Amphiphilic NLC-Gel formulation loaded with Sebacoyl dinalbuphine ester and Nalbuphine for localized postoperative pain management.

Opioids are powerful analgesics; however, their significant systemic adverse effects and the need for frequent administration restrict their use. Nalbuphine (NA) is a κ-agonist narcotic with limited adverse effects, but needs to be frequently administrated due to its short elimination half-life. Whereas sebacoyl dinalbuphine ester (SDE) is a NA prodrug, which can effectively prolong the analgesic effect, but lacks immediate pain relief. Therefore, in this study, a rapid and sustained local delivery formulation to introduce NA and SDE directly into surgical sites was developed. An amphiphilic nanostructured lipid carrier (NLC) poloxamer 407 (P407) gel (NLC-Gel) was developed to permit concurrent delivery of hydrophobic SDE from the NLC core and hydrophilic NA from P407, offering a dual rapid and prolonged analgesic effect. Benefiting from the thermal-sensitive characteristic of P407, the formulation can be injected in liquid phase and instantly transit into gel at wound site. NLC-Gel properties, including particle size, drug release, rheology, and stability, were assessed. In vivo evaluation using a rat spinal surgery model highlighted the effect of the formulation through pain behavior test and hematology analysis. NLC-Gels demonstrated an analgesic effect comparable with that of commercial intramuscular injected SDE formulation (IM SDE), with only 15 % of the drug dosage. The inclusion of supplemental NA in the exterior gel (PA12-Gel + NA) provided rapid drug onset owing to swift NA dispersion, addressing acute pain within hours along with prolonged analgesic effects. Our findings suggest that this amphiphilic formulation significantly enhanced postoperative pain management in terms of safety and efficacy.

Massively parallel CRISPR-assisted homologous recombination enables saturation editing of full-length endogenous genes in yeast.

Performing saturation editing of chromosomal genes will enable the study of genetic variants in situ and facilitate protein and cell engineering. However, current in vivo editing of endogenous genes either lacks flexibility or is limited to discrete codons and short gene fragments, preventing a comprehensive exploration of genotype-phenotype relationships. To enable facile saturation editing of full-length genes, we used a protospacer adjacent motif-relaxed Cas9 variant and homology-directed repair to achieve above 60% user-defined codon replacement efficiencies in Saccharomyces cerevisiae genome. Coupled with massively parallel DNA design and synthesis, we developed a saturation gene editing method termed CRISPR-Cas9- and homology-directed repair-assisted saturation editing (CHASE) and achieved highly saturated codon swapping of long genomic regions. By applying CHASE to massively edit a well-studied global transcription factor gene, we found known and unreported genetic variants affecting an industrially relevant microbial trait. The user-defined codon editing capability and wide targeting windows of CHASE substantially expand the scope of saturation gene editing.

Twice-daily rivaroxaban after percutaneous left atrial appendage closure for atrial fibrillation.

Background and aim: Rivaroxaban is an emerging oral anticoagulant for postoperative anticoagulation after percutaneous left atrial appendage closure (LAAC). Because a once-daily dosing regimen of rivaroxaban causes fluctuations in the drug plasma concentration, we studied the feasibility and safety of twice-daily rivaroxaban as a postoperative anticoagulation regimen for patients with atrial fibrillation (AF) undergoing LAAC. Methods: This study involved patients with AF who underwent LAAC and took rivaroxaban postoperatively. A total of 326 patients who received a standard total dose (15 or 20 mg) of rivaroxaban based on their creatinine clearance rate were divided into the twice-daily (BID) rivaroxaban group (n = 208) and once-daily (QD) rivaroxaban group (n = 118) according to their anticoagulation strategy. Transesophageal echocardiography was recommended at 3-6 months postoperatively to check for device-related thrombosis (DRT). Clinical outcomes were evaluated during postoperative anticoagulation. Results: The median CHA2DS2-VASc score (4 [3, 5] vs. 4 [3, 5], p = 0.28) and HAS-BLED score (2 [2, 3] vs. 2 [2, 3], p = 0.48) were not significantly different between the groups. During the anticoagulation period (4.1 ± 0.7 vs. 4.1 ± 0.9 months, p = 0.58), 148 (71.2%) patients in the BID group and 75 (63.6%) in the QD group underwent follow-up transesophageal echocardiography. There were no statistically significant differences between the two groups in terms of DRT (1.4% vs. 2.7%, p = 0.60), minor bleeding (8.2% vs. 11.0%, p = 0.39), thromboembolic events (1.0% vs. 0.8%, p = 1.00), major bleeding (0.5% vs. 0.8%, p = 1.00), or death. Conclusion: A short course of twice-daily rivaroxaban following LAAC is a feasible alternative regimen with a low rate of major bleeding events, DRT, and thromboembolic events for patients with AF.

Highly efficient visible-light driven dye degradation via 0D BiVO4 nanoparticles/2D BiOCl nanosheets p-n heterojunctions.

The construction of hybrid heterojunction photocatalysts is an effective strategy to improve the utilization of photogenerated carriers and photocatalytic activity. To enhance the separation distance of photogenerated carriers and accelerate the effective separation at the heterojunction of the interface, a unique 0D-2D hierarchical nanostructured p-n heterojunction was successfully fabricated in this work. BiOCl (BOC) nanosheets (p-type) were in situ grown on BiVO4 (BVO) nanoparticles (n-type) using the microemulsion-calcination method for highly efficient visible-light-driven organic dye degradation. Compared with pure BVO (the degradation rate of rhodamine B (RhB): about 32.0% in 55 min, the mineralization rate: 24.9% in 120 min), the RhB degradation rate can reach about 99.5% in 55 min and the mineralization rate of 62.1% in 120 min by utilizing BVO/25%BOC heterojunction photocatalyst under visible light irradiation. Various characterizations demonstrate that the formation of BVO/BOC p-n heterojunction greatly facilitates photogenerated carriers separation efficiency. Meanwhile, the results of the scavenging experiments and electron spin resonance tests indicate that ·O2- and h+ are the prominent active species for Rh B degradation. In addition, possible degradation pathways for Rh B were proposed using LC-MS tests. This work proves that building low dimensional p-n heterojunction photocatalysts is a promising strategy for developing photocatalysts with high efficiency.

Predicting long-term time to cardiovascular incidents using myocardial perfusion imaging and deep convolutional neural networks.

Myocardial perfusion imaging (MPI) is a clinical tool which can assess the heart's perfusion status, thereby revealing impairments in patients' cardiac function. Within the MPI modality, the acquired three-dimensional signals are typically represented as a sequence of two-dimensional grayscale tomographic images. Here, we proposed an end-to-end survival training approach for processing gray-scale MPI tomograms to generate a risk score which reflects subsequent time to cardiovascular incidents, including cardiovascular death, non-fatal myocardial infarction, and non-fatal ischemic stroke (collectively known as Major Adverse Cardiovascular Events; MACE) as well as Congestive Heart Failure (CHF). We recruited a total of 1928 patients who had undergone MPI followed by coronary interventions. Among them, 80% (n = 1540) were randomly reserved for the training and 5- fold cross-validation stage, while 20% (n = 388) were set aside for the testing stage. The end-to-end survival training can converge well in generating effective AI models via the fivefold cross-validation approach with 1540 patients. When a candidate model is evaluated using independent images, the model can stratify patients into below-median-risk (n = 194) and above-median-risk (n = 194) groups, the corresponding survival curves of the two groups have significant difference (P < 0.0001). We further stratify the above-median-risk group to the quartile 3 and 4 group (n = 97 each), and the three patient strata, referred to as the high, intermediate and low risk groups respectively, manifest statistically significant difference. Notably, the 5-year cardiovascular incident rate is less than 5% in the low-risk group (accounting for 50% of all patients), while the rate is nearly 40% in the high-risk group (accounting for 25% of all patients). Evaluation of patient subgroups revealed stronger effect size in patients with three blocked arteries (Hazard ratio [HR]: 18.377, 95% CI 3.719-90.801, p < 0.001), followed by those with two blocked vessels at HR 7.484 (95% CI 1.858-30.150; p = 0.005). Regarding stent placement, patients with a single stent displayed a HR of 4.410 (95% CI 1.399-13.904; p = 0.011). Patients with two stents show a HR of 10.699 (95% CI 2.262-50.601; p = 0.003), escalating notably to a HR of 57.446 (95% CI 1.922-1717.207; p = 0.019) for patients with three or more stents, indicating a substantial relationship between the disease severity and the predictive capability of the AI for subsequent cardiovascular inciidents. The success of the MPI AI model in stratifying patients into subgroups with distinct time-to-cardiovascular incidents demonstrated the feasibility of proposed end-to-end survival training approach.

Cepharanthine synergizes with photodynamic therapy for boosting ROS-driven DNA damage and suppressing MTH1 as a potential anti-cancer strategy.

OBJECTIVE: Photodynamic therapy (PDT) primarily treats skin diseases or cancer by generating reactive oxygen species (ROS) to damage cellular DNA, yet drug resistance limits its application. To tackle this problem, the present study was carried out to improve the efficacy of chlorin e6 (Ce6)-PDT using Cepharanthine (CEP) as well as to reveal the potential molecular mechanism. MATERIALS AND METHODS: Lewis lung cancer cell line (LLC) was utilized as the cancer cell model. chlorin e6 (Ce6) acted as the photosensitizer to induce PDT. The in vitro anti-cancer efficacy was measured by CCK-8, Annexin-V/PI staining, and migration assay. The Ce6 uptake was observed using flow cytometry and confocal microscopy. The ROS generation was detected by the DCFH-DA probe. The analysis of MutT Homolog 1 (MTH1) expression, correlation, and prognosis in databases was conducted by bioinformatic. The MTH1 expression was detected through western blots (WB). DNA damage was assayed by WB, immunofluorescent staining, and comet assay. RESULTS: Ce6-PDT showed robust resistance in lung cancer cells under certain conditions, as evidenced by the unchanged cell viability and apoptosis. The subsequent findings confirmed that the uptake of Ce6 and MTH1 expression was enhanced, but ROS generation with laser irradiation was not increased in LLC, which indicated that the ROS scavenge may be the critical reason for resistance. Surprisingly, bioinformatic and in vitro experiments identified that MTH1, which could prevent the DNA from damage of ROS, was highly expressed in lung cancer and thereby led to the poor prognosis and could be further up-regulated by Ce6 PDT. CEP exhibited a dose-dependent suppressive effect on the lung cancer cells. Further investigations presented that CEP treatment boosted ROS production, thereby resulting in DNA double-strand breakage (DDSB) with activation of MTH1, indicating that CEP facilitated Ce6-PDT-mediated DNA damage. Finally, the combination of CEP and Ce6-PDT exhibited prominent ROS accumulation, MTH1 inhibition, and anti-lung cancer efficacy, which had synergistic pro-DNA damage properties. CONCLUSION: Collectively, highly expressed MTH1 and the failure of ROS generation lead to PDT resistance in lung cancer cells. CEP facilitates ROS generation of PDT, thereby promoting vigorous DNA damage, inactivating MTH1, alleviating PDT resistance, and ameliorating the anti-cancer efficacy of Ce6-PDT, provides a novel approach for augmented PDT.

Toxic effects and molecular mechanisms of estuarian crustaceans (Scylla paramamosain) exposed to five commonly used benzophenones.

Benzophenones (BPs) are commonly used in personal care products like sunscreens and are increasingly being released into the environment, raising concerns about their potential ecotoxic effects. BPs as emerging environmental contaminants, little is known about their toxic effects on estuarine organisms. This study firstly investigated the toxic effects of five commonly used BPs on mud crabs (Scylla paramamosain). The crabs were exposed to varying concentrations of BPs for 14 days. The results showed that BPs caused damage to antioxidant systems in crabs. Transcriptome sequencing revealed that BP-3 and BP-1 had a greater impact on the crabs compared to the other BPs. Specifically, BP-1 and BP-3 caused severe damage to organelles and ribosomes. BP affected catalytic activity and hydrolase activity, BP-2 affected phosphoenolpyruate carboxykinase activity, and BP-4 affected tRNA aminoacylation and hydrolase activity. These findings can enhance our understanding of the ecotoxicity of BPs and may help to protect estuarine ecosystems.

The protective effect of LCZ696 in coxsackievirus B3-induced acute viral myocarditis mice.

AIMS: Acute viral myocarditis (AVMC) is the aetiology of heart failure (HF) with few specific treatments. The improvement of left ventricular ejection fraction (LVEF) is a critical predictor for the prognosis of AVMC. LCZ696 is a drug used in HF to improve LVEF, with a few research on AVMC. In this research, we evaluated the effects and mechanism of LCZ696 in improving LVEF in AVMC. METHODS: Eighty 4-week-old male BALB/c mice were randomly divided into four groups of 20: Sham; Sham + LCZ696 (60 mg/kg/d); AVMC; AVMC + LCZ696. The above experiments were repeated by CVB3-infected HL-1 and Mdivi-1 to down-regulated dynamin-related protein 1(Drp1). Adeno-associated virus 9 (AAV9) with enhanced green fluorescent proteins (GFP) was injected to produce Drp1-overexpression mice and set up four groups: AVMC group, AVMC + AAV group, AVMC + LCZ696 group, and AVMC + LCZ696 + AAV group (n = 20 in each group). LVEF was evaluated by echocardiography at a similar heart rate (HR) at d7, Drp1 (p-Drp1), inflammation and apoptosis by histology and Western blot (WB), and mitochondrial by electron microscopy. RESULTS: Cardiac function were injured in AVMC that LCZ696 reversed (LVEF, %: Sham: 68.99 ± 9.67; Sham + LCZ696: 71.96 ± 6.20; AVMC: 30.95 ± 6.40*; AVMC + LCZ696: 68.99 ± 9.67*#, *P < 0.05 vs. Sham, #P < 0.05 vs. AVMC). LCZ696 attenuated p-Drp1 expression, inflammation, apoptosis, and mitochondrial fission (p-Drp1/Drp1: Sham: 1; Sham + LCZ696: 1.37 ± 0.22; AVMC: 2.29 ± 0.36*; AVMC+LCZ696: 1.43 ± 0.08*#, *P < 0.05 vs. Sham, #P < 0.05 vs. AVMC). Some of the above results were repeated in CVB3-infected HL-1 cells and Mdivi-1. AAV increased Drp1 expression and mitochondrial fission, inflammatory, and apoptosis. Compared with the AVMC + AAV group, the LVEF increased from 24.44 ± 0.03% to 32.33 ± 0.05% in the AVMC + LCZ696 + AAV group(P < 0.05), p-Drp1/Drp1 decreased from 0.54 ± 0.12 to 0.42 ± 0.09*, and IL-6, c-IL-1ß, and c-caspase-3/caspase-3 decreased from 1.07 ± 0.22 to 0.72 ± 0.08*, from 1.03 ± 0.14 to 0.79 ± 0.09*, and from 4.69 ± 0.29 to 0.92 ± 0.13*, respectively (*P < 0.05). CONCLUSIONS: LCZ696 has a protective effect on AVMC by improving LVEF and reducing inflammation and apoptosis, which may be due to the inhibition of Drp1-mediated mitochondrial fission.

Body composition of Miao nationality in Guizhou and Western Hu'nan / 解剖学报

Objective To explore the influence of regional differences on the body composition of the Miao nationality. Methods The bioelectrical impedance method was used to measure 17 body composition indexes of 357 adults of Miao Nationality in Guizhou (162 males and 195 females) and 471 adults of Miao Nationality in Western Hu'nan (210 males and 261 females). The correlation analysis between body composition and age, One-Way ANOVA and principal component analysis were carried out. Results The visceral fat grade and trunk fat percentage of Miao men in Guizhou and Miao in Western Hu'nan were positively correlated with age, and total muscle mass and trunk muscle mass were negatively correlated with age. The visceral fat grade and trunk fat rate of Miao women in the two regions were positively correlated with age, and the presumed bone mass and trunk muscle mass were negatively correlated with age. The index values of weight, muscle mass, estimated bone mass, water content, visceral fat grade, limb and trunk muscle mass in Guizhou Miao and Miao men in Western Hu'nan were all larger than women, and the body fat rate, limb and trunk fat mass were all smaller than women. The body fat percentages, limbs and trunk fat percentages of Guizhou Miao men and women were similar to those of Xiangxi Miao, and the muscle mass, limbs and trunk muscle mass were less than that of Xiangxi Miao. Conclusion There are obvious regional differences in muscle mass between the Miao nationality in Guizhou and the Miao nationality in Western Hu'nan.

Percutaneous left atrial appendage closure using the LAmbre device in patients with atrial fibrillation and left atrial appendage thrombus.

Background: Left atrial appendage closure (LAAC) is considered a valid alternative for the prevention of thromboembolic stroke in patients with persistent left atrial appendage thrombus (LAAT) despite adequate anticoagulation. However, the data on LAAC using the LAmbre device for patients with LAAT is limited. This study was performed to explore efficacy and safety as well as to share the experience of the modified LAAC procedure with the LAmbre device. Materials and methods: A total of 7 patients with persistent LAAT despite adequate anticoagulation underwent modified LAAC with the LAmbre device between November 2019 and April 2022. Transesophageal echocardiography was performed 3 months postoperatively to detect device-related thrombosis and peridevice leak. The patients' clinical events were evaluated during the perioperative and follow-up periods. Results: The median age, CHA2DS2-VASc score, and HAS-BLED score of all patients were 71 [53-73], 3 [2-4], and 2 [2-3], respectively. In the procedure, a cerebral protection system was used in two patients. LAAC with the LAmbre device was successfully performed in all patients without perioperative events. During the median follow-up of 383 [325-865] days, postoperative transesophageal echocardiography was performed in six (85.7%) patients. Device-related thrombosis was detected in one (16.7%) patient, and no significant peridevice leak was observed. No thromboembolic event or bleeding event occurred in any patients. Conclusion: LAAC with the LAmbre device is effective and safe when performed by experienced operators in highly selected patients with LAAT after adequate anticoagulation.

Combined atrial fibrillation ablation and left atrial appendage closure: Watchman vs. LAmbre devices.

Background: Left atrial appendage closure (LAAC) combined with radiofrequency catheter ablation is an emerging one-stop hybrid procedure for non-valvular atrial fibrillation (AF). This study was performed to compare the efficacy and safety of the Watchman device vs. the LAmbre device for this combined procedure. Methods: Two hundred and thirty two patients with AF who underwent the combined procedure were enrolled and divided into two subgroups depending on the device choice: the Watchman-combined group (n = 118) and the LAmbre-combined group (n = 114). The periprocedural and follow-up adverse events in both groups were documented. Results: The mean CHA2DS2-VASc score and HAS-BLED score in the Watchman-combined group and LAmbre-combined group were 3.7 ± 1.5 vs. 3.8 ± 1.5 and 2.5 ± 1.1 vs. 2.3 ± 1.1, respectively (all P > 0.05). Successful LAAC was achieved in all patients. The rate of major periprocedural complications and AF recurrence at 6 months post-procedure were similar between the Watchman-combined group and LAmbre-combined group (0.8 vs. 0.9%, P = 1.00; 22.0 vs. 15.8%, P = 0.23). During 2.6 ±0 .7 vs.1.6 ± 1.6 years follow-up, the rate of major clinical adverse events, including stroke and major bleeding, were comparable between the Watchman-combined group and the LAmbre-combined group (2.6 vs. 1.1% per 100 patient-years, P = 0.33). The intraprocedural peri-device leakage (PDL) rate was similar between the Watchman-combined group and the LAmbre-combined group (5.1 vs. 6.1%, P = 0.73), but the PDL rate was significantly higher at 3-6 months transesophageal echocardiography (TEE) follow-up than the intraprocedural PDL rate in both groups (21.6 vs. 5.1%; 36.6 vs. 6.1%, respectively), with a more obvious increase in minimal PDL rate in the LAmbre-combined group than the Watchman-combined group (36.6 vs. 21.6%, P < 0.05). Conclusion: The Watchman and LAmbre devices were comparable in efficacy and safety for the combined procedure. The minimal PDL rate at short-term TEE follow-up was higher in the LAmbre-combined group than the Watchman-combined group.

[Circular RNA in Diabetes and its Complications].

Circular RNA (circRNA) is a special group of non-coding RNA.Unlike linear RNA,circRNA is a closed circular structure formed by reverse splicing of pre-mRNA,with highly stable expression and wide distribution in eukaryotes.CircRNA has multiple functions by acting as microRNA sponges or binding with RNA-binding proteins.They can be used as biomarkers for many diseases.The latest research shows that circRNA plays a role in the pathogenesis of diabetes mellitus and the related chronic complications.In this review,we summarized the current progress and underlying mechanisms of circRNA in diabetes mellitus and the related complications.

Determination of fipronil and its metabolites in environmental water samples by meltblown nonwoven fabric-based solid-phase extraction combined with gas chromatography-electron capture detection.

In this study, a new method for the determination of fipronil and its three metabolites in environmental water samples was developed based on meltblown nonwoven fabric solid-phase extraction combined with gas chromatography-electron capture detection. As the core material of medical masks, meltblown nonwoven fabric is made of polypropylene superfine fibers which are randomly distributed and bonded together with a relatively large specific surface area and good permeability. Polypropylene as a high molecular hydrocarbon-based polymer has the characteristics of good hydrophobicity and lipophilicity, which can be applied for the separation and enrichment of hydrophobic substances in food, environment, and biological samples. The meltblown nonwoven fabric is soft and can fill the solid-phase extraction cartridge tightly. This aspect also makes it suitable to be used as an ideal solid-phase extraction sorbent. A series of parameters influencing the extraction efficiency were investigated, and under the optimized conditions, fipronil and its three metabolites had a good linear relationship in the range of 0.2-100 µg/L with a correlation coefficient R2 of more than 0.999. The recoveries at three spiked concentrations were in the range of 99.2-107.3% with the relative standard deviations less than 9.8% (intra-day) and 8.1% (inter-day). The limit of detection for the four target analytes was in the range of 0.02-0.06 µg/L. Finally, this method was successfully applied in the analysis of fipronil and its three metabolites in various types of environmental water samples.

Determining protein polarization proteome-wide using physical dissection of individual Stentor coeruleus cells.

Cellular components are non-randomly arranged with respect to the shape and polarity of the whole cell.1-4 Patterning within cells can extend down to the level of individual proteins and mRNA.5,6 But how much of the proteome is actually localized with respect to cell polarity axes? Proteomics combined with cellular fractionation7-11 has shown that most proteins localize to one or more organelles but does not tell us how many proteins have a polarized localization with respect to the large-scale polarity axes of the intact cell. Genome-wide localization studies in yeast12-15 found that only a few percent of proteins have a localized position relative to the cell polarity axis defined by sites of polarized cell growth. Here, we describe an approach for analyzing protein distribution within a cell with a visibly obvious global patterning-the giant ciliate Stentor coeruleus.16,17 Ciliates, including Stentor, have highly polarized cell shapes with visible surface patterning.1,18 A Stentor cell is roughly 2 mm long, allowing a "proteomic dissection" in which microsurgery is used to separate cellular fragments along the anterior-posterior axis, followed by comparative proteomic analysis. In our analysis, 25% of the proteome, including signaling proteins, centrin/SFI proteins, and GAS2 orthologs, shows a polarized location along the cell's anterior-posterior axis. We conclude that a large proportion of all proteins are polarized with respect to global cell polarity axes and that proteomic dissection provides a simple and effective approach for spatial proteomics.

Effect of water-oil ratio on the photocatalytic performance of visible light-active BiVO4 nanoparticles prepared by inverse microemulsion-calcination method.

Nowadays, nano-photocatalysts (NPs) have become the research focus in the field of photocatalysis due to their excellent photocatalytic activity, and microemulsion is an effective method to prepare high-efficiency nano-photocatalysts. Here, BiVO4 NPs with high efficiency under visible light were prepared by a combination of reverse microemulsion method and calcination method. XRD, SEM, TEM, XPS, DRS, PL, BET and other characterization tests were used to comprehensively explore the influence of water-oil ratio on the physicochemical properties of the catalysts. The results show that BiVO4 NPs of monoclinic scheelite with high crystallization degree can be obtained by this method. The microscopic morphology, specific surface area and total pore volume of BiVO4 NPs are significantly affected by the water-oil ratio. It is difficult to obtain BiVO4 NPs with small particle size and uniform dispersion under the condition of too low or too high water-oil ratio. Meanwhile, the photogenerated carrier recombination efficiency of the catalyst is significantly improved, thus reducing the photocatalytic activity of the catalyst. Strikingly, the BiVO4 NPs obtained under the condition of water-oil ratio is 20 exhibited well-dispersed nanospheres with diameters ranging from 80 to 100 nm. It has the highest photocatalytic activity due to its high crystallinity, large specific surface area and total pore volume and relatively low photogenerated carrier recombination efficiency. Under visible light irradiation, the degradation efficiency of RhB can reach 97.69% in 100 min, and the rate constant is 0.03253 min-1.

[Mechanism of polyphyllin A in inhibition of proliferation and induction of apoptosis of gastric cancer cells by directly targeting ETS-1].

The present study investigated the mechanism of polyphyllin A(PPA) in inhibiting gastric cancer(GC) cells. GC cells(SGC7901 and MGC803 cell lines) were treated with PPA at different concentrations. The effect of PPA on the proliferation of GC cells was detected by MTT assay, real-time cell analysis(RTCA) assay, and clone-forming assay, respectively. Reactive oxygen species(ROS) of GC cells was detected by flow cytometry. The change of mitochondrial membrane potential was detected by JC-1 assay. The expression and phosphorylation levels of apoptosis-related proteins(caspase-9, caspase-3, and PARP) and proteins related to the signaling pathway(ETS-1, CIP2 A, and Akt) were detected by Western blot. The binding sites of PPA to ETS-1 were analyzed by molecular docking. The affinity of PPA and ETS-1 was detected by drug affinity responsive target stability(DARTS) assay. PPA had a significant inhibitory effect on the proliferation and colony formation of GC cells at a low concentration. The PPA groups showed increased ROS and decreased mitochondrial membrane potential. PPA down-regulated the precursor expression of caspase-9 and caspase-3 and promoted the cleavage of PARP, suggesting that PPA induced the apoptosis of GC cells through the mitochondrial pathway. PPA significantly reduced expression levels of CIP2 A and the phosphorylation of downstream Akt. Molecular docking showed that PPA bound to the ETS domain of ETS-1, the transcription factor of CIP2 A, and formed hydrogen bonds with Pro319 and Asp317. DARTS assay further confirmed that PPA significantly prevented the hydrolysis of ETS-1 by pronase, which was inductive of the direct binding effect of PPA and ETS-1. PPA inhibits the proliferation and induces the apoptosis of GC cells by directly targeting ETS-1 to down-regulate the ETS-1/CIP2 A/Akt signaling pathway.

[Mechanism of polyphyllin â targeting EGFR to affect proliferation and apoptosis of human breast cancer cells].

This study aims to investigate the molecular mechanism of polyphyllin Ⅰ(PPⅠ) inhibiting proliferation of human breast cancer cells. Human breast cancer BT474 and MDA-MB-436 cells were treated with different concentrations of PPⅠ, and then the effect of PPⅠ on cell proliferation was detected by MTT assay, trypan blue dye exclusion assay, real-time cell analysis, and clone forming assay, respectively. The apoptosis was detected by Annexin V-FITC/PI staining and then analyzed by flow cytometry. The change of mitochondrial membrane potential was detected by flow cytometry after fluorescent probe JC-1 staining. Western blot was used to detect protein expression and phosphorylation. Molecular docking was performed to detect the binding between PPⅠ and EGFR. The affinity between PPⅠ and EGFR was determined by drug affinity responsive target stability assay. The results indicated that PPⅠ inhibited the proliferation and colony formation of BT474 and MDA-MB-436 cells in a time-and concentration-dependent manner. The PPⅠ treatment group showed significantly increased apoptosis rate and significantly decreased mitochondrial membrane potential. PPⅠ down-regulated the expression of pro-caspase-3 protein, promoted the cleavage of PARP, and significantly reduced the phosphorylation levels of EGFR, Akt, and ERK. Molecular docking showed that PPⅠ bound to the extracellular domain of EGFR and formed hydrogen bond with Gln366 residue. Drug affinity responsive target stability assay confirmed that PPⅠ significantly prevented pronase from hydrolyzing EGFR, indicating that PPⅠ and EGFR have a direct binding effect. In conclusion, PPⅠ inhibited the proliferation and induced apoptosis of breast cancer cells by targeting EGFR to block its downstream signaling pathway. This study lays a foundation for the further development of PPⅠ-targeted drugs against breast cancer.

Determination of fipronil and its metabolites in edible oil by pollen based solid-phase extraction combined with gas chromatography-electron capture detection.

In this study, a convenient and economic method for the determination of fipronil and its three metabolites in edible oil was developed based on pollen grain solid-phase extraction (SPE). As a natural material, pollen grains exhibit well absorption capacity for some polar compounds due to their special functional structures. Their stable composition and appropriate particle size also make them suitable for SPE. In the present study, natural pine pollen grains without broken wall were used as sorbent for selective isolation and enrichment of fipronil and its three metabolites from edible oils based on hydrogen bond interaction. Several parameters influencing the extraction recoveries were investigated. By coupling with gas chromatography-electron capture detection (GC-ECD), a new method for analysis of fipronil and its metabolites in edible oils was established. The linearity range was 2-200 ng/g with correlation coefficient R2 more than 0.999. The recoveries in edible oils at three spiked concentrations were in the range of 80.1-96.0% with the RSDs less than 10.6% (intra-day) and 11.5% (inter-day). The limit of detection (LOD) for four target analytes were in the range of 0.2-0.6 ng/g, which was comparable to the previous reported methods. Finally, the established method was successfully applied to detect fipronil and its metabolites in several oil samples with different brands from local market.

Application of a novel nylon needle filter-based solid-phase extraction device to determination of 1-hydroxypyrene in urine.

In this work, a simple and miniaturized solid-phase extraction device was constructed by connecting a commercial nylon needle filter to a syringe, which was applied for extracting 1-hydroxypyrene from a urine sample via hydrophobic and hydrogen bond interactions. The nylon membrane in the needle filter acted as the solid-phase extraction adsorbent, meanwhile, it filtered the particles in the urine sample. To obtain high extraction efficiency, key parameters influencing extraction recovery were investigated. The entire pretreatment process was accomplished within 5 min under the optimal conditions. By coupling high-performance liquid chromatography-ultraviolet, a rapid, low-cost, and convenient nylon needle filter-based method was established for the analysis of 1-hydroxypyrene in a complex urine matrix. Within the linearity range of 0.2-1000 µg/L, the method exhibited a satisfactory correlation coefficient (R = 0.9999). The limit of detection was 0.06 µg/L, and the recoveries from urine sample spiked with three concentrations (5, 20, and 100 µg/L) ranged from 105.8% to 113.1% with the relative standard deviations less than 6.7% (intra-day, n = 6) and 8.9% (inter-day, n = 4). Finally, the proposed method was successfully applied for detecting 1-hydroxypyrene in urine samples from college students, smokers, gas station workers, and chip factory workers. The detected concentration in actual urine samples ranged from 0.46 to 5.26 µg/L. Taken together, this simple and cost-effective nylon needle filter-based solid-phase extraction device showed an excellent application potential for pretreating hydrophobic analytes from aqueous samples.