- Email: [email protected]
High-intensity pulsed ultrasound and electric field permit localized pharmacotherapy targeting articular cartilage
S522
Abstracts / Osteoarthritis and Cartilage 24 (2016) S63eS534
C2, up to 900 mm depth into AC (Figure 1). Regarding tissue damage, no significant differences (p > 0.05) were found between control groups C1 and C2 at any depth. Furthermore, FESEM and histological analysis showed no evidence of structural changes induced by the treatment.
901 HIGH-INTENSITY PULSED ULTRASOUND AND ELECTRIC FIELD PERMIT LOCALIZED PHARMACOTHERAPY TARGETING ARTICULAR CARTILAGE rez y, H.J. Nieminen y, z, M. Finnila € z, A. Salmi y, A. García Pe €rvi y, T. Paulin y, K.P. Pritzker x, k, S. Saarakkala z, ¶, E. Lampsija € m y. y Univ. of Helsinki, Helsinki, Finland; z Univ. of Oulu, Oulu, E. Hæggstro Finland; x Univ. of Toronto, Toronto, ON, Canada; k Mount Sinai Hosp., Toronto, ON, Canada; ¶ Oulu Univ. Hosp., Oulu, Finland Purpose: A major challenge in the development of effective pharmacotherapy to treat osteoarthritis (OA) is the limited uptake and retention of drug into articular cartilage (AC). The development of methods to deliver drug locally into AC could permit to provide the target tissue with sufficient drug availability without increasing systemic exposure. Our previous research showed that MHz sonication enhances diffusion of kDa-sized molecules into AC. However, the clinical suitability of this approach could be limited by the slow delivery rate (hours). This study, therefore, investigated whether high intensity pulsed ultrasound (HIPU) and electric field (E-field) could offer a fast means to deliver a drug surrogate, i.e., Methylene blue (MB), into articular cartilage without inducing tissue damage. Methods: We conducted ten experiments in-vitro. For each experiment, one cylindrical plug (Ø ¼ 12.8 mm, 6 mm thick) of bovine AC was obtained from the femoral condyle; one condyle per animal (n ¼ 10, age < 3 years; weight ¼ 262 ± 46 kg; mean ± SD). The cylindrical plugs were split into four quarters and grouped as it follows: (i) Treatment group (T1; n ¼ 10), i.e., simultaneous immersion in MB and exposure to HIPU/E-field; (ii) Control 1 (C1; n ¼ 10), i.e.,first exposed to HIPU/E-field and then immersed in MB; (iii) Control 2 (C2; n ¼ 10), i.e., immersed in MB (not sonicated); and (iv) Control 3 (C3; n ¼ 10), i.e., neither immersed in MB nor sonicated. A solution of MB 0.005% w/v in 1X Phosphate Buffer Saline (PBS) served as contrast agent. The sonication treatment consisted of 1000 high-intensity ultrasound/E-field pulses (treatment time in minutes: 4.68 ± 0.40; pulse repetition frequency (PRF): 3.57 ± 0.03 Hz, mean ± SD). The required HIPU and E-field were generated by a custombuilt spark gap device comprising a spark gap, an air gap switch, a high voltage generator, and a capacitors bank. The breakdown voltage of the air gap switch (~4.5 kV) and the total capacitance (166.67 nF) yielded an average spark energy of ~1.7 J, as estimated from E ¼ 1⁄2 CV^2. The spark gap consisted of two electrodes detached from commercial spark plugs (Model: SGO005; McCulloch, Husqvarna, Sweden) with 600 mm electrode gap. Delivery of MB into AC was estimated from Napierian optical absorbance in transmission light microscopy of 150 mm-thick sample slices. To investigate the potential effect of the treatment on the structural properties of the AC matrix, six additional cylindrical plugs from different animals were split into 4 quarters and each quarter grouped into treatment groups (DTa and DTb) and control groups (DCa and DCb). Treatment samples DTa and DTb were simultaneously treated with HIPU/E-field while immersed in PBS. Control samples DCa and DCb were not sonicated. Then, groups DTa and DCa underwent histological imaging under field emission scanning electron microscopy (FESEM), and groups DTb and DCb underwent histological analysis and digital densitometry to assess proteoglycan content. Results: Based on optical absorbance, MB concentration was significantly higher (p < 0.01) in the treated group T1 as compared to controls C1 and
Figure 1. Light absorbance as a function of the depth into AC and one set of slices from the different groups. Substantial higher light absorbance (related to MB concentration) was evidenced in the treatment group as compared to controls. Conclusions: Our results show that HIPU/E-field offers a fast and effective means to deliver agents into AC in a non-destructive fashion as determined by FESEM and histology. As no difference in optical absorbance between groups C2 and C3, and no evidence of AC damage induced by the treatment were found, changes in AC permeability is an unlikely mechanism for the enhanced MB diffusion in the treatment group. However, further study of the short-term and long-term effects of the proposed treatment is required. The proposed method could enable development of local pharmacological OA therapies. 902 MICROLOCALIZED TRANSPORT OF AGENTS INTO ARTICULAR CARTILAGE USING HIGH INTENSITY ULTRASOUND €rvi y, z, A. Salmi y, A. Airaksinen y, H.J. Nieminen y, z, E. Lampsija €m y. y Univ. of Helsinki, S. Saarakkala x, K.P. Pritzker k, E. Hæggstro Helsinki, Finland; z Univ. of Oulu, Oulu, Finland; x Univ. of Oulu; Oulu Univ. Hosp., Oulu, Finland; k Univ. of Toronto; Mount Sinai Hosp., Toronto, ON, Canada Purpose: Drug delivery into articular cartilage is limited to systematic delivery (e.g., oral administration and intra-muscular injections) and localized administration (e.g., topical administration and intra-articular injections). With these standard approaches the spatial accuracy for localizing drugs is ~10 cm. While not yet clinically available, microlocalized delivery (50e100 enhancement, i.e., delivery into small volume on the order of ~mm) could permit drug therapies targeted to specific cartilage lesions reducing toxicity in uninvolved cartilage as well as systemic toxicity. This would be a paradigm shift for developing prospective drug therapies for treating osteoarthritis and cartilage injuries. Previously, we demonstrated the capability of MHz highintensity ultrasound (HIU) to deliver MW ~2.9 kDa agent half-way into bovine AC without histologically discernible damage in 2.5 hrs. The current study investigates whether focused HIU permits microlocalized delivery of agents into AC within a clinically relevant time frame (minutes) with agents (methylene blue, i.e., MB, MW ~320 Da) equivalent by MW to some current drugs (diacerein, MW ~368 Da; glucosamine, MW ~179 Da).
Abstracts / Osteoarthritis and Cartilage 24 (2016) S63eS534
C2, up to 900 mm depth into AC (Figure 1). Regarding tissue damage, no significant differences (p > 0.05) were found between control groups C1 and C2 at any depth. Furthermore, FESEM and histological analysis showed no evidence of structural changes induced by the treatment.
901 HIGH-INTENSITY PULSED ULTRASOUND AND ELECTRIC FIELD PERMIT LOCALIZED PHARMACOTHERAPY TARGETING ARTICULAR CARTILAGE rez y, H.J. Nieminen y, z, M. Finnila € z, A. Salmi y, A. García Pe €rvi y, T. Paulin y, K.P. Pritzker x, k, S. Saarakkala z, ¶, E. Lampsija € m y. y Univ. of Helsinki, Helsinki, Finland; z Univ. of Oulu, Oulu, E. Hæggstro Finland; x Univ. of Toronto, Toronto, ON, Canada; k Mount Sinai Hosp., Toronto, ON, Canada; ¶ Oulu Univ. Hosp., Oulu, Finland Purpose: A major challenge in the development of effective pharmacotherapy to treat osteoarthritis (OA) is the limited uptake and retention of drug into articular cartilage (AC). The development of methods to deliver drug locally into AC could permit to provide the target tissue with sufficient drug availability without increasing systemic exposure. Our previous research showed that MHz sonication enhances diffusion of kDa-sized molecules into AC. However, the clinical suitability of this approach could be limited by the slow delivery rate (hours). This study, therefore, investigated whether high intensity pulsed ultrasound (HIPU) and electric field (E-field) could offer a fast means to deliver a drug surrogate, i.e., Methylene blue (MB), into articular cartilage without inducing tissue damage. Methods: We conducted ten experiments in-vitro. For each experiment, one cylindrical plug (Ø ¼ 12.8 mm, 6 mm thick) of bovine AC was obtained from the femoral condyle; one condyle per animal (n ¼ 10, age < 3 years; weight ¼ 262 ± 46 kg; mean ± SD). The cylindrical plugs were split into four quarters and grouped as it follows: (i) Treatment group (T1; n ¼ 10), i.e., simultaneous immersion in MB and exposure to HIPU/E-field; (ii) Control 1 (C1; n ¼ 10), i.e.,first exposed to HIPU/E-field and then immersed in MB; (iii) Control 2 (C2; n ¼ 10), i.e., immersed in MB (not sonicated); and (iv) Control 3 (C3; n ¼ 10), i.e., neither immersed in MB nor sonicated. A solution of MB 0.005% w/v in 1X Phosphate Buffer Saline (PBS) served as contrast agent. The sonication treatment consisted of 1000 high-intensity ultrasound/E-field pulses (treatment time in minutes: 4.68 ± 0.40; pulse repetition frequency (PRF): 3.57 ± 0.03 Hz, mean ± SD). The required HIPU and E-field were generated by a custombuilt spark gap device comprising a spark gap, an air gap switch, a high voltage generator, and a capacitors bank. The breakdown voltage of the air gap switch (~4.5 kV) and the total capacitance (166.67 nF) yielded an average spark energy of ~1.7 J, as estimated from E ¼ 1⁄2 CV^2. The spark gap consisted of two electrodes detached from commercial spark plugs (Model: SGO005; McCulloch, Husqvarna, Sweden) with 600 mm electrode gap. Delivery of MB into AC was estimated from Napierian optical absorbance in transmission light microscopy of 150 mm-thick sample slices. To investigate the potential effect of the treatment on the structural properties of the AC matrix, six additional cylindrical plugs from different animals were split into 4 quarters and each quarter grouped into treatment groups (DTa and DTb) and control groups (DCa and DCb). Treatment samples DTa and DTb were simultaneously treated with HIPU/E-field while immersed in PBS. Control samples DCa and DCb were not sonicated. Then, groups DTa and DCa underwent histological imaging under field emission scanning electron microscopy (FESEM), and groups DTb and DCb underwent histological analysis and digital densitometry to assess proteoglycan content. Results: Based on optical absorbance, MB concentration was significantly higher (p < 0.01) in the treated group T1 as compared to controls C1 and
Figure 1. Light absorbance as a function of the depth into AC and one set of slices from the different groups. Substantial higher light absorbance (related to MB concentration) was evidenced in the treatment group as compared to controls. Conclusions: Our results show that HIPU/E-field offers a fast and effective means to deliver agents into AC in a non-destructive fashion as determined by FESEM and histology. As no difference in optical absorbance between groups C2 and C3, and no evidence of AC damage induced by the treatment were found, changes in AC permeability is an unlikely mechanism for the enhanced MB diffusion in the treatment group. However, further study of the short-term and long-term effects of the proposed treatment is required. The proposed method could enable development of local pharmacological OA therapies. 902 MICROLOCALIZED TRANSPORT OF AGENTS INTO ARTICULAR CARTILAGE USING HIGH INTENSITY ULTRASOUND €rvi y, z, A. Salmi y, A. Airaksinen y, H.J. Nieminen y, z, E. Lampsija €m y. y Univ. of Helsinki, S. Saarakkala x, K.P. Pritzker k, E. Hæggstro Helsinki, Finland; z Univ. of Oulu, Oulu, Finland; x Univ. of Oulu; Oulu Univ. Hosp., Oulu, Finland; k Univ. of Toronto; Mount Sinai Hosp., Toronto, ON, Canada Purpose: Drug delivery into articular cartilage is limited to systematic delivery (e.g., oral administration and intra-muscular injections) and localized administration (e.g., topical administration and intra-articular injections). With these standard approaches the spatial accuracy for localizing drugs is ~10 cm. While not yet clinically available, microlocalized delivery (50e100 enhancement, i.e., delivery into small volume on the order of ~mm) could permit drug therapies targeted to specific cartilage lesions reducing toxicity in uninvolved cartilage as well as systemic toxicity. This would be a paradigm shift for developing prospective drug therapies for treating osteoarthritis and cartilage injuries. Previously, we demonstrated the capability of MHz highintensity ultrasound (HIU) to deliver MW ~2.9 kDa agent half-way into bovine AC without histologically discernible damage in 2.5 hrs. The current study investigates whether focused HIU permits microlocalized delivery of agents into AC within a clinically relevant time frame (minutes) with agents (methylene blue, i.e., MB, MW ~320 Da) equivalent by MW to some current drugs (diacerein, MW ~368 Da; glucosamine, MW ~179 Da).