Danielle M. Minteer

Adipose-Derived Mesenchymal Stem Cells: Biology and Potential Applications

Adipose tissue is derived from the mesoderm during embryonic development and is present in every mammalian species, located throughout the body. Adipose tissue serves as an endocrine organ, functioning to maintain energy metabolism through the storage of lipids. While two types of adipose tissue exist (brown and white), white adipose yields the commonly studied adipose-derived stem cells (ASCs). Adipose-derived stem cells provide a promising future in the field of tissue engineering and regenerative medicine. Due to their wide availability and ability to differentiate into other tissue types of the mesoderm-including bone, cartilage, muscle, and adipose-ASCs may serve a wide variety of applications. Adipose stem cells have been utilized in studies addressing osteoarthritis, diabetes mellitus, heart disease, and soft tissue regeneration and reconstruction after mastectomy and facial repair. Various delivery systems and scaffolds to incorporate adipose stem cells have also been established. Adipose stem cells have been studied in vitro and in vivo. Much information in vitro has been obtained on adipose stem cell potency and biology as a function of donor gender, body mass index, and anatomical location. Further in vitro studies have examined the various cell populations within the heterogeneous population within the stromal vascular fraction (SVF) from which ASCs are obtained. While many animal models are used to investigate adipose tissue, preclinical in vivo experiments are most widely conducted in the mouse model. Common analyses of animal studies utilizing ASCs include pre-labeling cells and immunostaining cells.

Adipose Stem Cells: Biology, Safety, Regulation, and Regenerative Potential

This article discusses adipose-derived stem cell (ASC) biology, describes the current knowledge in the literature for the safety and regulation of ASCs, and provides a brief overview of the regenerative potential of ASCs. It is not an exhaustive listing of all available clinical studies or every study applying ASCs in tissue engineering and regenerative medicine, but is an objective commentary of these topics.

Initial observations on using magnesium metal in peripheral nerve repair

Biodegradable magnesium metal filaments placed inside biodegradable nerve conduits might provide the physical guidance support needed to improve the rate and extent of regeneration of peripheral nerves across injury gaps. In this study, we examined basic issues of magnesium metal resorption and biocompatibility by repairing sub-critical size gap injuries (6 mm) in one sciatic nerve of 24 adult male Lewis rats. Separated nerve stumps were connected with poly(caprolactone) nerve conduits, with and without magnesium filaments (0.25 mm diameter, 10 mm length), with two different conduit filler substances (saline and keratin hydrogel). At 6 weeks after implantation, magnesium degradation was examined by micro-computed tomography and histological analyses. Magnesium degradation was significantly greater when the conduits were filled with an acidic keratin hydrogel than with saline (p < 0.05). But magnesium filaments in some animals remained intact for 6 weeks. Using histological and immunocytochemical analyses, good biocompatibility of the magnesium implants was observed at 6 weeks, as shown by good development of regenerating nerve mini-fascicles and only mild inflammation in tissues even after complete degradation of the magnesium. Nerve regeneration was not interrupted by complete magnesium degradation. An initial functional evaluation, determination of size recovery of the gastrocnemius muscle, showed a slight improvement due to magnesium with the saline but not the keratin filler, compared with respective control conduits without magnesium. These results suggest that magnesium filament implants have the potential to improve repair of injured peripheral nerve defects in this rodent model.

Combining micro-computed tomography with histology to analyze biomedical implants for peripheral nerve repair ☆

BACKGROUND Biomedical implants used in tissue engineering repairs, such as scaffolds to repair peripheral nerves, can be too large to examine completely with histological analyses. Micro-computed tomography (micro-CT) with contrast agents allows ex vivo visualization of entire biomaterial implants and their interactions with tissues, but contrast agents can interfere with histological analyses of the tissues or cause shrinkage or loss of antigenicity. NEW METHOD Soft tissue, ex vivo micro-CT imaging using Lugols iodine was compatible with histology after using a rapid (48 h) method of removing iodine. RESULTS Adult normal and repaired rat sciatic nerves were infiltrated ex vivo with iodine, imaged with micro-CT and then the iodine was removed by incubating tissues in sodium thiosulfate. Subsequent paraffin sections of normal nerve tissues showed no differences in staining with hematoxylin and eosin or immunostaining with multiple antibodies. Iodine treatment and removal did not alter axonal diameter, nuclear size or relative area covered by immunostained axons (p>0.05). Combining imaging modalities allowed comparisons of macroscopic and microscopic features of nerve tissues regenerating through simple nerve conduits or nerve conduits containing a titanium wire for guidance. COMPARISON WITH EXISTING METHODS Quantification showed that treatment with iodine and sodium thiosulfate did not result in tissue shrinkage or loss of antigenicity. CONCLUSIONS Because this combination of treatments is rapid and does not alter tissue morphology, this expands the ex vivo methods available to examine the success of biomaterial implants used for tissue engineering repairs.

Bioreactors addressing diabetes mellitus.

The concept of bioreactors in biochemical engineering is a well-established process; however, the idea of applying bioreactor technology to biomedical and tissue engineering issues is relatively novel and has been rapidly accepted as a culture model. Tissue engineers have developed and adapted various types of bioreactors in which to culture many different cell types and therapies addressing several diseases, including diabetes mellitus types 1 and 2. With a rising world of bioreactor development and an ever increasing diagnosis rate of diabetes, this review aims to highlight bioreactor history and emerging bioreactor technologies used for diabetes-related cell culture and therapies.

Analysis of type II diabetes mellitus adipose-derived stem cells for tissue engineering applications.

To address the functionality of diabetic adipose-derived stem cells in tissue engineering applications, adipose-derived stem cells isolated from patients with and without type II diabetes mellitus were cultured in bioreactor culture systems. The adipose-derived stem cells were differentiated into adipocytes and maintained as functional adipocytes. The bioreactor system utilizes a hollow fiber–based technology for three-dimensional perfusion of tissues in vitro, creating a model in which long-term culture of adipocytes is feasible, and providing a potential tool useful for drug discovery. Daily metabolic activity of the adipose-derived stem cells was analyzed within the medium recirculating throughout the bioreactor system. At experiment termination, tissues were extracted from bioreactors for immunohistological analyses in addition to gene and protein expression. Type II diabetic adipose-derived stem cells did not exhibit significantly different glucose consumption compared to adipose-derived stem cells from patients without type II diabetes (p > 0.05, N = 3). Expression of mature adipocyte genes was not significantly different between diabetic/non-diabetic groups (p > 0.05, N = 3). Protein expression of adipose tissue grown within all bioreactors was verified by Western blotting.The results from this small-scale study reveal adipose-derived stem cells from patients with type II diabetes when removed from diabetic environments behave metabolically similar to the same cells of non-diabetic patients when cultured in a three-dimensional perfusion bioreactor, suggesting that glucose transport across the adipocyte cell membrane, the hindrance of which being characteristic of type II diabetes, is dependent on environment. The presented observation describes a tissue-engineered tool for long-term cell culture and, following future adjustments to the culture environment and increased sample sizes, potentially for anti-diabetic drug testing.

Short and long gap peripheral nerve repair with magnesium metal filaments: MAGNESIUM FILAMENTS IN PERIPHERAL NERVE REPAIR

A current clinical challenge is to replace autografts for repair of injury gaps in peripheral nerves, which can occur due to trauma or surgical interruption. Biodegradable metallic magnesium filaments, placed inside hollow nerve conduits, could support nerve repair by providing contact guidance support for axonal regeneration. This was tested by repairing sciatic nerves of adult rats with single magnesium filaments placed inside poly(caprolactone) nerve conduits. Controls were empty conduits, conduits containing titanium filaments and/or isografts from donor rats. With a nerve gap of 6 mm and 6 weeks post-repair, magnesium filaments had partially resorbed. Regenerating cells had attached to the filaments and axons were observed in distal stumps in all animals. Axon parameters were improved with magnesium compared to conduits alone or conduits with single titanium filaments. With a longer gap of 15 mm and 16 weeks post-repair, functional parameters were improved with isografts, but not with magnesium filaments or empty conduits. Magnesium filaments were completely resorbed and no evidence of scarring was seen. While axon outgrowth was not improved with the longer gap, histological measures of the tissues were improved with magnesium compared to empty conduits. Therefore, the use of magnesium filaments is promising because they are biocompatible and improve aspects of nerve regeneration.

The Influence of Fat Grafting on Skin Quality in Cosmetic Foot Grafting: A Randomized, Cross-Over Clinical Trial

BACKGROUND Pedal fat grafting is a cosmetic procedure to treat the functional and aesthetic sequelae of pedal fat pad atrophy. Fat grafting has been found to mitigate these symptoms, but the exact mechanism is unknown. OBJECTIVES The authors hypothesized that pedal fat grafting may improve skin quality, accounting for prolonged symptomatic improvement despite loss of grafted fat. METHODS Patients with pedal atrophy were enrolled in a randomized crossover clinical trial. Group 1 underwent fat grafting upon enrollment with 2-year follow-up. Group 2 was managed conservatively for 1 year then placed into the fat grafting group with 1-year follow-up. Patients underwent pedal ultrasounds to determine thicknesses of the fat pad and dermis, and photographs were taken to assess skin quality. RESULTS Three men and 20 women with an average age of 63 ± 6 years and an average BMI of 26.0 ± 4.6 kg/m2 were enrolled in the study. Twenty-six feet were injected in Group 1 and 17 were injected in Group 2. Group 1 dermal thickness increased at 6 months post-injection (P < 0.05). This increase persisted through 24 months. Group 2 dermal thickness decreased prior to injection (P < 0.05) but returned to baseline after injection and through 12-month follow-up (P < 0.05). Fat pad thickness returned to baseline by study completion in both groups (P < 0.05). CONCLUSIONS Pedal fat grafting yielded a significant, sustained increase in dermal thickness, though grafted fat was not retained. Fat grafting may improve skin quality, which could contribute to improved clinical outcomes despite loss of grafted fat. LEVEL OF EVIDENCE: 2

Abstract: Post-Operative Complications in the Massive Weight Loss Patient

INTRODUCTION: Patients with end-stage renal disease (ESRD) may require a panniculectomy in preparation for renal transplantation. ESRD is associated with increased cardiovascular risk factors, electrolyte imbalances, and chronic anemia. These factors may increase the risk of adverse outcomes in patients undergoing panniculectomy. Purpose of this study was to evaluate safety and perioperative complication rates in ESRD patients following panniculectomy.

Abstract P21: Magnesium and Biological Hydrogel Degradation within a Polycaprolactone (PCL) Conduit for Peripheral Nerve Repair

ConClusion: Up until now, five patients in our center received a tracheal allotransplantation with withdrawal of immunosuppression. To reconstruct a long segment tracheal stenosis, it is of utmost importance to provide a vascularized inner mucosal lining. We modified the time points in our protocol to reduce the adverse effects of immunologic rejection. Currently we are fine-tuning the enzymatic treatment of the inner lining and cultivation of mucosa in the rabbit model. P21 Magnesium and Biological hydrogel degradation within a Polycaprolactone (Pcl) conduit for Peripheral nerve repair