Comparison of enzymatic and nonenzymatic isolation methods for endometrial stem cells

Abstract

Human endometrial stem cells (hESCs) are mesenchymal stem cells, which are responsible for the monthly renewal of the basal layer of the human endometrium by facilitating stromal and vascular regeneration. In this study, hESCs were isolated by using three different isolation methods including nonenzymatic and enzymatic digestion with trypsin and collagenase type 1. To determine the efficiency of these three methods, cells were characterized using a cell proliferation assay and mesenchymal and hematopoietic stem cell markers. Our results demonstrate that although the nonenzymatic isolation method gave rise to hESCs that had a higher proliferative rate, the mesenchymal stem cell profiles for hESCs isolated with three methods were similar, with no significant difference for the early passages. However, late passage hESCs isolated using trypsin showed a CD31 high CD44 low profile. Similarly, when hESCs isolated with the nonenzymatic method were kept until late passage, they demonstrated a CD31 high profile with a significant decrease in CD90, CD73, CD44, and CD105 surface expression levels. Only hESCs isolated with collagenase type 1 did not present a significant shift in their mesenchymal CD marker profile from early to late passages, suggesting that the long-term maintenance of mesenchymal markers could only be achieved in cell isolation with collagenase type 1.

Garlic (Allium sativum)-derived SEVs inhibit cancer cell proliferation and induce caspase mediated apoptosis

“…Cells were seeded with a concentration of of 5 × 10 3 cells/well in a 96 well plate and treated with escalating doses of A. sativum nanovesicles and garlic juice (5–50 μg/ml) for 24, 48 or 72 h in a humidified incubator (5% CO 2 in air at 37 °C). The cell viability was performed with WST-1 assay (BioVision, CA) as described previously 20 . The absorbance was evaluated with the spectrophotometer at 540 nm.…”

Mekani̇k Ve Enzi̇mati̇k Yöntem İle İzole Edi̇len Stromal Vasküler Fraksi̇yon’un Yara İyi̇leşmesi̇ Üzeri̇ndeki̇ Etki̇si̇ni̇n in Vi̇tro İncelenmesi̇

“…Hücre proliferasyon analizi için, iki farklı yöntem ile izole edilen hücre kokteyllerinden 3x10 3 hücre/kuyucuk olacak şekilde, 96 oyuklu hücre kültür kaplarına ekilerek, %10'luk fetal sığır serumu (Gibco, UK) ile desteklenmiş düşük glukozlu DMEM besiyerinde 24, 48 ve 72 saat boyunca muamele edilmiştir. Hücre proliferasyonu testi, literatürde belirtilen WST-1 testi (BioVision, CA) ile değerlendirilmiştir (9). WST-1 test sonuçları Absorbans değeri, 540 nm'de spektrotometre kullanılarak ölçümlenmiştir.…”

Role of Tissue Transglutaminase Catalytic and Guanosine Triphosphate-Binding Domains in Renal Cell Carcinoma Progression

“…CD marker analysis was performed as described previously. 82 In brief, cells fixed with 4% paraformaldehyde were incubated with FITC-or PEconjugated antibodies (CD44, CD73, CD106, CD11b, Sca-1, and CD45) for 1 h at room temperature. Following two sets of washes with PBS, the mesenchymal stem cell marker expression profile was determined using a Becton Dickinson FACSCalibur flow cytometer, and the data were analyzed with the CellQuest Pro program.…”

“…CD marker analysis was performed as described previously . In brief, cells fixed with 4% paraformaldehyde were incubated with FITC- or PE-conjugated antibodies (CD44, CD73, CD106, CD11b, Sca-1, and CD45) for 1 h at room temperature.…”