Quality Control in Human Adipose-Derived Stromal/Stem Cells and Tissue Engineering Fat Models for Aging Studies.

Adipose tissue is recognized not only as an endocrine organ but also as a reservoir for adipose-derived stromal/stem cells (ASCs). ASCs have stimulated the interest of both the scientific and medical communities due to their therapeutic potential and applications in tissue engineering and regenerative medicine. ASCs are leveraged for their multipotency and their paracrine function.

Functional Assays in 3D ObaCell® Fat-on-a-Chip Cultures: Lipolysis and Glucose Uptake Assays.

Advancements in three-dimensional in vitro cultures pose a need for modification of established two-dimensional culture functional assay methods. Application of three-dimensional in vitro models in drug screening and target validation, specifically in the development of compounds targeting adipose metabolic activity, requires optimization of current glucose uptake and lipolysis assay protocols to effectively measure adipocyte function in a three-dimensional platform. This chapter describes the establishment of three-dimensional cultures using Obatala Sciences' human-derived hydrogel, maintenance and treatment of the cultures, and evaluation of compound response via lipolysis and glucose uptake assays.

Quality Control Assessment of Human Adipose-Derived Hydrogels.

Decellularized human-adipose tissue (hDAT) can serve as an alternative to two-dimensional monolayer culture and current ECM hydrogels due to its unlimited availability and cytocompatibility. A major hurdle in the clinical translation and integration of hDAT and other hydrogels into current in vitro culture processes is adherence to current good manufacturing practices (cGMP). Transferring of innovative technologies, including hydrogels, requires the establishing standardized protocols for quality assurance and quality control (QA/QC) of the material.

Manufacturing of a Human Adipose-Derived Hydrogel.

Hydrogels are considered a viable in vitro alternative to monolayer cultures. They provide quintessential characteristics for in vitro studies including biocompatibility, biodegradability, viscoelasticity, hydrophilicity, and low toxicity. Furthermore, many provide necessary extracellular matrix proteins and architecture to support cell growth, proliferation, differentiation, and migration.

Adipose Tissue in Breast Cancer Microphysiological Models to Capture Human Diversity in Preclinical Models.

Female breast cancer accounts for 15.2% of all new cancer cases in the United States, with a continuing increase in incidence despite efforts to discover new targeted therapies. With an approximate failure rate of 85% for therapies in the early phases of clinical trials, there is a need for more translatable, new preclinical in vitro models that include cellular heterogeneity, extracellular matrix, and human-derived biomaterials.

3D bio scaffold support osteogenic differentiation of mesenchymal stem cells.

The regeneration of osteochondral lesions by tissue engineering techniques is challenging due to the lack of physicochemical characteristics and dual-lineage (osteogenesis and chondrogenesis). A scaffold with better mechanical properties and dual lineage capability is required for the regeneration of osteochondral defects. In this study, a hydrogel prepared from decellularized human umbilical cord tissue was developed and evaluated for osteochondral regeneration.

Biomechanical and Biological Characterization of XGel, a Human-Derived Hydrogel for Stem Cell Expansion and Tissue Engineering.

Hydrogels are 3D scaffolds used as alternatives to in vivo models for disease modeling and delivery of cells and drugs. Existing hydrogel classifications include synthetic, recombinant, chemically defined, plant- or animal-based, and tissue-derived matrices. There is a need for materials that can support both human tissue modeling and clinically relevant applications requiring stiffness tunability.

Non-homologous use of adipose-derived cell and tissue therapies: Osteoarthritis as a case study.

Adipose tissue is widely recognized as an abundant and accessible human tissue that serves as a source of cells and extracellular matrix scaffolds for regenerative surgical applications. Increasingly, orthopedic surgeons are turning to adipose tissue as a resource in their treatment of osteoarthritis and related conditions. In the U.

The Emerging Use of ASC/Scaffold Composites for the Regeneration of Osteochondral Defects.

Articular cartilage is composed of chondrocytes surrounded by a porous permeable extracellular matrix. It has a limited spontaneous healing capability post-injury which, if left untreated, can result in severe osteochondral disease. Currently, osteochondral (OC) defects are treated by bone marrow stimulation, artificial joint replacement, or transplantation of bone, cartilage, and periosteum, while autologous osteochondral transplantation is also an option; it carries the risk of donor site damage and is limited only to the treatment of small defects.

Adipose-derived cells: building blocks of three-dimensional microphysiological systems.

Microphysiological systems (MPS) created with human-derived cells and biomaterial scaffolds offer a potential in vitro alternative to in vivo animal models. The adoption of three-dimensional MPS models has economic, ethical, regulatory, and scientific implications for the fields of regenerative medicine, metabolism/obesity, oncology, and pharmaceutical drug discovery. Key opinion leaders acknowledge that MPS tools are uniquely positioned to aid in the objective to reduce, refine, and eventually replace animal experimentation while improving the accuracy of the finding's clinical translation.

Decellularized Human Umbilical Tissue-Derived Hydrogels Promote Proliferation and Chondrogenic Differentiation of Mesenchymal Stem Cells.

Tissue engineering is a promising approach for the repair and regeneration of cartilaginous tissue. Appropriate three-dimensional scaffolding materials that mimic cartilage are ideal for the repair of chondral defects. The emerging decellularized tissue-based scaffolds have the potential to provide essential biochemical signals and structural integrity, which mimics the natural tissue environment and directs cellular fate.

Human Adipose-Derived Stromal/Stem Cell Culture and Analysis Methods for Adipose Tissue Modeling In Vitro: A Systematic Review.

Human adipose-derived stromal/stem cells (hASC) are widely used for in vitro modeling of physiologically relevant human adipose tissue. These models are useful for the development of tissue constructs for soft tissue regeneration and 3-dimensional (3D) microphysiological systems (MPS) for drug discovery. In this systematic review, we report on the current state of hASC culture and assessment methods for adipose tissue engineering using 3D MPS.

Histology Scoring System for Murine Cutaneous Wounds.

Monitoring wound progression over time is a critical aspect for studies focused on in-depth molecular analysis or on evaluating the efficacy of potential novel therapies. Histopathological analysis of wound biopsies can provide significant insight into healing dynamics, yet there is no standardized and reproducible scoring system currently available. The purpose of this study was to develop and statistically validate a scoring system based on parameters in each phase of healing that can be easily and accurately assessed using either Hematoxylin & Eosin (H&E) or Masson's Trichrome (MT) staining.

Breast Cancer Reconstruction: Design Criteria for a Humanized Microphysiological System.

International regulatory agencies such as the Food and Drug Administration have mandated that the scientific community develop humanized microphysiological systems (MPS) as an alternative to animal models in the near future. While the breast cancer research community has long appreciated the importance of three-dimensional growth dynamics in their experimental models, there are remaining obstacles preventing a full conversion to humanized MPS for drug discovery and pathophysiological studies. This perspective evaluates the current status of human tissue-derived cells and scaffolds as building blocks for an "idealized" breast cancer MPS based on bioengineering design principles.

The effect of hypoxia on the proteomic signature of pig adipose-derived stromal/stem cells (pASCs).

Human adipose-derived stem cells (ASCs) have potential to improve wound healing; however, their equivalents from domestic animals have received less attention as an alternative cell-based therapy for animals or even humans. Hypoxia is essential for maintaining stem cell functionality in tissue-specific niches. However, a cellular response to low oxygen levels has not been demonstrated in pig ASCs.

Clinical Translational Potential in Skin Wound Regeneration for Adipose-Derived, Blood-Derived, and Cellulose Materials: Cells, Exosomes, and Hydrogels.

Acute and chronic skin wounds due to burns, pressure injuries, and trauma represent a substantial challenge to healthcare delivery with particular impacts on geriatric, paraplegic, and quadriplegic demographics worldwide. Nevertheless, the current standard of care relies extensively on preventive measures to mitigate pressure injury, surgical debridement, skin flap procedures, and negative pressure wound vacuum measures. This article highlights the potential of adipose-, blood-, and cellulose-derived products (cells, decellularized matrices and scaffolds, and exosome and secretome factors) as a means to address this unmet medical need.

Human Adipose Derived Cells in Two- and Three-Dimensional Cultures: Functional Validation of an In Vitro Fat Construct.

Obesity, defined as a body mass index of 30 kg/m or above, has increased considerably in incidence and frequency within the United States and globally. Associated comorbidities including cardiovascular disease, type 2 diabetes mellitus, metabolic syndrome, and nonalcoholic fatty liver disease have led to a focus on the mechanisms promoting the prevention and treatment of obesity. Commonly utilized models employ human or mouse preadipocyte cell lines in a 2-dimensional (2D) format.

Fat-On-A-Chip Models for Research and Discovery in Obesity and Its Metabolic Comorbidities.

The obesity epidemic and its associated comorbidities present a looming challenge to health care delivery throughout the world. Obesity is characterized as a sterile inflammatory process within adipose tissues leading to dysregulated secretion of bioactive adipokines such as adiponectin and leptin, as well as systemic metabolic dysfunction. The majority of current obesity research has focused primarily on preclinical animal models and two-dimensional cell culture models .

Safety of Human Adipose Stromal Vascular Fraction Cells Isolated with a Closed System Device in an Immunocompetent Murine Pressure Ulcer Model.

Pressure ulcers (PUs) result in part due to ischemia-reperfusion injury to the skin and present frequently in elderly or quadriplegic patients with reduced mobility. Despite the high economic and societal cost of this condition, PU therapy relies primarily on preventive strategies and invasive surgical intervention. A growing body of clinical literature suggests that localized injection of adipose-derived cells can accelerate and enhance the closure of PUs.

Safety and Efficacy of Human Adipose-Derived Stromal/Stem Cell Therapy in an Immunocompetent Murine Pressure Ulcer Model.

Pressure injuries/ulcers are frequent complications in elderly, paraplegic, and quadriplegic patients, which account for considerable cost to the international health care economy and remain refractory to current treatment options. Autologous or allogeneic adult stromal/stem cells represent an alternative therapeutic approach. The current study extends prior findings by exploring the safety and efficacy of human adipose-derived stromal/stem cell (ASC) therapy in an established immunocompetent murine skin pressure ulcer model where dermal fibroblast cells (DFCs) served as a control.

Arguments for a Different Regulatory Categorization and Framework for Stromal Vascular Fraction.

Although adipose tissue and cells show considerable promise for clinical translation in the emerging field of regenerative medicine, they present a challenge to the regulatory community both nationally and internationally. This commentary evaluates the status of adipose-derived therapeutics and considers regulatory approaches designed to maximize patient safety while advancing clinical translation in accordance with evidence-based medical science.

Human Platelet Lysate as a Functional Substitute for Fetal Bovine Serum in the Culture of Human Adipose Derived Stromal/Stem Cells.

Introduction: Adipose derived stromal/stem cells (ASCs) hold potential as cell therapeutics for a wide range of disease states; however, many expansion protocols rely on the use of fetal bovine serum (FBS) as a cell culture nutrient supplement. The current study explores the substitution of lysates from expired human platelets (HPLs) as an FBS substitute.

Methods: Expired human platelets from an authorized blood center were lysed by freeze/thawing and used to examine human ASCs with respect to proliferation using hematocytometer cell counts, colony forming unit-fibroblast (CFU-F) frequency, surface immunophenotype by flow cytometry, and tri-lineage (adipocyte, chondrocyte, osteoblast) differentiation potential by histochemical staining.

A Novel, Sterilized Microvascular Tissue Product Improves Healing in a Murine Pressure Ulcer Model.

Background: Processed microvascular tissue (PMVT), a human structural allograft, is derived from lyophilized human tissue containing microcirculatory cellular components. Since PMVT serves as a source of extracellular matrix (ECM), growth factors, cytokines, and chemokines modulating angiogenesis, inflammation, apoptosis, and endogenous cell recruitment, we hypothesized its application would accelerate wound regeneration in a validated pressure ulcer (PU) model developed in C57BL/6 mice using two 24-hour cycles of skin ischemia/reperfusion created by placement and removal of external magnets.

Methods: Two identical PU injuries (n = 50 female mice) were treated with (a) topical particulate PMVT, (b) injected rehydrated PMVT, or (c) saline control injection, and assessed daily for closure rates, scab formation/removal, and temperature.

Gender and age-related cell compositional differences in C57BL/6 murine adipose tissue stromal vascular fraction.

Adipose tissue is now recognized as a functional organ that contains cellular heterogeneity and diversity within anatomical depots. The stromal vascular fraction (SVF) of adipose contains endothelial progenitors, fibroblasts, lymphocytes, monocyte/macrophages, pericytes, pre-adipocytes, and stromal/stem cells, among others. In recent years, there has been a growing appreciation of the influence of age and gender in the field of stem cell biology.