The Application Study Of Ultrasound In Diagnosis And Therapy

PART Ⅰ: The clinical value of radial endorectal ultrasound in the assessment preoperative staging of rectal carcinomaObjective:To evaluate the clinical value of radial endorectal ultrasound (ERUS) in the assessment preoperative staging of rectal carcinoma.Methods:During the study period from February2010to September2011,110patients with rectal cancer were examined preoperatively using endorectal ultrasound (ERUS). ERUS is performed using a HITACHI900, HITACHI HI VISION PREIRUS US scanner, with a5-10MHz rigid rotating radial transducer which focal length is2-5cm. The size, shape, echo patten, infiltration depth, degree of circumferential involvement, extra-rectal invasion of lesions and lymph node involvement were observed. The results of ERUS staging were compared with histopathology findings of the resected specimens.Results:The accuracy of ERUS for T stage was91.4%. The accuracy of ERUS to diagnose stage T1, T2, T3, T4cancer were92.7%,88.2%,88.2%,96.4%, respectively; The sensitivity of ERUS to diagnose stage T1, T2, T3, T4cancer were92.3%,72.7%,85.4%,71.4%, respectively; specificity of ERUS to diagnose stage T1, T2, T3, T4cancer were92.9%,92.0%,90.3%,100.0%, respectively. Comparing the consistency of preoperative T-staging and postoperative pathological results, the Kappa value was0.75, the consistency was considerable. The sensitivity, specificity, and accuracy of ERUS in lymph node metastasis were74.2%,89.9%%, and85.5%, respectively. Comparing the consistency of preoperative N-staging and postoperative pathological results, the Kappa value was0.64, the consistency was considerable.Conclusion:With the simple operation, less pain, high accuracy, ERUS is currently the most practical and accurate tool to preoperatively stage rectal tumors in regard to tumor depth of invasion (T) and regional lymph node status (N). PART II: The value of elastosonography application in differentiating benign from malignant rectal neoplasms and preoperative staging of rectal carcinomaObjective:To investigate the value of elastosonography application in differentiating benign from malignant rectal neoplasms and preoperative staging of rectal carcinoma.Methods:Analysis the characteristic of elastosonography in85patients, tissue elasticity was scored from one(greatest elastic strain) to five (no strain) in elastosonography, perform an analysis according to the histological results, elastosonography from which the strain ratios(SR)were derived, with pathologic results as the reference standard. And a receiver-operating characterisitc(ROC) curve was used to identify the value of optimal operating point for differential diagnosis of rectal neoplasms.Results:(1) The strain ratio of the rectal adenomas and hemorrhoids were1.36±1.14,3.61±1.95, respectively; the strain ratio of the rectal cancer were T1:23.0±25.8、T2:39.9±45.7、 T3:61.0±51.2and T4:60.5±36.2, respectively. The strain ratio of rectal adenomas was no significantly different from the value of rectal hemorrhoids, p>0.05. The strain ratio of rectal cancer stage T1,T2had significantly different from the rectal cancer stage T3and T4, P<0.05.(2) The area under the curve of elastic5score method between benign and malignant lesions was0.943, which showed a high statistical significance(P<0.05).(3) According to ROC curve, the critical point of elastic5score method between benign and malignant lesions was3.5, with high sensitivity(81.5%) and specificity(100%). The critical point of strain ratio method between benign and malignant lesions was3.6.(4) Set the T1-2were early cancer, T3-4were advanced cancer, with sensitivity for ordinate,1-specific degrees for the abscissa, making elastic5score method ROC curve, the area under the curve was0.679, compare with AUC=0.5, which showed a no statistical significance. It meansed elastic ultrasound had no value in preoperative staging of rectal carcinoma.Conclusion:The elastosonography can provide a noninvasive, quantitative and convenient tool to differentiating benign from malignant rectal neoplasms, but no value to preoperative staging of rectal carcinoma. PART Ⅲ: Bioeffects of hyperthermia on human breast cancer MDA-MB-231cells In vitroObjective:To observe the changes in human breast cancer MDA-MB-231cells morphorlogy, membrane permeability, viability and cell migration after exposed to hyperthermia in different temperature and different time in vitro,and study the enhanced antitumor chemotherapy efficacy, mechanism and application value and safety.Methods:MDA-MB-231cells seeded in8well chamber slide. The cells were exposed by hyperthermia in a43℃,45℃,50℃water bath tank3min,10min,15min,30min, respectively.. The cells in control group were managed in the same environment but without hypertermia exposed. A Trypan blue assay evaluated cell viability; Scanning electron microscopy(SEM) were used to observe cell morphology. MDA-MB-231cells seeded in6well plate. The cells were exposed by hyperthermia in a43℃,45℃,50℃water bath tank3min,10min,15min,30min, respectively. The scratch wound assay was used to measure in vitro hyperthermia stimulated scratch wound closure rate;MTT assay was used to estimate the number of viable cells in drug screening trials. MDA-MB-231cells seeded in96-well plate. The different concentration gradient of Paclitaxel (PTX) and Colchicine (COL) were administered to the culture solution for culturing MDA-MB-231cell, as the experimental groups, and in the control group were administered with DMEM solution. The cells were exposed by hyperthermia in a 45℃water bath tank15min. The optic density(OD)of viable cell, cell inhibition rate(IR), the values of50%inhibition concentration(IC50) were assayed.Results:1. MDA-MB-231cell viability following hyperthermia in a43℃,45℃,50℃water bath tank3min,10min,15min,30min, respectively were counted using the trypanblue. The cell viability was control:98.9%±0.4;To43℃3min hyperthermia,the cell survival rate and the treatment time had no significant difference, but to the45℃,50℃hyperthermia treatment, a temperature and time dependent decrease in viability was observed.2.Scanning Electron Microscopy(SEM):The morphological alterations included the microvilli of the cell surfaces become stubby, plenty bubbles in cytoplasm, microvilli disappear and appear pores after hyperthermia exposures with different temperature and time in the experiment.3. Determining rate of scratch closure by image J analaysis. To43℃,45℃,50℃3min hyperthermia,the treatment time and temperature no significant effect on scratch wound closure rate. However, the scratch wound closure ratea significant decrease on10min,15min,30min,43℃,45℃,50℃hyperthermia. A time and temperature dependent decrease in would closure rate was observed between24hours and48hours.4.After5ng/ml PTX and COL treatment24hour,control and drug only groups showed no significant difference. Heat only and heat and drug groups showed more effective anti-tumor than the control and drug only groups. After the48hours and72hours treatment, the drug only, heat only and drug and heat groups showed significant difference compare the control.The heat and drug group showed significant difference compare with the others on anti-tumor effective, p<0.001.Conclusion:This study suggests hyperthermia may enhanced cell permeabilisation,significantly inhibit cell migration and cell proliferation and utilised for local drug delivery. PART IV: Biological effects of Pulsed HIFU:the mechanism and application on targeted drug delivery to tumors and on biosafety of ultrasoundObjective:To observe the changes in human breast cancer MDA-MB-231cells morphorlogy, viability and cell migration after exposed to pHIFU in different intensity in vitro, to observe the changes in human MDA-MB-231cells、MIA-Paca2cells、U87cells and SAOS-2cells cytoskeleton, and study the enhanced antitumor chemotherapy efficacy, mechanism and application value and safety.Methods:MDA-MB-231cells seeded in8well chamber slide. The cells were exposed by ultrasound, frequency of1MHz,25%duty cycle, with intensity output of1MHz,3MHz,5MHz,7MHz respectively for3min. The cells in control group were managed in the same environment but without ultrasound exposed. A Trypan blue assay evaluated cell viability; Scanning electron microscopy(SEM) were used to observe cell morphology. MDA-MB-231cells、MIA-Paca2cells、U87cells and SAOS-2cells were seeded in8well chamber slide. The cells were exposed by ultrasound, frequency of1MHz,25%duty cycle, with intensity output of1MHz,3MHz,5MHz,7MHz respectively for3min. The cells in control group were managed in the same environment but without ultrasound exposed. Laser scanning confocal microscope was used to examine the changes of cytoskeleton include F-actin and α-, β-tubulin. MTT assay was used to estimate the number of viable cells in drug screening trials.MDA-MB-231cells seeded in96-well plate. The different concentration gradient of Paclitaxel (PTX) and Colchicine (COL) were administered to the culture solution for culturing MDA-MB-231cell, as the experimental groups, and in the control group were administered with DMEM solution. The cells were exposed by1Mpa,5Mpa3min, respectively. The optic density(OD)of viable cell, cell inhibition rate(IR), the values of50%inhibition concentrationIC50) were assayed. Live cell spin disk confocal image system was used to detect cell migration ability after3minutes pHIFU exposed. MTT assay was used to estimate the number of viable cells in drug trials. MDA-MB-231cells seeded in96-well plate. The different concentration gradient of Paclitaxel (PTX) and Colchicine (COL) were administered to the culture solution for culturing MDA-MB-231cell, as the experimental groups, and in the control group were administered with DMEM solution. The cells were exposed by1Mpa、5Mpa pHIFU3minutes. The optic density(OD)of viable cell, cell inhibition rate(IR), the values of50%inhibition concentration(IC50) were assayed.Results:1. MDA-MB-231cell viability following3minutes of exposure at incremental powers include1MHz,3MHz,5MHz,7MHz were counted using the trypanblue. The cell viability was control:93.7%±4.15;1MHz:90.17%±9.79;3MHz:85.0%±6.84;5MHz:76.2%±3.67and7MHz:61.65%±0.2respectively, a power-dependent decrease in viability was observed.2.Scanning Electron Microscopy(SEM):The morphological alterations included the microvilli of the cell surfaces become stubby, plenty bubbles in cytoplasm, microvilli disappear and appear pores after ultrasound exposures with different intensity outputs in the experiment.3. Laser scanning confocal microscope showed microfilament, microtubule become stubby and irregular patches of actin and tubulin staining across the cytoplasm. Changes to the cell periphery and disruption to the actin and tubulin cytoskeleton were dose-dependent.4. Live cell spin disk confocal image system showed that MDA-MB-231cells migration ability decreased significantly after3minutes pHIFU exposed.5. After5ng/ml PTX and COL with1Mpa.5Mpa pHIFU treatment24hour, control and drug only groups,1MPa pHIFU only,1MPa and drug showed no significant difference. Drug and5MPa pHIFU and control showed had significant difference. p<0.001(PTX and5MPa pHIFU),p<0.01(COL and5MPa pHIFU).Conclusion:This study suggests pHIFU may enhanced cell permeabilisation, significantly inhibit cell migration and cell proliferation and utilised for local drug delivery.