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1.
Article in Chinese | WPRIM | ID: wpr-932390

ABSTRACT

Objective:To evaluate the feasibility and applicability of using phospholipid-hybridization method for preparing biomimetic microbubbles (Bio-MBs) ultrasound contrast agents.Methods:Leukocyte biomimetic microbubbles (MB leu), platelet biomimetic microbubbles (MB pla) and erythrocyte biomimetic microbubbles (MB ery) were prepared by multiple steps: film-hydration, phospholipid-hybridization, mechanical oscillation. The size and zeta potential of Bio-MBs were measured by dynamic light scattering. A laser scanning confocal microscopy experiment was performed to confirm the presence of membrane proteins on the shell of Bio-MBs. The fluorescence of FITC-labeled typical membrane protein was evaluated using a flow cytometer. Sodium dodecyl sulfate polyacrylamide gel electrophoresis was used to characterize the membrane protein. Biosafety of Bio-MBs was evaluated by CCK-8 counting kit, blood and major organs. The contrast enhancement effect and stability were observed in vitro and in vivo. An in vivo fluorescence imaging system was performed to evaluate the distribution of Bio-MBs. The application value of biomimetic microbubbles was measured by ultrasound molecular imaging by using ischemia-reperfusion rat models and acute hepatitis rat models. Results:Bio-MBs with spherical shape distributed homogenously, without obvious aggregation. The membrane proteins were successfully integrated into the shell of Bio-MBs.The diameter of three Bio-MBs was similar to that of control microbubbles (MB con) ( P>0.05), three Bio-MBs had a lower zeta potential than MB con ( P<0.05). The Bio-MBs had an appreciable performance in vitro and in vivo biosafety. The Bio-MBs retained the main proteins inherited from cell membrane. Contrast enhanced ultrasound imaging in vitro and in vivo showed that the Bio-MBs had a stable imaging ability.MB leu and MB pla have good targeted imaging effect in two disease models. Conclusions:A series of Bio-MBs ultrasound contrast agents, which have high stability, biosafety and targeted imaging efficiency, were successfully prepared by using phospholipid-hybridization method. This fabrication method for obtaining Bio-MBs can be applied to different clinical scenarios with different cell types in the future.

2.
Article in Chinese | WPRIM | ID: wpr-486857

ABSTRACT

Objective To investigate the therapeutic effect of high intensity focused ultrasound (HIFU) ablation combined with homemade liquid fluorocarbon nanoparticles on cervical cancer in nude mice.Methods The cell experiment was divided into three groups:a,control group;b,HIFU group;c, HIFU + PFB nanoparticle group,and the viability of cells was detected using CCK-8 reagent.The mice were also divided into three groups:A,0.9% NaCI group;B,HIFU + 0.9% NaCI group;C,HIFU + PFB nanoparticle group. The tumors were removed and underwent triphenyl tetrazolium chloride(TTC) staining,and the necrosis area was measured.Histopathological changes of the tumors were examined by light microscopy.Results After HIFU irradiation,the viability rate of group c was (40.5 ±9.7)%,it was lower than that of group b (77.7 ±8.5)% (P <0.05) and that of group a(100 ±4.8)% (P <0.05). TTC staining of tumor showed a large scale of necrotic tissue in group C.The necrosis ratio of the three groups was 0%,(34.14±12.2)% and (65.97 ±25.1)%,respectively (P <0.05).HE staining showed karyorrhexis or an absence of nuclei in group B and group C,which demonstrated the coagulation necrosis. Conclusions HIFU ablation combined with liquid fluorocarbon nanoparticles can effectively treat the xenograft model of the human cervical carcinoma in nude mice.

3.
Article in Chinese | WPRIM | ID: wpr-355266

ABSTRACT

<p><b>OBJECTIVE</b>To evaluate the feasibility of integrating cancer gene therapy with therapeutic effect evaluation using siRNA-loaded nano-scale microbubbles (siRNA-NBs).</p><p><b>METHODS</b>siRNA-NBs were prepared by hetero-assembly of polymeric siRNA micelles and liposomal microbubbles, and the particle sizes and surface potentials were examined with dynamic light scattering. The distributions of cy3-labled siRNA in the tumor tissues were evaluated using confocal laser scanning microscopy. A siRNA targeting the anti-apoptosis gene SIRT2 was designed and its gene silencing effects was tested in vivo using siRNA-NBs with ultrasound exposure. The therapeutic effect of the loaded siRNA-NBs was evaluated by contrast-enhanced ultrasonography.</p><p><b>RESULTS</b>The siRNA-NBs had a mean diameter of 400.7 ± 30.5 nm with a weak positive charge of +8.8 ± 0.8 mV. With ultrasound exposure, siRNA-NBs effectively delivered cy3-siRNA into the cytoplasm of cancer cells and caused SIRT2 suppression and cell apoptosis in tumor tissues, resulting in significantly suppressed tumor growth. In addition, contrast-enhanced ultrasonography of siRNA-NBs provided good imaging quality to allow real-time observation of blood supply during gene therapy.</p><p><b>CONCLUSIONS</b>As a novel ultrasound contrast agent, siRNA-NBs make possible the integration of tumor gene therapy and therapeutic effect evaluation for cancer.</p>


Subject(s)
Apoptosis , Contrast Media , Gene Silencing , Genetic Therapy , Humans , Liposomes , Micelles , Microbubbles , Neoplasms , Therapeutics , Particle Size , Polymers , RNA, Small Interfering , Sirtuin 2 , Genetics , Ultrasonics
4.
Chinese Journal of Ultrasonography ; (12): 1078-1081, 2012.
Article in Chinese | WPRIM | ID: wpr-430030

ABSTRACT

Objective To compare the efficiency of oscillation with sonication in preparing nano-scale microbubbles (NBs).Methods Nano-scale microbubbles were prepared using oscillation and sonication respectively,and then compared the NBs' size,size distribution,concentrations and time-consumption of the two methods.Results The sizes of nanobubbles prepared by sonication and oscillation were (373.88 ±18.43)nm and (360.74 ± 14.39)nm,respectively.There was no significant difference in size between the two methods (P =0.523).The polidispersity was larger in sonication before centrifugation,there was significant difference between the two methods (P <0.001).The concentration of nanobubbles prepared by oscillation was (1.48 ± 0.15) × 1010,which was higher than that by oscillation [(8.07 ± 0.62) × 108],there was significant difference between the two methods (P < 0.001).The consuming time was shorter in oscillation,the difference was significant when compared with sonication (P <0.001).Conclusions Both two methods can successfully prepare NBs.Compared with sonication,oscillation can effectively produce NBs with smaller polidispersity,higher concentration and shorter time-consumption.

5.
Article in Chinese | WPRIM | ID: wpr-429731

ABSTRACT

Objective To tested the passive targeting of nanobubbles penetrating tumor vascular endothelial cells gap.Methods Twenty female BALB/c nude mice subcutaneously bearing human ovary cancer SKOV3 were devided into two groups:group A (ultrasound imaging) and group B (frozen sections:B1 and B2).DiI labled nanobubbles and microbubbles were prepared and adjusted into the same concentrations.Group A:Microbubbles and nanobubbles of 35 μl were injected into the tail vein of every mouse respectively (1.5 h interval).Ultrasound imaging were acquired.Group B:Nanobubbles and microbubbles of 10 μl were injected into the tail vein of mice in Group B1 and Group B2 respectively.Heart perfusion by PBS or 0.9% normal saline was carried out 1.5 h after bubbles injection to clear the free bubbles in blood circulation.And the tumor and muscle of right lower limb were immediately cut off for frozen slices (3 μm),which were stained by Hoechst 33342 to mark the nucleus.Images were obtained with a confocal microscope.Results In vivo ultrasound imaging,the time to peak and clearance time of nanobubbles were longer than those of microbubbles,whereas the intensity of enhancement was lower than microbubbles.Frozen sections showed:with the confocal laser scanning microscopy imaging,quite a number of DiI-labeled nanobubbles existed in the intercellular space of SKOV3 tumor,whereas there were few nanobubbles in skeletal muscle sections.In the control,rare DiI-labeled microbubbles were observed in tumors and skeletal muscle.Conclusions Self-made lipid nanobubbles were small enough to pass through the tumor vascular endothelial gap,namely achieve the tumor passive targeting.

6.
Article in Chinese | WPRIM | ID: wpr-423555

ABSTRACT

Objective To prepare targeted nanoscale lipid ultrasound contrast agent and study the targeting function in vitro.Methods After the biotinylated monoclonal antibody Herceptin was prepared,the biotinylated degree and immunological activity were determined.Then biotinylated antibody was attached to the surface of nanoscale lipid ultrasound contrast agents by avidin-biotin system to prepare the targeted nanobubbles.The targeting function was studied by observing the combination ability of the targeted nanobubbles with SKOV3 cells in vitro,non-targeted nanobubbles as controls,and observing ultrasound imaging in vitro.Results About 16 biotin molecules were coupled to each antibody in average,and the immunological activity of the biotinylated antibody didn't decrease compared with the free one(P >0.05).SKOV3 cells were combined firmly and surrounded regularly by red dyed targeted nanobubbles,while control groups were negative.Ultrasound imaging could be significantly enhanced by targeted nanobubble binding to SKOV3 cell slides,the other two control groups were negative.Conclusions Nanoscale ultrasound contrast agent and antibodys can be combined firmly by avidin-biotin system to produce the targeted nanobubbles,which have strong targeting function in vitro and significantly enhanced ultrasound signal.

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