| This thesis researches on several effects induced by ultrasound and laser irradiations on biological samples, which mainly include two topics:(1) Thermodynamic and dynamic effects of photolyses of gas-liganded hemoglobins and the structure differences between the studied liganded hemoglobins, which are described in the second and third chapters separately. (2) Experimental and theoretical researches of thermal effects of biological tissues induced by moderate intensity focused ultrasound and enhanced by laser coordinated irradiations, which are presented in the fourth and fifth chapters separately. In addition, the Introduction (the first chapter) describes briefly the recent research progresses of related fields and the Summary (the sixth chapter) reports the final results and presents the prospects on the fields. Finally, the contents of both topics are described briefly as follows:1. The investigation of the thermodynamic and dynamic effects of photolysis of gas-liganded hemoglobinsHemoglobin (Hb) is an important oxygen-binding protein. The study of the molecular structures and internal energy changes during the photolysis and recombination processes of gas-liganded Hb is considered as a fundamental step to understand the molecular structures and function of hemoglobin.In chapter 2, the photolyses of the carboxy-hemeglobin (HbCO) for five kinds of mammals (human, bovine, pig, horse and rabbit) are studied. The thermodynamic and dynamic parameters of the photolyses including the quantum yields, and enthalpy and conformational volume changes are measured by using laser pumping-probing technique (LPPT) and photoacoustic calorimetry (PAC) respectively. In the PAC experiment, two different kinds of the receiving transducers are used to detect the enthalpy and conformational volume changes of two photolysis processes of HbCO. The PVDF transducer is used to detect the fast geminate recombination with the lifetime about 44 ns, while the 1.5MHz PZT transducer detects mainly the tertiary relaxation process with lifetime about 700 ns. The enthalpy and conformational volume changes of the two processes of HbCO photolysis for five kinds of mammals can be obtained by combining with the quantum yields measurement and the related theoretical calculations. The results indicate that the photolysis parameters for different mammals are different, but most of them are similar with each other.To study the influence of the living environments on the photolysis parameters, the photolyses of HbCO for high-altitude species (bar-headed goose) and its low altitude counterparts (Chinese goose and chicken) are investigated by the same methods. The bar-headed goose as a typical high-altitude species, has the capacity for oxygen uptake from a hypoxic environment, which means it may has a higher oxygen affinity than other species from lower-altitude environments. The study of the photolysis of gas-liganded Hb for bar-headed goose has important significance for the studies of the structures and functions of Hb.The measurement results show that the photolysis parameters of bar-headed goose and its low altitude counterparts have big differences, while the parameters of the bar-headed goose are considerably smaller than those of the others. It can be proposed that the association or dissociation of the Hb with liganded gases for bar-headed goose requires less energy and can be realized more easily. So the living environments may have influence on the photolysis parameters of HbCO.In chapter 3, to explain the possible mechanisms of these differences of the thermodynamic and dynamic parameters, the HbCO molecule structures of the samples have been analyzed and compared. At first, the structures of Hb and the allosteric mechanism in the binding and dissociating processes of Hb with ligand gases are presented. Then the amino acid sequences of Hb, the modifications of salt bridges and hydrogen bonds at subunit interfaces of Hb are compared and discussed briefly.By analyzing the differences of the Hb structures,20% of the amino acid residues are different among the five mammalian hemoglobins. The salt-bridges numbers in Hb of human and rabbit are more than the others, and the hydrogen bond numbers in five mammalian Hb are also different among them, while the differences of the structures of rabbit Hb are particularly obvious. So the differences of the amino acid residues, salt bridges and hydrogen bonds of Hb may be the possible factors to affect the differences of the photolysis parameters of the liganded Hb among five mammals.The Hb structures of bar-headed goose and its low altitude counterparts are also analyzed and compared. The results show that there are four residues different between the Hb of bar-headed goose and its low altitude counterparts (goose), which located at the surfaces of the proteins, and the variants are easily to be displayed in the photolysis processes. Since the salt-bridges numbers in Hb of bar-headed goose and goose are similar with each other, the salt bridge modifications are analyzed. The salt bridge modifications and involved structure changes during the HbCO photolysis of bar-headed goose are much smaller than its low altitude counterparts, which may explain why the enthalpy and conformational volume changes of bar-headed goose HbCO are smaller than its low altitude counterparts.2. Experimental and theoretical researches of laser-enhanced thermal effects of moderate intensity focused ultrasound on the biological tissuesThe thermal effect of the focused ultrasound is a hot point of the researches in recent years. In this thesis, the heating processes induced by moderate intensity focused ultrasound and enhanced by combined irradiations of laser pulses for bio-tissues are studied experimentally and theoretically.In chapter 4, a focused PZT-based transducer with the central frequency 1.3 MHz is employed to produce ultrasonic continuous waves, and a pulsed laser with the width 8 ns, repetition frequency 10 Hz, is used as the combined irradiation source. The temperature variation around the co-focal point is measured by a thermocouple with the diameter of 130μm. A focused transducer with broadband of 2-16 MHz is used to detect the ultrasonic harmonic and/or cavitation signals, which may be helpful to the further study of the mechanism of enhanced thermal effects. The bio-tissues used are fresh porcine liver, kidney, heart and fat tissue, while several damaged tissues and tissue-mimicking materials are also studied for comparison. The results indicate that the laser-enhanced thermal effects induced by the ultrasound appear more easily in the fresh bio-tissues.When the intensity of the ultrasound is large enough (1.25 MPa), the enhanced thermal effects and increases of the harmonic signals and broadband chaos indicate that the cavitation occurred in the samples. The synergetic irradiations of laser pulses produce easily additional cavitation bubbles, which induce more broadband noises at low frequency range, scattering and attenuations, and then enhance the temperature rises of the samples. For fresh bio-tissues, the laser induced changes of the micromorphology and microstructures of the tissues, which may benefit to provide nucleation sites. Thus, the cavitations induced by the transient photo-biological effects in bioactive tissues may occur more easily than other samplesIn chapter 5, the theoretical calculation of the thermal effects induced by ultrasound are studied based on the Westervelt Equation and Pennes Bioheat Transfer Equation, and numerically calculated by the COMSOL Multiphysics 5.1. Considering the cavitations do not occur when the intensity of the ultrasound is less than the cavitation threshold (1.25 MPa), the theoretical calculations are adjusted to fit the experimental results at low ultrasonic intensities. When the intensity of the ultrasound is larger than 1.25 MPa, the theoretical calculated results are less than those of the experiments, and the temperature differences between them may be induced by cavitations. To study additional thermal effects by ultrasound with much large intensity and enhanced by laser, the temperature elevation induced by a single cavitation bubble is calculated for the bio-tissues and tissue mimicking materials. By fitting the calculations with the experimental results, the corresponding numbers of bubbles induced by the cavitation effects can be estimated. Based on the calculations, the numbers of bubbles in both the fresh bio-tissues and the tissue-mimicking materials are increased with the raising of the ultrasound pressure. Based on the enhanced thermal effects induced by the combined laser, the number of bubbles induced by the combined laser can also be estimated, which is also increased with the raising of the ultrasound pressure. It is suggested that the cavitation enhanced by the combined laser in the fresh bio-tissues are more obvious than that in the tissue-mimicking materials.The combination treatments of ultrasound and laser can be recognized as a beneficial method to increase heating efficiencies, while the ultrasonic intensity can be reduced by combined with the laser to get the similar heating effects, which may provide auxiliary methods for medical therapies.In chapter 6, the summary of all contents and prospects for future work are presented. |
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