Optimizing adipogenic differentiation of adipose stem cells in two biocompatible hydrogels
Koskimäki, Sanna (2020)
2020
Bioteknologian tutkinto-ohjelma, luonnontieteiden kandidaatin tutkinto - Degree Programme in Biotechnology, Bachelor of Science
Lääketieteen ja terveysteknologian tiedekunta - Faculty of Medicine and Health Technology
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Hyväksymispäivämäärä
2020-05-07
Julkaisun pysyvä osoite on
https://urn.fi/URN:NBN:fi:tuni-202004284228
https://urn.fi/URN:NBN:fi:tuni-202004284228
Tiivistelmä
Adipose tissue serves as an important energy storage, but it also participates in several biological processes, including endocrine and paracrine regulation of energy metabolism and thermoregulation. Understanding these processes is vital for development of targeted therapeutics for diseases linked to them, such as diabetes and obesity. Moreover, regenerated adipose-like tissues are needed for soft tissue reconstruction in addition to in vitro models.
2D models fail to equate the features of native adipose tissue, which 3D models aim to mimic. Adipogenesis is the process during which stem cells differentiate into mature adipocytes. Aim of this study is to optimize adipogenic differentiation of adipose stem cells using two different hydrogels and varying cell densities. Hydrogels are polymer networks that have high water absorption capacity. Also, quantitative and qualitative analysis methods for adipogenic differentiation are optimized for 3D-cell culture.
In the study, adipogenic differentiation of human adipose-derived stem cells (hASCs) was tested in hydrazone crosslinked gellan gum/gelatin (1:1) hydrogel (gellan gum 40 mg/ml in αMEM and gelatin CDH 40 mg/ml in αMEM.) and in fibrin hydrogel (fibrinogen in DPBS 5 mg/ml and thrombin in EBM-2 2 iU/ml in a 1:1 mixture). The cells were seeded inside the hydrogel in a 48-well-plate format at densities of 0,5, 1,0, ja 2,0 x 106 cells/ml hydrogel and cultured in adipogenic differentiation medium (AM). Differentiation of hASCs was analyzed and quantified using AdipoRed staining and WST-1 cell proliferation assay after 14 or 21 days of culturing. 2D hASC culture with 1053 cells/cm2 was used as a differentiation control. Cell morphology and lipid droplet formation were observed during culture time.
In the study, we showed that the adipogenic differentiation of hASC is dependent on the cell number. Furthermore, quantification of lipid droplet formation, characteristic for differentiated adipocytes, can be performed in a hydrogel in 3D environment. Human adipose-derived stem cells differentiated in hydrogel and obtained features found in native, mature adipocytes, including intracellular lipid droplets and spherical shape. The simultaneous quantification of both lipid formation and cell viability can be performed in 3D hydrogel presented in this study.
2D models fail to equate the features of native adipose tissue, which 3D models aim to mimic. Adipogenesis is the process during which stem cells differentiate into mature adipocytes. Aim of this study is to optimize adipogenic differentiation of adipose stem cells using two different hydrogels and varying cell densities. Hydrogels are polymer networks that have high water absorption capacity. Also, quantitative and qualitative analysis methods for adipogenic differentiation are optimized for 3D-cell culture.
In the study, adipogenic differentiation of human adipose-derived stem cells (hASCs) was tested in hydrazone crosslinked gellan gum/gelatin (1:1) hydrogel (gellan gum 40 mg/ml in αMEM and gelatin CDH 40 mg/ml in αMEM.) and in fibrin hydrogel (fibrinogen in DPBS 5 mg/ml and thrombin in EBM-2 2 iU/ml in a 1:1 mixture). The cells were seeded inside the hydrogel in a 48-well-plate format at densities of 0,5, 1,0, ja 2,0 x 106 cells/ml hydrogel and cultured in adipogenic differentiation medium (AM). Differentiation of hASCs was analyzed and quantified using AdipoRed staining and WST-1 cell proliferation assay after 14 or 21 days of culturing. 2D hASC culture with 1053 cells/cm2 was used as a differentiation control. Cell morphology and lipid droplet formation were observed during culture time.
In the study, we showed that the adipogenic differentiation of hASC is dependent on the cell number. Furthermore, quantification of lipid droplet formation, characteristic for differentiated adipocytes, can be performed in a hydrogel in 3D environment. Human adipose-derived stem cells differentiated in hydrogel and obtained features found in native, mature adipocytes, including intracellular lipid droplets and spherical shape. The simultaneous quantification of both lipid formation and cell viability can be performed in 3D hydrogel presented in this study.
Kokoelmat
- Kandidaatintutkielmat [9002]