吉田 憲司

Choosing an appropriate dynamic range for acquiring radio-frequency (RF) data from a high-frequency ultrasound (HFU) system is challenging because the RF data amplitude typically covers several orders of magnitude between the sample surface and the deepest imaged regions. In addition, the saturated signal may decrease the accuracy of QUS methods because quantitative ultrasound (QUS) methods are sensitive to saturated data. In this study, the effects of saturation on QUS estimates of Nakagami shape parameter (Nakagami parameter) were quantified by analyzing data acquired from 20 dissected human lymph nodes with a single-element transducer operating at a center frequency of 26 MHz. Artificially saturated signals (x(sat)) were produced by applying artificial saturation methods to the original unsaturated signals (x(ori)). Saturation degree was quantified using an index termed Saturate-SNR (SSNR). Nakagami parameters were estimated from xsat over a wide range of SSNR values. Nakagami parameters of saturated signals were increased (0.18 when the signal decremented 2 bit) significantly with decreasing SSNR. Nakagami parameters were corrected by pretreatment that applied a smoothing spline to the saturated signal. The smoothing spline restoration method is tuned P parameter. The best correction occurred when P was 1 (i.e., cubic spline interpolation). The maximum Nakagami parameter error in the corrected with 6-bit signal was 0.10, which is less than the average difference of 0.12 that existed between non-metastatic and metastatic lymph nodes.

© 2015 IEEE. Choosing an appropriate dynamic range for acquiring radio-frequency (RF) data from a high-frequency ultrasound (HFU) system is challenging because signals can vary greatly in amplitude because of focusing and attenuation effects. In addition, quantitative ultrasound (QUS) methods are sensitive to saturated data. In this study, the effects of saturation on QUS estimates of effective scatterer diameter (ESD) and effective acoustic concentration (EAC) were quantified using data acquired from 69 dissected human lymph nodes with a single-element transducer operating at a center frequency of 26 MHz. Artificially saturated signals (xc) were produced by thresholding the original, unsaturated RF echo signals (x). Saturation degree was quantified using Saturate-SNR. ESD and EAC were estimated from xc over a wide range of Saturate-SNR values. The value of the ESD estimate was minimally affected when Saturate-SNR ranged from infinity to 3.99 dB. However, the value of the EAC estimate decreased significantly with decreasing Saturate-SNR. EAC estimates were corrected using a linear relationship between EAC values over a range of Saturate-SNR values and l1 - norm of x (i.e., the sum of absolute values of the true RF echo signal). To correct the estimates for ESD and EAC of saturated signals, the estimated original RF signal was derived from xc by a cubic spline. A linear-regression line was computed for the EAC values of a set of Saturate-SNR values vs. l1 - norm. The intersection of the regression line and l1- norm of the estimated original signal gave a corrected value for EAC. The maximum error in the corrected EAC estimate was 0.48 dB/mm3. The average differences in corrected ESD and EAC estimates of non-metastatic and metastatic lymph nodes were 7.80 μm and 4.18 dB/mm3, respectively. The error in the corrected EAC value was much smaller than the difference in the corrected values of non-metastatic and metastatic lymph nodes.

Early diagnosis of non-alcoholic steatohepatitis (NASH) is highly desired because NASH can lead to cirrhosis or even to hepatocellular carcinoma in some severe cases. Towards non-invasive diagnosis with ultrasound, we studied free fatty acids (FFAs) present in the liver, which is the organ most likely to be affected by the disease. As a preclinical study, we performed acoustic-impedance measurements of five kinds of FFAs in solvent or in cultured Huh7 cells. To measure the acoustic impedance, a concave transducer with an 80-MHz center frequency was incorporated in a scanning acoustic microscopy system. One-way ANOVA showed statistically-significant differences (p<0.05) in acoustic impedance among the FFAs in FFA solvent and with cultured Huh7 cells. These results suggest that each of the FFAs, especially PA, OA and PAOA could be distinguished from each other regardless of whether they were in solution or in absorbed by cells.

Targeted microbubbles (MBs) has been applied to the ultrasound molecular imaging. It is important to understand the sensitivity of bubbles to the target molecules. Thus, we aim to construct an evaluation system using quartz crystal microbalance (QCM) to measure quantitatively both the concentration of target molecular and the number density of specifically-absorbed MBs. To quantitatively counting absorbed MBs on QCM, the physical model for expressing the relationship between the impedance of QCM and absorbed MBs was proposed in our previous study. In this paper, we validates the model based on the comparison of model-based analysis of the number density of MBs with the value measured by using an optical microscope.

肝臓の繊維化に伴いミクロな領域で生じる組織性状変化を調べる目的で,肝線維症モデルのラット肝臓を対象に,超音波顕微鏡を用いて音速,減衰,音響インピーダンスの計測を行った.計測用に厚みが約10μmの薄切試料を作成し,中心周波数80MHz(方位分解能20μm)及び250MHz(方位分解能4μm)の超音波を用いて計測した.計測結果より求められるエコー信号の周波数スペクトルと理論的な計算結果をフィッティングすることで,音速,減衰,音響インピーダンスを推定した.線維組織の音速,減衰は他の部位や正常肝臓の値と比べて大きいことを確認した.一方,音響インピーダンスに関しては線維部分とその他部分で顕著な違いはみうけられなかった.

超音波を用いた高精度かつ定量的な組織性状診断を実現するためには,生体組織の構造及び組成と物性との関係を明確にする必要がある.組織固有の指標として音速と減衰という音響特性に着目し,計測を行っている.本研究では細胞1つ程度の領域での分解能を有する中心周波数250MHzの超音波振動子を用いてラット臓器の音響特性解析を行った.性状の異なる組織として硬変肝モデルの計測を行い,硬変肝の音響特性が正常肝とは異なることを示した.また同様に組織構造が特徴的な腎臓,脾臓に関しても音響特性の比較を行い,臓器間の音響特性の違いを示した.

一分子レベルでのDNA分子の観察を基に,低周波数超音波の照射により生じるDNAの二重鎖切断を定量的に評価した.特にキャビテーション現象に着目し,キャビテーション核の有無による二重鎖切断の頻度の違いについて検討した.キャビテーション核としてマイクロバブルである超音波診断用造影剤(Sonazoid^[○!R])を用いた.DNA含有液体にマイクロバブル懸濁液を添加し,超音波を照射した場合では,負のピーク圧力が50kPa以上においてDNA分子の切断頻度が上昇することを確認した.また,この音圧閾値はBlakeの閾値圧力よりも低いことを確認した.一方,マイクロバブル懸濁液を添加しなかった場合ではDNA分子の二重鎖切断は確認されなかった.本結果はDNA分子の二重鎖切断にキャビテーション現象が寄与していることを強く示している.

Early diagnosis for non-alcoholic steatohepatitis (NASH) is desired because it may progress to cirrhosis or finally hepatocellular carcinoma in the severe case. To propose the possible noninvasive diagnosis using ultrasound, we aim to summarize the change of the tissue property. As a basic study, this report demonstrates the acoustic impedance of mouse livers in case of the normal, the liver cirrhosis and the NASH model using bio-acoustic microscopy. To measure the acoustic impedance, a transducer with 80-MHz center frequency, which is incorporated in a bio-acoustic microscopy system, was employed. The value could be calculated based on the analysis of the echo amplitude, and 2D image of acoustic impedance can be obtained by scanning the transducer. The 2D image demonstrated that the tissue structure is homogeneous in normal liver. In contrast, the pattern of random granular texture like lipid droplet was found in NASH liver. Based on statistical analysis, it was found that the acoustic impedance in case of NASH is lowest among all models.

To realize a quantitative diagnosis method of liver fibrosis, we have been developing a modeling method for the probability density function of the echo amplitude. In our previous study, we proposed a multi-Rayleigh model with two or three Rayleigh distributions, which is considered as a heterogeneous medium consisting of normal and different tissue, and constructed the evaluation method using modified quantile-quantile probability plot (Q-Q plot). In this study, we examined a multi-Rayleigh model and generalized Nakagami model which is general technique, corresponding to the distribution of the echo amplitude from homogeneous and heterogeneous medium. In clinical data of liver fibrosis, we showed validity of proposed model.

By using two different analysis method, the scatter structure of rat liver was evaluated. Rat liver of fibrosis, NASH and control model were prepared. The self-made scanner system with 15-MHz ultrasound, which has good spatial resolution compared with commercial ultrasound scanner, were constructed. The single focused transducer with 15-MHz resonant frequency and 200 mu m lateral resolution was employed for the measurement. To quantitatively evaluate the scatter structure, two parameters were evaluated: the quantile-quantile (Q-Q) parameter and the scatterer size. As a result, there was no significant difference between the three different rat models.

Acoustic property of tissue such as speed of sound (SoS), attenuation provide beneficial information for the pathological diagnosis. The pathological change due to liver fibrosis have been investigated by using a 250-MHz central frequency transducer with fine spatial resolution of 4-mu m. The SoS and attenuation of rat liver in case of control and fibrosis models were measured by bioacoustics microscopy incorporating the 250-MHz transducer. In addition, rat spleen was measured to investigate whether tissue property in spleen was affected by liver fibrosis. The SoS and attenuation were calculated by analyzing the echo data in frequency domain, and their 2D images were obtained by scanning the transducer. After the measurement, the histology in the same cross section (H&E staining and Azan staining) was acquired for comparing with 2D image of acoustic properties. Rat livers and spleens of the control and fibrosis model were measured. In case of liver, there were the individual variability for SoS and attenuation. In a sample of fibrosis model, for example, the difference of SoS between normal and fiber portions were approximately 50 m/s. In contrast, there was no significant difference between them in another case. In addition, there was few difference between control and fibrosis models. In case of spleen, we clearly found the significant difference in SoS and attenuation between white pulp and red pulp. However, there was few difference between control and fibrosis models as well as the liver.

超音波を用いた高精度な組織性状診断を実現するためには,各種生体組織の有する構造および組成と物性との関係を明確にする必要がある.超音波顕微鏡を用いて生体組織の構造を視覚的に観察するとともに,同組織の音響的な性質を計測することで,光学顕微鏡を用いた病理検査では知ることのできない特徴を診断指標として得ることが可能である.本研究では,超音波顕微鏡で汎用的に使用されている周波数に比して高周波である250MHzでの生体組織の構造観察と音響特性計測を試みた.

標的指向性マイクロバブルは,標的部位に特異的に付着するため,分子イメージングにおける造影剤や超音波ドラッグデリバリーシステムにおける薬物運搬体への応用が期待されている.標的指向性マイクロバブルの実用化のため,水晶振動子微量天秤(Quartz Crystal Microbalance: QCM)を用いたマイクロバブルの特異的付着能力の評価方法を提案する.QCMの金電極上にBiotin化自己組織化単分子膜を用いてStreptavidin膜を形成することで標的部位を模擬し,作製したBiotin修飾マイクロバブルを特異的に付着させた.QCMのコンダクタンスが半値となるときの周波数f_1, f_2 (f_1<f_2)と定義し,バブルの特異的付着に伴うf_1, f_2の変化を測定した.Biotin非修飾マイクロバブルおよびバブルを含まない溶液を用いた結果との比較から,f_1の変化にマイクロバブルの影響が,f_2の変化にバブルの膜物質(リン脂質)の特異的な付着の影響がそれぞれ現れる可能性を示した.

超音波で駆動される壁面付着気泡について表面振動モードが発生する条件を実験的に評価することを月的にレーザドップラ振動計と高速度ビデオカメラを用いた観測システムを構築した.高速度カメラを用いた観測では気泡画像を基に視覚的に表面振動モードの発生を評価し,レーザドップラ振動計を用いた観測では振動変位波形の周波数解析を基に評価した.気泡サイズは29μm〜190μmであり、照射超音波の周波数は28kHz,38kHzもしくは81.3kHzである.両者の結果を比較することにより,気泡振動に含まれる分調波成分の有無が表面振動モード発生の評価指標として用いることができることを示し,分調波発生条件の気泡サイズ、照射音波の周波数に対する依存性を調べた.得られた結果を基に,表面振動モードの発生条件の実験式を導くことに成功した.

標的部位に特異的に吸着する標的指向性マイクロバブルは,超音波造影剤や超音波ドラッグデリバリーシステム(Drug Delivery System: DDS)における薬物運搬体への応用が期待されている.本報告では標的指向性マイクロバブルとしてBiotin修飾マイクロバブルを作製し,水晶共振子型質量センサ(Quartz Crystal Mlcrobalance: QCM)を用いてその標的指向性の評価を試みた.QCM表面上にBiotin化自己組織化単分子膜,さらにはStreptavidinを固定化し,Biotin-Avidin-Biotin結合を介して,バブルをQCM表面上に特異的に吸着させるバブル吸着によりQCMの共振特性が変化することが期待されるネットワークアナライザを用いてQCMの共振周波数の変化を計測することで,センサ表面へのStreptavidinの吸着,Streptavidin膜上へのBiotin修飾バブルの捕捉を確認した.コントロール実験として,Biotin修飾していないバブルを使用して同様の実験を行い,Biotin修飾バブルはBiotin非修飾バブルに比べて優位な周波数変化を示すことを明らかにした.

Double-strand brakes of T4-DNA molecules caused by ultrasound irradiation are monitored through the methodology of single giant DNA observation by fluorescence microscopy. The experimental results indicate the existence of the threshold power of ultrasound on the double-strand brakes. In addition, we have examined the effect of an anti-oxidative agent, 2-mercaptoethanol (2-Me), on the ultrasound damage of DNA. It becomes clear that 2-Me exhibits almost no protective effect.

ポリ乳酸を膜材質とする中空カプセルを作製し,超音波照射下におけるカプセルの破壊特性及び,内包物の放出を評価した.作製したカプセルの粒径は1〜45μmであり,ダブルエマルジョン法を用いて作製した.周囲膜内部には模擬薬物である蛍光色素(acid red 52)と空気が封入されており,三層構造を持つ.膜厚は100nm〜3μmであり,中空率は75〜97%である.実験容器内を脱気水で満たし,周波数700kHz,1,2MHzの連続正弦波,音圧400kPaでカプセルの破壊実験を行った.各周波数におけるカプセルの共振径はそれぞれ30,25,20μmであった.カプセル破壊後,内包物は60秒間にわたり放出がなされた.カプセル内部の気体の振動が内包物の放出に影響を与えると考えられる.

This paper demonstrates characteristics of an oscillation of an attached bubble in ultrasonic field, compared with a free bubble. First, we experimentally investigated the resonant frequency of the attached bubble with radius of 50 - 250 mu m, based on the analysis of steady or transient oscillation. The incident frequency f(d) of the ultrasound with sound pressure amplitude smaller than several kilo Pascal was 28 or 38 kHz. The displacement amplitude of bubble oscillation was measured by using laser Doppler vibrometer. It was found that the frequency of the attached bubble was smaller than that of a free bubble which was calculated by using theoretical model. Second, we estimated the sound pressure threshold where the non-spherical oscillation mode appeared. We measured sub-harmonic component (f(d)/2) in the frequency spectrum of the oscillation which generated by parametric instability. Based on the sub-harmonic generation, the appearance of the non-spherical oscillation mode was evaluated. As a result, the threshold pressure was one tenth of that in case of a free bubble

Hollow microcapsules have been considered for potential application as drug or gene carriers. Targeted drug or gene release to tumor vascular can be achieved by destroying the capsules by high-intensity ultrasound. This report investigates fabrication of microcapsules having biocompatible polylactic acid (PLA) shell and the mechanical property for the destruction under ultrasound irradiation. The PLA hollow microcapsules with the diameter of 1 to 35 mu m were fabricated by double emulsion method. Red fluorescent dye which is substitute for drug was enclosed into the capsules. The capsules have trilaminar structure. Inner air is surrounded by red fluorescent, and the PLA shell encloses them. Shell thickness of capsules was distributed from 100 nm to 3 mu m. Destruction of the fabricated capsules was investigated by using ultrasound pulse with the center frequencies from 500 kHz to 2 MHz. The behavior of capsule was observed by using an optical microscope under ultrasound irradiation. The frequency dependence of the capsule destruction was investigated and 35% of the capsules with the resonance size was destroyed with 400 kPa at 700 kHz. The capsule shell was ruptured by the ultrasound, and then the inner dye was released; the dye was completely released for 120 seconds under ultrasound irradiation.

We construct the experimental system with laser Doppler vibrometer (LDV) for measuring the vibration of a single bubble. With the use of this system, we observed the vibration of a bubble attached to rigid material in ultrasonic standing wave with the frequency of 27.8 kHz. The initial bubble radius was ranged from 45 mu m to 235 mu m. Compared to conventional methods e.g. the observation using a high speed camera, this system was effective to measure the vibration with very small amplitude of nanometer order. This advantage enabled us to measure the resonant characteristics in case that the bubble linearly responded to the incident sound. The experimentally measured resonant radius was about 90 mu m, which was smaller than the theoretical prediction of 100 mu m by using the model of a single free bubble. The difference possibly resulted from the effect of rigid material to which the bubble was attached. The results support that the LDV measurement system will be an effective tool to reveal the detail characteristics of bubble vibration.