Non-invasive monitoring of cancer cell death would permit rapid feedback on treatment response. O... more Non-invasive monitoring of cancer cell death would permit rapid feedback on treatment response. One technique showing such promise is quantitative ultrasound. High-frequency ultrasound spectral radiofrequency analysis was used to study cell death in breast cancer cell samples. Quantitative ultrasound parameters, including attenuation, spectral slope, spectral 0-MHz-intercept, midband fit, and fitted parameters displayed significant changes with paclitaxel-induced cell death, corresponding to observations of morphological changes seen in histology and electron microscopy. In particular, a decrease in spectral slope from 0.24±0.07 dB/MHz to 0.04±0.09 dB/MHz occurred over 24 hours of treatment time and was identified as an ultrasound parameter capable of differentiating post-mitotic arrest cell death from classical apoptosis. The formation of condensed chromatin aggregates of 1 micron or greater in size increased the number of intracellular scatterers, consistent with a hypothesis that...
2014 Ieee International Ultrasonics Symposium, Sep 3, 2014
Acoustic impedance maps can be used to gain an insight into the microstructure and physiological ... more Acoustic impedance maps can be used to gain an insight into the microstructure and physiological state of cancer cells. In this we correlated acoustic impedance maps with cells structures detected by fluorescence microscopy. Fluorescence microscope was used to image MCF-7 cells marked with Celltracker Orange and Hoechst for staining the cytoplasm and nucleus, respectively. A single element 375MHz center frequency transducer (SASAM) was used to generate acoustic impedance maps of inverted MCF-7 cells on a plastic substrate. The acoustic impedance of 20 single-live, 10 clustered-live and 7 clustered-fixed MCF7 cells were measured and compared. Fluorescent images and acoustic impedance maps suggest that there is an acoustic impedance mismatch between the nucleus and the cytoplasm. The average acoustic impedance values for single-live, clustered-live and clustered-fixed cancer cells were measured to be 1.600±0.006 MRayl, 1.609±0.009 MRayl and 1.572±0.011 MRayl.
Many types of cancer therapies target the tumor microenvironment, causing biochemical and morphol... more Many types of cancer therapies target the tumor microenvironment, causing biochemical and morphological changes in tissues. In therapies using ultrasound activated microbubbles, vascular collapse is typically reported. Red blood cells (RBCs) that leak out of the vasculature become exposed to the ceramide that is released from damaged endothelial cells. Ceramide can induce programmed cell death in RBCs (eryptosis), and is characterized by cell shrinkage, membrane blebbing and scrambling. Since the effect of eryptotic cells on generated photoacoustics (PA) signals has not been reported, we investigated the potential PA may have for cancer treatment monitoring by using PA spectral analysis to sense eryptosis. To induce eryptosis, C2-ceramide was added to RBC suspensions and that were incubated for 24 hours at 37 o C. A control and ceramide-induced sample was imaged in a vessel phantom using a high frequency PA system (VevoLAZR, 10 – 45 MHz bandwidth) irradiated with multiple wavelengths ranging from 680 to 900 nm. PA spectral parameters were measured and linked to changes in RBCs as it underwent eryptosis. These samples were examined using optical microscopy, a blood gas analyzer and an integrating sphere setup to measure optical properties (wavelengths 600 – 900 nm). The results of the experiment demonstrate how PA spectral analysis can be used to identify eryptosis at a depth of more than 1 cm into the phantom using ultrasound derived the y-intercept and mid bandfit (MBF) parameters at optical wavelengths of 800 – 900 nm. These parameters were correlated to the morphological and biochemical changes that eryptotic RBCs display. The results establish the potential of PA in cancer treatment monitoring through sensing treatment induced eryptosis.
Quantitative ultrasound parameters based on form factor models were investigated as potential bio... more Quantitative ultrasound parameters based on form factor models were investigated as potential biomarkers of cell death in breast tumor (MDA-231) xenografts treated with chemotherapy. Ultrasound backscatter radiofrequency data were acquired from MDA-231 breast cancer tumor-bearing mice (n = 20) before and after the administration of chemotherapy drugs at two ultrasound frequencies: 7 MHz and 20 MHz. Radiofrequency spectral analysis involved estimating the backscatter coefficient from regions of interest in the center of the tumor, to which form factor models were fitted, resulting in estimates of average scatterer diameter and average acoustic concentration (AAC). The ∆AAC parameter extracted from the spherical Gaussian model was found to be the most effective cell death biomarker (at the lower frequency range, r(2) = 0.40). At both frequencies, AAC in the treated tumors increased significantly (P = .026 and .035 at low and high frequencies, respectively) 24 hours after treatment compared with control tumors. Furthermore, stepwise multiple linear regression analysis of the low-frequency data revealed that a multiparameter quantitative ultrasound model was strongly correlated to cell death determined histologically posttreatment (r(2) = 0.74). The Gaussian form factor model-based scattering parameters can potentially be used to track the extent of cell death at clinically relevant frequencies (7 MHz). The 20-MHz results agreed with previous findings in which parameters related to the backscatter intensity (i.e., AAC) increased with cell death. The findings suggested that, in addition to the backscatter coefficient parameter ∆AAC, biological features including tumor heterogeneity and initial tumor volume were important factors in the prediction of cell death response.
Apoptosis is a form of programmed cell death characterized by a series of predictable morphologic... more Apoptosis is a form of programmed cell death characterized by a series of predictable morphological changes at the subcellular level, which modify the light-scattering properties of cells. We present a spectroscopic optical coherence tomography (OCT) technique to detect changes in subcellular morphology related to apoptosis in vitro and in vivo. OCT data were acquired from acute myeloid leukemia (AML) cells treated with cisplatin over a 48-h period. The backscatter spectrum of the OCT signal acquired from the cell samples was characterized by calculating its in vitro integrated backscatter (IB) and spectral slope (SS). The IB increased with treatment duration, while the SS decreased, with the most significant changes occurring after 24 to 48 h of treatment. These changes coincided with striking morphological transformations in the cells and their nuclei. Similar trends in the spectral parameter values were observed in vivo in solid tumors grown from AML cells in mice, which were treated with chemotherapy and radiation. Our results provide a strong foundation from which future experiments may be designed to further understand the effect of cellular morphology and kinetics of apoptosis on the OCT signal and demonstrate the feasibility of using this technique in vivo.
Photons Plus Ultrasound: Imaging and Sensing 2015, 2015
ABSTRACT A methodology for simulating photoacoustic (PA) images of samples with solid spherical a... more ABSTRACT A methodology for simulating photoacoustic (PA) images of samples with solid spherical absorbers acquired using linear array transducer is described. Two types of numerical phantoms (i.e. polystyrene beads suspended in agar medium) of two different size regimes were imaged with a 40 MHz linear array transducer utilizing this approach. The frequency domain features and statistics of the simulated signals were quantified for tissue characterization. The midband fit at 40 MHz was found to be about 35 dB higher for the sample with larger beads (radius ∼ 7.36 μm) than that of the sample with smaller particles (radius ∼ 1.77 μm). The scale parameter of the generalized gamma distribution function was found to be nearly 51 times greater for the former sample compared to the latter sample. The method developed here shows potential to be used as a fast simulation tool for the PA imaging of collection of absorbers mimicking biological tissue.
Biomedical Applications of Light Scattering VIII, 2014
ABSTRACT We present an in vivo implementation of a multi-parametric technique for detecting apopt... more ABSTRACT We present an in vivo implementation of a multi-parametric technique for detecting apoptosis using optical coherence tomography in a mouse tumor model. Solid tumors were grown from acute myeloid leukemia cells in the hind leg of SCID mice and treated with a single dose of cisplatin and dexamethasone to induce apoptosis. Both spectral features and speckle decorrelation times indicated good consistency between control mice and reasonable agreement with in vitro measurements. The integrated backscatter increased significantly in tumors responding to treatment while the spectral slope and decorrelation time did not show significant changes. This study demonstrates the feasibility of using spectroscopic OCT and dynamic light scattering for treatment monitoring in vivo.
2014 IEEE International Ultrasonics Symposium, 2014
ABSTRACT Ultrasound thermal therapy is used for noninvasive treatment of cancer. For accurate ult... more ABSTRACT Ultrasound thermal therapy is used for noninvasive treatment of cancer. For accurate ultrasound based temperature monitoring in thermal therapy, the temperature dependence of acoustic parameters is required. In this study, the temperature dependence of acoustic harmonics was investigated in water. The pressure amplitudes of the transmitted fundamental frequency (p1), and its harmonics (second (p2), third (p3), fourth (p4), and fifth (p5)) generated by nonlinear ultrasound propagation were measured by a calibrated hydrophone in water. The hydrophone was placed at the focal point of a focused 5-MHz transducer (f-number 4.5) to measure the acoustic pressure. Higher harmonics were generated by transmitting a 5-MHz 15-cycle pulse that resulted in a focal positive peak pressure of approximately 0.26 MPa in water. The water temperature was increased from 26 °C to 52 °C in increments of 2 °C. Due to this temperature elevation, the value of p1 decreased by 9%±1.5% (compared to its value at 26 °C) and values of p2, p3, p4, and p5 increased by %5±2%, 22%±8%, 44%±7%, and 55%±5%, respectively. The results indicate that the nonlinear harmonics are highly temperature dependent and their temperature sensitivity increase with the harmonic number. It is concluded that the nonlinear harmonics could potentially be used for ultrasound-based thermometry.
The photoacoustic signal generated from particles when irradiated by light is determined by attri... more The photoacoustic signal generated from particles when irradiated by light is determined by attributes of the particle such as the size, speed of sound, morphology and the optical absorption coefficient. Unique features such as periodically varying minima and maxima are observed throughout the photoacoustic signal power spectrum, where the periodicity depends on these physical attributes. The frequency content of the photoacoustic signals can be used to obtain the physical attributes of unknown particles by comparison to analytical solutions of homogeneous symmetric geometric structures, such as spheres. However, analytical solutions do not exist for irregularly shaped particles, inhomogeneous particles or particles near structures. A finite element model (FEM) was used to simulate photoacoustic wave propagation from four different particle configurations: a homogeneous particle suspended in water, a homogeneous particle on a reflecting boundary, an inhomogeneous particle with an ab...
Early alterations in textural characteristics of quantitative ultrasound spectral parametric maps... more Early alterations in textural characteristics of quantitative ultrasound spectral parametric maps, in conjunction with changes in their mean values, are demonstrated here, for the first time, to be capable of predicting ultimate clinical/pathologic responses of breast cancer patients to chemotherapy. Mechanisms of cell death, induced by chemotherapy within tumor, introduce morphological alterations in cancerous cells, resulting in measurable changes in tissue echogenicity. We have demonstrated that the development of such changes is reflected in early alterations in textural characteristics of quantitative ultrasound spectral parametric maps, followed by consequent changes in their mean values. The spectral/textural biomarkers derived on this basis have been demonstrated as non-invasive surrogates of breast cancer chemotherapy response. Particularly, spectral biomarkers sensitive to the size and concentration of acoustic scatterers could predict treatment response of patients with u...
Modulation of the tumour microvasculature has been demonstrated to affect the effectiveness of ra... more Modulation of the tumour microvasculature has been demonstrated to affect the effectiveness of radiation, stimulating the search for anti-angiogenic and vascular-disrupting treatment modalities. Microbubbles stimulated by ultrasound have recently been demonstrated as a radiation enhancer when used with different cancer models including PC3. Here, photoacoustics imaging technique was used to assess this treatment's effects on haemoglobin levels and oxygen saturation. Correlations between this modality and power doppler assessments of blood flow, and histology measurements of vascular integrity and cell death were also investigated. Xenograft prostate tumours in SCID mice were treated with 0, 2, or 8 Gy radiation combined with microbubbles exposed to 500 kHz ultrasound at a peak negative pressure of 0, 570, and 750 kPa. Tumours were assessed and levels of total haemoglobin, oxygen saturation were measured using photoacoustics before and 24 hours after treatment along with power do...
Non-invasive monitoring of cancer cell death would permit rapid feedback on treatment response. O... more Non-invasive monitoring of cancer cell death would permit rapid feedback on treatment response. One technique showing such promise is quantitative ultrasound. High-frequency ultrasound spectral radiofrequency analysis was used to study cell death in breast cancer cell samples. Quantitative ultrasound parameters, including attenuation, spectral slope, spectral 0-MHz-intercept, midband fit, and fitted parameters displayed significant changes with paclitaxel-induced cell death, corresponding to observations of morphological changes seen in histology and electron microscopy. In particular, a decrease in spectral slope from 0.24±0.07 dB/MHz to 0.04±0.09 dB/MHz occurred over 24 hours of treatment time and was identified as an ultrasound parameter capable of differentiating post-mitotic arrest cell death from classical apoptosis. The formation of condensed chromatin aggregates of 1 micron or greater in size increased the number of intracellular scatterers, consistent with a hypothesis that...
2014 Ieee International Ultrasonics Symposium, Sep 3, 2014
Acoustic impedance maps can be used to gain an insight into the microstructure and physiological ... more Acoustic impedance maps can be used to gain an insight into the microstructure and physiological state of cancer cells. In this we correlated acoustic impedance maps with cells structures detected by fluorescence microscopy. Fluorescence microscope was used to image MCF-7 cells marked with Celltracker Orange and Hoechst for staining the cytoplasm and nucleus, respectively. A single element 375MHz center frequency transducer (SASAM) was used to generate acoustic impedance maps of inverted MCF-7 cells on a plastic substrate. The acoustic impedance of 20 single-live, 10 clustered-live and 7 clustered-fixed MCF7 cells were measured and compared. Fluorescent images and acoustic impedance maps suggest that there is an acoustic impedance mismatch between the nucleus and the cytoplasm. The average acoustic impedance values for single-live, clustered-live and clustered-fixed cancer cells were measured to be 1.600±0.006 MRayl, 1.609±0.009 MRayl and 1.572±0.011 MRayl.
Many types of cancer therapies target the tumor microenvironment, causing biochemical and morphol... more Many types of cancer therapies target the tumor microenvironment, causing biochemical and morphological changes in tissues. In therapies using ultrasound activated microbubbles, vascular collapse is typically reported. Red blood cells (RBCs) that leak out of the vasculature become exposed to the ceramide that is released from damaged endothelial cells. Ceramide can induce programmed cell death in RBCs (eryptosis), and is characterized by cell shrinkage, membrane blebbing and scrambling. Since the effect of eryptotic cells on generated photoacoustics (PA) signals has not been reported, we investigated the potential PA may have for cancer treatment monitoring by using PA spectral analysis to sense eryptosis. To induce eryptosis, C2-ceramide was added to RBC suspensions and that were incubated for 24 hours at 37 o C. A control and ceramide-induced sample was imaged in a vessel phantom using a high frequency PA system (VevoLAZR, 10 – 45 MHz bandwidth) irradiated with multiple wavelengths ranging from 680 to 900 nm. PA spectral parameters were measured and linked to changes in RBCs as it underwent eryptosis. These samples were examined using optical microscopy, a blood gas analyzer and an integrating sphere setup to measure optical properties (wavelengths 600 – 900 nm). The results of the experiment demonstrate how PA spectral analysis can be used to identify eryptosis at a depth of more than 1 cm into the phantom using ultrasound derived the y-intercept and mid bandfit (MBF) parameters at optical wavelengths of 800 – 900 nm. These parameters were correlated to the morphological and biochemical changes that eryptotic RBCs display. The results establish the potential of PA in cancer treatment monitoring through sensing treatment induced eryptosis.
Quantitative ultrasound parameters based on form factor models were investigated as potential bio... more Quantitative ultrasound parameters based on form factor models were investigated as potential biomarkers of cell death in breast tumor (MDA-231) xenografts treated with chemotherapy. Ultrasound backscatter radiofrequency data were acquired from MDA-231 breast cancer tumor-bearing mice (n = 20) before and after the administration of chemotherapy drugs at two ultrasound frequencies: 7 MHz and 20 MHz. Radiofrequency spectral analysis involved estimating the backscatter coefficient from regions of interest in the center of the tumor, to which form factor models were fitted, resulting in estimates of average scatterer diameter and average acoustic concentration (AAC). The ∆AAC parameter extracted from the spherical Gaussian model was found to be the most effective cell death biomarker (at the lower frequency range, r(2) = 0.40). At both frequencies, AAC in the treated tumors increased significantly (P = .026 and .035 at low and high frequencies, respectively) 24 hours after treatment compared with control tumors. Furthermore, stepwise multiple linear regression analysis of the low-frequency data revealed that a multiparameter quantitative ultrasound model was strongly correlated to cell death determined histologically posttreatment (r(2) = 0.74). The Gaussian form factor model-based scattering parameters can potentially be used to track the extent of cell death at clinically relevant frequencies (7 MHz). The 20-MHz results agreed with previous findings in which parameters related to the backscatter intensity (i.e., AAC) increased with cell death. The findings suggested that, in addition to the backscatter coefficient parameter ∆AAC, biological features including tumor heterogeneity and initial tumor volume were important factors in the prediction of cell death response.
Apoptosis is a form of programmed cell death characterized by a series of predictable morphologic... more Apoptosis is a form of programmed cell death characterized by a series of predictable morphological changes at the subcellular level, which modify the light-scattering properties of cells. We present a spectroscopic optical coherence tomography (OCT) technique to detect changes in subcellular morphology related to apoptosis in vitro and in vivo. OCT data were acquired from acute myeloid leukemia (AML) cells treated with cisplatin over a 48-h period. The backscatter spectrum of the OCT signal acquired from the cell samples was characterized by calculating its in vitro integrated backscatter (IB) and spectral slope (SS). The IB increased with treatment duration, while the SS decreased, with the most significant changes occurring after 24 to 48 h of treatment. These changes coincided with striking morphological transformations in the cells and their nuclei. Similar trends in the spectral parameter values were observed in vivo in solid tumors grown from AML cells in mice, which were treated with chemotherapy and radiation. Our results provide a strong foundation from which future experiments may be designed to further understand the effect of cellular morphology and kinetics of apoptosis on the OCT signal and demonstrate the feasibility of using this technique in vivo.
Photons Plus Ultrasound: Imaging and Sensing 2015, 2015
ABSTRACT A methodology for simulating photoacoustic (PA) images of samples with solid spherical a... more ABSTRACT A methodology for simulating photoacoustic (PA) images of samples with solid spherical absorbers acquired using linear array transducer is described. Two types of numerical phantoms (i.e. polystyrene beads suspended in agar medium) of two different size regimes were imaged with a 40 MHz linear array transducer utilizing this approach. The frequency domain features and statistics of the simulated signals were quantified for tissue characterization. The midband fit at 40 MHz was found to be about 35 dB higher for the sample with larger beads (radius ∼ 7.36 μm) than that of the sample with smaller particles (radius ∼ 1.77 μm). The scale parameter of the generalized gamma distribution function was found to be nearly 51 times greater for the former sample compared to the latter sample. The method developed here shows potential to be used as a fast simulation tool for the PA imaging of collection of absorbers mimicking biological tissue.
Biomedical Applications of Light Scattering VIII, 2014
ABSTRACT We present an in vivo implementation of a multi-parametric technique for detecting apopt... more ABSTRACT We present an in vivo implementation of a multi-parametric technique for detecting apoptosis using optical coherence tomography in a mouse tumor model. Solid tumors were grown from acute myeloid leukemia cells in the hind leg of SCID mice and treated with a single dose of cisplatin and dexamethasone to induce apoptosis. Both spectral features and speckle decorrelation times indicated good consistency between control mice and reasonable agreement with in vitro measurements. The integrated backscatter increased significantly in tumors responding to treatment while the spectral slope and decorrelation time did not show significant changes. This study demonstrates the feasibility of using spectroscopic OCT and dynamic light scattering for treatment monitoring in vivo.
2014 IEEE International Ultrasonics Symposium, 2014
ABSTRACT Ultrasound thermal therapy is used for noninvasive treatment of cancer. For accurate ult... more ABSTRACT Ultrasound thermal therapy is used for noninvasive treatment of cancer. For accurate ultrasound based temperature monitoring in thermal therapy, the temperature dependence of acoustic parameters is required. In this study, the temperature dependence of acoustic harmonics was investigated in water. The pressure amplitudes of the transmitted fundamental frequency (p1), and its harmonics (second (p2), third (p3), fourth (p4), and fifth (p5)) generated by nonlinear ultrasound propagation were measured by a calibrated hydrophone in water. The hydrophone was placed at the focal point of a focused 5-MHz transducer (f-number 4.5) to measure the acoustic pressure. Higher harmonics were generated by transmitting a 5-MHz 15-cycle pulse that resulted in a focal positive peak pressure of approximately 0.26 MPa in water. The water temperature was increased from 26 °C to 52 °C in increments of 2 °C. Due to this temperature elevation, the value of p1 decreased by 9%±1.5% (compared to its value at 26 °C) and values of p2, p3, p4, and p5 increased by %5±2%, 22%±8%, 44%±7%, and 55%±5%, respectively. The results indicate that the nonlinear harmonics are highly temperature dependent and their temperature sensitivity increase with the harmonic number. It is concluded that the nonlinear harmonics could potentially be used for ultrasound-based thermometry.
The photoacoustic signal generated from particles when irradiated by light is determined by attri... more The photoacoustic signal generated from particles when irradiated by light is determined by attributes of the particle such as the size, speed of sound, morphology and the optical absorption coefficient. Unique features such as periodically varying minima and maxima are observed throughout the photoacoustic signal power spectrum, where the periodicity depends on these physical attributes. The frequency content of the photoacoustic signals can be used to obtain the physical attributes of unknown particles by comparison to analytical solutions of homogeneous symmetric geometric structures, such as spheres. However, analytical solutions do not exist for irregularly shaped particles, inhomogeneous particles or particles near structures. A finite element model (FEM) was used to simulate photoacoustic wave propagation from four different particle configurations: a homogeneous particle suspended in water, a homogeneous particle on a reflecting boundary, an inhomogeneous particle with an ab...
Early alterations in textural characteristics of quantitative ultrasound spectral parametric maps... more Early alterations in textural characteristics of quantitative ultrasound spectral parametric maps, in conjunction with changes in their mean values, are demonstrated here, for the first time, to be capable of predicting ultimate clinical/pathologic responses of breast cancer patients to chemotherapy. Mechanisms of cell death, induced by chemotherapy within tumor, introduce morphological alterations in cancerous cells, resulting in measurable changes in tissue echogenicity. We have demonstrated that the development of such changes is reflected in early alterations in textural characteristics of quantitative ultrasound spectral parametric maps, followed by consequent changes in their mean values. The spectral/textural biomarkers derived on this basis have been demonstrated as non-invasive surrogates of breast cancer chemotherapy response. Particularly, spectral biomarkers sensitive to the size and concentration of acoustic scatterers could predict treatment response of patients with u...
Modulation of the tumour microvasculature has been demonstrated to affect the effectiveness of ra... more Modulation of the tumour microvasculature has been demonstrated to affect the effectiveness of radiation, stimulating the search for anti-angiogenic and vascular-disrupting treatment modalities. Microbubbles stimulated by ultrasound have recently been demonstrated as a radiation enhancer when used with different cancer models including PC3. Here, photoacoustics imaging technique was used to assess this treatment's effects on haemoglobin levels and oxygen saturation. Correlations between this modality and power doppler assessments of blood flow, and histology measurements of vascular integrity and cell death were also investigated. Xenograft prostate tumours in SCID mice were treated with 0, 2, or 8 Gy radiation combined with microbubbles exposed to 500 kHz ultrasound at a peak negative pressure of 0, 570, and 750 kPa. Tumours were assessed and levels of total haemoglobin, oxygen saturation were measured using photoacoustics before and 24 hours after treatment along with power do...
Uploads
Papers by Michael Kolios