We studied the dependence of a perpendicular pressure to the luminal surface on the heating drug delivery performance using a laser-mediated thermal balloon with porcine carotid artery walls ex vivo. We proposed the combination use of our laser-mediated thermal balloon and drug coated balloon (DCB) to enhance a drug delivery performance. We prospected that the perpendicular pressure, which was applied directly to the luminal surface by balloon dilatation, would enhance the quantity of DCB drug delivered into artery wall. To simulate our laser thermal balloon heating, 63°C preheated artery samples were prepared by heated saline dropping for 15 s, and then these samples were dipped in 37°C saline for 15 s. Non-heated artery samples were prepared by dipping in 37°C saline for 30 s. The perpendicular pressure up to 10 atm corresponding to DCB dilatation pressure was added directly to these artery samples by fluorescence Rhodamine B solution for 30 s. The quantity of drug delivered was microscopically measured with fluorescence brightness in the cross-section of the drug delivered artery samples. We found drastic drug delivery increase at 8 atm using the pre-heated artery sample. Delamination of intima layer was observed by EVG stained cross-sectional specimens with 8 atm in the pre-heated artery sample. We think this drastic pressure dependence on the heating drug delivery performance might be corresponding to increase in permeability of drug into the artery wall originated to morphology change in intima.
To enhance drug delivery performance of drug eluting balloon (DEB) against re-stenosis, we have proposed a heating drug delivery during balloon dilatation using our laser driven short-term thermal angioplasty which may realize to suppress surrounding thermal injury. We studied an influence of vessel dilatation parameters on the heating drug delivery. These parameters were classified into two different forces, that is, circumferential tension and inter-luminal pressure. We think these parameters were not able to determine only by balloon pressure. The circumferential tension with 0—30 mN/mm2 was added to a porcine carotid artery using an automatic stage. Various temperature solutions with 37, and 70°C of hydrophobic fluorescent Rhodamine B with 3 μg/ml in concentration were dropped on pig carotid wall. We measured a defined drug delivery amount as well as delivery depth by a microscopic fluorescence measurement on the cross section of the solution delivered vessel. In the case of 37°C, we found the intima surface drug amount with 7 mN/mm2 was increased as 10-20 times as other tension cases. On the other hand, at 70°C, we found the optimum tension with 30 mN/mm2. We found the drug delivery enhancement might be related to the change of super microscopic surface structure of the vessel. We predict that the collagen thermal denaturation of the vessel wall might play important role to the drug delivery.
To enhance drug delivery performance of popular drug eluting balloon against re-stenosis after angioplasty, we have an idea regarding to adjacent use of our unique laser driven thermal balloon of which characteristics could realize short term and uniform temperature elevation to modify drug delivery characteristics. We have already reported a delivery enhancement effect using this idea, however, detailed characteristics have not been studied yet. We studied balloon dilatation in terms of vascular circumferential tension on the heating drug delivery performance using porcine carotid artery wall ex vivo. The extracted carotid artery was used and circumferential tension of 0-30 mN/mm2 was added. Heating drug delivery was performed on this carotid artery with the heated solution of hydrophobic fluorescent Rhodamine B with 3 μg/ml in concentration at 37 and 70°C. We obtained a defined drug delivery quantity as well as delivery depth by a microscopic fluorescence measurement on a cross section of the drug delivered vessel wall. In the cases of 70°C, we found the drug penetration increase against 3°C case. We predict that the collagen thermal denaturation of the vessel wall may play important role to this penetration. In the case of 3°C, we found the drug concentration on the intimal surface with 7 mN/mm2 was increased as 10-30 times as other tension values. We found surface grooves in this case using an electron micrography. Therefore, we think that the drug delivery enhancement might be related to the groove formations of the vessel wall.
We studied heating drug delivery to vascular wall with Rhodamine B ranging 50 to 70°C ex vivo study. Porcine carotid artery was dipped in the heated Rhodamine B solution in 15 s and then cooled by 37°C saline. Rhodamine B concentration distribution in the vascular wall cross-section was measured by a fluorescence microscope using 550 nm for excitation and 620 nm emission for fluorescence detection. The total amount of measured fluorescence in the vascular wall was calculated as a indication of delivered Rhodamine B quantity. The delivered Rhodamine B quantity was increased with increasing heating temperature with 50 to 70°C. In the cases of 60 to 70°C heating, the delivered Rhodamine B quantity was 3.1 to 23.3 fold by that of 37°C. Defined penetration depth of the delivered Rhodamine B in the vascular wall was also significantly increased with 65°C and 70°C heating. We also studied heating drug delivery to the vascular wall with fluorescence labeled Paclitaxel with 70°C in 15 s and 60 s heating ex vivo. In both contact duration, the delivered Paclitaxel quantity was increased. To understand these drug delivery enhancement effects, we investigated the vascular cross-sectional structure change by the heating. Some holes over 50 nm in diameter appeared on the internal elastic lamina with 70°C heating. We prospected that vascular surface structure change by the heating might enhance drug delivery to the vascular wall.
KEYWORDS: Cell death, Arteries, In vitro testing, In vivo imaging, Ranging, Injuries, Microscopes, Temperature metrology, Thermal effects, Biomedical optics
We studied the relations between the time history of smooth muscle cells (SMCs) death rate and heating condition in vitro to clarify cell death mechanism in heating angioplasty, in particular under the condition in which intimal hyperplasia growth had been prevented in vivo swine experiment. A flow heating system on the microscope stage was used for the SMCs death rate measurement during or after the heating. The cells were loaded step-heating by heated flow using a heater equipped in a Photo-thermo dynamic balloon. The heating temperature was set to 37, 50-60°C. The SMCs death rate was calculated by a division of PI stained cell number by Hoechst33342 stained cell number. The SMCs death rate increased 5-10% linearly during 20 s with the heating. The SMCs death rate increased with duration up to 15 min after 5 s heating. Because fragmented nuclei were observed from approximately 5 min after the heating, we defined that acute necrosis and late necrosis were corresponded to within 5 min after the heating and over 5 min after the heating, respectively. This late necrosis is probably corresponding to apoptosis. The ratio of necrotic interaction divided the acute necrosis rate by the late necrosis was calculated based on this consideration as 1.3 under the particular condition in which intimal hyperplasia growth was prevented in vivo previous porcine experiment. We think that necrotic interaction rate is larger than expected rate to obtain intimal hyperplasia suppression.
The purpose of this study was to investigate the artery dilatation performance of the short-duration heating balloon catheter in cadaver stenotic arteries. We designed a prototype short-duration heating balloon catheter that can heat artery media to around 60 °C in 15−25 s by a combination of laser-driven heat generation and continuous fluid irrigation in the balloon. We performed ex vivo short-duration heating dilatation in the cadaver atherosclerotic femoral arteries (initial percent diameter stenosis was 36-98%), with the maximum balloon temperature of 65±5 °C, laser irradiation duration of 25 s, and balloon dilatation pressure of 3.5 atm. The artery lumen configurations before and after the dilatations were assessed with a commercial IVUS system. After the short-duration heating dilatations, the percent diameter stenosis was reduced below 30% without any artery tears or dissections. We estimated that the artery media temperature was raised to around 60 °C in which plaque thickness was below 0.8 mm by a thermal conduction calculation. The estimated maximum temperature in artery adventitia and surrounding tissue was up to 45 °C. We found that the short-duration heating balloon could sufficiently dilate the cadaver stenotic arteries, without thermal injury in artery adventitia and surroundings.
We proposed a short-duration heating balloon angioplasty. We designed a prototype short-duration heating balloon
catheter that can heat artery media to 60-70°C within 15-25 s with a combination of laser-driven heat generation and
continuous fluid irrigation in the balloon. The purpose of this study was to investigate chronic artery lumen patency as
well as histological alteration of artery wall after the short-duration heating balloon dilatation with porcine healthy iliac
artery. The short-term heating balloon dilated sites were angiographically patent in acute (1 hour) and in chronic phases
(1 and 4 weeks). One week after the dilatation, smooth muscle cells (SMCs) density in the artery media measured from
H&E-stained specimens was approx. 20% lower than that in the reference artery. One and four weeks after the
dilatations, normal structure of artery adventitia was maintained without any incidence of thermal injury. Normal
lamellar structure of the artery media was also maintained. We found that the localized heating restricted to artery media
by the short-duration heating could maintain adventitial function and artery normal structure in chronic phase.
We investigated the influences on smooth muscle cells after our novel short-duration thermal angioplasty, Photo-thermo
Dynamic Balloon Angioplasty (PTDBA), to reveal the mechanism that can suppress neo-intimal hyperplasia after
PTDBA. We obtained the sufficient arterial dilatations by short-duration heating (≤15 s, <70°C) and low dilatation
pressure (<0.4 MPa) without arterial injuries in our previous in vivo studies. Smooth muscle cells, which play most
important role in chronic treatment effects, were heated during PTDBA and stretch-fixed after PTDBA. The dead cell
rate by heating, estimated by Arrhenius equation with A=2.5x1016 s-1 and Ea=1.17×105 J mol-1, was 15.7±2.2% after
PTDBA. The measured deformation rate of smooth muscle cells' nuclei was 1.6±0.1 after PTDBA in vivo. We found
that the expression of smooth muscle cells' growth factor after PTDBA was inhibited 0.52 fold compared to that after the
conventional balloon angioplasty in vivo. The measured neo-intimal hyperplasia occupancy rate was less than 20% after
PTDBA in vivo. We prospect that the inhibition of the growth factor's expression by stretch-fixing may result to
suppress the neo-intimal hyperplasia. In addition, the decrease of smooth muscle cells' density in the vessel media by
heating might be another reason for the neo-intimal hyperplasia suppression.
We investigated the effect on smooth muscle cells' proliferation with stretch-fixing in both in vitro and in vivo porcine
study to determine the optimum heat condition of novel short-term thermal angioplasty, Photo-thermo Dynamic Balloon
Angioplasty (PTDBA). With PTDBA, we have obtained the sufficient arterial dilatation by short-term heating (< 15 s, <
70 °C) and low dilatation pressure (< 0.4 MPa) without excessive neo-intimal hyperplasia on chronic phase. The smooth
muscle cells were found to be fixed with stretched shape in vascular wall after PTDBA in vivo. The deformation rate of
smooth muscle cells' nuclei was 1.6 ± 0.1 after PTDBA (15 s, 65 °C, 0.35 MPa). The smooth muscle cells, which were
extracted from porcine arteries, were cultured on the specially designed equipment to give stretch-fixing stimulus in
vitro. The cell proliferation was inhibited at 20 % stretching compared to 15 % stretching significantly (p < 0.05). The
immunostaining specimens of basic Fibroblast Growth Factor (bFGF) and its receptor FGFR-1 were made from the
porcine arteries in vivo. We found that the expressions of bFGF and FGFR-1 in the media were not observed after
PTDBA. We think that these results suggested the possibility for the inhibition of the excessive cell proliferation after
PTDBA.
We have been developing the novel short-term heating angioplasty in which sufficient artery lumen-dilatation was
attained with thermal softening of collagen fiber in artery wall. In the present study, we investigated on the relation
between the mechanical properties of heated artery and thermal denaturation fractures of arterial collagen in ex vivo. We
employed Lumry-Eyring model to estimate temperature- and
time-dependent thermal denaturation fractures of arterial
collagen fiber during heating. We made a kinetic model of arterial collagen thermal denaturation by adjustment of K
and k in this model, those were the equilibrium constant of reversible denaturation and the rate constant of irreversible
denaturation. Meanwhile we demonstrated that the change of reduced scattering coefficient of whole artery wall during
heating reflected the reversible denaturation of the collagen in artery wall. Based on this phenomenon, the K was
determined experimentally by backscattered light intensity measurement (at 633nm) of extracted porcine carotid artery
during temperature elevation and descending (25°C→80°C→25°C). We employed the value of according to our earlier
report in which the time-and temperature- dependent irreversible denaturation amount of the artery collagen fiber that
was assessed by the artery birefringence. Then, the time- and temperature- dependent reversible (irreversible)
denaturation fraction defined as the reversible ((irreversible) denatured collagen amount) / (total collagen amount) was
calculated by the model. Thermo-mechanical analysis of artery wall was performed to compare the arterial mechanical
behaviors (softening, shrinkage) during heating with the calculated denaturation fraction with the model. In any artery
temperature condition in 70-80°, the irreversible denaturation fraction at which the artery thermal shrinkage started was
estimated to be around 20%. On the other hand, the calculated irreversible denaturation fraction remained below 5% and
reversible denaturation fraction reached up to 20% while the artery softening occurred without shrinkage. We think that
our model of arterial collagen thermal denaturation might be reasonable to estimate the artery mechanical properties
during heating.
We have studied to develop the new thermal angioplasty methodology, photo-thermo dynamic balloon angioplasty
(PTDBA), which provides artery dilatation with short-term (<15s) and uniform heating through the balloon by the
combination of the efficient laser driven heat generation and fluid perfusion. Thermal denaturation degree of the
collagen in artery media may be the important factor to attain sufficient artery dilatation for the PTDBA. In order to
predict the optimum heating condition i.e. the balloon temperature and heating duration, we investigated the thermal
denaturation dynamics of artery collagen in ex vivo. The extracted fresh porcine carotid artery was used. The
temperature-dependent light scattering property and mechanical property of the artery specimen were simultaneously
measured during artery temperature rising by specially made setup to assess the denaturation of arterial collagen. The
change rate of the backscattered light intensity from the artery specimen; I(T)/I0 with 633nm was measured to evaluate
the artery scattering property change with the thermal denaturation. The artery specimen was heated from 25°C to 80°C
with constant temperature rising rate of 3°C/min. The measured
I(T)/I0 was suddenly increased over 48°C. This
boundary temperature might be the initiation temperature of the arterial collagen denaturation. We defined the variation
of the I(T)/I0 as the collagen denaturation ratio, and calculated the reactive enthalpy by the chemical equilibrium theory.
Since the calculated enthalpy was similar to the enthalpy in literature report, the variety of I(T)/I0 during the temperature
rising might be attributed to the collagen conformational change due to the denaturation. In terms of the artery internal
force measurement, the artery force was decreased with increasing of the artery temperature up to 65°C (i.e. softening),
and increased over 65°C (i.e. shrinkage). We confirmed that the changes of the backscattered light (at 633nm in
wavelength) from the artery might represent the artery collagen thermal denaturation degree.
We studied Ho:YAG laser irradiation in blood vessel as a vasodilator ex vivo. We thought that the Ho:YAG
laser-induced bubble expansion might be able to dilate the vessel because we found the vessel wall expansion after the
Ho:YAG laser irradiation, that is steady deformation, in the vessel ex vivo. There have been many reports regarding to
the Ho:YAG laser irradiation in the vessel. Most of studies concentrated on the interaction between Ho:YAG laser
irradiation and vessel wall to investigate side effect on Ho:YAG laser angioplasty. We proposed to use the Ho:YAG
laser-induced bubble expansion as a vasodilator. We studied vasodilation effect of the Ho:YAG laser-induced bubble
ex vivo. The flash lamp excited Ho:YAG laser surgical unit (IH102, NIIC, Japan) (&lgr;=2.1&mgr;m) was used. The laser
energy was delivered by a silica glass fiber (outer diameter: 1000&mgr;m, core diameter: 600&mgr;m). The laser-induced
bubble was generated in the extracted fresh porcine carotid artery with the warmed saline perfusion. The laser energy
at the fiber tip was ranging from 170-1300mJ per pulse. Number of the laser irradiation was ranged from 20pulses to
100pulses. The outer diameter of the vessel was observed. To examine the change in mechanical properties of the
vessel wall, the stress-strain curve of the laser-irradiated vessel was measured. Birefringence observation and
microscopic observation of staining specimen were performed. When the laser energy was set to 1300mJ per pulse,
the outer diameter of the vessel after the laser irradiation was expanded by 1.4 times comparing with that of before the
laser irradiation and the dilatation effect was kept even at 10minutes after the irradiation. The elasticity modulus of
the artery by collagen was changed by the laser irradiation. In the polarized microscopic observation, the brightness of
the intimal side of the vessel is increased comparing with that of the normal. We think this brightness increasing may
be attributed to birefringence change by the arrangement of stretched collagen fiber. We suppose it is likely to be able
to use the Ho:YAG laser irradiation as a temporary vasodilater tool in spite of further study should be performed.
In order to investigate the optimum operation parameters on novel short-term heating (<15s, approx. 70oC) balloon,
named Photo-thermo dynamic balloon (PTDB), we studied diameter and elasticity change of vascular wall after
dilatation ex vivo. We have been studying to develop the PTDB angioplasty in which we demonstrated sufficient
vascular dilatation with lower pressure by heat- induced denaturation of arterial collagen. And we have also
demonstrated the suppression of intimal hyperplasia in animal experiments. We need to understand the PTDB dilatation
mechanism to determine the optimum operation parameters. The prototype PTDB with diameter of 3mm was used in
our experiments. The internal diameters of extracted fresh porcine carotid arteries at pre- and post- PTDB dilatation
were measured. Balloon parameters were follows; pressure P=2atm, peak temperature in balloon T=60-80oC, and
heating duration t=4-30s. Morphological change in the media of dilated artery with PTDB were microscopically
examined with Weigert staining. Elastic properties were carried out by stress-strain measurements with calculation of
young's modulus. We found that PTDB dilatation provided the effect to prevent elastic recoil. We explained that the
reason of this effect might be arrangement of micro- structure in the media, i.e., heat-denatured collagen fibers sustained
the elastic recoil due to rubbery elastin fibers. The arterial elasticity was not significant different after PTDB dilatation.
It was suggested that there could be no compliance mismatch after PTDB dilatation in physiological range. We found
that a part of PTDB dilatation mechanism, in which the vascular wall structure played an important role. The optimum
operation parameters for PTDB might be determined in consideration of collagen denaturation progress and arterial
composition.
KEYWORDS: Birefringence, Collagen, Tissues, Scanning electron microscopy, Temperature metrology, Tissue optics, Microscopy, In vivo imaging, Arteries, Microscopes
Our photo thermal reaction heating architecture balloon realizes less than 10 s short term heating that can soften vessel wall collagen without damaging surrounding tissue thermally. New thermal balloon angioplasty, photo-thermo dynamic balloon angioplasty (PTDBA) has experimentally shown sufficient opening with 2 atm low pressure dilation and prevention of chronic phase restenosis and acute phase thrombus in vivo. Even though PTDBA has high therapeutic potential, the most efficient heating condition is still under study, because relationship of treatment and thermal dose to vessel wall is not clarified yet. To study and set the most efficient heating condition, we have been working on establishment of temperature history estimation method from our previous experimental results. Heating target of PTDBA, collagen, thermally denatures following rate process. Denaturation is able to be quantified with measured collagen birefringence value. To express the denaturation with equation of rate process, the following ex vivo experiments were performed. Porcine extracted carotid artery was soaked in two different temperature saline baths to enforce constant temperature heating. Higher temperature bath was set to 40 to 80 degree Celsius and soaking duration was 5 to 40 s. Samples were observed by a polarizing microscope and a scanning electron microscope. The birefringence was measured by polarizing microscopic system using Brace-Koehler compensator 1/30 wavelength. The measured birefringence showed temperature dependency and quite fit with the rate process equation. We think vessel wall temperature is able to be estimated using the birefringence changes due to thermal denaturation.
Access to the requested content is limited to institutions that have purchased or subscribe to SPIE eBooks.
You are receiving this notice because your organization may not have SPIE eBooks access.*
*Shibboleth/Open Athens users─please
sign in
to access your institution's subscriptions.
To obtain this item, you may purchase the complete book in print or electronic format on
SPIE.org.
INSTITUTIONAL Select your institution to access the SPIE Digital Library.
PERSONAL Sign in with your SPIE account to access your personal subscriptions or to use specific features such as save to my library, sign up for alerts, save searches, etc.