Spheroids derived from the stromal vascular fraction of adipose tissue self-organize in complex adipose organoids and secrete leptin
Journal
Stem Cell Research & Therapy
ISSN
1757-6512
Date Issued
2023
Author(s)
Fermín Robledo
Lila González-Hodar
Pablo Tapia
Ana-María Figueroa
Víctor Cortés
Type
Resource Types::text::journal::journal article
URL Institutional Repository
Abstract
<jats:title>Abstract</jats:title><jats:sec>
<jats:title>Background</jats:title>
<jats:p>Adipose tissue-derived stromal vascular fraction (SVF) harbors multipotent cells with potential therapeutic relevance. We developed a method to form adipose spheroids (AS) from the SVF with complex organoid structure and enhanced leptin secretion upon insulin stimulation.
</jats:p>
</jats:sec><jats:sec>
<jats:title>Methods</jats:title>
<jats:p>SVF was generated from the interscapular brown adipose tissue of newborn mice. Immunophenotype and stemness of cultured SVF were determined by flow cytometry and in vitro differentiation, respectively. Spheroids were generated in hanging drops and non-adherent plates and compared by morphometric methods. The adipogenic potential was compared between preadipocyte monolayers and spheroids. Extracellular leptin was quantified by immunoassay. Lipolysis was stimulated with isoprenaline and quantified by colorimetric methods. AS viability and ultrastructure were determined by confocal and transmission electron microscopy analyses.
</jats:p>
</jats:sec><jats:sec>
<jats:title>Results</jats:title>
<jats:p>Cultured SVF contained Sca1 + CD29 + CD44 + CD11b- CD45- CD90- cells with adipogenic and chondrogenic but no osteogenic potential. Culture on non-adherent plates yielded the highest quantity and biggest size of spheroids. Differentiation of AS for 15 days in a culture medium supplemented with insulin and rosiglitazone resulted in greater <jats:italic>Pparg</jats:italic>, <jats:italic>Plin1,</jats:italic> and <jats:italic>Lep</jats:italic> expression compared to differentiated adipocytes monolayers. AS were viable and maintained leptin secretion even in the absence of adipogenic stimulation. Glycerol release after isoprenaline stimulation was higher in AS compared to adipocytes in monolayers. AS were composed of outer layers of unilocular mature adipocytes and an inner structure composed of preadipocytes, immature adipocytes and an abundant loose extracellular matrix.</jats:p>
</jats:sec><jats:sec>
<jats:title>Conclusion</jats:title>
<jats:p>Newborn mice adipose SVF can be efficiently differentiated into leptin-secreting AS. Prolonged stimulation with insulin and rosiglitazone allows the formation of structurally complex adipose organoids able to respond to adrenergic lipolytic stimulation.</jats:p>
</jats:sec>
<jats:title>Background</jats:title>
<jats:p>Adipose tissue-derived stromal vascular fraction (SVF) harbors multipotent cells with potential therapeutic relevance. We developed a method to form adipose spheroids (AS) from the SVF with complex organoid structure and enhanced leptin secretion upon insulin stimulation.
</jats:p>
</jats:sec><jats:sec>
<jats:title>Methods</jats:title>
<jats:p>SVF was generated from the interscapular brown adipose tissue of newborn mice. Immunophenotype and stemness of cultured SVF were determined by flow cytometry and in vitro differentiation, respectively. Spheroids were generated in hanging drops and non-adherent plates and compared by morphometric methods. The adipogenic potential was compared between preadipocyte monolayers and spheroids. Extracellular leptin was quantified by immunoassay. Lipolysis was stimulated with isoprenaline and quantified by colorimetric methods. AS viability and ultrastructure were determined by confocal and transmission electron microscopy analyses.
</jats:p>
</jats:sec><jats:sec>
<jats:title>Results</jats:title>
<jats:p>Cultured SVF contained Sca1 + CD29 + CD44 + CD11b- CD45- CD90- cells with adipogenic and chondrogenic but no osteogenic potential. Culture on non-adherent plates yielded the highest quantity and biggest size of spheroids. Differentiation of AS for 15 days in a culture medium supplemented with insulin and rosiglitazone resulted in greater <jats:italic>Pparg</jats:italic>, <jats:italic>Plin1,</jats:italic> and <jats:italic>Lep</jats:italic> expression compared to differentiated adipocytes monolayers. AS were viable and maintained leptin secretion even in the absence of adipogenic stimulation. Glycerol release after isoprenaline stimulation was higher in AS compared to adipocytes in monolayers. AS were composed of outer layers of unilocular mature adipocytes and an inner structure composed of preadipocytes, immature adipocytes and an abundant loose extracellular matrix.</jats:p>
</jats:sec><jats:sec>
<jats:title>Conclusion</jats:title>
<jats:p>Newborn mice adipose SVF can be efficiently differentiated into leptin-secreting AS. Prolonged stimulation with insulin and rosiglitazone allows the formation of structurally complex adipose organoids able to respond to adrenergic lipolytic stimulation.</jats:p>
</jats:sec>
Cite this document
Robledo, F., González-Hodar, L., Tapia, P., Figueroa, A.-M., Ezquer, F., & Cortés, V. (2023). Spheroids derived from the stromal vascular fraction of adipose tissue self-organize in complex adipose organoids and secrete leptin. Stem Cell Research & Therapy, 14(1), 70. https://doi.org/10.1186/s13287-023-03262-2
Subjects
adipogenesis
;
adipose tissue
;
leptin
;
organoid
;
spheroid
;
adipose tissue
;
animals
;
cell differentiation
;
cells, cultured
;
insulins
;
isoproterenol
;
leptin
;
mice
;
obesity
;
organoids
;
rosiglitazone
;
stromal cells
;
stromal vascular fraction
;
beta1 integrin
;
cd11b antigen
;
complementary dna
;
genomic dna
;
glycerol
;
hermes antigen
;
insulin
;
isoprenaline
;
leptin
;
rosiglitazone
;
thy 1 membrane glycoprotein
;
insulin derivative
;
leptin
;
adipogenesis
;
adipose tissue
;
animal cell
;
animal experiment
;
animal tissue
;
article
;
bright field microscopy
;
cell aggregation
;
cell culture
;
cell differentiation
;
cell isolation
;
cell viability
;
chondrogenesis
;
colorimetry
;
confocal laser scanning microscopy
;
controlled study
;
dna extraction
;
enzyme linked immunosorbent assay
;
extracellular matrix
;
flow cytometry
;
fluorescence microscopy
;
hanging
;
human
;
immunoassay
;
immunophenotyping
;
in vitro study
;
light dark cycle
;
lipolysis
;
monolayer culture
;
morphometry
;
mouse
;
newborn
;
nonhuman
;
organoid
;
proadipocyte
;
rna extraction
;
spheroid cell
;
stromal vascular fraction
;
transmission electron microscopy
;
ultrastructure
;
adipose tissue
;
animal
;
metabolism
;
obesity
;
organoid
;
stroma cell