Which Culture System Is better for Chondrogenesis of Adipose-Derived Stem Cells ; Pellet or Micromass?

2020 ◽  
Author(s):  
Mahtab Teimouri ◽  
Batool Hashemibeni ◽  
Mohammad Mardani ◽  
Muhammadhosein Moradi
Keyword(s):  
Stem Cells ◽  
Collagen Type ◽  
Adipose Derived Stem Cells ◽  
Chain Reaction ◽  
Chondrogenic Medium ◽  
Expression Levels ◽  
Polymerase Chain ◽  
Cellular Condensation ◽  
Lower Expression ◽  
Gene Expression Levels

Background and Aims: The current study was conducted to compare the expression levels of collagen type Π and X during chondrogenesis of human adipose-derived mesenchymal stem cells (hADMSCs) pellet and micromass cultures.  Materials and Methods: Extracted hADMSCs were cultured until three passages and then transferred to pellet and micromass cultures in the experimental groups of day 7 and day14. For pellet and micromass cultures, aliquots of 5×105 cells/ml were centrifuged and respectively cultured in the conical tubes and droplets (12.5 µl) of the 24-well plates containing  chondrogenic medium. Realtime-polymerase chain reaction technique was performed for gene expression levels. Results: Increased expression of collagen type Π was shown in micromass day14 compared to micromass day 7, pellet day 14 (p<0.01) and pellet day 7 (p<0.001). Also, an increased expression of collagen type Π was seen in micromass day 7 and pellet day 14 compared to pellet day 7 (p< 0.05). Expression of collagen type X increased in pellet day 14 compared to micromass on days 7 and 14 (p<0.001, p<0.01) and pellet day14 compared to pellet day7 (p< 0.05). An increased expression of collagen type X was shown in pellet day 7 compared to micromass on days 7 and 14 (p<0.05). Conclusions: According to the results, higher expression of collagen type Π and lower expression of collagen type X in micromass cultures that are prepared by cell suspension play a better role during cellular condensation that leads to the formation of large nodules exhibiting cartilage-like morphology, suggests a higher efficiency for micromass cultures.

scholarly journals A Comparative Study of the Effect of Anatomical Site on Multiple Differentiation of Adipose-Derived Stem Cells in Rats

Cells ◽  
2021 ◽  
Vol 10 (9) ◽  
pp. 2469
Author(s):  
Hanan Hendawy ◽  
Masahiro Kaneda ◽  
Elsayed Metwally ◽  
Kazumi Shimada ◽  
Takashi Tanaka ◽  
...  
Keyword(s):  
Stem Cells ◽  
Polymerase Chain Reaction ◽  
Adipose Tissue ◽  
Cell Morphology ◽  
Anatomical Site ◽  
Adipose Derived Stem Cells ◽  
Chain Reaction ◽  
Differentiation Potential ◽  
Significant Difference ◽  
Polymerase Chain

Mesenchymal stem cells (MSCs) derived from adipose tissue are evolved into various cell-based regenerative approaches. Adipose-derived stem cells (ASCs) isolated from rats are commonly used in tissue engineering studies. Still, there is a gap in knowledge about how the harvest locations influence and guide cell differentiation. This study aims to investigate how the harvesting site affects stem-cell-specific surface markers expression, pluripotency, and differentiation potential of ASCs in female Sprague Dawley rats. ASCs were extracted from the adipose tissue of the peri-ovarian, peri-renal, and mesenteric depots and were compared in terms of cell morphology. MSCs phenotype was validated by cell surfaces markers using flow cytometry. Moreover, pluripotent gene expression of Oct4, Nanog, Sox2, Rex-1, and Tert was evaluated by reverse transcriptase-polymerase chain reaction (RT-PCR). ASCs multipotency was evaluated by specific histological stains, and the results were confirmed by quantitative polymerase chain reaction (RT-qPCR) expression analysis of specific genes. There was a non-significant difference detected in the cell morphology and immunophenotype between different harvesting sites. ASCs from multiple locations were significantly varied in their capacity to differentiate into adipocytes, osteoblastic cells, and chondrocytes. To conclude, depot selection is a critical element that should be considered when using ASCs in tissue-specific cell-based regenerative therapies research.

scholarly journals Serum lnc34a is a potential prediction biomarker for bone metastasis in hepatocellular carcinoma patients

BMC Cancer ◽  
2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Li Zhang ◽  
Hao Niu ◽  
Ping Yang ◽  
Jie Ma ◽  
Bao-Ying Yuan ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Bone Metastasis ◽  
Univariate Analysis ◽  
High Serum ◽  
Chain Reaction ◽  
Serum Samples ◽  
Early Screening ◽  
Expression Levels ◽  
Node Metastasis ◽  
Polymerase Chain

Abstract Background Early screening and intervention therapies are crucial to improve the prognosis of hepatocellular carcinoma (HCC) patients with bone metastasis. We aimed to identify serum lncRNA as a prediction biomarker in HCC bone metastasis. Methods The expression levels of lnc34a in serum samples from 157 HCC patients were detected by quantitative real-time polymerase chain reaction (PCR). Univariate analysis and multivariate analysis were performed to determine statistically significant variables. Results Expression levels of lnc34a in serum from HCC patients with bone metastasis were significantly higher than those without bone metastasis. The high expressions of lnc34a, vascular invasion and Barcelona Clinic Liver Cancer (BCLC) stage were associated with bone metastasis by analysis. Moreover, lnc34a expression was specifically associated with bone metastasis rather than lung or lymph node metastasis in HCC. Conclusions High serum lnc34a expression was a independent risk factor for developing bone metastasis in HCC.

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.

Electromagnetic fields enhance chondrogenesis of human adipose-derived stem cells in a chondrogenic microenvironment in vitro

2013 ◽  
Vol 114 (5) ◽  
pp. 647-655 ◽  
Author(s):  
Chung-Hwan Chen ◽  
Yi-Shan Lin ◽  
Yin-Chih Fu ◽  
Chih-Kuang Wang ◽  
Shun-Cheng Wu ◽  
...  
Keyword(s):  
Stem Cells ◽  
Electromagnetic Field ◽  
Cell Viability ◽  
Chondrogenic Differentiation ◽  
Collagen Type ◽  
Cartilage Tissue ◽  
Pulse Electromagnetic Field ◽  
Type I ◽  
Adipose Derived Stem Cells ◽  
Pemf Treatment

We tested the hypothesis that electromagnetic field (EMF) stimulation enhances chondrogenesis in human adipose-derived stem cells (ADSCs) in a chondrogenic microenvironment. A two-dimensional hyaluronan (HA)-coated well (2D-HA) and a three-dimensional pellet culture system (3D-pellet) were used as chondrogenic microenvironments. The ADSCs were cultured in 2D-HA or 3D-pellet, and then treated with clinical-use pulse electromagnetic field (PEMF) or the innovative single-pulse electromagnetic field (SPEMF) stimulation. The cytotoxicity, cell viability, and chondrogenic and osteogenic differentiations were analyzed after PEMF or SPEMF treatment. The modules of PEMF and SPEMF stimulations used in this study did not cause cytotoxicity or alter cell viability in ADSCs. Both PEMF and SPEMF enhanced the chondrogenic gene expression (SOX-9, collagen type II, and aggrecan) of ADSCs cultured in 2D-HA and 3D-pellet. The expressions of bone matrix genes (osteocalcin and collagen type I) of ADSCs were not changed after SPEMF treatment in 2D-HA and 3D-pellet; however, they were enhanced by PEMF treatment. Both PEMF and SPEMF increased the cartilaginous matrix (sulfated glycosaminoglycan) deposition of ADSCs. However, PEMF treatment also increased mineralization of ADSCs, but SPEMF treatment did not. Both PEMF and SPEMF enhanced chondrogenic differentiation of ADSCs cultured in a chondrogenic microenvironment. SPEMF treatment enhanced ADSC chondrogenesis, but not osteogenesis, when the cells were cultured in a chondrogenic microenvironment. However, PEMF enhanced both osteogenesis and chondrogenesis under the same conditions. Thus the combination of a chondrogenic microenvironment with SPEMF stimulation can promote chondrogenic differentiation of ADSCs and may be applicable to articular cartilage tissue engineering.

An Effective Cell Coculture Platform Based on the Electrospun Microtube Array Membrane for Nerve Regeneration

2017 ◽  
Vol 204 (3-4) ◽  
pp. 179-190 ◽  
Author(s):  
V. Chia-Hsuan Tseng ◽  
Chee Ho Chew ◽  
Wan-Ting Huang ◽  
Yang-Kao Wang ◽  
Ko-Shao Chen ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Chain Reaction ◽  
Tissue Culture Polystyrene ◽  
Coculture System ◽  
Submicron Scale ◽  
A Cell ◽  
Polymerase Chain ◽  
Transwell System ◽  
Lower Expression ◽  
Microtube Array

Recently, a novel substrate known as an electrospun polylactic acid (PLLA) microtube array membrane (MTAM) was successfully developed as a cell coculture platform. Structurally, this substrate is made up of one-to-one connected, ultrathin, submicron scale fibers that are arranged in an arrayed formation. Its unique structure confers several key advantages which are beneficial in a cell coculture system. In this study, the interaction between rat fetal neural stem cells (NSC) and astrocytes was examined by comparing the outcome of a typical Transwell-based coculture system and that of an electrospun PLLA MTAM-based coculture system. Compared to tissue culture polystyrene (TCP) and Transwell coculture inserts, a superior cell viability of NSC was observed when cultured in lumens of electrospun PLLA MTAM (with supportive immunostaining images). Reverse transcription polymerase chain reaction revealed a strong interaction between astrocytes and NSC through a higher expression of doublecortin and a lower expression of nestin. These data demonstrate that MTAM is clearly a better coculture platform than the traditional Transwell system.

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