Biodegradable silk catheters for the delivery of therapeutics across anatomical repair sites.

Biodegradable silk catheters for the delivery of therapeutics are designed with a focus on creating porous gradients that can direct the release of molecules away from the implantation site. Though suitable for a range of applications, these catheters are designed for drug delivery to transplanted adipose tissue in patients having undergone a fat grafting procedure. A common complication for fat grafts is the rapid reabsorption of large volume adipose transplants. In order to prolong volume retention, biodegradable catheters can be embedded into transplanted tissue to deliver nutrients, growth factors or therapeutics to improve adipocyte viability, proliferation, and ultimately extend volume retention. Two fabrication methods are developed: a silk gel-spinning technique, which uses a novel flash-freezing step to induce high porosity throughout the bulk of the tube, and a dip-coating process using silk protein solutions doped with a water soluble porogen. Increased porosity aids in the diffusion of drug through the silk tube in a controllable way. Additionally, we interface the porous tubes with ALZET osmotic pumps for implantation into a subcutaneous nude mouse model. The work described herein will discuss the processing parameters as well as the interfacing between pump and cargo therapeutic and the resulting release profiles. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 2018. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 107B: 501-510, 2019.

Controlled dexamethasone delivery via double-walled microspheres to enhance long-term adipose tissue retention.

Current materials used for adipose tissue reconstruction have critical shortcomings such as suboptimal volume retention, donor-site morbidity, and poor biocompatibility. The aim of this study was to examine a controlled delivery system of dexamethasone to generate stable adipose tissue when mixed with disaggregated human fat in an athymic mouse model for 6 months. The hypothesis that the continued release of dexamethasone from polymeric microspheres would enhance both adipogenesis and angiogenesis more significantly when compared to the single-walled microsphere model, resulting in long-term adipose volume retention, was tested. Dexamethasone was encapsulated within single-walled poly(lactic-co-glycolic acid) microspheres (Dex SW MS) and compared to dexamethasone encapsulated in a poly(lactic-co-glycolic acid) core surrounded by a shell of poly-l-lactide. The double-walled polymer microsphere system in the second model was developed to create a more sustainable drug delivery process. Dexamethasone-loaded poly(lactic-co-glycolic acid) microspheres (Dex SW MS) and dexamethasone-loaded poly(lactic-co-glycolic acid)/poly-l-lactide double-walled microspheres (Dex DW MS) were prepared using single and double emulsion/solvent techniques. In vitro release kinetics were determined. Two doses of each type of microsphere were examined; 50 and 27 mg of Dex MS and Dex DW MS were mixed with 0.3 mL of human lipoaspirate. Additionally, 50 mg of empty MS and lipoaspirate-only controls were examined. Samples were analyzed grossly and histologically after 6 months in vivo. Mass and volume were measured; dexamethasone microsphere-containing samples demonstrated greater adipose tissue retention compared to the control group. Histological analysis, including hematoxylin and eosin and CD31 staining, indicated increased vascularization (p < 0.05) within the Dex MS-containing samples. Controlled delivery of adipogenic factors, such as dexamethasone via polymer microspheres, significantly affects adipose tissue retention by maintaining healthy tissue formation and vascularization. Dex DW MS provide an improved model to former Dex SW MS, resulting in notably longer release time and, consequently, larger volumes of adipose retained in vivo. The use of microspheres, specifically double-walled, as vehicles for controlled drug delivery of adipogenic factors therefore present a clinically relevant model of adipose retention that has the potential to greatly improve soft tissue repair.

Dexamethasone Release from Within Engineered Cartilage as a Chondroprotective Strategy Against Interleukin-1α.

While significant progress has been made toward engineering functional cartilage constructs with mechanical properties suitable for in vivo loading, the impact on these grafts of inflammatory cytokines, chemical factors that are elevated with trauma or osteoarthritis, is poorly understood. Previous work has shown dexamethasone to be a critical compound for cultivating cartilage with functional properties, while also providing chondroprotection from proinflammatory cytokines. This study tested the hypothesis that the incorporation of poly(lactic-co-glycolic acid) (PLGA) (75:25) microspheres that release dexamethasone from within chondrocyte-seeded agarose hydrogel constructs would promote development of constructs with functional properties and protect constructs from the deleterious effects of interleukin-1α (IL-1α). After 28 days of growth culture, experimental groups were treated with IL-1α (10 ng/mL) for 7 days. Reaching native equilibrium moduli and proteoglycan levels, dexamethasone-loaded microsphere constructs exhibited tissue properties similar to microsphere-free control constructs cultured in dexamethasone-supplemented culture media and were insensitive to IL-1α exposure. These findings are in stark contrast to constructs containing dexamethasone-free microspheres or no microspheres, cultured without dexamethasone, where IL-1α exposure led to significant tissue degradation. These results support the use of dexamethasone delivery from within engineered cartilage, through biodegradable microspheres, as a strategy to produce mechanically functional tissues that can also combat the deleterious effects of local proinflammatory cytokine exposure.

Adipogenic factor-loaded microspheres increase retention of transplanted adipose tissue.

The aim of this study was to develop and test a controlled delivery system of two adipogenic factors (insulin and dexamethasone [Dex]), to generate stable adipose tissue when mixed with disaggregated human fat. Both drugs were encapsulated in poly(lactic-co-glycolic acid), (PLGA) microspheres (MS) and mixed with human lipoaspirate to induce adipogenesis in vivo. It was hypothesized that the slow release of insulin and Dex would enhance both adipogenesis and angiogenesis, thus retaining the fat graft volume in a nude mouse model. Insulin/Dex-loaded PLGA MS (Insulin/Dex MS) were prepared using both single and double emulsion/solvent extraction techniques. The bioactivity of the drugs was assessed by mixing the MS with human lipoaspirate and injecting subcutaneously into the dorsal aspect of an athymic mouse. Five doses of the drugs were examined and samples were analyzed grossly and histologically after 5 weeks in vivo. Mass and volume of the grafts were measured with the microsphere-containing samples, demonstrating increased mass and volume with increasing drug doses. Histological analysis, including H&E and CD31, indicated increased vascularization within the insulin/Dex MS-containing samples compared with the lipoaspirate-only samples. This study demonstrates that the controlled delivery of adipogenic factors such as insulin and Dex through polymer MS can significantly enhance tissue formation and vascularization, therefore presenting a potentially clinically relevant model of adipose retention.

Prevalence of endogenous CD34+ adipose stem cells predicts human fat graft retention in a xenograft model.

BACKGROUND: Fat grafting is a promising technique for soft-tissue augmentation, although graft retention is highly unpredictable and factors that affect graft survival have not been well defined. Because of their capacity for differentiation and growth factor release, adipose-derived stem cells may have a key role in graft healing. The authors' objective was to determine whether biological properties of adipose-derived stem cells present within human fat would correlate with in vivo outcomes of graft volume retention. METHODS: Lipoaspirate from eight human subjects was processed using a standardized centrifugation technique and then injected subcutaneously into the flanks of 6-week-old athymic nude mice. Graft masses and volumes were measured, and histologic evaluation, including CD31+ staining for vessels, was performed 8 weeks after transplantation. Stromal vascular fraction isolated at the time of harvest from each subject was analyzed for surface markers by multiparameter flow cytometry, and also assessed for proliferation, differentiation capacity, and normoxic/hypoxic vascular endothelial growth factor secretion. RESULTS: Wide variation in percentage of CD34+ progenitors within the stromal vascular fraction was noted among subjects and averaged 21.3 ± 15 percent (mean ± SD). Proliferation rates and adipogenic potential among stromal vascular fraction cells demonstrated moderate interpatient variability. In mouse xenograft studies, retention volumes ranged from approximately 36 to 68 percent after 8 weeks, with an overall average of 52 ± 11 percent. A strong correlation (r = 0.78, slope = 0.76, p < 0.05) existed between stromal vascular fraction percentage of CD34+ progenitors and high graft retention. CONCLUSION: Inherent biological differences in adipose tissue exist between patients. In particular, concentration of CD34+ progenitor cells within the stromal vascular fraction may be one of the factors used to predict human fat graft retention.

VEGF microsphere technology to enhance vascularization in fat grafting.

Vascularization is crucial for implantation of engineered tissues in reconstructive surgery. Polypeptides encapsulated in microspheres can be efficiently transported to their site of action and released in a sustained dosage. We evaluated the effect of delivering vascular endothelial growth factor (VEGF)-encapsulated microspheres in a lipoaspirate scaffold on vascularization and tissue survival. The VEGF-loaded (n=6) and empty (n=6) poly(lactic-co-glycolic acid) microspheres in human lipoaspirate and the human lipoaspirate alone (n=6) were injected subcutaneously into the flanks of athymic nude mice. Three mice from each group were killed, and grafts were explanted at weeks 3 and 6. Increases in mass and volume of VEGF samples, as well as decreases in empty and lipoaspirate-only samples, were observed at 3 and 6 weeks, reaching statistical significance at 6 weeks. Hematoxylin and eosin and CD31+ imaging demonstrated significantly greater vascularization in VEGF samples than in both the empty and lipoaspirate-only groups at both 3 and 6 weeks.

Adipose-derived stem cells for wound healing applications.

Nonhealing wounds remain a significant challenge for plastic surgeons. More than 600,000 people suffer from venous ulcers and 1.5 to 3 million people are being treated for pressure sores every year in the United States. The use of tissue engineering techniques such as stem-cell therapy and gene therapy to improve wound healing is a promising strategy. Adipose tissue represents a source of cells that may be able to enhance wound healing. Adipose-derived stem cells (ASCs) are adult stem cells that are easily harvested and of great interest for plastic surgeons. Specifically, ASCs secrete angiogenic growth factors that can induce tissue regeneration. This review describes innovative research strategies using ASCs therapies for treatment of chronic, nonhealing wounds.