A device for long-term perfusion, imaging, and electrical interfacing of brain tissue in vitro

Distributed microelectrode array (MEA) recordings from consistent, viable, ≥ 500 µm thick tissue preparations over time periods from days to weeks may aid in studying a wide range of problems in neurobiology that require in vivo-like organotypic morphology. Existing tools for electrically interfacing with organotypic slices do not address necrosis that inevitably occurs within thick slices with limited diffusion of nutrients and gas, and limited removal of waste. We developed an integrated device that enables long-term maintenance of thick, functionally active, brain tissue models using interstitial perfusion and distributed recordings from thick sections of explanted tissue on a perforated multi-electrode array. This novel device allows for automated culturing, in situ imaging, and extracellular multi-electrode interfacing with brain slices, 3 D cell cultures, and potentially other tissue culture models. The device is economical, easy to assemble, and integrable with standard electrophysiology tools. We found that convective perfusion through the culture thickness provided a functional benefit to the preparations as firing rates were generally higher in perfused cultures compared to their respective unperfused controls. This work is a step towards the development of integrated tools for days-long experiments with more consistent, healthier, thicker, and functionally more active tissue cultures with built-in distributed electrophysiological recording and stimulation functionality. The results may be useful for the study of normal processes, pathological conditions, and drug screening strategies currently hindered by the limitations of acute (a few hours long) brain slice preparations

In Vitro Kinetic Degradation and In Vivo Biocompatibility Evaluation of Polycaprolactone-Based Growth Factor Delivery Matrices in the Rotator Cuff

The recent years have seen a significant surge in the use of synthetic biodegradable polymers for growth factor delivery in the rotator cuff. While these polymers have been successfully applied in delivery of factors in other tissues, the anatomical complexity, hypovascularity, cellularity, and reduced clearance of degradation by-products in the rotator cuff, creates unique requirements in tailoring the physical dimensions, chemical constituents, drug release and degradation characteristics of biomaterials for implantation. In this study, we investigate poly-lactic acid co-epsilon-caprolactone (30:70 LA:CL ratio) at 35-45kDa range and varying polymeric films casting concentrations (5-20%) as potential growth factor delivery matrices in the rotator cuff. Matrices were fabricated of 300µm thickness and 3x3mm surface area to facilitate model protein encapsulation and controlled release, and smooth translation of the matrix under the bony acromion after implantation in the rotator cuff. The matrix with the highest casting concentration (20wt%) showed unique, highly regular, and controlled release of the protein payload compared to the lower- casting concentrations (15 and 10wt%) and - molecular weights (35kDa) matrices. All films were found to lose molecular weight rapidly during the first 4 weeks due to the preferential hydrolysis of lactide-rich regions within the polymer, and then maintain a relatively stable molecular weight between week 4 and 8 due to the emergence of highly-crystalline caprolactone-rich regions. Nevertheless, the cleaved lactide-chains were not small enough to exit through the polymeric matrix as was evident from the maintenance of bulk matrix weight, form, and polymer dispersity index. This resulted in recrystallization of the cleaved chains in the presence of water molecules increasing the crystallinity of the matrix as was evident from the H-NMR and thermal analysis. Kinetic analysis revealed an inverse-linear relationship between polymer casting concentration and polymer break down. The ‘context-dependent’ biocompatibility evaluation was carried in a clinically-relevant rat model of acute rotator cuff repair model to address the unique features of both the tissue and the biomaterial being investigated. The matrices were found to remodel locally without undergoing catastrophic breakdown or causing excessive inflammatory reaction at the tissue site during the study period and is anticipated to completely degrade within 6 months of implantation. Our study is significant as it provides a systematic assessment of polymer properties that can be modifies to engineer morphogen release, degradation rates and mechanisms for biologic delivery in the rotator cuff. It also provides a pilot assessment of in situ biocompatibility of the polymeric matrix in the complex rotator cuff tissue

Pegylated insulin-like growth factor-1 biotherapeutic delivery promotes rotator cuff regeneration in a rat model.

Tears in the rotator cuff are challenging to repair because of the complex, hypocellular, hypovascular, and movement-active nature of the tendon and its enthesis. Insulin-like Growth Factor-1 (IGF-1) is a promising therapeutic for this repair. However, its unstable nature, short half-life, and ability to disrupt homeostasis has limited its clinical translation. Pegylation has been shown to improve the stability and sustain IGF-1 levels in the systemic circulation without disrupting homeostasis. To provide localized delivery of IGF-1 in the repaired tendons, we encapsulated pegylated IGF-1 mimic and its controls (unpegylated IGF-1 mimic and recombinant human IGF-1) in polycaprolactone-based matrices and evaluated them in a pre-clinical rodent model of rotator cuff repair. Pegylated-IGF-1 mimic delivery reestablished the characteristic tendon-to-bone enthesis structure and improved tendon tensile properties within 8 weeks of repair compared to controls, signifying the importance of pegylation in this complex tissue regeneration. These results demonstrate a simple and scalable biologic delivery technology alternative to tissue-derived grafts for soft tissue repair