. Methods: The first part, 120 cases of patients were selected according to the height and gender; recording the patient's optimum LDLT and measurement the transverse diameter of the cricoid cartilage (TD-C) by ultrasound and CT MPR, and then obtained the TD-C range measurement by ultrasound and CT MPR corresponding to different types of LDLT. The second part, total of 102 patients were divided into the ultrasound group and the CT MPR group. In the ultrasound group, TD-C was measured by ultrasound, the corresponding size for intubation was selected based on the conclusions derived from the first part. In the CT MPR group, TD-C was measured by CT MPR, the corresponding size of LDLT based on the conclusions derived from the first part. Results: In the first part, 120 patients were no significant difference in the basic characteristics (P > 0.05). The accuracy of selecting the LDLT by conventional experience, namely height and gender was 58.3%. Ultrasonic measurement TD-C range was as follows: 32 Fr <15.88, 35 Fr: 15.88-16.80, 37 Fr: 16.75-17.81, and 39 Fr > 17.80. CT MPR measurement TD-C range was as follows: 32 Fr <15.74, 35 Fr: 15.74-16.65, 37 Fr: 16.56-17.68, and 39 Fr > 17.65. In the second part, there was no significant difference in the basic characteristics between the two groups (P > 0.05). The accuracy of intubation in the ultrasound group was 90.2% and the corresponding in the CT MPR group was 94.1% (P > 0.05). Conclusions: The accuracy of selecting the LDLT based on TD-C is significantly higher than conventional experience; it can significantly reduce the post-operative complications and there was no statistical significance in the accuracy of LDLT selected for TD-C measurement by ultrasound vs. CT, and both of them could be safely used for the evaluation before intubation under anesthesia in thoracic surgery.
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BACKGROUND: Taxol is an efficient anticancer drug accumulated in Taxus species. Pseudotaxus chienii is an important member of Taxaceae, however, the level of six taxoids in P. chienii is largely unknown. RESULTS: High accumulation of 10-DAB, taxol, and 7-E-PTX suggested that P. chienii is a good taxol-yielding species for large-scale cultivation. By the omics approaches, a total of 3,387 metabolites and 61,146 unigenes were detected and annotated. Compared with a representative Taxus tree (Taxus yunnanensis), most of the differentially accumulated metabolites and differential expressed genes were assigned into 10 primary and secondary metabolism pathways. Comparative analyses revealed the variations in the precursors and intermediate products of taxol biosynthesis between P. chienii and T. yunnanensis. Taxusin-like metabolites highly accumulated in P. chienii, suggesting a wider value of P. chienii in pharmaceutical industry. CONCLUSIONS: In our study, the occurrence of taxoids in P. chienii was determined. The differential expression of key genes involved in the taxol biosynthesis pathway is the major cause of the differential accumulation of taxoids. Moreover, identification of a number of differentially expressed transcription factors provided more candidate regulators of taxol biosynthesis. Our study may help to reveal the differences between Pseudotaxus and Taxus trees, and promote resource utilization of the endangered and rarely studied P. chienii.
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Vías Biosintéticas , Metaboloma , Metabolómica , Paclitaxel/biosíntesis , Plantas Medicinales/metabolismo , Especificidad de la Especie , Taxaceae/metabolismo , Especies en Peligro de Extinción , Variación GenéticaRESUMEN
OBJECTIVE: This paper retrospectively reviews our experience with endovascular and hybrid treatments for subclavian artery aneurysms (SAA). METHODS: Seventeen patients with SAAs were treated at our center between July 2011 and October 2018. Clinical and follow-up data were extracted from the hospital records and retrospectively reviewed. We routinely use endovascular treatments and stenting or axillary-axillary bypass to treat SAA if vertebral artery blood flow requires restoration. Patients were followed up at 3, 6, and 12 months after their operations and yearly thereafter. RESULTS: Median follow-up was 30.5 months. In 6 patients, the SAAs involved the ipsilateral vertebral artery. Simple coil embolization was performed for 1 patient; endovascular covered stenting and coil embolization for 4 patients; and hybrid treatment for 1 patient. In 11 patients, the SAAs did not involve the ipsilateral vertebral artery. Hybrid treatment was performed for 1 patient; thoracic aortic stent implantation with coil embolization for 1 patient; and covered stent placement in the subclavian artery for 9 patients. Among the 9 patients who were symptomatic at presentation, 8 had relief of symptoms. Leakage was observed in 1 patient, but it stopped spontaneously. Stent occlusion occurred in 18.8% (3/16), but all were asymptomatic, and no interventions were necessary. CONCLUSION: Endovascular and hybrid treatments appear to be effective for SAAs with few complications and good clinical outcomes. KEY WORDS: Covered stent, Coil embolization, Endovascular treatment, Hybrid, Subclavian artery aneurysm.
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Aneurisma , Procedimientos Endovasculares , Aneurisma/cirugía , Humanos , Estudios Retrospectivos , Arteria Subclavia/cirugía , Resultado del TratamientoRESUMEN
KEY MESSAGE: We employed both metabolomic and transcriptomic approaches to explore the accumulation patterns of physalins, flavonoids and chlorogenic acid in Physalis angulata and revealed the genes associated with the biosynthesis of bioactive compounds under methyl-jasmonate (MeJA) treatment. Physalis angulata L. is an annual Solanaceae plant with a number of medicinally active compounds. Despite the potential pharmacological benefits of P. angulata, the scarce genomic information regarding this plant has limited the studies on the mechanisms of bioactive compound biosynthesis. To facilitate the basic understanding of the main chemical constituent biosynthesis pathways, we performed both metabolomic and transcriptomic approaches to reveal the genes associated with the biosynthesis of bioactive compounds under methyl-jasmonate (MeJA) treatment. Untargeted metabolome analysis showed that most physalins, flavonoids and chlorogenic acid were significantly upregulated. Targeted HPLC-MS/MS analysis confirmed variations in the contents of two important representative steroid derivatives (physalins B and G), total flavonoids, neochlorogenic acid, and chlorogenic acid between MeJA-treated plants and controls. Transcript levels of a few steroid biosynthesis-, flavonoid biosynthesis-, and chlorogenic acid biosynthesis-related genes were upregulated, providing a potential explanation for MeJA-induced active ingredient synthesis in P. angulata. Systematic correlation analysis identified a number of novel candidate genes associated with bioactive compound biosynthesis. These results may help to elucidate the regulatory mechanism underlying MeJA-induced active compound accumulation and provide several valuable candidate genes for further functional study.
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Acetatos/farmacología , Ciclopentanos/farmacología , Regulación de la Expresión Génica de las Plantas/efectos de los fármacos , Oxilipinas/farmacología , Physalis/efectos de los fármacos , Physalis/metabolismo , Proteínas de Plantas/metabolismo , Flavonoides/biosíntesis , Flavonoides/química , Metaboloma , Estructura Molecular , Reguladores del Crecimiento de las Plantas/farmacología , Proteínas de Plantas/genética , Raíces de Plantas/genética , Raíces de Plantas/metabolismo , ARN de Planta/genética , TranscriptomaRESUMEN
Taxus stem barks can be used for extraction of paclitaxel. However, the composition of taxoids across the whole stem and the stem tissue-specificity of paclitaxel biosynthesis-related enzymes remain largely unknown. We used cultivated Taxus media trees for analyses of the chemical composition and protein of major stem tissues by an integrated metabolomic and proteomic approach, and the role of TmMYB3 in paclitaxel biosynthesis was investigated. The metabolomic landscape analysis showed differences in stem tissue-specific accumulation of metabolites. Phytochemical analysis revealed that there is high accumulation of paclitaxel in the phloem. Ten key enzymes involved in paclitaxel biosynthesis were identified, most of which are predominantly produced in the phloem. The full-length sequence of TmMYB3 and partial promoter sequences of five paclitaxel biosynthesis-related genes were isolated. Several MYB recognition elements were found in the promoters of TBT, DBTNBT and TS. Further in vitro and in vivo investigations indicated that TmMYB3 is involved in paclitaxel biosynthesis by activating the expression of TBT and TS. Differences in the taxoid composition of different stem tissues suggest that the whole stem of T. media has potential for biotechnological applications. Phloem-specific TmMYB3 plays a role in the transcriptional regulation of paclitaxel biosynthesis, and may explain the phloem-specific accumulation of paclitaxel.
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Paclitaxel/biosíntesis , Floema/metabolismo , Proteínas de Plantas/metabolismo , Tallos de la Planta/metabolismo , Taxus/metabolismo , Factores de Transcripción/metabolismo , Regulación de la Expresión Génica de las Plantas/genética , Redes y Vías Metabólicas , Metabolómica , Proteínas de Plantas/fisiología , Regiones Promotoras Genéticas , Proteómica , Factores de Transcripción/fisiologíaRESUMEN
BACKGROUND: Trees of the genus Taxus are highly valuable medicinal plants with multiple pharmacological effects on various cancer treatments. Paclitaxel from Taxus trees is an efficient and widely used anticancer drug, however, the accumulation of taxoids and other active ingredients can vary greatly among Taxus species. In our study, the metabolomes of three Taxus species have been investigated. RESULTS: A total of 2246 metabolites assigned to various primary and secondary metabolic pathways were identified using an untargeted approach. Analysis of differentially accumulated metabolites identified 358 T. media-, 220 T. cuspidata-, and 169 T. mairei-specific accumulated metabolites, respectively. By searching the metabolite pool, 7 MEP pathway precursors, 11 intermediates, side chain products and derivatives of paclitaxel, and paclitaxel itself were detected. Most precursors, initiated intermediates were highly accumulated in T. mairei, and most intermediate products approaching the end point of taxol biosynthesis pathway were primarily accumulated in T. cuspidata and T. media. Our data suggested that there were higher-efficiency pathways to paclitaxel in T. cuspidata and T. media compared with in T. mairei. As an important class of active ingredients in Taxus trees, a majority of flavonoids were predominantly accumulated in T. mairei rather than T. media and T. cuspidata. The variations in several selected taxoids and flavonoids were confirmed using a targeted approach. CONCLUSIONS: Systematic correlativity analysis identifies a number of metabolites associated with paclitaxel biosynthesis, suggesting a potential negative correlation between flavonoid metabolism and taxoid accumulation. Investigation of the variations in taxoids and other active ingredients will provide us with a deeper understanding of the interspecific differential accumulation of taxoids and an opportunity to accelerate the highest-yielding species breeding and resource utilization.
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Flavonoides/metabolismo , Metaboloma , Taxoides/metabolismo , Taxus/metabolismo , Redes y Vías Metabólicas , Metabolómica , Especificidad de la EspecieRESUMEN
BACKGROUND: Taxol is an efficient anticancer drug; however, the accumulation of taxoids can vary hugely among Taxus species. The mechanism underlying differential accumulation of taxoids is largely unknown. Thus, comparative analysis of the transcriptomes in three Taxus species, including T. media, T. mairei and T. cuspidata, was performed. RESULTS: KEGG enrichment analysis revealed that the diterpenoid biosynthesis and cytochrome P450 pathways were significantly enriched in different comparisons. Differential expressions of these taxol biosynthesis related genes might be a potential explanation for the interspecific differential accumulation of taxol and its derivatives. Besides, the sequences of several MEP pathway-associated genes, such as DXS, DXR, MCT, CMK, MDS, HDS, HDR, IPPI, and GGPPS, were re-assembled based on independent transcriptomes from the three Taxus species. Phylogenetic analysis of these MEP pathway-associated enzymes also showed a high sequence similarity between T. media and T. cuspidata. Moreover, 48 JA-related transcription factor (TF) genes, including 10 MYBs, 5 ERFs, 4 RAPs, 3 VTCs, and 26 other TFs, were analyzed. Differential expression of these JA-related TF genes suggested distinct responses to exogenous JA applications in the three Taxus species. CONCLUSIONS: Our results provide insights into the expression pattern and sequence similarity of several taxol biosynthesis-related genes in three Taxus species. The data give us an opportunity to reveal the mechanism underlying the variations in the taxoid contents and to select the highest-yielding Taxus species.
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Perfilación de la Expresión Génica/métodos , Paclitaxel/biosíntesis , Taxus/genética , Taxus/metabolismo , Transcriptoma/genética , Taxoides/metabolismoRESUMEN
This study was to investigate the effect of vinegar processing on esculentosides in n-BuOH fraction and the contents of the main toxic components esculentoside B (EsB) and esculentoside C (EsC) in Phytolaccae Radix pieces. n-BuOH fraction of Phytolaccae Radix pieces was processed with vinegar according to the processing method in Chinese Pharmacopoeia. HPLC-MS-MS was adopted to analyze the esculentosides composition changes in n-BuOH fraction before and after vinegar processing. HPLC-ELSD was used to detect EsC and EsB contents in raw and vinegar processed Phytolaccae Radix pieces, and investigate the content changes before and after vinegar processing. Results showed that the esculentosides contents in n-BuOH fraction were significantly decreased except esculentoside A (EsA); there were significant changes in saponins compositions, but no new compounds were generated in n-BuOH fraction after vinegar processing. The contents of EsC and EsB were 0.12% and 0.20% respectively in raw Phytolaccae Radix, and decreased to 0.048% and 0.094% accordingly after vinegar processing. It showed that vinegar processing could significantly change the composition of esculentosides in n-BuOH fraction from Phytolaccae Radix and reduce the contents of toxic components EsC and EsB, indicating the scientificity of vinegar processing for Phytolaccae Radix.
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Ácido Acético/química , Medicamentos Herbarios Chinos/toxicidad , Ácido Oleanólico/análogos & derivados , Phytolaccaceae/química , Química Farmacéutica , Cromatografía Líquida de Alta PresiónRESUMEN
The aim of the present study was to determine the toxic targets of proteins from Croton tiglium L. and to investigate the potential mechanism of their toxicity. The toxic targets were determined by oral medication and intraperitoneal injection. The median lethal dose of oral medication in mice was calculated using Bliss software (2,752.8-3,407.5 mg/kg), and that of intraperitoneal injection was 195.8272.69 mg/kg. The results of histopathological examination demonstrated that the kidney was primarily impaired by intraperitoneal injection, with slight degeneration of renal tubular epithelial cells. As to oral medication, the digestive tract was primarily injured, which manifested as congestion, bleeding, serious edema and other symptoms. Oral administration of the proteins caused gastrointestinal edema by increasing the intestinal permeability. Severe edema was associated with the inflammatory response, therefore the association between the toxicity of the proteins and inflammation was investigated. The proinflammatory effects of the crude proteins on the release of inflammatory mediator prostaglandin E2 (PGE2) were evaluated through intraperitoneal injection and the production of proinflammatory cytokines in RAW264.7 macrophages. Maximum PGE2 was released in the mice in vivo following intraperitoneal injection with 400 mg crude protein/kg body weight. Proinflammatory cytokines in macrophages, including tumor necrosis factorα and interleukin1ß, were produced in dose and timedependent manners in vitro. furthermore, the expressions of cell signaling molecules were detected by western blotting. The inflammatory response induced by crude protein in macrophages was associated with the mitogenactivated protein kinase (MAPK) signaling pathway mainly including p38MAPK, extracellular signalregulated kinase 1/2 and cJun Nterminal kinase 1/2/3 and the activated p38MAPK signaling pathway. However, extracellular signalregulated kinase 1/2 and cJun Nterminal kinases 13 exhibited no significant response.
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Croton/química , Citocinas/metabolismo , Mediadores de Inflamación/metabolismo , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Extractos Vegetales/farmacología , Proteínas de Plantas/farmacología , Animales , Biomarcadores , Relación Dosis-Respuesta a Droga , Femenino , Mucosa Intestinal/efectos de los fármacos , Mucosa Intestinal/metabolismo , Mucosa Intestinal/patología , Dosificación Letal Mediana , Macrófagos/efectos de los fármacos , Macrófagos/metabolismo , Masculino , Ratones , Permeabilidad/efectos de los fármacos , Extractos Vegetales/administración & dosificación , Extractos Vegetales/toxicidad , Proteínas de Plantas/administración & dosificación , Proteínas de Plantas/toxicidad , Células RAW 264.7 , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismoRESUMEN
By using a typical component in traditional Chinese medicine Pericarpium Arecae (PA), quantitative analysis of multi-components by single-marker (QAMS) was performed to determine the contents of four alkaloids. With a column packed with strong cation exchange bonded silica particles, the alkaloids were well separated, showing linear relationships within certain ranges. The limit of detection, limit of quantitation, precision, stability, repeatability and recovery all met requirements. By employing arecoline as internal standard, relative correction factors for arecaidine, guvacine and guvacoline at five concentrations were detected with three HPLC systems and three HPLC columns. The peaks of arecaidine, guvacine and guvacoline were positioned, during which the columns with the same packing materials from different manufacturers significantly affected relative retention values and retention time differences of the alkaloids. However, the columns, from different batches, managed to give relative retention values satisfying the requirements of HPLC peak positioning. The Thermo Fisher Scientific column packed with strong cation exchange bonded silica particles was finally selected by considering resolution and peak time. Compared with the external standard method, QAMS detected the alkaloid contents in 12 PA samples more accurately and reliably. The results provide valuable evidence for content determination and quality control of alkaloids in PA.
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Alcaloides/análisis , Areca/química , Cromatografía Líquida de Alta Presión/métodos , Arecolina/análogos & derivados , Arecolina/análisis , Límite de Detección , Ácidos Nicotínicos/análisis , Reproducibilidad de los ResultadosRESUMEN
To research the intestinal toxicity of n-BuOH fraction in Phytolacca Radix before and after being processed with vinegar. Toxic n-BuOH fractions were separated from Phytolacca Radix. In the animal model, the level of intestinal edema, water content of intestine and stool, IC50 values of HT-29 and IEC-6 were detected with MTT method to compare the changes in toxicity of n-BuOH fractions from Phytolacca Radix before and after being processed with vinegar. n-BuOH fractions of Phytolacca Radix could cause intestinal edema in mice, increase the edema of duodenum, jejunum and the water content in stool, inhibit the proliferation of HT-29 cells and IEC-6 cells, indicating its intestinal toxicity, with HT-29 IC50 at 14.59 mgâ¢L⻹ and IEC-6 IC50 at 43.77 mgâ¢L⻹. After being processed with vinegar, the level of intestinal edema, edema of duodenum and jejunum and the water content in stool and inhibition ratio of cells line were reduced, with HT-29 IC50 at 58.51 mgâ¢L⻹ and IEC-6 IC50 at 84.37 mgâ¢L⻹. After being processed with vinegar, the toxicity of n-BuOH fractions from Phytolacca Radix decreased obviously.
Asunto(s)
Medicamentos Herbarios Chinos/administración & dosificación , Medicamentos Herbarios Chinos/aislamiento & purificación , Edema/tratamiento farmacológico , Phytolacca/química , Ácido Acético/química , Animales , Química Farmacéutica , Medicamentos Herbarios Chinos/química , Edema/inmunología , Humanos , Intestinos/efectos de los fármacos , Intestinos/inmunología , Masculino , Ratones , Ratones Endogámicos ICR , Raíces de Plantas/químicaRESUMEN
This study was to investigate the mechanism of gingerols antagonizing the inflammatory effect of toxic raphides from Pinella pedatisecta. Mice peritonitis models induced by toxic raphides from P. pedatisecta were applied to observe the effect of gingerols on inflammatory mediators PGE2 in the exudates of abdominal inflammation in mice; rats peritoneal macrophage in vitro culture models were adopted to study the anti-inflammatory effects of gingerol against toxic raphides, with TNF-α and IL-1ß in supernatant as indexes. Scanning electron microscopy was used to observe the changes in surface morphology of macrophages treated by raphides and gingerols. Macrophages-neutrophils co-cultured models were used to study the antagonism of gingerols against the effect of toxic raphides' stimulation on neutrophils migration. Results showed that gingerols could significantly inhibit the production of PGE2 in the exudates of abdominal inflammation induced by toxic raphides from P. pedatisecta in mice. Gingerols could significantly inhibit the toxic raphides from P. pedatisecta to induce the release of inflammatory factors, with certain dose dependence. Scanning electron microscopy showed that gingerols could significantly inhibit phagocytosis of macrophages, cytomembrane injury, and neutrophils migration induced by toxic raphides from P. pedatisecta. The results showed that the antagonism mechanism of gingerols against the toxic raphides from P. pedatisecta may be associated with inhibiting the pro-inflammatory toxicity including macrophage activation, inflammatory factors release, and neutrophils migration.