| 1. |
- El-Habta, Roine, et al.
(författare)
-
The adipose tissue stromal vascular fraction secretome enhances the proliferation but inhibits the differentiation of myoblasts
- 2018
-
Ingår i: Stem Cell Research & Therapy. - : BioMed Central. - 1757-6512. ; 9
-
Tidskriftsartikel (refereegranskat)abstract
- Background: Adipose tissue is an excellent source for isolation of stem cells for treating various clinical conditions including injuries to the neuromuscular system. Many previous studies have focused on differentiating these adipose stem cells (ASCs) towards a Schwann cell-like phenotype (dASCs), which can enhance axon regeneration and reduce muscle atrophy. However, the stromal vascular fraction (SVF), from which the ASCs are derived, also exerts broad regenerative potential and might provide a faster route to clinical translation of the cell therapies for treatment of neuromuscular disorders.Methods: The aim of this study was to establish the effects of SVF cells on the proliferation and differentiation of myoblasts using indirect co-culture experiments. A Growth Factor PCR Array was used to compare the secretomes of SVF and dASCs, and the downstream signaling pathways were investigated.Results: SVF cells, unlike culture-expanded dASCs, expressed and secreted hepatocyte growth factor (HGF) at concentrations sufficient to enhance the proliferation of myoblasts. Pharmacological inhibitor studies revealed that the signal is mediated via ERK1/2 phosphorylation and that the effect is significantly reduced by the addition of 100 pM Norleual, a specific HGF inhibitor. When myoblasts were differentiated into multinucleated myotubes, the SVF cells reduced the expression levels of fast-type myosin heavy chain (MyHC2) suggesting an inhibition of the differentiation process.Conclusions: In summary, this study shows the importance of HGF as a mediator of the SVF effects on myoblasts and provides further evidence for the importance of the secretome in cell therapy and regenerative medicine applications.
|
|
| 2. |
- Sloniecka, Marta, et al.
(författare)
-
Acetylcholine decreases formation of myofibroblasts and excessive extracellular matrix components production in an in vitro human corneal fibrosis model
- 2018
-
Ingår i: Investigative Ophthalmology and Visual Science. - : The Association for Research in Vision and Ophthalmology, Inc.. - 0146-0404 .- 1552-5783. ; 59:9
-
Tidskriftsartikel (övrigt vetenskapligt/konstnärligt)abstract
- Purpose : Acetylcholine (ACh) is a neurotransmitter present in corneal stroma and produced by keratocytes. It has been shown to play a role in processes important for wound healing. Based on literature and our previous studies, we hypothesize that ACh regulates expression of extracellular matrix (ECM) components that are overexpressed during fibrosis, such as collagens, proteoglycans, fibronectin and metalloproteinases, in a protective manner during corneal fibrosis, i.e. decreasing their expression.Methods : Primary keratocytes were isolated from healthy human corneas obtained from the local cornea bank and grown in presence of 10% fetal bovine serum in order to obtain corneal fibroblasts. A corneal fibrosis in vitro model, in which fibroblasts are stimulated with transforming growth factor beta 1 (TGF-β1) and stable vitamin C, was used throughout this study. Contractile ability of myofibroblasts was tested using a cell contraction assay. Gene expression of ECM components (collagen I, collagen III, collagen V and lumican), markers of fibrosis (α-smooth muscle actin [α-SMA] and fibronectin), and metalloproteinases (MMP2, MMP9 and MMP12) were assessed by qRT-PCR. Intracellular production and secretion of pro-collagen I and lumican was determined by ELISA. α-SMA protein expression was assessed by western blot.Results : ACh decreased the contractile ability of the newly formed myofibroblasts. ACh significantly decreased gene expression of collagen I, collagen III and collagen V in myofibroblasts. Moreover, ACh treated cells produced and secreted less pro-collagen I. Gene expression of lumican was unaffected by ACh treatment up to day 2 but significantly decreased by day 4. However, no differences in lumican protein level, both intracellular and secreted, were found. ACh downregulated expression of both α-SMA and fibronectin genes. Additionally, α-SMA protein expression was also diminished in ACh treated cells. Furthermore, ACh treatment resulted in downregulation of MMP2, MMP9 and MMP12 genes.Conclusions : Our results are consistent with our hypothesis that ACh regulates expression of various collagens, lumican, fibronectin and metalloproteinases during corneal fibrosis in vitro, in a way that it diminishes their expression, both on RNA and protein levels. In conclusion, ACh seems to provide protection against formation of fibrosis in human cornea.
|
|
| 3. |
- Sloniecka, Marta, et al.
(författare)
-
Substance P Promotes Fibrosis in Human Corneal Stroma
- 2018
-
Ingår i: Wound Repair and Regeneration. - : Wound Healing Society. - 1067-1927 .- 1524-475X. ; 26:1, s. A22-A22
-
Tidskriftsartikel (övrigt vetenskapligt/konstnärligt)abstract
- Substance P (SP) is a neuropeptide which has been shown to be present in human corneal cells, keratocytes. Many studies suggest its role in various cellular processes important in wound healing such as proliferation or migration. We hypothesize that SP regulates expression of keratocyte markers, extracellular matrix (ECM) components and fibrotic markers that are overexpressed during fibrosis, in both primary keratocytes and myofibroblasts. Primary keratocytes, which were isolated from healthy human corneas obtained from the local cornea bank, and an in vitro corneal fibrosis model (myofibroblasts) were used throughout this study. The effect of SP on keratocyte and myofibroblast contractile abilities was assessed by cell contraction assay. Gene expression of keratocyte markers (keratocan and aldehyde dehydrogenase 3 family, member A1 [ALDH3A1]), ECM components (collagen I, collagen III, collagen V and lumican), and markers of fibrosis (a-smooth muscle actin [a-SMA] and fibronectin), was determined by qRT-PCR. Treatment of keratocytes with SP resulted in decreased expression of keratocan gene but increased ALDH3A expression. SP increased expression of fibrotic markers, a-SMA and fibronectin. Moreover, collagen I, collagen III and collagen V genes were also up-regulated by SP. Expression of lumican was unaffected by SP. Furthermore, keratocytes treated with SP showed increased contractile abilities. Similar effects of SP were observed in the corneal fibrosis model. SP decreased keratocan, but increased ALDH3A1 gene expression. a-SMA, fibronectin, collagen I, collagen III and collagen V genes were up-regulated. Expression of lumican was unaffected. Contractile abilities of myofibroblasts increased upon SP treatment. In conclusion, SP is able to regulate keratocyte marker genes and to increase expression of various ECM genes and fibrotic markers in both keratocytes and myofibroblasts. This suggests that SP might promote fibrosis in human cornea.
|
|
