MCPNET: Development of an interpretable deep learning model based on multiple conformations of the compound for predicting developmental toxicity.

The development of deep learning models for predicting toxicological endpoints has shown great promise, but one of the challenges in the field is the accuracy and interpretability of these models. The bioactive conformation of a compound plays a critical role for it to bind in the target. It is a big issue to figure out the bioactive conformation in deep learning without the co-crystal structure or highly precise molecular simulations. In this study, we developed a deep learning framework of Multi-Conformation Point Network (MCPNET) to construct classification and regression models, respectively, based on electrostatic potential distributions on vdW surfaces around multiple conformations of the compound using a dataset of compounds with developmental toxicity in zebrafish embryo. MCPNET applied 3D multi-conformational surface point cloud to extract the molecular features for model training, which may be critical for capturing the structural diversity of compounds. The models achieved an accuracy of 85 % on the classification task and R2 of 0.66 on the regression task, outperforming traditional machine learning models and other deep learning models. The key feature of our model is its interpretability with the component visualization to identify the factors contributing to the prediction and to understand the compound action mechanism. MCPNET may predict the conformation quietly close to the bioactive conformation of a compound by attention-based multi-conformation pooling mechanism. Our results demonstrated the potential of deep learning based on 3D molecular representations in accurately predicting developmental toxicity. The source code is publicly available at https://github.com/Superlit-CC/MCPNET.

Synthesis of 2,3-Fused Quinazolinones via the Radical Cascade Pathway and Reaction Mechanistic Studies by DFT Calculations.

An efficient radical cascade cyclization of unactivated alkenes toward the synthesis of a series of ring-fused quinazolinones has been developed in moderate to excellent yields using commercially available ethers, alkanes, and alcohols, respectively, under a base-free condition in a short time without a transition metal as catalyst. Notably, the transformations can be carried out with the advantages of a broad substrate scope and high atomic economy. Density functional theory calculations and wavefunction analyses were performed to elucidate the radical reaction mechanism.

Reliable CA-(Q)SAR generation based on entropy weight optimized by grid search and correction factors.

Chromosome aberration (CA) is a serious genotoxicity of a compound, leading to carcinogenicity and developmental side effects. In the present manuscript, we developed a QSAR model for CA prediction using artificial intelligence methodologies. The reliable QSAR model was constructed based on an enlarged data set of 3208 compounds by optimizing machine learning and deep learning algorithms based on hyperparametric iterations and using multiple descriptors of molecular fingerprint in combination with drug-like molecular properties (MP) screened by entropy weight methodology on the open-source Python platform. Furthermore, molecular similarity for returning search and molecular connection index for additional descriptor were additionally introduced to differentiate the compounds with high similarity for correct CA prediction for QSAR model generation. The final generated CA-(Q)SAR model exhibited good prediction accuracy of 80.6%. The bias of the final model is about 0.9793. On the basis of generated QSAR model, data analyses were further performed to analyze the typical structure features in numerical intervals (MPI) of molecular properties MW, XlogP, and TPSA, respectively, for potential CA or non-CA toxicity with a normalized occurrence probability (NOP) more than 70%, which may provide useful clues for drug design of leads or candidate devoid of CA genotoxicity.

[Role of DNMT3a in Hydroquinone-Induced Hematopoietic Stem Cell Toxicity].

OBJECTIVE: To investigate the regulatory effect and mechanism of DNA methyltransferase 3A (DNMT3a) in hydroquinone-induced hematopoietic stem cell toxicity. METHODS: Cells (HSPC-1) were divided into 4 groups, that is A: normal HSPC-1; B: HQ-intervented HSPC-1; C: group B + pcDNA3 empty vector; D: group B + pcDNA3- DNMT3a. RT-qPCR and Western blot were used to detect the expression levels of DNMT3a and PARP-1 mRNA and protein, respectively. Cell morphology was observe; Cell viability and apoptosis rate of HSPC-1 were detected by MTT and flow cytometry, respectively. RESULTS: Compared with group A, the expression levels of DNMT3a mRNA and protein in HSPC-1 of group B were decreased, while PARP-1 mRNA and protein were increased (P<0.05); there was no significant difference in the above indexes between group C and group B; compared with group B, the expression levels of DNMT3a mRNA and protein showed increased, while PARP-1 mRNA and protein were decreased significantly in cells of group D transfected with DNMT3a (P<0.05). Cells in each group were transfected with DNMT3a and cultured for 24 h, HSPC-1 in group A showed high density growth and mononuclear fusion growth, while the number of HSPC-1 in group B and C decreased and grew slowly. Compared with group B and C, the cell growth rate of group D was accelerated. The MTT analysis showed that cell viability of HSPC-1 in group B were lower than that of group A at 24 h, 48 h and 72 h (P<0.05); after transfected with DNMT3a, the cell viability of HSPC-1 in group D were higher than that of group B at 24 h, 48 h and 72 h (P<0.05). The apoptosis rate of cells in group B was significantly higher than that of group A (P<0.001), while the apoptosis rate in group D was lower than that of group B (P<0.001). CONCLUSION: DNMT3a may be involved in the damage of hematopoietic stem cells induced by hydroquinone, which may be related to the regulation of PARP-1 activity by hydroquinone-inhibited DNMT3a.

Novel combined shield design for eye and face protection from COVID-19.

The World Health Organization emphasized the importance of goggles and face shields for protection of medical personnel at the outbreak of the COVID-19 pandemic. Unsurprisingly, almost all countries suffered from a critical supply shortage of goggles and face shields, as well as many other types of personal protective equipment (PPE), for a long period, owing to the lack of key medical material supplies and the inefficiency of existing fabrication methods arising from the need to avoid crowds during the outbreak of COVID-19. In this paper, we propose a novel combined shield design for eye and face protection that can be rapidly fabricated using three-dimensional printing technology. The designed prototype eye-face shield is accessible to the general public, offering more possibilities for yield improvement in PPE during emergent infectious disease events such as COVID-19.

The Geriatric Nutritional Risk Index as a Prognostic Factor in Patients with Advanced Non-Small-Cell Lung Cancer.

The Geriatric Nutritional Risk Index (GNRI) is widely applied as a prognostic factor in different cancers. We aimed to analyze the prognostic value of the GNRI in 257 patients diagnosed with advanced non-small-cell lung cancer (NSCLC). Patients with GNRI >98, 92-98, and <92 were grouped into normal, low risk and moderate/high risk groups, respectively. There were 45.1% patients at risk for malnutrition. Kaplan-Meier survival curves indicated that patients with lower GNRI scores had a poorer overall survival (OS). Two-year OS for normal, low risk and moderate/high risk groups were 57.4%, 42.3% and 15.8%, respectively. In multivariate survival analysis, GNRI (<92), body mass index (BMI, ≥24 kg/m2), combined therapy, hemoglobin and neutrophil-to-lymphocyte ratio (NLR) were independent prognostic factors of OS. Stratifying by age groups, GNRI (<92), hemoglobin and NLR were independent prognostic factors of OS in patients aged <65 years. GNRI (<92), smoking, BMI (≥24 kg/m2) and platelet-to-lymphocyte ratio were independent prognostic factors of OS in patients aged ≥65 years. In conclusion, GNRI was a significant prognostic factor in advanced NSCLC patients regardless of age. A decreased GNRI may be considered as a clinical trigger for nutritional support in advanced NSCLC patients, though additional studies are still required to confirm the best cut-point.

Preparation of gold nanorod-incorporated monolith for solid phase extraction of polycyclic aromatic hydrocarbons.

An organic-inorganic hybrid monolithic column doped with gold nanorods (AuNRs) was prepared and evaluated for solid phase extraction (SPE) of polycyclic aromatic hydrocarbons (PAHs). Excellent dispersibility of AuNRs in binary green porogen system consisting of 1-hexyl-3-methylimidazolium tetrafluoroborate and deep eutectic solvents (DESs) was confirmed by energy dispersive spectrometry (EDS). The particle size of the resulting AuNRs (70-90 nm) was thoroughly examined by a transmission electron microscopy (TEM). The redox system including ammonium persulfate (APS) and tetramethylethylenediamine (TEMED) was used to initiate in situ polymerization at 4 °C to prepare the hybrid monolith. The mesoporous structure of the AuNR hybrid monoliths was confirmed by scanning electron microscopy (SEM) and nitrogen adsorption. With enrichment factors (EFs) of 150- to 292-fold, the developed method was successfully applied to the determination of 10 PAHs in wastewater samples. The recoveries at a spiked level were in the range 84.9 to 99.5% with limit of detections (LODs) and relative standard deviations (RSDs) ranging from 0.02 to 0.10 µg L-1 and 1.5 to 4.2%, respectively. The correlation coefficients (R2) for the calibration function obtained were better 0.9991 for the target compounds. Compared to the AuNR-free monolith, the extraction efficiency of the AuNR-incorporated monolith is more than two times higher. The results indicated that the doping of AuNRs is an effective approach to obtain the hybrid monolithic column with good separation ability for PAHs. Graphical abstract.

Body mass index, waist circumference, and risk of hearing loss: a meta-analysis and systematic review of observational study.

BACKGROUND: Emerging evidence implicates excess weight as a potential risk factor for hearing loss. However, this association remained inconclusive. Therefore, we aimed to systematically and quantitatively review the published observational study on the association between body mass index (BMI) or waist circumference (WC) and hearing loss. METHODS: The odds ratios (ORs) or relative risks (RRs) with their 95% confidence intervals (CIs) were pooled under a random-effects model. Fourteen observational studies were eligible for the inclusion in the final analysis. RESULTS: In the meta-analysis of cross-sectional studies, the ORs for prevalent hearing loss were 1.10 (95% CI 0.88, 1.38) underweight, 1.14 (95% CI 0.99, 1.32) for overweight, OR 1.40 (95% CI 1.14, 1.72) for obesity, 1.14 (95% CI 1.04, 1.24) for each 5 kg/m2 increase in BMI, and 1.22 (95% CO 0.88. 1.68) for higher WC. In the meta-analysis of longitudinal studies, the RRs were 0.96 (95% CI 0.52, 1.79) for underweight, 1.15 (95% CI 1.04, 1.27) for overweight, 1.38 (95% CI 1.07, 1.79) for obesity, 1.15 (95% CI 1.01, 1.30) for each 5 kg/m2 increase in BMI, and 1.11 (95% CI 1.01, 1.22) for higher WC. CONCLUSIONS: In summary, our findings add weight to the evidence that elevated BMI and higher WC may be positively associated with the risk of hearing loss.

[Effects of WNK1 on Human Chronic Myeloid Leukemia K562 Cells via MAPK7 Phosphorylation and Its Relative Mechanism].

OBJECTIVE: To investigate the biological effects and mechanism of WNK1 on K562 cells through regulating MAPK7. METHODS: Cells were routinely cultured in vitro, the expression of WNK1 and MAPK7 in different blood tumor cell lines was analyzed by RT-qPCR and Western blot analysis.K562 cells were transfected with siRNA-WNK1 lentivirus.The effect of WNK1 on K562 cell proliferation was analyzed by using CCK-8 reagent.And K562 cell apoptosis was analyzed by using flow cytometry. The expression level of phosphorylated MAPK7 protein and total MAPK7 protein in K562 cells was analyzed by Western blot. RESULTS: The mRNA and protein of WNK1 were highly expressed in HL60, THP-1, U266 and K562 cells, however, the expressions were the highest in K562 cells (P<0.05), while the changes of mRNA and protein expressions of MAPK7 were not significantly in HL60, THP-1, U266 and K562 cells (P>0.05), but the phosphorylated MAPK7 expression was the highest in K562 cells (P<0.05). Proliferation of K562 cells transfected by WNK siRNA was significantly suppressed, and apoptosis was obviously increased (P<0.05). And the pMAPK7 protein expression in K562 cells transfected by WNK1 siRNA significantly decreased (P<0.05), however, the total MAPK7 protein expression in K562 cells showed no obvious change (P>0.05). CONCLUSION: WNK1 is highly expressed in K562 cells, which can promote the proliferation of K562 cells and reduce apoptosis probably by promoting phosphorylation of its downstream MAPK7.

The association between milk consumption and the metabolic syndrome: a cross-sectional study of the residents of Suzhou, China and a meta-analysis.

The association between milk consumption and the metabolic syndrome remains inconclusive, and data from Chinese populations are scarce. We conducted a cross-sectional study to investigate the association between milk consumption and the metabolic syndrome and its components among the residents of Suzhou Industrial Park, Suzhou, China. A total of 5149 participants were included in the final analysis. A logistic regression model was applied to estimate the OR and 95 % CI for the prevalence of the metabolic syndrome and its components according to milk consumption. In addition, the results of our study were further meta-analysed with other published observational studies to quantify the association between the highest v. lowest categories of milk consumption and the metabolic syndrome and its components. There was no significant difference in the odds of having the metabolic syndrome between milk consumers and non-milk consumers (OR 0·86, 95 % CI 0·73, 1·01). However, milk consumers had lower odds of having elevated waist circumference (OR 0·78, 95 % CI 0·67, 0·92), elevated TAG (OR 0·83, 95 % CI 0·70, 0·99) and elevated blood pressure (OR 0·85, 95 % CI 0·73, 0·99). When the results were pooled together with other published studies, higher milk consumption was inversely associated with the risk of the metabolic syndrome (relative risk 0·80, 95 % CI 0·72, 0·88) and its components (except elevated fasting blood glucose); however, these results should be treated with caution as high heterogeneity was observed. In summary, the currently available evidence from observational studies suggests that higher milk consumption may be inversely associated with the metabolic syndrome.

Lactoferrin Alleviates the Progression of Atherosclerosis in ApoE-/- Mice Fed with High-Fat/Cholesterol Diet Through Cholesterol Homeostasis.

Lactoferrin (LF) is a multifunctional glycoprotein and has beneficial effects on the regulation of lipid metabolism. However, whether LF supplementation alleviates the development of atherosclerosis (AS) remains unclear. In the present study, all of 48 male Apolipoprotein E-/- mice were fed with high-fat diet with 1.25% added cholesterol and divided to four treatment groups with either distilled water (HFCD), LF solutions at 2 mg/mL (low LF), 10 mg/mL (middle LF or MLF), or 20 mg/mL (high LF or HLF) for 12 weeks. Oral glucose tolerance tests (OGTT) were performed at weeks 0, 4, 8, and 12. At the end of the experiment, lipids in serum, liver, and feces were determined. The livers, whole aortas, and aortic sinuses were pathologically examined. The protein expression of factors related to cholesterol synthesis, absorption, and excretion were detected through western blot. No significant difference in body weight, food intake, and OGTT was observed among the four groups. Compared with the HFCD group, the MLF and HLF groups had significantly decreased serum and hepatic cholesterol levels and significantly increased fecal cholesterol contents. LF alleviated the hepatic steatosis and lipid droplet, especially in the MLF group. LF also significantly decreased the average lesion areas in the whole aorta, especially in the MLF group. On the other hand, LF downregulated hepatic protein expression of HMG-CoA reductase (the rate-limiting enzyme in cholesterol synthesis) and upregulated cholesterol 7-alpha hydroxylase (the rate-limiting enzyme in bile acid synthesis from cholesterol). LF also downregulated the intestinal expression of Niemann-Pick C1-like 1 protein, which is known to bind to a critical mediator of cholesterol absorption. In conclusion, LF supplementation alleviates the AS in mice on HFCD likely by reducing the synthesis and absorption of cholesterol and increasing cholesterol excretion.

Fasting blood glucose levels and prognosis in patients with non-small-cell lung cancer: a prospective cohort study in China.

PURPOSE: Non-small-cell lung cancer (NSCLC) is the most diagnosed lung cancer and is associated with poor prognosis. This study aimed to analyze whether fasting blood glucose (FBG) levels could provide prognostic information in Chinese patients with NSCLC, using the Suzhou Lung Cancer Survival study. PATIENTS AND METHODS: A prospective cohort study of adult patients with primary NSCLC was performed. The patients who were hospitalized between January 2016 and April 2018 in two hospitals affiliated with Soochow University were recruited. Patient information, including lifestyle habits and clinical and laboratory data, were collected through face-to-face interviews and evaluation of medical records. Follow-up was initiated from the date of patient enrollment until May 8, 2018 or until patient death. The long-term survival of patients was assessed every 6 months. Patient vital status was confirmed by using hospital records, telephone interview, or local death registration system. Cox proportional hazards regression was used to estimate hazard ratio and 95% confidence interval (CI) for death, with adjustment for cancer stage, medical treatments, smoking, and other potential confounders. RESULTS: A total of 387 patients were included in the analysis, and the numbers (percentages) of patients with stages I, II, III, and IV NSCLC were 53 (13.7%), 41 (10.6%), 64 (16.5%), and 215 (55.6%), respectively. The median duration of follow-up was 19.1 months. Compared with patients in the second tertile of FBG, the HRs for mortality were 2.16 (95% CI: 1.26-3.73) and 1.87 (95% CI: 1.03-3.42) for those in the lowest one and diabetic group, respectively. Subgroup analysis according to various patient characteristics confirmed these associations. CONCLUSION: Diabetes and low FBG could be important predictors of death in patients with NSCLC. Maintaining appropriate blood glucose levels may improve prognosis in patients with NSCLC.

Post-diagnostic C-reactive protein and albumin predict survival in Chinese patients with non-small cell lung cancer: a prospective cohort study.

Non-small cell lung cancer (NSCLC) is the most commonly diagnosed lung cancer and is associated with poor prognosis. This study aimed to analyze if serum C-reactive protein (CRP), albumin (Alb), and CRP/Alb ratio could provide prognostic information in patients with NSCLC. 387 patients with primary NSCLC were included in this analysis. Cox proportional hazards regression was used to estimate hazard ratio (HR) and 95% confidence interval (CI) of death with adjustment for some potential confounders. The multivariate regression analyses revealed the statistically significant associations of decreased survival of patients with NSCLC with elevated CRP, decreased Alb, and elevated CRP/Alb ratio. The HRs of mortality were 1.56 (95% CI: 0.80-3.04) and 2.64 (95% CI: 1.35-5.16) for patients in the second and the highest tertiles of CRP (P-trend = 0.003). For albumin, the HR was 0.50 (95% CI: 0.29-0.85) for the normal group. The CRP/Alb ratio strongly predicted the survival of patients in the highest tertile with a fourfold risk of dying compared with those in the lowest tertile (HR = 4.14, 95% CI: 2.15-7.98). The subgroup analysis according to various patient characteristics confirmed these associations. In conclusion, serum CRP, albumin, and CRP/Alb ratio are predictive of survival for Chinese patients with NSCLC.

Irradiated VEGF164-modified tumor cell vaccine protected mice from the parental tumor challenge.

Vascular endothelial growth factor (VEGF) is an important regulating molecule of angiogenesis in tumor formation and progression. Cancer cells always secrete VEGF to stimulate angiogenesis that facilitate growth and invasion of the tumor. In this study, we established a VEGF164 overexpressing LL/2 lung cancer cell model and found that the postirradiated VEGF164-modified tumor cells protected the host against the challenge with LL/2 wild-type tumor cells. Histochemical assay showed that there were large areas of tumor necrosis with macrophage infiltration in the mice vaccinated with the VEGF164-modified tumor vaccine. T-cells isolated from the vaccinated mice showed cytotoxicity against the parental tumor cells in a dose-dependent manner. Meanwhile, sera from the mice vaccinated with LL/2-VEGF164 showed higher titers of antibodies against parental tumor cells compared with the nonvaccinated groups. Our results indicated that VEGF164-modified tumor vaccine could modulate host antitumor immune response and hold therapeutic potential for cancer.

Macromolecular crowding of molecular imprinting: A facile pathway to produce drug delivery devices for zero-order sustained release.

This paper reported the facile fabrication of drug delivery devices for zero-order sustained release by molecular crowding strategy of molecularly imprinting technology. Crowding-assisted molecularly imprinting polymers (MIPs) matrices were prepared by free-radical precipitation polymerization using aminoglutethimide (AG) as a model drug. The crowding effect was achieved by adding polystyrene as a macromolecular co-solute in pre-polymerization mixture. The MIP prepared under the non-MMC condition and the two corresponding non-imprinted particles were tested as controlled vehicles. The release profiles presented zero-order behaviors from two crowding-assisted polymers, the duration of approximately 18h for the crowding-assisted MIP and 10h for the crowding-assisted NIP, respectively while AG were all very rapid released from the other two controlled particles (85% occurring in the first hour). The BET surface area and pore volume of the crowding-assisted MIP were about ten times than those of the controlled MIP. The value of imprinting factor is 6.02 for the crowding-assisted MIP and 1.19 for the controlled MIP evaluated by the equilibrium adsorption experiment. Furthermore, the values of effective diffusivity (Deff) obtained from crowding-assisted MIP (10(-17)cm(2)/s) was about two orders of magnitude smaller than those from the controlled MIP, although the values of free drug diffusivity (D) were all found in the order of 10(-13)cm(2)/s. Compared with the commercial AG tablet, the MMC-assisted MIP gave a markedly high relative bioavailability of 266.3%, whereas the MMC-assisted NIP gave only 57.7%. The results indicated that the MMC condition can modulate the polymer networks approaciate to zero-order release of the drug and maintain the molecular memory pockets, even if under the poor polymerization conditions of MIPs preparation.

Effective induction therapy with subcutaneous administration of bortezomib for newly diagnosed POEMS syndrome: a case report and a review of the literature.

POEMS syndrome is characterized by polyneuropathy, organomegaly, endocrinopathy, monoclonal gammopathy and skin changes. Bortezomib is an important component of the chemotherapy regimen associated with multiple myeloma, and has been previously applied to POEMS syndrome. We present a 56-year-old Chinese man who was given subcutaneous administration of bortezomib as part of the BDex (bortezomib-dexamethasone) regimen for his POEMS syndrome. The peripheral neuropathy and laboratory-test results of the patient improved dramatically with 4 cycles of treatment, resulting in a complete response. In addition, the treatment was well tolerated and adequate peripheral blood hematopoietic stem cells were collected for an ensuing autologous stem cell transplant.

Molecularly imprinted nanoparticles with nontailing peaks in capillary electrochromatography.

The combination of microparticles of molecularly imprinted polymers (MIPs) with partial filling capillary electrochromatography (CEC) has previously been demonstrated for the enantiomer separation. In this paper, precipitation polymerization was used to prepare d-zopiclone imprinted nanoparticles (50-80 nm) by a strategy of the dilution of pre-polymerization mixtures. The influence of some important parameters on the preparation of MIPs nanoparticles, including template to monomer ratio, type and amount of cross-linking monomer, and functional monomer composition ratio were investigated. In addition, the effect of separation condition, e.g., organic modifier content, pH value and salt concentration of buffer, on the electrochromatographic behavior of the MIP nanoparticles were studied. In spite of lower selectivity factor (1.11), high column performance (theoretical plates 41,400) of template was obtained and the resolution of enantiomers separation was 4.75 under the optimized conditions. Compared to the previously reported MIP microparticles, the MIP nanoparticles showed good peak symmetry and an ability of high speed separation (<15 min) in CEC mode.

Graphene oxide used as a carrier for adriamycin can reverse drug resistance in breast cancer cells.

This study evaluates the reversal effects of graphene oxide (GO) used as a carrier for adriamycin (ADR) in cancer drug resistance, and provides a preliminary investigation into the reversal mechanism. ADR was loaded onto the GO surface (ADR-GO) by physical mixing and drug loading content was found to be high, up to 93.6%. In vitro releases of ADR from ADR-GO were studied using a dialysis method, and they exhibited a significant pH-sensitive property. Cell experiments showed that GO significantly enhanced the accumulation of ADR in MCF-7/ADR cells (an ADR resistant breast cancer cell line) and exhibited much higher cytotoxicity than free ADR, suggesting that ADR-GO could effectively reverse ADR resistance of MCF-7/ADR, with the reversal index reaching 8.35. Microscopy studies found that GO could effectively carry drug molecules into cells in both endocytosis-dependent and independent manners. In conclusion, use of GO as a carrier for chemotherapeutic agents is favorable for the treatment of drug resistant cancers.

[Effect of diagnostic ultrasound-mediated microbubble contrast and urokinase on augmentation thrombolysis and optimization of the major parameters: an in vitro study].

OBJECTIVE: To investigate the efficacy of diagnostic ultrasound and microbubble contrast (MB) on enhancing thrombolysis in combination with urokinase (UK) and to determine the optimal combination for thrombolysis in vitro. METHODS: Four types of standardized red thrombus were prepared in vitro, including 3-hour-old (3 h), 6-hour-old (6 h), 12-hour-old (12 h), and 24-hour-old (24 h). The major parameters for the designed experiments included transmit powers of ultrasound (factor A, 5%, 25%, 50%, 100%), MB volumes (factor B, 50 microL, 100 microL, 200 microL, 400 microL), UK concentrations (factor C, 100 U/mL, 200 U/mL, 400 U/mL, 800 U/mL), and lysis time (factor D, 10 min, 20 min, 30 min, 40 min). An orthogonal array experimental design (OAD) based on four levels L16 (4(5)) of the above four parameters was employed to optimize the thrombolysis conditions. During the procedure of thrombolysis, the diagnostic ultrasound frequency was fixed at 1.82 MHz. The histopathological changes measured by HE staining and scanning electron microscope (SEM) were carried out to observe the clots before and after thrombolysis. The loss of clot weight before and after treatment was measured to determine the lysis efficiency (LE). Analysis of variance (ANOVA) was performed to assess the LE according to the L16 (4(5)) matrix. RESULTS: The HE staining and SEM observation of thrombolysis under the following experimental conditions of 5% ultrasound transmit power, 400 microL MB volume, 800 U/mL UK concentration, and 40 min lysis time showed remarkable disaggregation of fibrin nets. The above four factors had significant impact on thrombus (all P < 0.05), among which UK concentrations (factor C) was the most significant one. The optimal scheme was determined as a C4-D4-A1-B4 mode, with UK concentration 800 U/mL, lysis time 40 min, transmit power 5%, and MB volume 400 microL, respectively. The LE curves for 3h clots were superior to the others. The lysis efficiencies for the clots showed significant differences among different type of thrombus (all P < 0.05). CONCLUSION: 1.82 MHz diagnostic ultrasound and microbubble contrast can be applied to augment thrombolysis in vitro even with a transmit power as low as 5%. Under the condition of fixed ultrasound frequency, the LE of thrombus increase with increased UK concentrations, lysis time and MB volumes, and decrease with increased thrombus ages.