Establishment of an Immortalized Endometriotic Stromal Cell Line from Human Ovarian Endometrioma

We show here that mouse pluripotent hematopoietic stem cells can be maintained in vitro on stroma for at least 3 weeks at levels close to those found in bone marrow. The extent of stem cell maintenance is affected by the nature of the stromal cells. The stromal cell line S17 supported stem cells significantly better than heterogeneous, primary stromal layers or the stromal cell line Strofl-1. Stem cells cultured on S17 repopulated all hematopoietic lineages in marrow-ablated hosts for at least 10 months, indicating that this culture system maintained primitive stem cells with extensive proliferative capacity. Furthermore, we demonstrate that, while pluripotent stem cells express c-kit, this receptor appears to play only a minor role in stem cell maintenance in vitro. The addition of an antibody that blocks the interaction of c-kit with its ligand essentially abrogated myelopoiesis in cultures. However, the level of stem cells in antibody-treated cultures was similar to that found in untreated cultures. Thus, it seems likely that the maintenance of primitive stem cells in vitro depends on yet unidentified stromal cell-derived factor(s).

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Improved hematopoiesis in anemic Sl/Sld mice by splenectomy and therapeutic transplantation of a hematopoietic microenvironment

Blood ◽

10.1182/blood.v74.3.1144.bloodjournal7431144 ◽

1989 ◽

Vol 74 (3) ◽

pp. 1144-1151

Author(s):

P Anklesaria ◽

TJ FitzGerald ◽

K Kase ◽

A Ohara ◽

JS Greenberger

Keyword(s):

Bone Marrow ◽

Cell Line ◽

Hind Limb ◽

Stromal Cell ◽

Bone Marrow Stromal Cell ◽

Volume Distribution ◽

Distribution Analysis ◽

Marrow Stromal Cell ◽

Stromal Cell Line ◽

The Right

The ability of a clonal hematopoiesis-supportive bone-marrow stromal cell line GBlneor to engraft and alter the microenvironment-induced anemia of Sl/Sld mice was studied. Prior to stromal cell transplantation, Sl/Sld mice received 1 Gy total body irradiation (TBI) and 13 Gy to the right hind limb. Two months after intravenous (IV) injection of 5 x 10(5) GBlneor cells, 54.4% +/- 17.0% donor origin (G418r) colony-forming cells were recovered from the right hind limb of Sl/Sld mice. Long-term bone marrow cultures (LTBMCs) established from GBlneor-transplanted mice produced 189.5 CFU-GEMM-forming progenitors/flask over 10 weeks compared with 52.7 +/- 6.2 CFU-GEMM forming progenitors/flask from irradiated nontransplanted Sl/Sld mice. A partial correction of macrocytic anemia was detected 2 months after GBlneor transplantation in splenectomized, irradiated Sl/Sld mice (HgB 7.2 +/- 0.4 g/dL; MCV 68.3 +/- 7.0 fL) compared to splenectomized, irradiated, nontransplanted Sl/Sld mice (HgB 5.5 +/- 1.1 g/dL; MCV 76 +/- 8.5 fL) or control Sl/Sld mice (HgB 5.4 +/- 0.5 g/dL; MCV 82.4 +/- 1.3 fL). Mean RBC volume distribution analysis showed a 2.5-fold increase in percentage of peripheral blood RBCs with MCV less than or equal to 45 fL and confirmed reduction of the MCV in splenectomized- GBlneor-transplanted mice compared to control Sl/Sld mice. A hematopoiesis-suppressive clonal stromal cell line derived from LTBMCs of Sl/Sld mice (Sldneor) engrafted as effectively (43.5% +/- 1.2% G418r CFU-F/limb) as did GBlneor cells (38.3% +/- 0.16% G418r CFU-F/limb) to the irradiated right hind limbs of C57Bl/6 mice. LTBMCs established after 2 or 6 months from Sldneor-transplanted mice showed decreased hematopoiesis (182 +/- 12 [2 months] and 3494.3 +/- 408.1 [6 months] CFU-GEMM forming progenitors/flask over 10 weeks) compared to those established from GBlneor-transplanted mice (5980 +/- 530 [2 months] and 7728 +/- 607, [6 months] CFU-GEMM progenitors forming/flask). Thus, transplantation of clonal bone-marrow stromal cell lines in vivo can stably transfer their physiologic properties to normal or mutant mice.

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Expression of adipogenesis markers in a murine stromal cell line treated with 15-deoxy Δ12,14-prostaglandin J2, interleukin-11, 9-cis retinoic acid and vitamin K2

Prostaglandins Leukotrienes and Essential Fatty Acids ◽

10.1054/plef.2001.0314 ◽

2001 ◽

Vol 65 (4) ◽

pp. 215-221 ◽

Cited By ~ 15

Author(s):

V. Garcia Palacios ◽

I. Morita ◽

S. Murota

Keyword(s):

Retinoic Acid ◽

Cell Line ◽

Stromal Cell ◽

Vitamin K2 ◽

Stromal Cell Line ◽

Interleukin 11 ◽

Prostaglandin J2

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Establishment of an immortalized stromal cell line derived from human Endometriotic lesion

Reproductive Biology and Endocrinology ◽

10.1186/s12958-020-00669-x ◽

2020 ◽

Vol 18 (1) ◽

Author(s):

Zhi-Xiong Huang ◽

Rong-Feng Wu ◽

Xiao-Mei Mao ◽

Shao-Min Huang ◽

Tian-Tian Liu ◽

...

Keyword(s):

Inflammatory Response ◽

Life Span ◽

Cell Line ◽

Nude Mouse ◽

Stromal Cells ◽

Colony Formation ◽

Stromal Cell ◽

Primary Cells ◽

Immortalized Cells ◽

Stromal Cell Line

Abstract Background Endometriosis is a benign gynecological disease with obviously feature of estrogen-dependence and inflammatory response. The applications of primary endometriotic stromal cells in research of endometriosis are restricted for short life span, dedifferentiation of hormone and cytokine responsiveness. The objective of this study was to establish and characterize immortalized human endometriotic stromal cells (ihESCs). Methods The endometriotic samples were from a patient with ovarian endometriosis and the primary endometriotic stromal cells were isolated from the endometriotic tissues. The primary cells were infected by lentivirus to establish telomerase reverse transcriptase (hTERT)-induced immortalized cells. Quantification of mRNA and proteins was examined by quantitative real-time polymerase chain reaction (qRT-PCR) and Western Blot. CCK-8 assay and EdU labeling assay were assigned to assess the growth of ihESCs. Karyotype assay was performed to detect the chromosomes of ihESCs. Colony formation assay and nude mouse tumorigenicity assay were used to evaluate colony-formation and tumorigenesis abilities. Results ihESCs continuously overexpressed hTERT via infection of lentivirus and significant extended the life span reaching 31 passages. The morphology, proliferation and karyotype of ihESCs remained unchanged. The expression of epithelial-mesenchymal transition (EMT) markers, estrogen-metabolizing proteins and estrogen/progesterone receptors (ERs and PRs) were unaltered. Furthermore, the treatment of estrogen increased the proliferation and EMT of ihESCs. Lipopolysaccharides (LPS) and IL-1β remarkably induced inflammatory response. The clonogenesis ability of ihESCs was consistent with primary cells, which were much lower than Ishikawa cells. In addition, nude mouse tumorigenicity assay demonstrated that ihESCs were unable to trigger tumor formation. Conclusion This study established and characterized an immortalized endometriotic stromal cell line that exhibited longer life span and kept the cellular morphology and physiological function as the primary cells. The immortalized cells remained normal feedback to estrogen and inflammatory response. Moreover, the immortalized cells were not available with tumorigenic ability. Therefore, ihESCs would be serviceable as in vitro cell tool to investigate the pathogenesis of endometriosis.

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In Vitro Murine Hematopoiesis Supported by Signaling from a Splenic Stromal Cell Line

Stem Cells International ◽

10.1155/2018/9896142 ◽

2018 ◽

Vol 2018 ◽

pp. 1-9 ◽

Cited By ~ 1

Author(s):

Hong Kiat Lim ◽

Pravin Periasamy ◽

Helen C. O’Neill

Keyword(s):

Bone Marrow ◽

Cell Line ◽

Stromal Cell ◽

Hematopoietic Cells ◽

Model Systems ◽

Hematopoietic Stem ◽

Cell Production ◽

Stromal Cell Line ◽

Reticular Cells

There are very few model systems which demonstrate hematopoiesis in vitro. Previously, we described unique splenic stromal cell lines which support the in vitro development of hematopoietic cells and particularly myeloid cells. Here, the 5G3 spleen stromal cell line has been investigated for capacity to support the differentiation of hematopoietic cells from progenitors in vitro. Initially, 5G3 was shown to express markers of mesenchymal but not endothelial or hematopoietic cells and to resemble perivascular reticular cells in the bone marrow through gene expression. In particular, 5G3 resembles CXCL12-abundant reticular cells or perivascular reticular cells, which are important niche elements for hematopoiesis in the bone marrow. To analyse the hematopoietic support function of 5G3, specific signaling pathway inhibitors were tested for the ability to regulate cell production in vitro in cocultures of stroma overlaid with bone marrow-derived hematopoietic stem/progenitor cells. These studies identified an important role for Wnt and Notch pathways as well as tyrosine kinase receptors like c-KIT and PDGFR. Cell production in stromal cocultures constitutes hematopoiesis, since signaling pathways provided by splenic stroma reflect those which support hematopoiesis in the bone marrow.

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