Machine Learning Radiomics-Based Prediction of Non-sentinel Lymph Node Metastasis in Chinese Breast Cancer Patients with 1-2 Positive Sentinel Lymph Nodes: A Multicenter Study.

RATIONALE AND OBJECTIVES: This study aimed to construct a machine learning radiomics-based model using dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) images to evaluate non-sentinel lymph node (NSLN) metastasis in Chinese breast cancer (BC) patients who underwent total mastectomy (TM) and had 1-2 positive sentinel lymph nodes (SLNs). MATERIALS AND METHODS: In total, 494 patients were retrospectively enrolled from two hospitals, and were divided into the training (n = 286), internal validation (n = 122), and external validation (n = 86) cohorts. Features were extracted from DCE-MRI images for each patient and screened. Six ML classifies were trained and the best classifier was evaluated to calculate radiomics (Rad)-scores. A combined model was developed based on Rad-scores and clinical risk factors, then the calibration, discrimination, reclassification, and clinical usefulness were evaluated. RESULTS: 14 radiomics features were ultimately selected. The random forest (RF) classifier showed the best performance, with the highest average area under the curve (AUC) of 0.833 in the validation cohorts. The combined model incorporating RF-based Rad-scores, tumor size, lymphovascular invasion, and proportion of positive SLNs resulted in the best discrimination ability, with AUCs of 0.903, 0.890, and 0.836 in the training, internal validation, and external validation cohorts, respectively. Furthermore, the combined model significantly improved the classification accuracy and clinical benefit for NSLN metastasis prediction. CONCLUSION: A RF-based combined model using DCE-MRI images exhibited a promising performance for predicting NSLN metastasis in Chinese BC patients who underwent TM and had 1-2 positive SLNs, thereby aiding in individualized clinical treatment decisions.

Evaluating clinical pharmacists' involvement in postoperative acute pain services: a multicenter survey in Guangdong Province, China.

Objective: Postoperative pain management is an important part of surgical pharmacy. Postoperative acute pain services in China are in their initial stages. This survey aimed to investigate the attitudes, involvement, and knowledge of clinical pharmacists in China regarding postoperative acute pain services. The results can provide valuable information to guide clinical pharmacists in developing targeted strategies to improve their postoperative acute pain service capabilities. Methods: A questionnaire was distributed to the pharmacy departments of 133 grade A tertiary hospitals in Guangdong province, and the responses were collected electronically. Results: 123 completed questionnaires were collected from clinical pharmacists. Although 95.93% of clinical pharmacists believed they should participate in postoperative pain services, only 62.6% reported substantial involvement. Overall satisfaction with the postoperative pain service was 93.5%. Understanding of non-steroidal anti-inflammatory drugs and opioid analgesics by clinical pharmacists was comparable (p > 0.05). Furthermore, 98.37% of clinical pharmacists desired systematic learning in postoperative pain management, and 40.65% expressed a strong need. Conclusion: Clinical pharmacists in China demonstrate a positive attitude toward participating in postoperative acute pain services. However, the actual level of involvement was concerning, and the lack of systematic training and well-established work protocols may be contributing factors. Efforts should be made to establish comprehensive and standardized processes and work protocols for postoperative acute pain services and provide systematic and hierarchical professional training to enhance clinical pharmacists' capabilities in postoperative acute pain services.

Research on Hydrodynamic Characteristics of Electronic Paper Pixels Based on Electrowetting.

In this paper, we propose a driving waveform with a complex ramp pulse for an electrowetting display system. The relationship between the contact angle and viscosity of inks was calculated based on the fluid-motion characteristics of different viscosities. We obtained the suitable range of viscosity and voltage in the liquid-oil-solid three-phase contact display system. We carried out model simulation and driving waveform design. The result shows that the driving waveform improves the response speed and aperture ratio of electrowetting. The aperture ratio of electrowetting pixels is increased to 68.69%. This research is of great significance to optimizing the structure of fluid material and the design of driving waveforms in electrowetting displays.

Development and validation of convolutional neural network-based model to predict the risk of sentinel or non-sentinel lymph node metastasis in patients with breast cancer: a machine learning study.

Background: For patients with sentinel lymph node (SLN) metastasis and low risk of residual non-SLN (NSLN) metastasis, axillary lymph node (ALN) dissection could lead to overtreatment. This study aimed to develop and validate an automated preoperative deep learning-based tool to predict the risk of SLN and NSLN metastasis in patients with breast cancer (BC) using dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) images. Methods: In this machine learning study, we retrospectively enrolled 988 women with BC from three hospitals in Zhejiang, China between June 1, 2013 to December 31, 2021, June 1, 2017 to December 31, 2021, and January 1, 2019 to June 30, 2023, respectively. Patients were divided into the training set (n = 519), internal validation set (n = 129), external test set 1 (n = 296), and external test set 2 (n = 44). A convolutional neural network (CNN) model was proposed to predict the SLN and NSLN metastasis and was compared with clinical and radiomics approaches. The performance of different models to detect ALN metastasis was measured by the area under the curve (AUC), accuracy, sensitivity, and specificity. This study is registered at ChiCTR, ChiCTR2300070740. Findings: For SLN prediction, the top-performing model (i.e., the CNN algorithm) achieved encouraging predictive performance in the internal validation set (AUC 0.899, 95% CI, 0.887-0.911), external test set 1 (AUC 0.885, 95% CI, 0.867-0.903), and external test set 2 (AUC 0.768, 95% CI, 0.738-0.798). For NSLN prediction, the CNN-based model also exhibited satisfactory performance in the internal validation set (AUC 0.800, 95% CI, 0.783-0.817), external test set 1 (AUC 0.763, 95% CI, 0.732-0.794), and external test set 2 (AUC 0.728, 95% CI, 0.719-0.738). Based on the subgroup analysis, the CNN model performed well in tumour group smaller than 2.0 cm, with the AUC of 0.801 (internal validation set) and 0.823 (external test set 1). Of 469 patients with BC, the false positive rate of SLN prediction declined from 77.9% to 32.9% using CNN model. Interpretation: The CNN model can predict the SLN status of any detectable lesion size and condition of NSLN in patients with BC. Overall, the CNN model, employing ready DCE-MRI images could serve as a potential technique to assist surgeons in the personalized axillary treatment of in patients with BC non-invasively. Funding: National Key Research and Development projects intergovernmental cooperation in science and technology of China, National Natural Science Foundation of China, Natural Science Foundation of Zhejiang Province, and Zhejiang Medical and Health Science Project.

Multi-algorithms analysis for pre-treatment prediction of response to transarterial chemoembolization in hepatocellular carcinoma on multiphase MRI.

OBJECTIVES: This study compared the accuracy of predicting transarterial chemoembolization (TACE) outcomes for hepatocellular carcinoma (HCC) patients in the four different classifiers, and comprehensive models were constructed to improve predictive performance. METHODS: The subjects recruited for this study were HCC patients who had received TACE treatment from April 2016 to June 2021. All participants underwent enhanced MRI scans before and after intervention, and pertinent clinical information was collected. Registry data for the 144 patients were randomly assigned to training and test datasets. The robustness of the trained models was verified by another independent external validation set of 28 HCC patients. The following classifiers were employed in the radiomics experiment: machine learning classifiers k-nearest neighbor (KNN), support vector machine (SVM), the least absolute shrinkage and selection operator (Lasso), and deep learning classifier deep neural network (DNN). RESULTS: DNN and Lasso models were comparable in the training set, while DNN performed better in the test set and the external validation set. The CD model (Clinical & DNN merged model) achieved an AUC of 0.974 (95% CI: 0.951-0.998) in the training set, superior to other models whose AUCs varied from 0.637 to 0.943 (p < 0.05). The CD model generalized well on the test set (AUC = 0.831) and external validation set (AUC = 0.735). CONCLUSIONS: DNN model performs better than other classifiers in predicting TACE response. Integrating with clinically significant factors, the CD model may be valuable in pre-treatment counseling of HCC patients who may benefit the most from TACE intervention.

CYLD deubiquitinates plakoglobin to promote Cx43 membrane targeting and gap junction assembly in the heart.

During heart maturation, gap junctions assemble into hemichannels and polarize to the intercalated disc at cell borders to mediate electrical impulse conduction. However, the molecular mechanism underpinning cardiac gap junction assembly remains elusive. Herein, we demonstrate an important role for the deubiquitinating enzyme cylindromatosis (CYLD) in this process. Depletion of CYLD in mice impairs the formation of cardiac gap junctions, accelerates cardiac fibrosis, and increases heart failure. Mechanistically, CYLD interacts with plakoglobin and removes lysine 63-linked polyubiquitin chains from plakoglobin. The deubiquitination of plakoglobin enhances its interaction with the desmoplakin/end-binding protein 1 complex localized at the microtubule plus end, thereby promoting microtubule-dependent transport of connexin 43 (Cx43), a key component of gap junctions, to the cell membrane. These findings establish CYLD as a critical player in regulating gap junction assembly and have important implications in heart development and diseases.

[Serum Lipid Levels and Their Prognostic Significance in Patients with Multiple Myeloma].

OBJECTIVE: To investigate the serum lipid levels and their prognostic significance in patients with multiple myeloma (MM). METHODS: A total of 87 newly diagnosed MM patients and 87 healthy controls in our hospital from January 2012 to April 2021 were selected. Serum lipid levels were compared between MM patients and healthy controls. The differences of serum lipid levels in patients among two groups of sex, age, hemoglobin (Hb), albumin (ALB), platelet (PLT), ß2-microglobulin (ß2-MG) and bone marrow plasma cell ratio (BMPC), different immune types, different ISS stages, before and after chemotherapy were analyzed. Univariate and COX multivariate regression analysis were used to analyze the influence of clinical parameters such as serum lipid indexes on prognosis of MM. RESULTS: The serum levels of total cholesterol (TC), low density lipoprotein cholesterol (LDL-C), high density lipoprotein cholesterol (HDL-C), apolipoprotein A1 (Apo A1) and apolipoprotein B (Apo B) in MM patients were significantly lower than those in healthy controls (P<0.05). Anemia, low protein and low PLT in patients were related to low cholesterol. The levels of TC, LDL-C, HDL-C, Apo A1 and Apo B in patients with low Hb and ALB were significantly lower than those in patients with high Hb and ALB (P<0.05). The Apo B level of low PLT patients was significantly lower than that of high PLT patients (P<0.05). The levels of TC, LDL-C, HDL-C, Apo A1 and Apo B in patients with different immune types were significantly different, the above indexes of IgA type were significantly lower than IgG type(P<0.05), IgG type were significantly lower than light chain type(P<0.05), double clone type were significantly lower than light chain type (P<0.05). The levels of TC, LDL-C, and Apo B in patients with different ISS stages were significantly different, stage Ⅱ were lower than those of stage Ⅰ (P>0.05), stage Ⅲ were significantly lower than those of stage Ⅱ and stageⅠ(P<0.05). The levels of TC, TG, LDL-C, HDL-C, Apo A1 and Apo B in patients after chemotherapy were significantly higher than those before chemotherapy (P<0.05). Univariate analysis showed that Hb, PLT, ß2-MG, BMPC, LDL-C and Apo B affected the prognosis of MM. Multivariate analysis showed that BMPC and Apo B were independent factors affecting the prognosis of MM. CONCLUSION: The serum cholesterol level is decreased in MM patients, and hypocholesterolemia is related to the classification and staging of the disease. With the improvement of the disease, the serum cholesterol level is increased, and low serum Apo B level predicts a poor prognosis.

Home Dust Mites Promote MUC5AC Hyper-Expression by Modulating the sNASP/TRAF6 Axis in the Airway Epithelium.

House dust mites (HDMs) are a common source of respiratory allergens responsible for allergic asthma and innate immune responses in human diseases. Since HDMs are critical factors in the triggering of allergen-induced airway mucosa from allergic asthma, we aimed to investigate the mechanisms of Toll-like receptors (TLR) in the signaling of the HDM extract that is involved in mucus hypersecretion and airway inflammation through the engagement of innate immunity. Previously, we reported that the somatic nuclear autoantigenic sperm protein (sNASP)/tumor necrosis factor receptor-associated factor 6 (TRAF6) axis controls the initiation of TLRs to maintain the homeostasis of the innate immune response. The present study showed that the HDM extract stimulated the biogenesis of Mucin 5AC (MUC5AC) in bronchial epithelial cells via the TLR2/4 signaling pathway involving MyD88 and TRAF6. Specifically, sNASP binds to TRAF6 in unstimulated bronchial epithelial cells to prevent the activation of TRAF6-depenedent kinases. Upon on HDMs' stimulation, sNASP is phosphorylated, leading to the activation of TRAF6 downstream of the p38 MAPK and NF-κB signaling pathways. Further, NASP-knockdown enhanced TRAF6 signaling and MUC5AC biogenesis. In the HDM-induced mouse asthma model, we found that the HDM extract promoted airway hyperresponsiveness (AHR), MUC5AC, and allergen-specific IgE production as well as IL-5 and IL-13 for recruiting inflammatory cells. Treatment with the PEP-NASP peptide, a selective TRAF6-blocking peptide, ameliorated HDM-induced asthma in mice. In conclusion, this study indicated that the sNASP/TRAF6 axis plays a regulatory role in asthma by modulating mucus overproduction, and the PEP-NASP peptide might be a potential target for asthma treatment.

Ureteral calculi lithotripsy for single ureteral calculi: can DNN-assisted model help preoperatively predict risk factors for sepsis?

OBJECTIVES: To explore the utility of radiomics and deep learning model in assessing the risk factors for sepsis after flexible ureteroscopy lithotripsy (FURL) or percutaneous nephrolithotomy (PCNL) in patients with ureteral calculi. METHODS: This retrospective analysis included 847 patients with treatment-naive proximal ureteral calculi who received FURL or PCNL. All participants were preoperatively conducted non-contrast computed tomography scans, and relevant clinical information was meanwhile collected. After propensity score matching, the radiomics model was established to predict the onset of sepsis. A deep learning model was also adapted to further improve the prediction accuracy. Performance of these trained models was verified in another independent external validation set including 40 cases of ureteral calculi patients. RESULTS: The overall incidence of sepsis after FURL or PCNL was 5.9%. The least absolute shrinkage and selection operator (LASSO) regression analysis revealed 26 predictive variables, with an overall AUC of 0.881 (95% CI, 0.813-0.931) and an AUC of 0.783 (95% CI, 0.766-0.801) in external validation cohort. Judicious adaption of a deep neural network (DNN) model to our dataset improved the AUC to 0.920 (95% CI, 0.906-0.933) in the internal validation. To eliminate the overfitting, external validation was carried out for DNN model (AUC = 0.874 (95% CI, 0.858-0.891)). CONCLUSIONS: The DNN was more effective than the LASSO model in revealing risk factors for sepsis after FURL or PCNL in single ureteral calculi patients, and females are more susceptible to sepsis than males. Deep learning models have the potential to act as gatekeepers to facilitate patient stratification. KEY POINTS: • Both the least absolute shrinkage and selection operator (LASSO) and deep neural network (DNN) models were shown to be effective in sepsis prediction. • The DNN model achieved superior prediction capability, with an AUC of 0.920 (95% CI, 0.906-0.933). • DNN-assisted model has potential to serve as a gatekeeper to facilitate patient stratification.

Extracellular domain of semaphorin 5A serves a tumor­suppressing role by activating interferon signaling pathways in lung adenocarcinoma cells.

Semaphorin 5A (SEMA5A), which was originally identified as an axon guidance molecule in the nervous system, has been subsequently identified as a prognostic biomarker for lung cancer in nonsmoking women. SEMA5A acts as a tumor suppressor by inhibiting the proliferation and migration of lung cancer cells. However, the regulatory mechanism of SEMA5A is not clear. Therefore, the purpose of the present study was to explore the roles of different domains of SEMA5A in its tumor­suppressive effects in lung adenocarcinoma cell lines. First, it was revealed that overexpression of full length SEMA5A or its extracellular domain significantly inhibited the proliferation and migration of both A549 and H1299 cells using MTT, colony formation and gap closure assays. Next, microarray analyses were performed to identify genes regulated by different domains of SEMA5A. Among the differentially expressed genes, the most significant function of these genes that were enriched was the 'Interferon Signaling' pathway according to Ingenuity Pathway Analysis. The activation of the 'Interferon Signaling' pathway was validated by reverse transcription­quantitative PCR and western blotting. In summary, the present study demonstrated that the extracellular domain of SEMA5A could upregulate genes in interferon signaling pathways, resulting in suppressive effects in lung adenocarcinoma cells.

HIV-1 exposure promotes PKG1-mediated phosphorylation and degradation of stathmin to increase epithelial barrier permeability.

Exposure of mucosal epithelial cells to the human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein gp120 is known to disrupt epithelial cell junctions by impairing stathmin-mediated microtubule depolymerization. However, the pathological significance of this process and its underlying molecular mechanism remain unclear. Here we show that treatment of epithelial cells with pseudotyped HIV-1 viral particles or recombinant gp120 protein results in the activation of protein kinase G 1 (PKG1). Examination of epithelial cells by immunofluorescence microscopy reveals that PKG1 activation mediates the epithelial barrier damage upon HIV-1 exposure. Immunoprecipitation experiments show that PKG1 interacts with stathmin and phosphorylates stathmin at serine 63 in the presence of gp120. Immunoprecipitation and immunofluorescence microscopy further demonstrate that PKG1-mediated phosphorylation of stathmin promotes its autophagic degradation by enhancing the interaction between stathmin and the autophagy adaptor protein p62. Collectively, these results suggest that HIV-1 exposure exploits the PKG1/stathmin axis to affect the microtubule cytoskeleton and thereby perturbs epithelial cell junctions. Our findings reveal a novel molecular mechanism by which exposure to HIV-1 increases epithelial permeability, which has implications for the development of effective strategies to prevent mucosal HIV-1 transmission.

Comparison of reduced field-of-view diffusion-weighted imaging (DWI) and conventional DWI techniques in the assessment of Cervical carcinoma at 3.0T: Image quality and FIGO staging.

OBJECTIVE: To evaluate imaging quality (IQ) and International Federation of Gynecology and Obstetrics (FIGO) staging of reduced field-of-view (r-FOV) diffusion-weighted imaging (DWI) in cervical carcinoma (CC). MATERIALS AND METHODS: Sixty patients with pathologically proven CC who underwent both pre-treatment r-FOV DWI and full field-of-view (f-FOV) DWI on a 3.0T MRI scanner were retrospectively reviewed. The subjective qualitative image scores were compared using the Wilcoxon signed-rank test. Objective quality values and apparent diffusion coefficient (ADC) were estimated by paired t-test or Wilcoxon signed-rank test for the two DWI sequences according to Normality test. Spearman rank correlation analysis was used to evaluate the relationship between pathological results and mean ADC value. RESULTS: The subjective IQ scores for r-FOV DWI were significantly higher than those for f-FOV DWI (P < 0.001). Similarly, the contrast-to-noise (CNR) value of r-FOV DWI was superior to that of f-FOV DWI (10.30 ±â€¯3.676, 8.91 ±â€¯3.008, P = 0.021). However, the signal-to-noise ratio (SNR) value of r-FOV DWI was considerably lower than that of f-FOV DWI (27.80 ±â€¯6.056, 33.67 ±â€¯7.833, P＜0.001). No significant difference was found between mean ADC values of f-FOV DWI and r-FOV DWI. There was a significant tendency for a negative correlation between the ADC values and FIGO stages of CC for both two sequences (r=-0. 436, P＜0.01; r=-0.470, P＜0.01, respectively). CONCLUSIONS: The rFOV DWI sequence provided significantly better IQ and lesion conspicuity than the fFOV DWI sequence. In addition, rFOV sequences can be used in evaluation of FIGO staging of cervical cancer.

Prediction of the World Health Organization Grade of rectal neuroendocrine tumors based on CT histogram analysis.

OBJECTIVES: To investigate the diagnostic value of contrast-enhanced computed tomography (CECT) histogram analysis in predicting the World Health Organization (WHO) grade of rectal neuroendocrine tumors (R-NETs). MATERIALS AND METHODS: A total of 61 (35 G1, 12 G2, 10 G3, and 4 NECs) patients who underwent preoperative CECT and treated with surgery to be confirmed as R-NETs were included in this study from January 2014 to May 2019. We depicted ROIs and measured the CECT texture parameters (mean, median, 10th, 25th, 75th, 90th percentiles, skewness, kurtosis, and entropy) from arterial phase (AP) and venous phase (VP) images by two radiologists. We calculated intraclass correlation coefficient (ICC) and compared the histogram parameters between low-grade (G1) and higher grade (HG) (G2/G3/NECs) by applying appropriate statistical method. We obtained the optimal parameters to identify G1 from HG using receiver operating characteristic (ROC) curves. RESULTS: The capability of AP and VP histogram parameters for differentiating G1 from HG was similar in several histogram parameters (mean, median, 10th, 25th, 75th, and 90th percentiles) (all p < 0.001). Skewness, kurtosis, and entropy on AP images showed no significant differences between G1 and HG (p = 0.853, 0.512, 0.557, respectively). Entropy on VP images was significantly different (p = 0.017) between G1 and HG, however, skewness and kurtosis showed no significant differences (p = 0.654, 0.172, respectively). ROC analysis showed a good predictive performance between G1 and HG, and the 75th (AP) generated the highest area under the curve (AUC = 0.871), followed by the 25th (AP), mean (VP), and median (VP) (AUC = 0.864). Combined the size of tumor and the 75th (AP) generated the highest AUC. CONCLUSIONS: CECT histogram parameters, including arterial and venous phases, can be used as excellent indicators for predicting G1 and HG of rectal neuroendocrine tumors, and the size of the tumor is also an important independent predictor.

A Novel Clinical-Radiomics Model Pre-operatively Predicted the Stone-Free Rate of Flexible Ureteroscopy Strategy in Kidney Stone Patients.

Purpose: The purpose of the study is to develop and validate a novel clinical-radiomics nomogram model for pre-operatively predicting the stone-free rate of flexible ureteroscopy (fURS) in kidney stone patients. Patients and Methods: Altogether, 2,129 fURS cases with kidney stones were retrospectively analyzed, and 264 patients with a solitary kidney stone were included in a further study. For lower calyx calculi, a radiomics model was generated in a primary cohort of 99 patients who underwent non-contrast-enhanced computed tomography (NCCT). Radiomics feature selection and signature building were conducted by using the least absolute shrinkage and selection operator (LASSO) method. Multivariate logistic regression analysis was employed to build a model incorporating radiomics and potential clinical factors. Model performance was evaluated by its discrimination, calibration, and clinical utility. The model was internally validated in 43 patients. Results: The overall success rate of fURS was 72%, while the stone-free rate (SFR) for lower calyx calculi and non-lower calyx calculi was 56.3 and 90.16%, respectively. On multivariate logistic regression analysis of the primary cohort, independent predictors for SFR were radiomics signature, stone volume, operator experience, and hydronephrosis level, which were all selected into the nomogram. The area under the curve (AUC) of clinical-radiomics model was 0.949 and 0.947 in the primary and validation cohorts, respectively. Moreover, the calibration curve showed a satisfactory predictive accuracy, and the decision curve analysis indicated that the nomogram has superior clinical application value. Conclusion: In this novel clinical-radiomics model, the radiomics scores, stone volume, hydronephrosis level, and operator experience were crucial for the flexible ureteroscopy strategy.

Effects of methylthiouracil on the proliferation and apoptosis of rat bone marrow stromal cells.

The aim of the present study was to investigate the effects of methylthiouracil (MTU) on the proliferation and apoptosis of rat bone marrow stromal cells (BMSCs). Rat BMSCs were isolated, cultured in vitro and treated with various concentrations of MTU. Cell growth curves were determined using the Cell Counting Kit-8 method and the effect of MTU on BMSCs in a logarithmic growth phase was observed. BMSC apoptosis following MTU treatment was detected by flow cytometry. The experimental results demonstrated that the proliferation-inhibition effect was gradually enhanced with increasing MTU concentrations and the extension of treatment time. Statistically significant differences were observed between the treatment and the control groups (P<0.05). In addition, the BMSC apoptosis rate gradually increased with increasing drug concentrations and treatment time extension; statistically significant differences were observed between the treatment and the control groups (P<0.05). Therefore, the results of the present study demonstrated that MTU inhibited the proliferation of BMSCs and promoted apoptosis, indicating the cytotoxic effects of MTU on BMSCs.

Synthesis, crystal structures and DNA-cleaving activities of [Cemp]2[MCl4] (Cemp = N-carbethoxymethyl-1,10-phenanthrolinium, M = Cu(II), Zn(II), Co(II), Ni(II) and Mn(II)).

N-Carbethoxymethyl-1,10-phenanthrolinium bromide (CempBr) and its five ionic metal complexes, [Cemp]2[MCl4] where M = Cu(II) (1), Zn(II) (2), Co(II) (3), Ni(II) (4) and Mn(II) (5) were synthesized and fully characterized. Complexes 1-5 have similar structures, and consist of isolated [Cemp](+) cations and [MCl4](2-) anions in which there are no obvious interactions between the oxygen or nitrogen donor atoms in [Cemp](+) and the metal center in [MCl4](2-). Agarose gel electrophoresis studies on the cleavage of plasmid pBR322 DNA by complexes 1-5 indicated that complex 1 was capable of efficiently cleaving DNA under physiological conditions, most probably via an oxidative mechanism. Kinetic assay of complex 1 afforded the maximal catalytic rate constant kmax of 0.55 h(-1) and Michaelis constant KM of 47.6 µM, respectively, which gives about 1.5×10(7)-fold rate acceleration over uncatalyzed cleavage of supercoiled DNA. Ethidium bromide displacement experiments indicated that complex 1 had a binding affinity of (1.58±1.12)×10(6) M(-1) toward calf-thymus DNA, 20-100-fold higher than those shown by CempBr and complexes 2-5. The high cleaving efficacy of complex 1 is thought to be due to the efficient catalysis of the copper(II)-coordinated center and the efficient binding of the quaternized 1,10-phenanthroline cation to DNA.

Synthesis, characterization and potent DNA-cleaving activity of copper(II)-complexed berberine carboxylate.

9-O-(4-carboxybenzyl)berberine (CBB) 1 was synthesized from the reaction of berberrubine with methyl 4-(bromomethyl)benzoate and subsequent hydrolysis. Its Cu(II) complex 2 was prepared from the reaction with Cu(NO(3))(2)·3H(2)O, and established as [Cu(CBB)(2)](NO(3))(2)·2H(2)O by means of (1)H NMR, UV, IR, elemental analysis and TGA measurements. Agarose gel electrophoresis study on the cleavage of plasmid pBR322 DNA indicated that complex 2 was capable of efficiently cleaving DNA under physiological conditions, most probably via an oxidative mechanistic pathway involving the formation of singlet oxygen as the reactive species. Kinetic assay afforded the maximal first-order rate constant k(max) of 2.41 h(-1) and Michaelis constant K(M) of 2.64 mM, respectively, representing ca. 10(8)-fold acceleration in the cleavage. This catalytic efficacy is attributed to the Cu(II)-assisted formation of dimeric species, in which the two berberine subunits cooperatively bind to DNA, whereas the carboxylate-coordinated Cu(II) moiety functions as the cleavage-active center.

Protective effects of gypenosides against fatty liver disease induced by high fat and cholesterol diet and alcohol in rats.

In the present study, the protective effects of gypenosides from Gynostemma pentaphyllum on fatty liver disease (FLD) were examined in rats treated with high fat and cholesterol diet and alcohol. Male SD rats were divided into seven groups: control, model, lovastatin, silymarin, gypenosides high-, medium- and low-treatment groups. The latter 6 groups were fed high-fat and cholesterol diet and administered alcohol intragastricly once a day. Body weight was measured every week for 10 weeks, and the hepatic index was measured after 10 weeks. Compared with model group, levels of serum triglyceride (TG), total cholesterol (TC), free fatty acid (FFA), and low density lipoprotein cholesterol (LDL-C) level, malondialdehyde (MDA), serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activity, and hepatocyte apoptosis were significantly decreased in gypenosides groups; while serum high density lipoprotein cholesterol (HDL-C), superoxide dismutase (SOD) activity in both serum and hepatic tissue and mRNA and protein level of peroxisome proliferator-activated receptor α (PPAR-α) were significantly increased. Moreover, hepatic steatosis and mitochondrial damage were improved. These results suggested that gypenosides could prevent liver fatty degeneration in fatty liver disease through modulating lipid metabolism, ameliorating liver dysfunction and reducing oxidative stress.

[Effect of berberine on HL-60 cell proliferation, apoptosis and vascular endothelial growth factor receptor 2 expression].

This study was aimed to investigate the effect of berberine on the proliferation and apoptosis of HL-60 cells, and the expression of vascular endothelial growth factor receptor 2 (VEGFR2) in HL-60 cells. Berberine (6 - 96 µg/ml) was added to the HL-60 cell line culture medium, the CCK-8 method was used to reveal the inhibitory effect on cell proliferation, the flow cytometry was used to determine the apoptosis rate and cell cycle in HL-60 cells treated with berberine. The expression of VEGFR2 mRNA and protein were examined by RT-PCR and Western blot respectively. The results showed that the berberine inhibited the proliferation of HL-60 cells and induced their apoptosis in dose- and time-dependent manners. With the increased concentration of berberine, the percentage of HL-60 cells in G(1) phase of cycle increased significantly, and the percentage of HL-60 cells in S phase decreased significantly. The expression of mRNA and protein of VEGFR2 decreased with the increased concentration of berberine. It is concluded that the berberine can inhibit HL-60 cell proliferation and induce HL-60 cell apoptosis. The expression of mRNA and proteins of VEGFR2 decreased after treatment with berberine.

[Inhibition of vegfr-2 gene expression by RNA interference].

This study was aimed to explore a novel potential gene target for therapy of malignant tumor. The recombinant expression plasmids of VEGF/VEGFR-2 were designed, constructed and then transfected into A549 cells by using lipofectamine. The expressions of VEGF/VEGFR-2 mRNA and protein were detected by RT-PCR and Western blot respectively. The cell proliferation was assayed by CCK-8 and the cell apoptosis was detected using Hoechst Staining. The results indicated that as compared with the blank control, pGenesil-1 and scramble groups, the interference effect of pGenesil-1-vegfr-2-shRNA-1 vector group was more obvious. As the expression of endogenous vegfr-2 mRNA decreased, the expression of VEGFR-2 protein decreased correspondingly. The proliferation of A549 cells was inhibited significantly by RNAi at 72 hours (p<0.01). The apoptosis of A549 cells was induced at 48 hours after being transfected with pGenesil-1-vegfr-2-shRNA-1 and the typical apoptosis morphology could be seen by fluorescence microscopy. It is concluded that the expression of vegfr-2 gene is inhibited effectively by vegfr-2 specific shRNA. The proliferation of A549 cells is inhibited significantly and the apoptosis of cells is induced. This result showed that VEGF/VEGFR-2 signaling pathway can be an effective target for the prevention and treatment of malignant tumor.