Cryopreservation of lipoaspirates: in vitro measurement of the viability of adipose-derived stem cell and lipid peroxidation.
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ABSTRACT:
SUBMITTER: Kim DY
PROVIDER: S-EPMC7949318 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:BackgroundFabrication of three-dimensional stem cell spheroids have been studied to improve stem cell function, but the hypoxic core and limited penetration of nutrients and signaling cues to the interior of the spheroid were challenges. The incorporation of polymers such as silica and gelatin in spheroids resulted in relatively relaxed assembly of composite spheroids, and enhancing transport of nutrient and biological gas. However, because of the low surface area between cells and since the polymers were heterogeneously distributed throughout the spheroid, these polymers cannot increase the cell to extracellular matrix interactions needed to support differentiation.MethodsWe developed the stem cell spheroids that incorporate poly(ι-lactic acid) single-segmented fibers synthesized by electrospinning and physical and chemical fragmentation. The proper mixing ratio was 2000 cells/μg fibers (average length of the fibers was 50 μm - 100 μm). The SFs were coated with polydopamine to increase cell binding affinity and to synthesize various-sized spheroids. The function of spheroids was investigated by in vitro analysis depending on their sizes. For statistical analysis, Graphpad Prism 5 software (San Diego, CA, USA) was used to perform one-way analysis of variance ANOVA with Tukey's honest significant difference test and a Student's t-test (for two variables) (P < 0.05).ResultsSpheroids of different sizes were created by modulating the amount of cells and fibers (0.063 mm2-0.322 mm2). The fibers in the spheroid were homogenously distributed and increased cell viability, while cell-only spheroids showed a loss of DNA contents, internal degradation, and many apoptotic signals. Furthermore, we investigated stemness and various functions of various-sized fiber-incorporated spheroids. In conclusion, the spheroid with the largest size showed the greatest release of angiogenic factors (released VEGF: 0.111 ± 0.004 pg/ng DNA), while the smallest size showed greater effects of osteogenic differentiation (mineralized calcium: 18.099 ± 0.271 ng/ng DNA).ConclusionThe spheroids incorporating polydopamine coated single-segmented fibers showed enhanced viability regardless of sizes and increased their functionality by regulating the size of spheroids which may be used for various tissue reconstruction and therapeutic applications.
| S-EPMC8074457 | biostudies-literature
Project description:Disruption of redox homeostasis is a key phenotype of many pathological conditions. Though multiple oxidizing compounds such as hydrogen peroxide are widely recognized as mediators and inducers of oxidative stress, increasingly, attention is focused on the role of lipid hydroperoxides as critical mediators of death and disease. As the main component of cellular membranes, lipids have an indispensible role in maintaining the structural integrity of cells. Excessive oxidation of lipids alters the physical properties of cellular membranes and can cause covalent modification of proteins and nucleic acids. This review discusses the synthesis, toxicity, degradation, and detection of lipid peroxides in biological systems. Additionally, the role of lipid peroxidation is highlighted in cell death and disease, and strategies to control the accumulation of lipid peroxides are discussed.
| S-EPMC5319403 | biostudies-literature
Project description:Adipose-derived stem cells (ADSC) are an abundant population of adult stem cells with the potential to differentiate into several specialized tissue types, including neural and neural crest-derived cells. This study sought to determine if ADSC express keratocyte-specific phenotypic markers when cultured under conditions inducing differentiation of corneal stromal stem cells to keratocytes.Human subcutaneous adipose tissue was obtained by lipoaspiration. ADSC were isolated by collagenase digestion and differential centrifugation. Side population cells in ADSC were demonstrated using fluorescence-activated cell sorting after staining with Hoechst 33342. Differentiation to keratocyte phenotype was induced in fibrin gels or as pellet cultures with serum-free or reduced-serum media containing ascorbate. Keratocyte-specific gene expression was characterized using western blotting, quantitative RT-PCR, and immunostaining.ADSC contained a side population and exhibited differentiation to adipocytes and chondrocytes indicating adult stem-cell potential. Culture of ADSC in fibrin gels or as pellets in reduced-serum medium with ascorbate and insulin induced expression of keratocan, keratan sulfate, and aldehyde dehydrogenase 3 family, member A1 (ALDH3A1), products highly expressed by differentiated keratocytes. Expression of differentiation markers was quantitatively similar to corneal stromal stem cells and occurred in both serum-free and serum containing media.ADSC cultured under keratocyte-differentiation conditions express corneal-specific matrix components. Expression of these unique keratocyte products suggests that ADSC can adopt a keratocyte phenotype and therefore have potential for use in corneal cell therapy and tissue engineering.
| S-EPMC3002955 | biostudies-literature
Project description:Osteoarthritis is characterized by loss of articular cartilage also due to reduced chondrogenic activity of mesenchymal stem cells (MSCs) from patients. Adipose tissue is an attractive source of MSCs (ATD-MSCs), representing an effective tool for reparative medicine, particularly for treatment of osteoarthritis, due to their chondrogenic and osteogenic differentiation capability. The treatment of symptomatic knee arthritis with ATD-MSCs proved effective with a single infusion, but multiple infusions could be also more efficacious. Here we studied some crucial aspects of adipose tissue banking procedures, evaluating ATD-MSCs viability, and differentiation capability after cryopreservation, to guarantee the quality of the tissue for multiple infusions. We reported that the presence of local anesthetic during lipoaspiration negatively affects cell viability of cryopreserved adipose tissue and cell growth of ATD-MSCs in culture. We observed that DMSO guarantees a faster growth of ATD-MSCs in culture than trehalose. At last, ATD-MSCs derived from fresh and cryopreserved samples at -80°C and -196°C showed viability and differentiation ability comparable to fresh samples. These data indicate that cryopreservation of adipose tissue at -80°C and -196°C is equivalent and preserves the content of ATD-MSCs in Stromal Vascular Fraction (SVF), guaranteeing the differentiation ability of ATD-MSCs.
| S-EPMC5153503 | biostudies-literature
Project description:Extensive research has shown that increased production of reactive oxygen species (ROS) results in tissue injury under a variety of pathological conditions and chronic degenerative diseases. While ROS are highly reactive and can incite significant injury, polyunsaturated lipids in membranes and lipoproteins are their main targets. ROS-triggered lipid-peroxidation reactions generate a range of reactive carbonyl species (RCS), and these RCS spread and amplify ROS-related injury. Several RCS generated in oxidizing lipids, such as 4-hydroxy trans-2-nonenal (HNE), 4-oxo-2-(E)-nonenal (ONE), acrolein, malondialdehyde (MDA) and phospholipid aldehydes have been shown to be produced under conditions of oxidative stress and contribute to tissue injury and dysfunction by depleting glutathione and other reductants leading to the modification of proteins, lipids, and DNA. To prevent tissue injury, these RCS are metabolized by several oxidoreductases, including members of the aldo-keto reductase (AKR) superfamily, aldehyde dehydrogenases (ALDHs), and alcohol dehydrogenases (ADHs). Metabolism via these enzymes results in RCS inactivation and detoxification, although under some conditions, it can also lead to the generation of signaling molecules that trigger adaptive responses. Metabolic transformation and detoxification of RCS by oxidoreductases prevent indiscriminate ROS toxicity, while at the same time, preserving ROS signaling. A better understanding of RCS metabolism by oxidoreductases could lead to the development of novel therapeutic interventions to decrease oxidative injury in several disease states and to enhance resistance to ROS-induced toxicity.
| S-EPMC4414726 | biostudies-literature
Project description:Unlabelled: Adipose-derived mesenchymal stem cells (AD-MSCs) have been shown to ameliorate hyperglycemia in diabetic animals and individuals. However, little is known about whether AD-MSCs affect lipid metabolism. Here we have demonstrated for the first time that AD-MSC infusion can significantly suppress the increase in body weight and remarkably improve dyslipidemia in db/db obese mice and diet-induced obesity mice. Induction of white fat tissue "browning" and activation of adenosine monophosphate-activated protein kinase and its downstream hormone-sensitive lipase in adipose tissue contribute to the antiobesity and lipid-lowering effects. Thus, AD-MSC infusion holds great therapeutic potential for dyslipidemia and associated cardiovascular diseases.SignificanceMesenchymal stem cells (MSCs) are considered one of the most promising types of stem cells for translational application because of their rich tissue sources, multilineage differentiation capacity, and easy amplification in vitro and unique immunobiological properties. This study demonstrated that adipose-derived MSCs (AD-MSCs) infusion can significantly suppress the increase in body weight and remarkably improve dyslipidemia in obese mice. Induction of white fat tissue "browning" and activation of adenosine monophosphate-activated protein kinase and its downstream hormone-sensitive lipase in adipose tissue were demonstrated to contribute to the antiobesity and lipid-lowering effects. Thus, AD-MSC infusion holds great therapeutic potential for dyslipidemia.
| S-EPMC4996437 | biostudies-literature
Project description:Tooth morphogenesis requires sequential and reciprocal interactions between the cranial neural crest-derived mesenchymal cells and the stomadial epithelium, which regulate tooth morphogenesis and differentiation. We show how mesenchyme-derived single stem cell populations can be induced to transdifferentiate in vitro in a structure similar to a dental bud. The presence of stem cells in the adipose tissue has been previously reported. We incubated primary cultures of human adipose tissue-derived stem cells in a dental-inducing medium and cultured the aggregates in three-dimensional conditions. Four weeks later, cells formed a three-dimensional organized structure similar to a dental bud. Expression of dental tissue-related markers was tested assaying lineage-specific mRNA and proteins by RT-PCR, immunoblot, IHC, and physical-chemical analysis. In the induction medium, cells were positive for ameloblastic and odontoblastic markers as both mRNAs and proteins. Also, cells expressed epithelial, mesenchymal, and basement membrane markers with a positional relationship similar to the physiologic dental morphogenesis. Physical-chemical analysis revealed 200-nm and 50-nm oriented hydroxyapatite crystals as displayed in vivo by enamel and dentin, respectively. In conclusion, we show that adipose tissue-derived stem cells in vitro can transdifferentiate to produce a specific three-dimensional organization and phenotype resembling a dental bud even in the absence of structural matrix or scaffold to guide the developmental process.
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Project description:Cryopreservation procedures negatively affect the quality traits of sperm, causing certain changes at structural and molecular levels due to thermal, mechanical, osmotic, and oxidative damage. The objective of this study was to examine the potential of canine adipose-derived mesenchymal stem cells (Ad-MSCs) for providing protection to the dog sperm against cryo-damage. Canine Ad-MSCs were selected on the basis of the significantly higher gene expression for different proteins actively involved in the cell repair including annexin 1 (ANX1), histone H3 (H3) and high mobility group B (HMGB) protein compared to skin fibroblasts. Semen was collected from four healthy dogs by digital manipulation. The washed pooled ejaculates were diluted with buffer 2 (extender) supplemented without Ad-MSCs (Control), with 2.5 × 106 Ad-MSCs/mL (Group 1) or with 5 × 106 Ad-MSCs/mL (Group 2). Group 1 exhibited significantly higher post-thaw motility, live sperm, intact plasma membrane and normal acrosomes than the other groups. Additionally, Group 1 showed significantly higher expression levels of genes related to the repair of membranes (ANX1, dysferlin; DYSF, and fibronectin; FN1) and chromatin material (H3 and HMGB). Protein expression of ANX1, H 3, and FN1 was also statistically more in Group 1 than in Control. The results confirm that canine Ad-MSCs can effectively preserve the quality of frozen-thawed sperm by a reduction in cryoinjury. At an appropriate concentration, Ad-MSCs significantly improve the quality of post-thaw dog sperm.
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Project description:A hallmark of ageing is the redistribution of body fat. Particularly, subcutaneous fat decreases paralleled by a decrease of skin collagen I are typical for age-related skin atrophy. In this paper, we hypothesize that collagen I may be a relevant molecule stimulating the differentiation of adipose-derived stem cells (ASCs) into adipocytes augmenting subcutaneous fat. In this context lipogenesis, adiponectin, and collagen I receptor expression were determined. Freshly isolated ASCs were characterized by stemness-associated surface markers by FACS analysis and then transdifferentiated into adipocytes by specific medium supplements. Lipogenesis was evaluated using Nile Red staining and documented by fluorescence microscopy or quantitatively measured by using a multiwell spectrofluorometer. Expression of adiponectin was measured by real-time RT-PCR and in cell-free supernatants by ELISA, and expression of collagen I receptors was observed by western blot analysis. It was found that supports coated with collagen I promote cell adhesion and lipogenesis of ASCs. Interestingly, a reverse correlation to adiponectin expression was observed. Moreover, we found upregulation of the collagen receptor, discoidin domain-containing receptor 2; receptors of the integrin family were absent or downregulated. These findings indicate that collagen I is able to modulate lipogenesis and adiponectin expression and therefore may contribute to metabolic dysfunctions associated with ageing.
| S-EPMC6523348 | biostudies-literature
Project description:A great variety of compounds are formed during lipid peroxidation of polyunsaturated fatty acids of membrane phospholipids. Among them, bioactive aldehydes, such as 4-hydroxyalkenals, malondialdehyde (MDA) and acrolein, have received particular attention since they have been considered as toxic messengers that can propagate and amplify oxidative injury. In the 4-hydroxyalkenal class, 4-hydroxy-2-nonenal (HNE) is the most intensively studied aldehyde, in relation not only to its toxic function, but also to its physiological role. Indeed, HNE can be found at low concentrations in human tissues and plasma and participates in the control of biological processes, such as signal transduction, cell proliferation, and differentiation. Moreover, at low doses, HNE exerts an anti-cancer effect, by inhibiting cell proliferation, angiogenesis, cell adhesion and by inducing differentiation and/or apoptosis in various tumor cell lines. It is very likely that a substantial fraction of the effects observed in cellular responses, induced by HNE and related aldehydes, be mediated by their interaction with proteins, resulting in the formation of covalent adducts or in the modulation of their expression and/or activity. In this review we focus on membrane proteins affected by lipid peroxidation-derived aldehydes, under physiological and pathological conditions.
| S-EPMC3761222 | biostudies-literature