Development of a nomogram-based model combining intra- and peritumoral ultrasound radiomics with clinical features for differentiating benign from malignant in Breast Imaging Reporting and Data System category 3-5 nodules.

Background: The differences in benign and malignant breast tumors are not only within the nodules but also involve changes in the surrounding tissues. Radiomics can reveal many details that are not discernible to the naked eye. This study aimed to distinguish between benign and malignant breast nodules using an ultrasound-based intra- and peritumoral radiomics model. Methods: This study retrospectively collected the information from 379 patients with Breast Imaging Reporting and Data System (BI-RADS) category 3-5 nodules and clear pathological diagnosis of breast nodules screened by routine ultrasound examination in the Sixth People's Hospital Affiliated to Medical College of Shanghai Jiao Tong University from January 2017 to December 2022. The largest dimension of the lesion on the 2D ultrasound image was selected to outline the area of interest which was conformally and outwardly expanded automatically by 5 mm to extract intra- and peritumor radiomics features. The included cases were randomly divided into training sets and test sets in a ratio of 7:3. The optimal features of the included models were retained by statistical and machine learning methods of dimensionality reduction, and logistic regression was used as the classifier to build an intratumoral model and a combined intratumoral-peritumoral radiomics model, respectively; through single-factor and multifactor logistic regression, the optimal features that could predict benign and malignant breast tumors were screened. The clinical and imaging models were established by selecting independent risk factors as clinical and imaging features through univariate and multifactorial logistic regression. Results: Among 379 BI-RADS category 3-5 breast nodules, there were 124 malignant nodules and 255 benign nodules; patients were aged 14 to 88 (46.22±15.51) years, and the age differences, radiomics score, and mass diameter between the training and test sets were not statistically significant (P>0.05). The intra- and peritumor radiomics model had an area under the curve (AUC) of 0.840 [95% confidence interval (CI): 0.766-0.914] in the test set. The model with intra- and peritumoral ultrasound radiomics features combined with clinical features had an AUC value of 0.960 (95% CI: 0.920-0.999). Conclusions: The nomogram, developed using intratumoral and peritumoral radiomics features combined with clinical risk features, demonstrated superior performance in distinguishing between benign and malignant BI-RADS 3-5 lesions.

Transiently impaired endothelial function during thyroid hormone withdrawal in differentiated thyroid cancer patients.

Purpose: Endothelial dysfunction, which was associated with chronic hypothyroidism, was an early event in atherosclerosis. Whether short-term hypothyroidism following thyroxine withdrawal during radioiodine (RAI) therapy was associated with endothelial dysfunction in patients with differentiated thyroid cancer (DTC) was unclear. Aim of the study was to assess whether short-term hypothyroidism could impair endothelial function and the accompanied metabolic changes in the whole process of RAI therapy. Methods: We recruited fifty-one patients who underwent total thyroidectomy surgery and would accept RAI therapy for DTC. We analyzed thyroid function, endothelial function and serum lipids levels of the patients at three time points: the day before thyroxine withdrawal(P1), the day before 131I administration(P2) and 4-6 weeks after RAI therapy(P3). A high-resolution ultrasound named flow-mediated dilation (FMD) was used to measure endothelial function of the patients. Results: We analyzed the changes of FMD, thyroid function and lipids at three time points. FMD(P2) decreased significantly compared to FMD(P1) (P1vsP2, 8.05 ± 1.55vs 7.26 ± 1.50, p<0.001). There was no significant difference between FMD(P3) and FMD(P1) after restoring TSH (thyroid stimulating hormone) suppression therapy (P1 vs P3, 8.05 ± 1.55 vs 7.79 ± 1.38, p=0.146). Among all parameters, the change of low-density lipoprotein (ΔLDL) was the only factor correlated negatively with the change of FMD (ΔFMD) throughout the RAI therapy process (P1-2, r=-0.326, p=0.020; P2-3, r=-0.306, p=0.029). Conclusion: Endothelial function was transiently impaired in DTC patients at short-term hypothyroidism state during the RAI therapy, and immediately returned to the initial state after restoring TSH suppression therapy.

Efficacy of low-intensity pulsed ultrasound in the treatment of COVID-19 pneumonia.

PURPOSE: As a public health emergency of international concern, coronavirus disease 2019 (COVID-19) still lacks specific antiviral drugs, and symptomatic treatment is currently the mainstay. The overactivated inflammatory response in COVID-19 patients is associated with a high risk of critical illness or even death. Low-intensity pulsed ultrasound (LIPUS) can mitigate inflammation and inhibit edema formation. We aimed to investigate the efficacy of LIPUS therapy for COVID-19 pneumonia. MATERIALS AND METHODS: 62 patients were randomly assigned to a treatment group (LIPUS treatment area - Group 1; self-control area - Group 2) and an external control group (Group 3). The primary outcomes were the volume absorption rate (VAR) and the area absorption rate (AAR) of lung inflammation in CT images. RESULTS: After an average duration of treatment 7.2 days, there were significant differences in AAR and VAR between Group 1 and Group 2 (AAR 0.25 vs 0.12, p=0.013; VAR 0.35 vs 0.11, p=0.005), and between Group 1 and Group 3 (AAR 0.25 vs 0.11, p=0.047; VAR 0.35 vs 0.19, p=0.042). Neither AAR nor VAR was statistically different between Group 2 and Group 3. After treatment, C-reactive protein, interleukin-6, leukocyte, and fingertip arterial oxygen saturation (SaO2) improved in Group 1, while in Group 3 only fingertip SaO2 increased. CONCLUSION: LIPUS therapy reduced lung inflammation and serum inflammatory factor levels in hospitalized COVID-19 patients, which might be a major advancement in COVID-19 pneumonia therapy.

Efficacy of low-intensity pulsed ultrasound for the treatment of viral pneumonia: study protocol for a randomized controlled trial.

BACKGROUND: Viral pneumonia has always been a problem faced by clinicians because of its insidious onset, strong infectivity, and lack of effective drugs. Patients with advanced age or underlying diseases may experience more severe symptoms and are prone to severe ventilation dysfunction. Reducing pulmonary inflammation and improving clinical symptoms is the focus of current treatment. Low-intensity pulsed ultrasound (LIPUS) can mitigate inflammation and inhibit edema formation. We aimed to investigate the efficacy of therapeutic LIPUS in improving lung inflammation in hospitalized patients with viral pneumonia. METHODS: Sixty eligible participants with clinically confirmed viral pneumonia will be assigned to either (1) intervention group (LIPUS stimulus), (2) control group (null stimulus), or (3) self-control group (LIPUS stimulated areas versus non-stimulated areas). The primary outcome will be the difference in the extent of absorption and dissipation of lung inflammation on computed tomography. Secondary outcomes include changes in lung inflammation on ultrasonography images, pulmonary function, blood gas analysis, fingertip arterial oxygen saturation, serum inflammatory factor levels, the sputum excretion volume, time to the disappearance of pulmonary rales, pneumonia status score, and course of pneumonia. Adverse events will be recorded. DISCUSSION: This study is the first clinical study of the efficacy of therapeutic LIPUS in the treatment of viral pneumonia. Given that the current clinical recovery mainly depends on the body's self-limiting and conventional symptomatic treatment, LIPUS, as a new therapy method, might be a major advance in the treatment of viral pneumonia. TRIAL REGISTRATION: ChiCTR2200059550 Chinese Clinical Trial Registry, May 3, 2022.

Three-dimensional pelvic ultrasound is a practical tool for the assessment of anal fistula.

OBJECTIVE: This study aims to investigate the diagnostic value of three-dimensional pelvic ultrasound in the preoperative assessment of anal fistula compared with findings of MRI and surgery. METHODS: A total of 67 patients (62 males) with suspected anal fistula were analyzed retrospectively. Preoperative three-dimensional pelvic ultrasound and magnetic resonance imaging were performed in all patients. The number of internal openings and the type of fistula were recorded. The accuracy of three-dimensional pelvic ultrasound was determined by comparing these parameters with surgical outcomes. RESULTS: At surgery, 5 (6%) were extrasphincteric, 10 (12%) were suprasphincteric, 11 (14%) were intersphincteric, and 55 (68%) were transsphincteric. There was no significant difference in the accuracy of pelvic 3D US and MRI, based on internal openings (97.92%, 94.79%), anal fistulas (97.01%, 94.03%), and those under Parks classification (97.53%, 93.83%). CONCLUSION: Three-dimensional pelvic ultrasound is a reproducible and accurate method for determining the type of fistula and detecting internal openings and anal fistulas.

Nomogram based on preoperative conventional ultrasound and shear wave velocity for predicting central lymph node metastasis in papillary thyroid carcinoma.

OBJECTIVE: To establish a nomogram for predicting cervical lymph node metastasis (CLNM) based on the preoperative conventional ultrasound (US) and shear wave velocity (SWV) features of papillary thyroid carcinoma (PTC). METHODS: A total of 101 patients with pathologically confirmed thyroid nodules were enrolled. These patients were divided into the CLNM-positive (n = 40) and CLNM-negative groups (n = 61). All patients underwent the preoperative conventional US and shear wave elastography (SWE) evaluation, and the US parameters and SWV data were collected. The association between SWV ratio and CLNM was compared to assess the diagnostic efficacy of SWV ratio alone as opposed to SWV ratio in combination with the conventional US for predicting CLNM. RESULTS: There were significant differences in shape, microcalcification, capsule contact, SWV mean, and SWV ratio between the CLNM-positive and CLNM-negative groups (P < 0.05). Logistic regression analysis showed that taller-than-wide shape, microcalcification, capsule contact, and SWV ratio > 1.3 were risk factors for CLNM; Logistic(P)=-6.93 + 1.647 * (microcalcification)+1.138 * (taller-than-wide-shape)+1.612 * (capsule contact)+2.933 * (SWV ratio > 1.3). The area under the curve (AUC) of the receiver operating characteristic (ROC) of the model for CLNM prediction was 0.87, with 81.19% accuracy, 77.5% sensitivity, and 85.25% specificity. CONCLUSION: The nomogram based on conventional US imaging in combination with SWV ratio has the potential for preoperative CLNM risk assessment. This nomogram serves as a useful clinical tool for active surveillance and treatment decisions.

Long noncoding RNA DLEU2 promotes growth and invasion of hepatocellular carcinoma by regulating miR-30a-5p/PTP4A1 axis.

Increasing data indicate that long noncoding RNA (lncRNA) DLEU2 is implicated in carcinogenesis in multiple malignancies including hepatocellular carcinoma (HCC). However, the role and molecular mechanism by which lncRNA DLEU2 contributes to HCC remain unknown. The association of lncRNA DLEU2 with clinicopathological characteristics and prognosis in patients with HCC was analyzed by qRT-PCR, and public TCGA dataset. CCK-8, colony formation and Transwell assays were performed to verify the role of lncRNA DLEU2 in HCC. RNA immunoprecipitation (RIP), luciferase gene report and qRT-PCR assays were employed to uncover lncRNA DLEU2-spevific binding with miR-30a-5p. The effect of lncRNA DLEU2 and (or) miR-30a-5p on PTP4A1 expression was examined by Western blot analysis. As a consequence, we found that lncRNA DLEU2 was upregulated in HCC tissue samples and associated with distant metastasis and poor survival in patients with HCC. Knockdown of lncRNA DLEU2 impaired HCC cell proliferation, colony formation and invasion, but ectopic expression of lncRNA DLEU2 abolished these effects. Furthermore, lncRNA DLEU2 harbored a negative correlation and specific binding with miR-30a-5p in HCC cells. Knockdown of lncRNA DLEU2 upregulated miR-30a-5p, but downregulated its target PTP4A1, and miR-30a-5p abrogated lncRNA DLEU2-induced tumor-promoting effects and PTP4A1 upregulation. Taken together, our findings demonstrate that lncRNA DLEU2 promotes growth and invasion of HCC cells by regulating miR-30a-5p/ PTP4A1 axis.

A Prediction Equation to Estimate Vascular Endothelial Function in Different Body Mass Index Populations.

Objective: Vascular endothelial dysfunction is considered an early predictor of endothelial injury and the initiating factor of atherosclerosis (AS). Brachial artery flow-mediated dilation (FMD) can detect endothelial injury early and provide important prognostic information beyond traditional cardiovascular (CV) risk factors. This study aimed to find the influencing factors of FMD and develop a simple prediction model in populations with different body mass indices (BMIs). Methods: In total, 420 volunteers with different BMIs were recruited in our study. Subjects were randomly assigned to the derivation and validation cohorts (the ratio of the two was 1:2) with simple random sampling. The former was used for influencing factors searching and model construction of FMD and the latter was used for verification and performance evaluation. Results: The population was divided into two groups, i.e., 140 people in the derivation group and 280 people in the verification group. Analyzing in the training data, we found that females had higher FMD than males (p < 0.05), and FMD decreased with age (p < 0.05). In people with diabetes, hypertension or obesity, FMD was lower than that in normal individuals (p < 0.05). Through correlation analysis and linear regression, we found the main influencing factors of FMD: BMI, age, waist-to-hip radio (WHR), aspartate aminotransferase (AST) and low-density lipoprotein (LDL). And we developed a simple FMD prediction model: FMD = -0.096BMI-0.069age-4.551WHR-0.015AST-0.242LDL+17.938, where R2 = 0.599, and adjusted R2 = 0.583. There was no statistically significant difference between the actual FMD and the predicted FMD in the verification group (p > 0.05). The intra-class correlation coefficient (ICC) was 0.77. In a Bland-Altman plot, the actual FMD and the predicted FMD also showed good agreement. This prediction model had good hints in CV risk stratification (area under curve [AUC]: 0.780, 95 % confidence intervals [95% CI]: 0.708-0.852, p < 0.001), with a sensitivity and specificity of 73.8 and 72.1%, respectively. Conclusions: Males, older, obesity, hypertension, diabetes, smoking, etc. were risk factors for FMD, which was closely related to CV disease (CVD). We developed a simple equation to predict FMD, which showed good agreement between the training and validation groups. And it would greatly simplify clinical work and may help physicians follow up the condition and monitor therapeutic effect. But further validation and modification bears great significance.

Low-frequency ultrasound combined with microbubbles improves gene transfection in prostate cancer cells in vitro and in vivo.

OBJECTIVES: The aim of this study is to explore whether low-frequency ultrasound combined with microbubbles improves pEGFP genes transfection into human prostate cancer cells. METHODS: Ultrasound with frequency of 80 kHz and duty cycle of 50% was adopted in the study; in in vitro experiments, cell lysis, and membrane damage were evaluated after ultrasound exposure; and the membrane continuity and transfection efficiency were observed by transmission electron microscope and laser scanner, respectively. Human prostate cancer xenograft models were exposed to ultrasound and transfection efficiency and histological examination were analyzed. RESULTS: Compared with the control group, ultrasound combined with microbubbles significantly improves gene transfection efficiency (P < .05). In in vitro study, ultrasound combined with microbubbles resulted in cell lysis and the interruption of cell membrane continuity, and its average transfection efficiency was 9.9%; the green fluorescence intensity was 15.2% in the ultrasound combined with microbubbles group in vivo; both values were higher than that in the control group (P < .05). CONCLUSION: Low-frequency ultrasound combined with microbubbles could be used as a method to promote gene transfection in prostate cancer cells.

Marriage of Virus-Mimic Surface Topology and Microbubble-Assisted Ultrasound for Enhanced Intratumor Accumulation and Improved Cancer Theranostics.

The low delivery efficiency of nanoparticles to solid tumors greatly reduces the therapeutic efficacy and safety which is closely related to low permeability and poor distribution at tumor sites. In this work, an "intrinsic plus extrinsic superiority" administration strategy is proposed to dramatically enhance the mean delivery efficiency of nanoparticles in prostate cancer to 6.84% of injected dose, compared to 1.42% as the maximum in prostate cancer in the previously reported study. Specifically, the intrinsic superiority refers to the virus-mimic surface topology of the nanoparticles for enhanced nano-bio interactions. Meanwhile, the extrinsic stimuli of microbubble-assisted low-frequency ultrasound is to enhance permeability of biological barriers and improve intratumor distribution. The enhanced intratumor enrichment can be verified by photoacoustic resonance imaging, fluorescence imaging, and magnetic resonance imaging in this multifunctional nanoplatform, which also facilitates excellent anticancer effect of photothermal treatment, photodynamic treatment, and sonodynamic treatment via combined laser and ultrasound irradiation. This study confirms the significant advance in nanoparticle accumulation in multiple tumor models, which provides an innovative delivery paradigm to improve intratumor accumulation of nanotherapeutics.

Serum Creatinine-to-Cystatin C Ratio in the Progression Monitoring of Non-alcoholic Fatty Liver Disease.

BACKGROUND: The simultaneous assessment of visceral adiposity and muscle mass might be useful to monitor the risk of non-alcoholic fatty liver disease (NAFLD) progression in large population. We aimed to investigate the value of serum creatinine-to-cystatin C ratio (CCR) in evaluating these two parameters and predicting liver steatosis and fibrosis. METHODS: 154 overweight/obese inpatients (49 males, 105 females) scheduled for bariatric surgery and 49 non-overweight/obese volunteers (18 males, 31 females) responded to the hospital advertisement were involved in the cross-sectional study. Liver steatosis and fibrosis were diagnosed with transient elastography (TE). The psoas muscle area (PMA) and visceral fat area (VFA) were measured using magnetic resonance imaging. RESULTS: The body mass index, insulin resistance, and lipid profiles showed significant differences between the CCR tertiles. Multiple regression analyses revealed that the CCR was significantly associated with the controlled attenuation parameter (ß = -0.30, P = 0.006 in males; ß = -0.19, P = 0.017 in females) and liver stiffness measurements in males (ß = -0.246, P = 0.044). A low CCR was associated with moderate-to-severe steatosis (P < 0.001), significant liver fibrosis (P < 0.01), and excellent predictive power for these two conditions (P < 0.01). The CCR had a negative correlation with the VFA/PMA ratio (r = -0.584, P < 0.001 in males; r = -0.569, P < 0.001 in females). CONCLUSIONS: The CCR is a serum marker for muscle-adjusted visceral fat mass, and a low CCR is associated with an increased risk of progressive NAFLD.

CircANKRD52 Promotes the Tumorigenesis of Hepatocellular Carcinoma by Sponging miR-497-5p and Upregulating BIRC5 Expression.

CircRNAs participate in the pathogenesis of a variety of cancers. Previous studies showed that baculoviral IAP repeat containing 5 (BIRC5) can promote tumor progression. But, the mechanisms by which circRNAs regulate BIRC5 expression in hepatocellular carcinoma (HCC) remain unknown. The clinical prognosis of BIRC5 or miR-497-5p expression in patients with HCC was assessed by TCGA RNA-seq dataset. hsa_circ_0026939 (circANKRD52) or BIRC5 was identified to bind with miR-497-5p by luciferase gene report, RIP and circRIP assays. MTT, colony formation, Transwell assays and a xenograft tumor model were used to estimate the role of miR-497-5p or circANKRD52 in HCC cells. As a result, we found that elevated expression of BIRC5 or decreased expression of miR-497-5p was linked to poor survival in HCC. Restored expression of miR-497-5p repressed cell proliferation, colony formation and invasiveness by targeting BIRC5, but its inhibitor showed the opposite results. Furthermore, circANKRD52 possessed a tumor-promoting effect by acting as a sponge of miR-497-5p and thereby upregulated BIRC5 in HCC cells. In conclusion, our findings demonstrated that circANKRD52 enhances the tumorigenesis of HCC by sponging miR-497-5p and upregulating BIRC5 expression.

Relationship Between Morphologic Characteristics of Ultrasonic Calcification in Thyroid Nodules and Thyroid Carcinoma.

The aim of this study was to investigate the relationship between morphologic characteristics of the calcifications detected by ultrasound in thyroid nodules and thyroid carcinoma. Morphologic characteristics of the calcifications on pre-operative ultrasound examinations of thyroids were compared with post-operative pathologic diagnoses in 543 patients undergoing thyroid surgery. Calcifications were divided into microcalcifications (≤2 mm) and macrocalcifications (>2 mm), and the latter were divided into eggshell calcifications in a row, eggshell discontinuous calcifications, irregular calcifications and multilayer-like calcifications, labeled types I-V. We found that thyroid microcalcifications and partial macrocalcifications, such as eggshell discontinuous calcifications, and multilayer-like calcifications were associated with thyroid carcinoma. In conclusion, microcalcifications were more commonly found in malignant thyroid nodules, particularly in papillary thyroid carcinoma. Eggshell discontinuous macrocalcifications and multilayer-like macrocalcifications also occurred mainly in malignant nodules.

Vascular endothelial growth factor suppresses dendritic cells function of human prostate cancer.

PURPOSE: Prostate cancer (PCa) patients often have dendritic cell (DC) function defects, but the mechanism is not clear. The aim of this study was to detect the effect of vascular endothelial growth factor (VEGF) in mature DCs. PATIENTS AND METHODS: In this study, we chose 30 PCa patients, 10 prostatic intraepithelial neoplasia (PIN) patients and 30 benign prostatic hyperplasia (BPH) patients, and compared the composition of peripheral blood T cells, the composition and function of local dendritic cells in prostate tissue, and the density of local VEGF. RESULTS: The results showed that the numbers of total DCs, mature and functional DCs, and CD4+ T cells were inhibited in PCa, and the inhibitory effect was enhanced with increased malignancy. In addition, the infiltration density of VEGF-positive cells was increased in PCa, and this increase was associated with an increased malignant degree of PCa. The inhibition of tumor immunity in patients with PCa is achieved by inhibiting the function of dendritic cells. CONCLUSION: VEGF plays an important role in the inhibition of the maturation and function of dendritic cells, and this inhibition is gradually increased with an increasing malignant degree of PCa.

Low-frequency ultrasound-induced VEGF suppression and synergy with dendritic cell-mediated anti-tumor immunity in murine prostate cancer cells in vitro.

High tumor vascular endothelial growth factor (VEGF) levels are associated with poor treatment outcomes in prostate cancer (PCa), and immune deficiency in the PCa microenvironment, especially suppression of dendritic cell (DC) proliferation, has been confirmed. In this study, we (1) investigated whether VEGF participates in DC suppression in murine PCa cells (RM-1), (2) down-regulated VEGF expression using low-frequency ultrasound and microbubbles (UM), and (3) further explored any synergistic effect on immunological activation. DCs from the bone marrow of BALB/c mice were stimulated by the addition of cytokines (granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin-4 (IL-4)), and we analyzed their proliferation status via flow cytometric recognition of the surface antigen markers CD11c and CD83. The results demonstrated that co-culture with RM-1 cells markedly inhibited expression of the general marker CD11c and the mature marker CD83; UM weakened this inhibition by down-regulating VEGF expression. T lymphocytes were extracted from murine spleens, and CD4 and CD8a were identified as the biomarkers of activated cells participating in the anti-tumor immune response. When DCs, T lymphocytes and RM-1 cells were co-cultured, cell migration and invasion assays and cytoactive detection showed that UM could not only directly suppress PCa cell evolution but also promote activation of anti-tumor immunocytes in the VEGF-inhibited microenvironment.

Low-frequency ultrasound-mediated microvessel disruption combined with docetaxel to treat prostate carcinoma xenografts in nude mice: A novel type of chemoembolization.

The aim of the present study was to investigate whether low-frequency ultrasound (US)-mediated microvessel disruption combined with docetaxel (DTX) can be used as a novel type of chemoembolization. Mice were assigned to four groups: i) The USMB group, treated with low-frequency US combined with microbubbles (USMB); ii) the DTX group, treated with DTX; iii) the USMB + DTX group, treated with combined therapy; and iv) the control group, which was untreated. Immediately after the first treatment, the average peak intensity (API) on contrast-enhanced US was calculated, and tumors were excised for hematoxylin and eosin (HE) staining. At 2 weeks post-treatment, the tumor volumes and wet weights were calculated, and tumors were excised for immunohistochemistry to calculate apoptotic index (AI), proliferative index (PI) and microvessel density (MVD) values. Immediately after the first treatment, in the DTX and control groups, the tumors demonstrated abundant perfusion enhancement, while in the USMB + DTX and USMB groups, blood perfusion of the tumors was interrupted. Compared with that of the control group, the API was significantly lower in the USMB + DTX USMB groups (all P<0.001). HE staining showed that tumor microvasculature was disrupted into flaky hematomas and severely dilated microvessels in the USMB + DTX and USMB groups. In the DTX and control groups, there was no distinct evidence of the disruption and dilation of blood microvessels. At the end of the treatment, the mean tumor inhibition ratio was 73.33, 46.67 and 33.33% for the USMB + DTX, DTX and USMB groups, respectively. The USMB + DTX group had the highest AI, and the lowest PI and MVD compared with the other groups, although the difference between the USMB + DTX and DTX groups with regard to PI and MVD was not significant (USMB + DTX vs. DTX group, P=0.345 and P=0.059, respectively). In conclusion, as a novel type of chemoembolization, USMB combined with DTX is more effective than USMB or DTX alone in inhibiting tumor growth via the enhancement of apoptosis, and the suppression of proliferation and angiogenesis.

Liposome-mediated transfection of wild-type P53 DNA into human prostate cancer cells is improved by low-frequency ultrasound combined with microbubbles.

Prostate cancer is a common type of cancer in elderly men. The aim of the present study was to evaluate the effects of ultrasound exposure in combination with SonoVue microbubbles on liposome-mediated transfection of wild-type P53 genes into human prostate cancer cells. PC-3 human prostate cancer cells were exposed to ultrasound; duty cycle was controlled at 20% (2 sec on, 8 sec off) for 5 min with and without SonoVue microbubble echo-contrast agent using a digital sonifier (frequency, 21 kHz; intensity, 46 mW/cm2). The cells were divided into eight groups, as follows: Group A (SonoVue + wild-type P53), group B (ultrasound + wild-type P53), group C (SonoVue + ultrasound + wild-type P53), group D (liposome + wild-type P53), group E (liposome + SonoVue + wild-type P53), group F (liposome + wild-type P53 + ultrasound), group G (liposome + wild-type P53 + ultrasound + SonoVue) and the control group (wild-type P53). Following treatment, a hemocytometer was used to measure cell lysis, reverse transcription-quantitative polymerase chain reaction and western blotting were performed to detect P53 gene transfection efficiency, Cell Counting Kit-8 was employed to reveal cell proliferation and Annexin V/propidium iodide staining was used to determine cell apoptosis. Cell lysis was minimal in each group. Wild-type P53 gene and protein expression were significantly increased in the PC-3 cells in group G compared with the control and all other groups (P<0.01). Cell proliferation was significantly suppressed in group G compared with the control group and all other groups (P<0.01). Cell apoptosis levels in group G were significantly improved compared with the control group and all other groups (P<0.01). Thus, the results of the present study indicate that the use of low-frequency and low-energy ultrasound in combination with SonoVue microbubbles may be a potent physical method for increasing liposome gene delivery efficiency.

Upregulation of ULK1 expression in PC-3 cells following tumor protein P53 transfection by sonoporation.

The aim of the present study was to investigate whether ultrasound combined with microbubbles was able to enhance liposome-mediated transfection of genes into human prostate cancer cells, and to examine the association between autophagy and tumor protein P53 (P53). An MTT assay was used to evaluate cell viability, while flow cytometry and fluorescence microscopy were used to measure gene transfection efficiency. Autophagy was observed using transmission electron microscopy. Reverse transcription-polymerase chain reaction (RT-PCR) and western blot analysis were used to assess the expression of autophagy-associated genes. The results of the present study revealed that cell viability was significantly reduced following successfully enhanced transfection of P53 by ultrasound combined with microbubbles. In addition, serine/threonine-protein kinase ULK1 levels were simultaneously upregulated. Castration-resistant prostate cancer is difficult to treat and is investigated in the present study. P53 has a significant role in a number of key biological functions, including DNA repair, apoptosis, cell cycle, autophagy, senescence and angiogenesis. Prior to the present study, to the best of our knowledge, increased transfection efficiency and reduced side effects have been difficult to achieve. Ultrasound is considered to be a 'gentle' technique that may be able to achieve increased transfection efficiency and reduced side effects. The results of the present study highlight a potential novel therapeutic strategy for the treatment of prostate cancer.

Low-frequency and low-intensity ultrasound-mediated microvessel disruption enhance the effects of radiofrequency ablation on prostate cancer xenografts in nude mice.

The aim of the present study was to examine the impact of low-frequency, low-intensity ultrasound (US)-stimulated microbubbles (USMB) on radiofrequency ablation (RFA) in the treatment of nude mice with human prostate cancer xenografts. The tumor­bearing nude mice were divided into three groups: The USMB+RFA group was treated with USMB immediately followed by RFA, the RFA group was treated with RFA alone, and the control group remained untreated. The animals underwent enhanced US to calculate the tumor volumes, ablation volumes and ablation rates. Subsequently, the tumors were excised for hematoxylin and eosin staining, to identify necrosis in the tumors following the treatments, and immunohistochemical staining, to analyze the apoptotic index (AI), proliferative index (PI) and microvessel density (MVD) at 1, 4 and 7 days post-treatment. Each group contained five mice at each time­point. Necrosis was apparent in the center of the tumors in the treatment groups. Ablation lesion volumes of the USMB+RFA group were larger than those in the RFA group at 1 and 4 days post­treatment (P=0.002 and P=0.022, respectively), and the ablation rates of the USMB+RFA group were significantly higher, compared with the RFA group at the three time­points (all P<0.001). There were fewer apoptotic cells and more proliferative cells in the RFA group, compared with the control group 1,4 and 7 days post­treatment (all P<0.05). The AI of the USMB+RFA group was higher than that of the control group and lower than that of the RFA group 1 day post-treatment (P=0.034 and P=0.016, respectively). The PI of the USMB+RFA group was lower than that of the control group and higher than that of the RFA group 4 and 7 days post-treatment (all P<0.05). No significant differences were observed in MVD among the three groups throughout the experiment. In conclusion, exposure to USMB prior to RFA produced larger volumes of ablation, compared with treatment with RFA alone, and increased AI and reduced PI in the residual carcinoma cells induced by RFA.

Optimization of low-frequency low-intensity ultrasound-mediated microvessel disruption on prostate cancer xenografts in nude mice using an orthogonal experimental design.

The present study aimed to provide a complete exploration of the effect of sound intensity, frequency, duty cycle, microbubble volume and irradiation time on low-frequency low-intensity ultrasound (US)-mediated microvessel disruption, and to identify an optimal combination of the five factors that maximize the blockage effect. An orthogonal experimental design approach was used. Enhanced US imaging and acoustic quantification were performed to assess tumor blood perfusion. In the confirmatory test, in addition to acoustic quantification, the specimens of the tumor were stained with hematoxylin and eosin and observed using light microscopy. The results revealed that sound intensity, frequency, duty cycle, microbubble volume and irradiation time had a significant effect on the average peak intensity (API). The extent of the impact of the variables on the API was in the following order: Sound intensity; frequency; duty cycle; microbubble volume; and irradiation time. The optimum conditions were found to be as follows: Sound intensity, 1.00 W/cm2; frequency, 20 Hz; duty cycle, 40%; microbubble volume, 0.20 ml; and irradiation time, 3 min. In the confirmatory test, the API was 19.97±2.66 immediately subsequent to treatment, and histological examination revealed signs of tumor blood vessel injury in the optimum parameter combination group. In conclusion, the Taguchi L18 (3)6 orthogonal array design was successfully applied for determining the optimal parameter combination of API following treatment. Under the optimum orthogonal design condition, a minimum API of 19.97±2.66 subsequent to low-frequency and low-intensity mediated blood perfusion blockage was obtained.