scholarly journals Prospects on the Potential In Vitro Regenerative Features of Mechanically Treated-Adipose Tissue for Osteoarthritis Care

2021 ◽  
Author(s):  
G. Desando ◽  
I. Bartolotti ◽  
L. Cattini ◽  
M. Tschon ◽  
L. Martini ◽  
...  
Keyword(s):  
Adipose Tissue ◽  
Stromal Vascular Fraction ◽  
In Vitro Study ◽  
Invasive Technique ◽  
Cell Populations ◽  
Cell Recovery ◽  
Functional Changes ◽  
Osteochondral Repair ◽  
Mechanical Approach

AbstractGathering a better grasp on the adipose stromal vascular fraction (SVF) is demanding among clinicians for osteoarthritis (OA) care because of its promising but multifaceted clinical outcomes. The aim of this preclinical in vitro study was to test whether the mechanical approach with Hy-Tissue SVF system, a class IIa CE marked device of adipose tissue micro-fragmentation, influences the biological features and functions of SVF. We compared mechanical generated-SVF (mSVF) with the enzymatic generated-SVF (eSVF) by testing cell survival, phenotype, differentiation, and paracrine properties using ELISA assays. Both adipose SVF showed 80% viable cells and enrichment for CD-44 marker. The mSVF product preserved the functions of cell populations within the adipose tissue; however, it displayed lowered nucleated cell recovery and CFU-F than eSVF. As for multipotency, mSVF and eSVF showed similar differentiation commitment for osteochondral lineages. Both adipose SVF exhibited an increased release of VEGF, HGF, IGF-1 and PDGF-bb, involved in pathways mediating osteochondral repair and cell migration. Both mSVF and eSVF also displayed high release for the anti-inflammatory cytokine IL-10. After in vitro culture, supernatants from both mSVF and eSVF groups showed a low release of cytokines except for IL-10, thereby giving evidence of functional changes after culture expansion. In this study, mSVF showed active cell populations in the adipose tissue comparable to eSVF with excellent survival, differentiation and paracrine properties under a new mechanical adipose tissue micro-fragmentation system; thereby suggesting its potential use as a minimally invasive technique for OA treatment. Graphical abstract

scholarly journals Generation of immune cell containing adipose organoids for in vitro analysis of immune metabolism

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Jacqueline Taylor ◽  
Julia Sellin ◽  
Lars Kuerschner ◽  
Lennart Krähl ◽  
Yasmin Majlesain ◽  
...  
Keyword(s):  
Adipose Tissue ◽  
Adipocyte Differentiation ◽  
Immune Cell ◽  
Stromal Vascular Fraction ◽  
Cell Populations ◽  
Endocrine Organ ◽  
Vitro Analysis ◽  
In Vitro Analysis ◽  
Better Than

AbstractAdipose tissue is an organized endocrine organ with important metabolic and immunological functions and immune cell-adipocyte crosstalk is known to drive various disease pathologies. Suitable 3D adipose tissue organoid models often lack resident immune cell populations and therefore require the addition of immune cells isolated from other organs. We have created the first 3D adipose tissue organoid model which could contain and maintain resident immune cell populations of the stromal vascular fraction (SVF) and proved to be effective in studying adipose tissue biology in a convenient manner. Macrophage and mast cell populations were successfully confirmed within our organoid model and were maintained in culture without the addition of growth factors. We demonstrated the suitability of our model for monitoring the lipidome during adipocyte differentiation in vitro and confirmed that this model reflects the physiological lipidome better than standard 2D cultures. In addition, we applied mass spectrometry-based lipidomics to track lipidomic changes in the lipidome upon dietary and immunomodulatory interventions. We conclude that this model represents a valuable tool for immune-metabolic research.

Abstract 15440: Dynamics of Adipocyte and Endothelial Progenitor Cell Pools in Expanding Adipose Tissue

Circulation ◽  
2014 ◽  
Vol 130 (suppl_2) ◽  
Author(s):  
Candice R Holden ◽  
Marcin Wysoczynski ◽  
Brian Sansbury ◽  
Jason Hellmann ◽  
Nagma Zafar ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Adipose Tissue ◽  
Stem Cell ◽  
Progenitor Cells ◽  
Progenitor Cell ◽  
Stromal Vascular Fraction ◽  
Cell Populations ◽  
Adipose Organ ◽  
The Impact

Objective: Obesity is a major risk factor for the development of several chronic diseases including type 2 diabetes and cardiovascular disease. Proper fat storage in white adipose tissue (WAT) is required to maintain insulin sensitivity and to preserve (cardio)vascular health. We hypothesize that endothelial and adipocyte progenitor cell populations (EPCs and APCs, respectively) must be appropriately balanced for physiological, as opposed to pathological, remodeling of WAT. Methods and Results: To determine the impact of nutrient excess on stem/progenitor cells in epididymal WAT, male C57BL/6J mice were placed on a high fat diet (HFD; 60% fat) for 12 weeks and changes in WAT stem cell populations were measured in the stromal vascular fraction by flow cytometry. Although the APC (CD24+/CD29+/Sca+/CD14-/CD45-) population, which has the capacity to differentiate into adipocytes both in vitro and in vivo , was not significantly changed with diet, Flk+/Sca+ EPCs were diminished, promoting a 4-fold decrease in the EPC/APC ratio (p <0.05, n = 6/group). To determine whether this deficit may be due to poor stem cell recruitment, mice were irradiated, and the bone marrow was repopulated with GFP+ donor marrow. The transplanted mice were then placed on a low fat diet (LFD; 10% fat) or HFD for 12 weeks, and WAT progenitor cells were again measured. Greater than 95% of the putative APCs in the WAT of HF-fed mice were GFP+ (p<0.0001, n=7-8/group), indicating a bone marrow-derived origin. Unexpectedly, less than 1% of the EPCs were GFP+ (p<0.001, n=7-8/group), which suggests that EPCs present in WAT are not derived from bone marrow in adult mice. Confocal analysis of WAT from HF-fed, bone marrow-transplanted mice showed little evidence of significant APC differentiation into triglyceride-laden adipocytes, suggesting that conditions associated with nutrient excess may impair the ability of the adipose organ to store fat properly. Conclusions: These results demonstrate that putative APCs, and not EPCs, in epididymal WAT are derived from bone marrow. Furthermore, our data suggest that conditions of nutrient excess promote an imbalance in EPCs and APCs, the stoichiometry of which may be critical for the development of new adipocytes and for proper storage of fat.

scholarly journals Simple and Rapid Non-Enzymatic Procedure Allows the Isolation of Structurally Preserved Connective Tissue Micro-Fragments Enriched with SVF

Cells ◽  
2020 ◽  
Vol 10 (1) ◽  
pp. 36
Author(s):  
Alice Busato ◽  
Francesco De Francesco ◽  
Reetuparna Biswas ◽  
Silvia Mannucci ◽  
Giamaica Conti ◽  
...  
Keyword(s):  
Adipose Tissue ◽  
Connective Tissue ◽  
Stromal Vascular Fraction ◽  
Enzymatic Digestion ◽  
Free Cell ◽  
Stem Cell Markers ◽  
Free Cells ◽  
Mechanical Approach ◽  
Enzymatic Procedure

The stromal vascular fraction (SVF) consists of a heterogeneous population of stem and stromal cells, generally obtained from adipose tissue by enzymatic digestion. For human cell-based therapies, mechanical process methods to obtain SVF represent an advantageous approach because they have fewer regulatory restrictions for their clinical use. The aim of this study was to characterize a novel commercial system for obtaining SVF from adipose tissue by a mechanical approach without substantial manipulations. Lipoaspirate samples collected from 27 informed patients were processed by a simple and fast mechanical system (by means of Hy-Tissue SVF). The Hy-Tissue SVF product contained a free cell fraction and micro-fragments of stromal connective tissue. The enzymatic digestion of the micro-fragments increased the yield of free cells (3.2 times) and CFU-F (2.4 times). Additionally, 10% of free cells from SVF were positive for CD34+, suggesting the presence of endothelial cells, pericytes, and potential adipose-derived stem cells (ADSC). Moreover, the SVF cells were able to proliferate and differentiate in vitro toward adipocytes, osteocytes, and chondrocytes. The immunophenotypic analysis of expanded cells showed positivity for typical mesenchymal stem cell markers. The Hy-Tissue SVF system allows the isolation of stromal vascular fraction, making this product of potential interest in regenerative medicine.

scholarly journals Long-Term Severe In Vitro Hypoxia Exposure Enhances the Vascularization Potential of Human Adipose Tissue-Derived Stromal Vascular Fraction Cell Engineered Tissues

2021 ◽  
Vol 22 (15) ◽  
pp. 7920
Author(s):  
Myroslava Mytsyk ◽  
Giulia Cerino ◽  
Gregory Reid ◽  
Laia Gili Sole ◽  
Friedrich S. Eckstein ◽  
...  
Keyword(s):  
Adipose Tissue ◽  
Therapeutic Potential ◽  
Stromal Vascular Fraction ◽  
Human Adipose Tissue ◽  
Bioreactor Culture ◽  
Proliferating Cells ◽  
Angiogenic Potential ◽  
Engineered Tissues

The therapeutic potential of mesenchymal stromal/stem cells (MSC) for treating cardiac ischemia strongly depends on their paracrine-mediated effects and their engraftment capacity in a hostile environment such as the infarcted myocardium. Adipose tissue-derived stromal vascular fraction (SVF) cells are a mixed population composed mainly of MSC and vascular cells, well known for their high angiogenic potential. A previous study showed that the angiogenic potential of SVF cells was further increased following their in vitro organization in an engineered tissue (patch) after perfusion-based bioreactor culture. This study aimed to investigate the possible changes in the cellular SVF composition, in vivo angiogenic potential, as well as engraftment capability upon in vitro culture in harsh hypoxia conditions. This mimics the possible delayed vascularization of the patch upon implantation in a low perfused myocardium. To this purpose, human SVF cells were seeded on a collagen sponge, cultured for 5 days in a perfusion-based bioreactor under normoxia or hypoxia (21% and <1% of oxygen tension, respectively) and subcutaneously implanted in nude rats for 3 and 28 days. Compared to ambient condition culture, hypoxic tension did not alter the SVF composition in vitro, showing similar numbers of MSC as well as endothelial and mural cells. Nevertheless, in vitro hypoxic culture significantly increased the release of vascular endothelial growth factor (p < 0.001) and the number of proliferating cells (p < 0.00001). Moreover, compared to ambient oxygen culture, exposure to hypoxia significantly enhanced the vessel length density in the engineered tissues following 28 days of implantation. The number of human cells and human proliferating cells in hypoxia-cultured constructs was also significantly increased after 3 and 28 days in vivo, compared to normoxia. These findings show that a possible in vivo delay in oxygen supply might not impair the vascularization potential of SVF- patches, which qualifies them for evaluation in a myocardial ischemia model.

scholarly journals Derivation, characterization, and in vitro cell regeneration of canine white adipose tissue-derived mesenchymal stem cells obtained from a mesenteric region

2021 ◽  
Vol 82 (1) ◽  
Author(s):  
Anirban Mandal ◽  
Ajeet Kumar Jha ◽  
Dew Biswas ◽  
Shyamal Kanti Guha
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Adipose Tissue ◽  
White Adipose Tissue ◽  
Stromal Vascular Fraction ◽  
Cell Regeneration ◽  
Wound Healing Assay ◽  
Chain Reaction ◽  
Polymerase Chain

Abstract Background The study was conducted to assess the characterization, differentiation, and in vitro cell regeneration potential of canine mesenteric white adipose tissue-derived mesenchymal stem cells (AD-MSCs). The tissue was harvested through surgical incision and digested with collagenase to obtain a stromal vascular fraction. Mesenchymal stem cells isolated from the stromal vascular fraction were characterized through flow cytometry and reverse transcription-polymerase chain reaction. Assessment of cell viability, in vitro cell regeneration, and cell senescence were carried out through MTT assay, wound healing assay, and β-galactosidase assay, respectively. To ascertain the trilineage differentiation potential, MSCs were stained with alizarin red for osteocytes, alcian blue for chondrocytes, and oil o red for adipocytes. In addition, differentiated cells were characterized through a reverse transcription-polymerase chain reaction. Results We observed the elongated, spindle-shaped, and fibroblast-like appearance of cells after 72 h of initial culture. Flow cytometry results showed positive expression for CD44, CD90, and negative expression for CD45 surface markers. Population doubling time was found 18–24 h for up to the fourth passage and 30±0.5 h for the fifth passage. A wound-healing assay was used to determine cell migration rate which was found 136.9 ± 4.7 μm/h. We observed long-term in vitro cell proliferation resulted in MSC senescence. Furthermore, we also found that the isolated cells were capable of differentiating into osteogenic, chondrogenic, and adipogenic lineages. Conclusions Mesenteric white adipose tissue was found to be a potential source for isolation, characterization, and differentiation of MSCs. This study might be helpful for resolving the problems regarding the paucity of information concerning the basic biology of stem cells. The large-scale use of AD-MSCs might be a remedial measure in regenerative medicine.

scholarly journals Divergent functions of endotrophin on different cell populations in adipose tissue

2016 ◽  
Vol 311 (6) ◽  
pp. E952-E963 ◽  
Author(s):  
Yueshui Zhao ◽  
Xue Gu ◽  
Ningyan Zhang ◽  
Mikhail G. Kolonin ◽  
Zhiqiang An ◽  
...  
Keyword(s):  
Adipose Tissue ◽  
Lipid Accumulation ◽  
Lysyl Oxidase ◽  
Stromal Vascular Fraction ◽  
Cell Types ◽  
Marker Genes ◽  
New Perspective ◽  
Collagen Genes ◽  
Different Cell Types

Endotrophin is a cleavage product of collagen 6 (Col6) in adipose tissue (AT). Previously, we demonstrated that endotrophin serves as a costimulator to trigger fibrosis and inflammation within the unhealthy AT milieu. However, how endotrophin affects lipid storage and breakdown in AT and how different cell types in AT respond to endotrophin stimulation remain unknown. In the current study, by using a doxycycline-inducible mouse model, we observed significant upregulation of adipogenic genes in the white AT (WAT) of endotrophin transgenic mice. We further showed that the mice exhibited inhibited lipolysis and accelerated hypertrophy and hyperplasia in WAT. To investigate the effects of endotrophin in vitro, we incubated different cell types from AT with conditioned medium from endotrophin-overexpressing 293T cells. We found that endotrophin activated multiple pathological pathways in different cell types. Particularly in 3T3-L1 adipocytes, endotrophin triggered a fibrotic program by upregulating collagen genes and promoted abnormal lipid accumulation by downregulating hormone-sensitive lipolysis gene and decreasing HSL phosphorylation levels. In macrophages isolated from WAT, endotrophin stimulated higher expression of the collagen-linking enzyme lysyl oxidase and M1 proinflammatory marker genes. In the stromal vascular fraction isolated from WAT, endotrophin induced upregulation of both profibrotic and proinflammatory genes. In conclusion, our study provides a new perspective on the effect of endotrophin in abnormal lipid accumulation and a mechanistic insight into the roles played by adipocytes and a variety of other cell types in AT in shaping the unhealthy microenvironment upon endotrophin treatment.

Abstract 585: Endothelial Cells from Adipose Tissue of Obese Humans Display Mesenchymal Characteristics

2016 ◽  
Vol 36 (suppl_1) ◽  
Author(s):  
Bronson A Haynes ◽  
Eric J Lehrer ◽  
Giann J Bhatt ◽  
Ryan W Huyck ◽  
Ashley N James ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Adipose Tissue ◽  
Prolonged Exposure ◽  
Endothelial Permeability ◽  
Fold Increase ◽  
Atp Production ◽  
Functional Changes ◽  
Twist 1

The mechanisms underlying vascular dysfunction in adipose tissue (AT) in obesity are not clearly understood. Our hypothesis is that in response to pro-inflammatory cytokines (PIC) present in obese AT, endothelial cells (EC) can de-differentiate and acquire a mesenchymal-like phenotype (EndoMT) that leads to endothelial dysfunction. To test our hypothesis, we measured endothelial and mesenchymal markers of CD31 + CD34 + EC isolated from omental (OM) and subcutaneous (SC) AT of bariatric subjects (BAMVEC) using RT-PCR and western blot. Permeability and oxidative metabolism were determined by ECIS and Seahorse analyzer XF e 24, respectively. BAMVEC isolated from both OM and SC fat showed very low protein expression of vWF and VE-Cadherin (EC markers) and abundantly expressed αSMA and the EMT transcription factor twist-1. To determine effects of PIC on EndoMT, commercially available primary endothelial cells from AT (HAMVEC) were treated in vitro with PIC (2.5ng/mL TNFα, IFNγ and TGFβ) for 1, 3 or 6 days. We found progressive down-regulation by >2-fold (p<0.001) of the EC markers vWF, VE-Cadherin, and Occludin compared to controls. As early as 1 day of PIC treatment twist-1 (p<0.001) and snail1 (p<0.05) showed an increase by >2-fold. Similarly, OM and SC BAMVEC expressed >2-fold increase in the mesenchymal genes twist-1, FSP1, αSMA, and snail1 compared to untreated HAMVEC. Metabolically, BAMVEC had increased ATP production and maximal respiration compared to HAMVEC suggesting increased oxidative phosphorylation, a marker of mesenchymal-like cells. PIC stimulation of HAMVEC yielded significant increases in endothelial permeability and motility (p<0.001). Notably, there were no significant differences in any of the markers between OM and SC BAMVEC. These results show that EC in obese AT exhibit a mesenchymal-like phenotype which may account for functional changes such as increased permeability and migration and are not depot specific. Using primary EC from human AT we showed that prolonged exposure to PIC induces a phenotype similar to CD31+CD34+ EC from obese AT. This supports the concept that AT inflammation can promote EC de-differentiation in vivo and our in vitro model is suitable for future studies to uncover the relevant mechanisms.

scholarly journals Activation of antilipolytic α2-adrenergic receptors by epinephrine during exercise in human adipose tissue

1999 ◽  
Vol 277 (4) ◽  
pp. R1076-R1083 ◽  
Author(s):  
Vladimir Stich ◽  
Isabelle de Glisezinski ◽  
Francois Crampes ◽  
Hana Suljkovicova ◽  
Jean Galitzky ◽  
...  
Keyword(s):  
Adipose Tissue ◽  
In Vitro Study ◽  
Exercise Bout ◽  
Human Adipose Tissue ◽  
Ringer Solution ◽  
Plasma Norepinephrine ◽  
Glycerol Concentration ◽  
Isolated Adipocytes ◽  
Exercise Induced

The involvement of the antilipolytic α2-adrenergic pathway and the specific role of epinephrine in the control of lipolysis during exercise in adipose tissue (AT) were investigated in healthy male subjects (age: 24.1 ± 2.2 yr; body mass index: 23.0 ± 1.6). An in vitro study carried out on isolated adipocytes showed that the weak lipolytic effect of epinephrine was potentiated after blockade of α2-adrenergic receptor (AR) by an α2-AR antagonist and reached that of isoproterenol, a β-AR agonist. The effect of the nonselective α2-AR antagonist phentolamine on the response of the extracellular glycerol concentration (EGC) in AT during two successive bouts of aerobic exercise (50% maximum O2 uptake, 60 min duration) was evaluated using the microdialysis method. The metabolic responses measured in perfused probes with Ringer solution were compared with those obtained in perfused probes with Ringer plus 0.1 mmol/l phentolamine. Plasma norepinephrine level was not different during the two exercise bouts, whereas that of epinephrine was 2.5-fold higher during the second exercise. EGC in AT was twofold higher in the second compared with the first exercise, and the same response pattern was found for plasma glycerol. The exercise-induced increase in EGC was higher in the probe perfused with phentolamine compared with the control probe in both bouts of exercise. However, the potentiating effect of phentolamine on EGC was significant during the second exercise bout but did not reach a significant level during the first. These results suggest that epinephrine is involved in the control of lipid mobilization through activation of antilipolytic α2-AR in human subcutaneous AT during exercise.

scholarly journals Effects of Berries on High-Fat Diet-Induced Inflammation

2020 ◽  
Vol 4 (Supplement_2) ◽  
pp. 449-449
Author(s):  
Patricia Perez ◽  
Desiree Wanders ◽  
Hannah Land ◽  
Kathryn Chiang ◽  
Rami Najjar ◽  
...  
Keyword(s):  
Gene Expression ◽  
Adipose Tissue ◽  
Inflammatory Markers ◽  
In Vitro Study ◽  
Vitro Study ◽  
In Vivo Study ◽  
High Fat ◽  
Anti Inflammatory

Abstract Objectives Studies suggest that inflammation mediates the link between obesity and its comorbidities including type 2 diabetes and cardiovascular disease. Hence, there is a demand for effective alternative or complementary approaches to treat obesity-associated inflammation. The objective of this study was to determine whether consumption of blackberries (BL) and raspberries (RB) alone or in combination reduce obesity-induced inflammation. Methods In Vitro Study: RAW 264.7 macrophages were pretreated with either BL, RB, or BL + RB, each at a final concentration of 200 µg/mL for 2 h. LPS (1 ng/mL) was then added to the media for 16 h. mRNA expression of inflammatory cytokines was measured. In Vivo Study: Five-week-old mice were acclimated to a low-fat low-sucrose (LFLS) diet for one week after which mice were randomized 10 per group to one of five groups: 1) LFLS, 2) high-fat high-sucrose (HFHS), 3) HFHS + 10% BL, 4) HFHS + 10% RB, or 5) HFHS + 5% BL + 5% RB. Expression of inflammatory markers was measured in the liver as well as epididymal and inguinal white adipose tissue. Results In Vitro Study: Each berry alone and in combination suppressed the LPS-induced increase in inflammatory markers, with the combination (BL + RB) having the greatest effect. The combination suppressed LPS-induced expression of Ccl2, Tnfa, F4/80, and Il6 by 3.7−, 5.3−, 5.3−, and 4.4-fold, respectively. In Vivo Study: Gene expression analysis indicated that berry consumption had no significant effect on proinflammatory (Ccl2, Il1b, Tnfa, Il6, Itgam) or anti-inflammatory (Adipoq, Arg1, Mgl1) markers in adipose tissue depots or liver. However, relatively low gene expression of inflammatory markers in the tissues indicates that the mice fed the HFHS diet failed to develop a robust inflammatory state. Conclusions BL and RB have direct anti-inflammatory effects on immune cells. Initial analysis indicates that consumption of BL and RB has no significant effects on markers of inflammation in a diet-induced mouse model of obesity. However, it is possible that the relatively low levels of inflammation in these mice masked the anti-inflammatory potential of BL and RB. Ongoing analysis will provide additional insights into the effects of BL and RB on inflammation in these tissues. Funding Sources Lewis Foundation Award.

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