Single Particle Analysis-Enhanced DNA Walking Machine for Sensitive miRNA Detection.

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.

Long-term outcomes of angioplasty for pediatric renovascular hypertension: A single-center experience.

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.

Single-Nanoparticle Differential Immunoassay for Multiplexed Gastric Cancer Biomarker Monitoring.

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.

Role of female-predominant MYB39-bHLH13 complex in sexually dimorphic accumulation of taxol in Taxus media.

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.

Prediction of Left Double-Lumen Tube Size by Measurement of Cricoid Cartilage Transverse Diameter by Ultrasound and CT Multi-Planar Reconstruction.

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.

Omic analysis of the endangered Taxaceae species Pseudotaxus chienii revealed the differences in taxol biosynthesis pathway between Pseudotaxus and Taxus yunnanensis trees.

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.

Comparative metabolomic analysis reveals the variations in taxoids and flavonoids among three Taxus species.

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.

Transcriptome analyses provide insights into the expression pattern and sequence similarity of several taxol biosynthesis-related genes in three Taxus species.

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.

Toxic proteins from Croton tiglium L. exert a proinflammatory effect by inducing release of proinflammatory cytokines and activating the p38-MAPK signaling pathway.

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.

[Antagonism mechanism of gingerols against inflammatory effect of toxic raphides from Pinella pedatisecta].

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.

Insult of gastroesophageal reflux on airway: clinical significance of pharyngeal nozzle.

At the very time of global paying the highest attention to the worst insults of smoking as well as haze on the airway, everybody knows both are exogenous and noticeable. However, people mostly, including many medical personnel, do not know how badly the gastroesophageal reflux (GER) insults on our own airway. Symptoms of GER are commonly seen as heartburn and regurgitation, which can be mostly tolerated. However, when the up going gastric content reversely passes the esophagus and then the distal pharynx, where it appears a beak like stricture, serving as a nozzle, so as to produce numerous micro-particles and reach the oro-nasal cavity and also the airway causing allergic rhinitis and asthmatic attacks, even pulmonary parenchyma lesions. It will reduce life quality or even jeopardize life. The point that the endogenous insult appears in the respiratory system, but originates from the digestive tract is not well known and often undiagnosed and not correctly treated. The GER induced airway challenge is a treatable and preventive entity, as soon as a diagnosis is made, a good relief could be expected by means of life style adjustment, medicine, or fixation of the patulous cardia through radiofrequency or fundoplication. The author Dr. Zhonggao Wang had suffered it for long and symptoms disappeared for 8 years after anti-reflux surgery. Here is a presentation of Dr. Zhonggao Wang and his team's work and would call attention to the public so as to recognize this relatively unknown entity - a treatable condition occurring from human itself, but not from outside surroundings as smoking or haze does.

The laparoscopic nissen fundoplication eliminates obstructive sleep apnea syndrome due to gastroesophageal reflux disease.

Gastroesophageal reflux disease may produce esophageal syndromes, such as heartburn and regurgitation. It is a common clinical presentation with extraesophageal manifestations, such as asthma, arrhythmia, snoring, and sleep disturbance, which could make identifying it more difficult than the usual esophageal symptoms. The aim of this study is to characterize the extraesophageal manifestations in patients with gastroesophageal reflux disease and investigate the effect of laparoscopic Nissen fundoplication. We describe the case of a 38-year-old male patient with a history of sleep disturbance attributable to gastroesophageal reflux disease, which resolved on successful laparoscopic Nissen fundoplication treatment. The long-standing sleep apnea obviously improved after laparoscopic Nissen fundoplication treatment. To our knowledge, this is a rare case of successful laparoscopic Nissen fundoplication treatment of a patient with extraesophageal manifestations induced by gastroesophageal reflux disease. The results indicate an underlying mechanism for extraesophageal manifestations and the success of laparoscopic Nissen fundoplication treatment.

A preliminary investigation of laparoscopic fundoplication treatment on gastroesophageal reflux disease-related respiratory symptoms.

BACKGROUND: Extraesophageal syndromes are more difficult to identify and treat than the usual esophageal symptoms. The current study explores the efficacy of laparoscopic Nissen fundoplication (LNF) on gastroesophageal reflux disease (GERD)-related respiratory symptoms (RSs) during a 12-month follow-up observation. METHODS: From April 2008 to September 2009, LNF was performed on 198 patients (107 men and 91 women) with GERD-related RSs according to underlying esophageal motility. A questionnaire form ranging from 0 to 5 was the basic requirement for recording pretreatment and posttreatment and for making detailed evaluation of the symptoms. All scores of GERD-related RSs, such as heartburn, regurgitation, coughing, breathe holding, wheezing, shortness of breath, and choking, significantly decreased at the 12th month. RESULTS: All the patients who participated in the current study were adults and elders aged 22 to 84 years with a mean age of 49 ± 12.89. The median length of stay was 4.3 days with a range of 2 to 8 days. The median score of heartburn, regurgitation, coughing, wheezing, shortness of breath, choking, and chest pain decreased from 4.92 ± 1.99, 4.98 ± 1.81, 7.23 ± 1.87, 7.50 ± 1.88, 5.83 ± 2.13, 5.94 ± 2.22, and 4.92 ± 1.88 to 1.62 ± 2.33, 0.64 ± 1.43, 2.79 ± 2.82, 2.53 ± 2.96, 1.37 ± 2.10, 1.28 ± 2.09, and 1.57 ± 2.55 (P<0.01), respectively. A total of 173 patients had various relieved symptom scores, 16 patients (8.1%) had different scale recurrence of symptoms after laparoscopic fundoplication treatment, and 13 patients had to retreat to omeprazole as an auxiliary medical therapy. Three other patients rejected any therapy, and no deaths occurred. A single patient converted from laparoscopic surgery to open surgery. Several short-term symptoms included retrosternal uneasiness or pain (n = 63; 31.8%), dysphagia (n = 45; 22.7%), abdominal distension (n = 87; 43.9%), and diarrhea (n = 23; 11.6%). Early dysphagia lasting <6 weeks was common, and 45 patients (22.7%) underwent an early esophagogastroduodenoscopy or contrast swallow. Five patients (2.5%) who had prolonged dysphagia during the 6-month clinical review required esophageal dilatation, and the outcomes were successful. CONCLUSIONS: LNF can be an effective means for treating RSs in patients with GERD.

Radiofrequency treatment on respiratory symptoms due to gastroesophageal reflux disease.

BACKGROUND: Diagnosis and treatment for respiratory symptoms (RSs) of gastroesophageal reflux disease (GERD) is more difficult than that for common esophageal symptoms. The goal of this study was to evaluate the efficacy and safety of radiofrequency (RF) treatment on RSs of GERD in a preliminary 12-month follow-up observation. METHODS: From April 2006 to October 2008, 505 GERD patients with mainly respiratory presentations such as wheezing, chronic cough or hoarseness, were treated by endoscopic RF. A questionnaire was completed before and after treatment, using a six-point scale ranging from 0 to 5 to assess symptom severity and frequency. The symptom score was the sum of frequency and severity. RESULTS: Symptom scores were significantly improved at the end of the follow-up period. The mean heartburn score decreased from 5.31 to 1.79. The mean regurgitation score decreased from 5.02 to 1.64; mean cough score decreased from 6.77 to 2.85; mean wheezing score decreased from 7.83 to 3.07; and mean hoarseness score decreased from 5.13 to 1.81 (P < 0.01). No major complications or deaths occurred. Minor complications included temporary post-procedural retrosternal unease or pain (n = 106; 21.0%), mild fever (n = 86; 17.0%), transient nausea/vomiting (n = 97; 19.2%), and transient dysphagia (n = 42; 9.3%). Thirty-five (6.9%) patients had recurrence of symptoms. Endoscopic RF treatment was repeated in six patients, and laparoscopic fundoplication was performed in seven. CONCLUSION: Endoscopic RF is an effective and safe means to treat RSs in patients with GERD.