scholarly journals P049 Development of human embryonic stem cell-derived intestinal organoids for in vitro studies on intestinal inflammation and fibrosis

2021 ◽  
Vol 15 (Supplement_1) ◽  
pp. S158-S158
Author(s):  
L Kandilogiannakis ◽  
E Filidou ◽  
I Drygiannakis ◽  
G Tarapatzi ◽  
K Arvanitidis ◽  
...  
Keyword(s):  
Mrna Expression ◽  
Collagen Type ◽  
Embryonic Stem ◽  
Collagen Type I ◽  
Type I ◽  
Maturation Process ◽  
Type Iii ◽  
Intestinal Organoids ◽  
Tnf Α

Abstract Background Organoids are self-renewing, 3D structures, consisting of different cell types, with histology and physiology features very close to the physiology of the studied organ. Specifically, human Intestinal Organoids (HIOs) develop epithelial crypts consisting of all subtypes of intestinal epithelial cells which are surrounded by mesenchymal cells. Our aim was to develop 3D HIOs from human embryonic stem cells (hESCs) and examine the expression of fibrotic and mesenchymal factors during their maturation process. Additionally, we investigated the effect of the pro-inflammatory cytokines, IL-1α and TNF-α on the expression of fibrotic and inflammatory mediators in HIOs. Methods The human ESC line (H1) was cultured and then differentiated towards HIOs using commercially available kit. HIOs were characterized by immunofluorescence in all differentiation stages. In order to examine their maturation process, we compared the mRNA expression of fibrotic and mesenchymal markers from passages 1–10. In order to examine their functionality, HIOs from different passages were stimulated with 5ng/ml IL-1α and 50ng/ml TNF-α for 12 hours, total RNA was collected and the fibrotic and inflammatory mRNA expression was examined. The mRNA transcripts of CD90, collagen type I, III, fibronectin, CXCL8, CXCL10 and CXCL11 were measured by reverse transcription quantitative PCR. Results HIOs were successfully developed as they were stained positive for all tested markers throughout their developmental process. Regarding their maturation process, we observed high expression of CD90, collagen type I, type III and fibronectin that was gradually decreased during passages. As for the fibrotic and inflammatory responses from HIOs, we found that the IL-1α and TNF-α stimulation resulted in statistically significant upregulation of the fibrotic factors, fibronectin, collagen type I and type III in culture passages 2 and 4, but had no effect in culture passages 8 and 10. Similarly, IL-1α and TNF-α stimulation led to the statistically significant induction of the inflammatory chemokines CXCL8, CXCL10 and CXCL11 in culture passages 2 and 4, while no effect was observed in culture passages 8 and 10. Conclusion Our findings indicate that HIOs contain a functional mesenchymal component that is gradually diminished during passages. Inflammatory and fibrotic responses of HIOs seem to depend on the fitness of their mesenchyme. IBD studies using HIOs as in vitro models should be performed on early passages, when HIO’s mesenchymal component is still functional.

scholarly journals Collagen Type I Containing Hybrid Hydrogel Enhances Cardiomyocyte Maturation in a 3D Cardiac Model

Polymers ◽  
2019 ◽  
Vol 11 (4) ◽  
pp. 687 ◽  
Author(s):  
Sam G. Edalat ◽  
Yongjun Jang ◽  
Jongseong Kim ◽  
Yongdoo Park
Keyword(s):  
Growth Factor ◽  
Collagen Type ◽  
Transforming Growth Factor ◽  
Embryonic Stem ◽  
Collagen Type I ◽  
Transforming Growth Factor Β1 ◽  
Type I ◽  
Hybrid Hydrogel ◽  
Cardiac Model

In vitro maturation of cardiomyocytes in 3D is essential for the development of viable cardiac models for therapeutic and developmental studies. The method by which cardiomyocytes undergoes maturation has significant implications for understanding cardiomyocytes biology. The regulation of the extracellular matrix (ECM) by changing the composition and stiffness is quintessential for engineering a suitable environment for cardiomyocytes maturation. In this paper, we demonstrate that collagen type I, a component of the ECM, plays a crucial role in the maturation of cardiomyocytes. To this end, embryonic stem-cell derived cardiomyocytes were incorporated into Matrigel-based hydrogels with varying collagen type I concentrations of 0 mg, 3 mg, and 6 mg. Each hydrogel was analyzed by measuring the degree of stiffness, the expression levels of MLC2v, TBX18, and pre-miR-21, and the size of the hydrogels. It was shown that among the hydrogel variants, the Matrigel-based hydrogel with 3 mg of collagen type I facilitates cardiomyocyte maturation by increasing MLC2v expression. The treatment of transforming growth factor β1 (TGF-β1) or fibroblast growth factor 4 (FGF-4) on the hydrogels further enhanced the MLC2v expression and thereby cardiomyocyte maturation.

scholarly journals Effect of acute resistance exercise and sex on human patellar tendon structural and regulatory mRNA expression

2009 ◽  
Vol 106 (2) ◽  
pp. 468-475 ◽  
Author(s):  
Bridget E. Sullivan ◽  
Chad C. Carroll ◽  
Bozena Jemiolo ◽  
Scott W. Trappe ◽  
S. Peter Magnusson ◽  
...  
Keyword(s):  
Resistance Exercise ◽  
Mrna Expression ◽  
Patellar Tendon ◽  
Type I Collagen ◽  
Collagen Type ◽  
Collagen Type I ◽  
Tissue Inhibitors Of Metalloproteinases ◽  
Type I ◽  
Collagen Type Iii ◽  
Type Iii

Tendon is mainly composed of collagen and an aqueous matrix of proteoglycans that are regulated by enzymes called matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs). Although it is known that resistance exercise (RE) and sex influence tendon metabolism and mechanical properties, it is uncertain what structural and regulatory components contribute to these responses. We measured the mRNA expression of tendon's main fibrillar collagens (type I and type III) and the main proteoglycans (decorin, biglycan, fibromodulin, and versican) and the regulatory enzymes MMP-2, MMP-9, MMP-3, and TIMP-1 at rest and after RE. Patellar tendon biopsy samples were taken from six individuals (3 men and 3 women) before and 4 h after a bout of RE and from a another six individuals (3 men and 3 women) before and 24 h after RE. Resting mRNA expression was used for sex comparisons (6 men and 6 women). Collagen type I, collagen type III, and MMP-2 were downregulated ( P < 0.05) 4 h after RE but were unchanged ( P > 0.05) 24 h after RE. All other genes remained unchanged ( P > 0.05) after RE. Women had higher resting mRNA expression ( P < 0.05) of collagen type III and a trend ( P = 0.08) toward lower resting expression of MMP-3 than men. All other genes were not influenced ( P > 0.05) by sex. Acute RE appears to stimulate a change in collagen type I, collagen type III, and MMP-2 gene regulation in the human patellar tendon. Sex influences the structural and regulatory mRNA expression of tendon.

scholarly journals In vitro fibrillogenesis of tropocollagen type III in collagen type I affects its relative fibrillar topology and mechanics

2017 ◽  
Vol 7 (1) ◽  
Author(s):  
Meisam Asgari ◽  
Neda Latifi ◽  
Hossein K. Heris ◽  
Hojatollah Vali ◽  
Luc Mongeau
Keyword(s):  
Collagen Type ◽  
Collagen Type I ◽  
Type I ◽  
Type Iii

Orthosilicic Acid Increases Collagen Type I mRNA Expression in Human Bone-Derived Osteoblasts In Vitro

2003 ◽  
Vol 254-256 ◽  
pp. 869-872 ◽  
Author(s):  
Meera Q. Arumugam ◽  
D.C. Ireland ◽  
Roger A. Brooks ◽  
Neil Rushton ◽  
William Bonfield
Keyword(s):  
Mrna Expression ◽  
Collagen Type ◽  
Collagen Type I ◽  
Human Bone ◽  
Orthosilicic Acid ◽  
Type I

scholarly journals Collagen metabolism in cultured osteoblasts from osteogenesis imperfecta patients

1992 ◽  
Vol 286 (1) ◽  
pp. 73-77 ◽  
Author(s):  
M Mörike ◽  
R E Brenner ◽  
G B Bushart ◽  
W M Teller ◽  
U Vetter
Keyword(s):  
Osteogenesis Imperfecta ◽  
Type I Collagen ◽  
Collagen Type ◽  
Bone Cells ◽  
Collagen Type I ◽  
Collagen Metabolism ◽  
Type I ◽  
Type Iii ◽  
Type Iv

Collagen produced in vitro by bone cells isolated from 19 patients with different forms of osteogenesis imperfecta (OI) was analysed. Clinically, four patients were classified as OI type I, 10 patients as OI type III and five patients as OI type IV. Bone cells of 12 of the 19 OI patients produced structurally abnormal type I collagen. Electrophoretically uniformly slower migrating collagen type I alpha-chains were found in one case of OI type I, in seven cases of OI type III and in one case of OI type IV; two cultures of OI type III produced two different populations of collagen type I alpha-chains, and one culture of OI type IV showed reduction-sensitive dimer formation of alpha 1(I) chains, resulting from the inadequate incorporation of a cysteine residue into the triple helical domain of alpha 1(I). Quantitative analysis of collagen metabolism led to the distinction of two groups of cultured OI osteoblasts. In osteoblasts of OI type I, mainly production of collagen was decreased, whereas secretion, processing and pericellular accumulation of (pro)collagen type I was similar to that in control osteoblasts. In contrast, in osteoblasts of OI types III and IV, production as well as secretion, processing and pericellular accumulation of (pro)collagen type I were significantly decreased. Low levels of type I collagen were found irrespective of the presence or absence of structural abnormalities of collagen type I in all OI types.

Marked attenuation of production of collagen type I from cardiac fibroblasts by dehydroepiandrosterone

2005 ◽  
Vol 288 (6) ◽  
pp. E1222-E1228 ◽  
Author(s):  
Tomoyuki Iwasaki ◽  
Koji Mukasa ◽  
Masato Yoneda ◽  
Satoshi Ito ◽  
Yoshihiko Yamada ◽  
...  
Keyword(s):  
Mrna Expression ◽  
Collagen Type ◽  
Cardiac Fibroblasts ◽  
Collagen Type I ◽  
Protein Accumulation ◽  
Transcriptional Level ◽  
Type I ◽  
Procollagen Type ◽  
Procollagen Type I

Dehydroepiandrosterone (DHEA) is a type of adrenal steroid. The concentrations of DHEA and its sulfate (DHEA-S) in serum reach a peak between the ages of 25 and 30 yr and thereafter decline steadily. It was reported that DHEA-S concentration in humans is inversely related to death from cardiovascular diseases. In this study, we examined the effects of DHEA on regulation of collagen mRNA and collagen synthesis in cultured cardiac fibroblasts. Treatment with DHEA (10−6 M) resulted in a significant decrease in procollagen type I mRNA expression compared with controls. This was accompanied by a significant decrease in procollagen type I protein accumulation in the medium and also a significant decrease in procollagen type I protein synthesis in the cellular matrix. Furthermore, to confirm in vitro results, we administered DHEA to Sprague-Dawley rats, which were treated with angiotensin II for 8 wk to induce cardiac damage. Procollagen type I mRNA expression was significantly decreased and cardiac fibrosis significantly inhibited in DHEA-treated rat hearts without lowering the systolic blood pressure. These results strongly indicate that DHEA can directly attenuate collagen type I synthesis at the transcriptional level in vivo and in vitro in cardiac fibroblasts.

The Effect Orthosilicic Acid on Collagen Type I, Alkaline Phosphatase and Osteocalcin mRNA Expression in Human Bone-Derived Osteoblasts In Vitro

2006 ◽  
Vol 309-311 ◽  
pp. 121-124 ◽  
Author(s):  
Meera Q. Arumugam ◽  
D.C. Ireland ◽  
Roger A. Brooks ◽  
Neil Rushton ◽  
William Bonfield
Keyword(s):  
Alkaline Phosphatase ◽  
Mrna Expression ◽  
Messenger Rna ◽  
Collagen Type ◽  
Collagen Type I ◽  
Orthosilicic Acid ◽  
Type I ◽  
Mrna Levels ◽  
Polymerase Chain

The object of this study was to investigate the effect of the concentration of orthosilicic acid (0, 0.5, 1, 5 and 10µM) on gene expression in human osteoblast cells isolated from trabecular bone. This was measured using reverse transcriptase-polymerase chain reaction (RT-PCR) to quantify messenger RNA (mRNA) levels for collagen type I, alkaline phosphatase and osteocalcin. Results showed that while collagen type I mRNA expression was increased by the addition of up to 10µM orthosilicic acid, ALP message was suppressed over time and osteocalcin levels were decreased.

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