Myofibroblast Differentiation of Normal Human Keratocytes and hTERT, Extended-Life Human Corneal Fibroblasts

2003 ◽  
Vol 44 (5) ◽  
pp. 1850 ◽  
Author(s):  
James V. Jester ◽  
Jiying Huang ◽  
Stephen Fisher ◽  
Jennifer Spiekerman ◽  
Jin Ho Chang ◽  
...  
Keyword(s):  
Myofibroblast Differentiation ◽  
Corneal Fibroblasts ◽  
Normal Human ◽  
Extended Life ◽  
Human Keratocytes

scholarly journals FAK Inhibition Attenuates Corneal Fibroblast Differentiation In Vitro

Biomolecules ◽  
2021 ◽  
Vol 11 (11) ◽  
pp. 1682
Author(s):  
Vincent Yeung ◽  
Sriniwas Sriram ◽  
Jennifer A. Tran ◽  
Xiaoqing Guo ◽  
Audrey E. K. Hutcheon ◽  
...  
Keyword(s):  
Wound Healing ◽  
Transforming Growth Factor ◽  
Smooth Muscle Actin ◽  
Myofibroblast Differentiation ◽  
Corneal Scarring ◽  
Corneal Fibroblast ◽  
Corneal Fibroblasts ◽  
Corneal Wound ◽  
Tgf Β1

Corneal fibrosis (or scarring) occurs in response to ocular trauma or infection, and by reducing corneal transparency, it can lead to visual impairment and blindness. Studies highlight important roles for transforming growth factor (TGF)-β1 and -β3 as modulators in corneal wound healing and fibrosis, leading to increased extracellular matrix (ECM) components and expression of α-smooth muscle actin (αSMA), a myofibroblast marker. In this study, human corneal fibroblasts (hCF) were cultured as a monolayer culture (2D) or on poly-transwell membranes to generate corneal stromal constructs (3D) that were treated with TGF-β1, TGF-β3, or TGF-β1 + FAK inhibitor (FAKi). Results show that hCF 3D constructs treated with TGF-β1 or TGF-β3 impart distinct effects on genes involved in wound healing and fibrosis—ITGAV, ITGB1, SRC and ACTA2. Notably, in the 3D construct model, TGF-β1 enhanced αSMA and focal adhesion kinase (FAK) protein expression, whereas TGF-β3 did not. In addition, in both the hCF 2D cell and 3D construct models, we found that TGF-β1 + FAKi attenuated TGF-β1-mediated myofibroblast differentiation, as shown by abrogated αSMA expression. This study concludes that FAK signaling is important for the onset of TGF-β1-mediated myofibroblast differentiation, and FAK inhibition may provide a novel beneficial therapeutic avenue to reduce corneal scarring.

scholarly journals Nitric oxide attenuated transforming growth factor-β induced myofibroblast differentiation of human keratocytes

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Joo-Hee Park ◽  
Martha Kim ◽  
Bora Yim ◽  
Choul Yong Park
Keyword(s):  
Nitric Oxide ◽  
Growth Factor ◽  
Sodium Nitrite ◽  
Transforming Growth Factor ◽  
Smooth Muscle Actin ◽  
Corneal Opacity ◽  
Myofibroblast Differentiation ◽  
No Donor ◽  
Tgf Β1 ◽  
Human Keratocytes

AbstractNitric oxide (NO) has the potential to modulate myofibroblast differentiation. In this study, we investigated the effect of exogenous NO on the myofibroblast differentiation of human keratocytes using sodium nitrite as a NO donor. Myofibroblasts were induced by exposing resting keratocytes to transforming growth factor (TGF)-β1. N-cadherin and α-smooth muscle actin (αSMA) were used as myofibroblast markers. Both resting keratocytes and -stimulated keratocytes were exposed to various concentrations of sodium nitrite (1 μM to 1000 mM) for 24 to 72 h. Exposure to sodium nitrite did not alter keratocytes’ viability up to a 10 mM concentration for 72 h. However, significant cytotoxicity was observed in higher concentrations of sodium nitrite (over 100 mM). The expression of αSMA and N-cadherin was significantly increased in keratocytes by TGF-β1 stimulation after 72 h incubation. The addition of sodium nitrite (1 mM) to TGF-β1-stimulated keratocytes significantly decreased αSMA and N cadherin expression. Smad3 phosphorylation decreased after sodium nitrite (1 mM) exposure in TGF-β1-stimulated keratocytes. The effect of NO was reversed when NO scavenger, 2-4-carboxyphenyl-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (cPTIO) was added in the culture medium. Application of sodium nitrite resulted in significant decrease of corneal opacity when measured at 2 weeks after the chemical burn in the mouse. These results verified the potential therapeutic effect of NO to decrease myofibroblast differentiation of human keratocytes and corneal opacity after injury.

scholarly journals TGF-β1 stimulates HDAC4 nucleus-to-cytoplasm translocation and NADPH oxidase 4-derived reactive oxygen species in normal human lung fibroblasts

2017 ◽  
Vol 312 (6) ◽  
pp. L936-L944 ◽  
Author(s):  
Weichao Guo ◽  
Shigeki Saito ◽  
Cecilia G. Sanchez ◽  
Yan Zhuang ◽  
Rafael E. Gongora Rosero ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Nuclear Export ◽  
Human Lung ◽  
Fiber Formation ◽  
Lung Fibroblasts ◽  
Myofibroblast Differentiation ◽  
Oxygen Species ◽  
Human Lung Fibroblasts ◽  
Nadph Oxidase 4 ◽  
Normal Human

Myofibroblasts are important mediators of fibrogenesis; thus blocking fibroblast-to-myofibroblast differentiation (FMD) may be an effective strategy to treat pulmonary fibrosis (PF). Previously, we reported that histone deacetylase 4 (HDAC4) activity is necessary for transforming growth factor-β1 (TGF-β1)-induced human lung FMD. Here, we show that TGF-β1 increases NADPH oxidase 4 (NOX4) mRNA and protein expression in normal human lung fibroblasts (NHLFs) and causes nuclear export of HDAC4. Application of the NOX family inhibitor diphenyleneiodonium chloride reduces TGF-β1-induced HDAC4 nuclear export, expression of the myofibroblast marker α-smooth muscle actin (α-SMA), and α-SMA fiber formation. Inhibition of HDAC4 nucleus-to-cytoplasm translocation using leptomycin B (LMB) had little effect on α-SMA expression but blocked α-SMA fiber formation. A coimmunoprecipitation assay showed that HDAC4 associates with α-SMA. Moreover, LMB abolishes TGF-β1-induced α-SMA fiber formation and cell contraction. Relevant to human pulmonary fibrosis, idiopathic PF specimens showed significantly higher NOX4 RNA expression and scant HDAC4 staining within nuclei of fibroblast foci myofibroblasts. Taken together, these results indicate that reactive oxygen species promote TGF-β1-mediated myofibroblast differentiation and HDAC4 nuclear export. The physical association of HDAC4 with α-SMA suggests that HDAC4 has a role in regulating the α-SMA cytoskeleton arrangement.

CD-34 Expression by Cultured Human Keratocytes Is Downregulated during Myofibroblast Differentiation Induced by TGF-β1

2004 ◽  
Vol 45 (9) ◽  
pp. 2985 ◽  
Author(s):  
Edgar M. Espana ◽  
Tetsuya Kawakita ◽  
Chia-Yang Liu ◽  
Scheffer C. G. Tseng
Keyword(s):  
Myofibroblast Differentiation ◽  
Cd 34 ◽  
Tgf Β1 ◽  
Human Keratocytes

scholarly journals Analysis of Genomic Integrity and p53-Dependent G1Checkpoint in Telomerase-Induced Extended-Life-Span Human Fibroblasts

1999 ◽  
Vol 19 (3) ◽  
pp. 2373-2379 ◽  
Author(s):  
Homayoun Vaziri ◽  
Jeremy A. Squire ◽  
Tej K. Pandita ◽  
Grace Bradley ◽  
Robert M. Kuba ◽  
...  
Keyword(s):  
Ionizing Radiation ◽  
Life Span ◽  
P53 Protein ◽  
Human Fibroblasts ◽  
Strand Break ◽  
Human Cells ◽  
Normal Human ◽  
Normal Human Cells ◽  
Extended Life

ABSTRACT Life span determination in normal human cells may be regulated by nucleoprotein structures called telomeres, the physical ends of eukaryotic chromosomes. Telomeres have been shown to be essential for chromosome stability and function and to shorten with each cell division in normal human cells in culture and with age in vivo. Reversal of telomere shortening by the forced expression of telomerase in normal cells has been shown to elongate telomeres and extend the replicative life span (H. Vaziri and S. Benchimol, Curr. Biol. 8:279–282, 1998; A. G. Bodnar et al., Science 279:349–352, 1998). Extension of the life span as a consequence of the functional inactivation of p53 is frequently associated with loss of genomic stability. Analysis of telomerase-induced extended-life-span fibroblast (TIELF) cells by G banding and spectral karyotyping indicated that forced extension of the life span by telomerase led to the transient formation of aberrant structures, which were subsequently resolved in higher passages. However, the p53-dependent G1 checkpoint was intact as assessed by functional activation of p53 protein in response to ionizing radiation and subsequent p53-mediated induction of p21Waf1/Cip1/Sdi1. TIELF cells were not tumorigenic and had a normal DNA strand break rejoining activity and normal radiosensitivity in response to ionizing radiation.

scholarly journals Autophagy is deregulated in cancer-associated fibroblasts from oral cancer and is stimulated during the induction of fibroblast senescence by TGF-β1

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
May Leng Tan ◽  
E. Kenneth Parkinson ◽  
Lee Fah Yap ◽  
Ian C. Paterson
Keyword(s):  
Cell Migration ◽  
Oral Cancer ◽  
Squamous Cell ◽  
Functional Role ◽  
Squamous Cell Carcinomas ◽  
Myofibroblast Differentiation ◽  
Cancer Associated Fibroblasts ◽  
Fibroblast Activation ◽  
Normal Human ◽  
Tgf Β1

AbstractMany of the characteristics ascribed to cancer-associated fibroblasts (CAFs) are shared by activated, autophagic and senescent fibroblasts. Whilst most oral squamous cell carcinomas (OSCCs) are genetically unstable (GU-OSCC), genetically stable variants (GS-OSCC) have been described and, notably, CAF activation (myofibroblast differentiation) and senescence are characteristics particularly associated with GU-OSCCs. However, it is not known whether autophagy is disrupted in these cells or whether autophagy regulates the development of the myofibroblast and senescent phenotypes. In this study, we show that senescent CAFs from GU-OSCCs contained more autophagosomes than normal human oral fibroblasts (NHOFs) and CAFs from GS-OSCCs possibly due to autophagic impairment. Further, we show that deregulation of autophagy in normal fibroblasts, either by inhibition with autophagy inhibitor, SAR405, or activation with TGF-β1, induced fibroblast activation and senescence: In response to TGF-β1, autophagy was induced prior to the development of the activated and senescent phenotypes. Lastly, we show that both SAR405- and TGF-β1-treated NHOFs enhance OSCC cell migration but only TGF-β1-treated cells increase OSCC invasion through Matrigel, indicating that TGF-β1 has additional effects that are independent of fibroblast activation/senescence. These results suggest a functional role for autophagy in the development of myofibroblast and CAF phenotypes.

scholarly journals Glucosamine impedes transforming growth factor β1-mediated corneal fibroblast differentiation by targeting Krüppel-like factor 4

2019 ◽  
Vol 26 (1) ◽  
Author(s):  
Ying-Jen Chen ◽  
Shih-Ming Huang ◽  
Ming-Cheng Tai ◽  
Jiann-Torng Chen ◽  
Chang-Min Liang
Keyword(s):  
Growth Factor ◽  
Transforming Growth Factor ◽  
Smooth Muscle Actin ◽  
Disease Model ◽  
Immunoblot Analysis ◽  
Transforming Growth Factor Β1 ◽  
Myofibroblast Differentiation ◽  
Alpha Smooth Muscle Actin ◽  
Corneal Fibroblasts ◽  
Tgf Β1

Abstract Background Transforming growth factor (TGF) family members play important roles in the regulation of corneal integrity, and the pathogenesis of corneal fibrosis. Currently, there are no effective agents targeting TGF-β signaling to diminish corneal fibrosis. Glucosamine (GlcN), which is widely used in the treatment of osteoarthritis, abrogates the morphologic effects of TGF-β2 on retinal pigmented epithelial cells in a mouse disease model. Here, we sought to determine whether GlcN would exert beneficial effects against TGF-β1-induced corneal fibrosis. Methods In human corneal fibroblasts (HCFs) treated with GlcN, the expression of Krüppel-like factor 4 (KLF4) and its downstream signaling effects were determined in the presence and absence of TGF-β1 using immunoblot analysis. We further explored GlcN inhibition of fibroblast-to-myofibroblast differentiation via KLF4 siRNA. The effect of cycloheximide on KLF4 protein levels with or without GlcN administration was assessed to determine whether GlcN affects the stability of the KLF4 protein. Results In HCFs, GlcN induced the expression of KLF4, which regulated the maturation and maintenance of the ocular surface. GlcN partially suppressed the TGF-β1-induced expression of alpha-smooth muscle actin (α-SMA) and reduced the collagen contraction capacity in HCFs, suggesting a decrease in fibroblast-to-myofibroblast differentiation. This effect appeared to be mediated through suppression of Smad2 phosphorylation and ERK-dependent signaling. The levels of KLF4 mRNA were increased by GlcN and decreased by TGF-β1 and the TGF-β1-induced α-SMA mRNA expression was upregulated when the KLF4 gene was silenced. GlcN also appeared to stabilize the KLF4 protein, reducing its turnover in corneal fibroblasts. Conclusion These findings shed light on a novel mechanism by which GlcN suppresses TGF-β1-induced fibroblast-to-myofibroblast differentiation through the upregulation of KLF4 expression. Current strategies for treating corneal fibrosis were not effective. Elevating KLF4 levels through the use of GlcN might provide an effective alternative to alleviate the development and progression of corneal fibrosis.

Ultrastructural modification of cellular interactions in cancer tumor

1978 ◽  
Vol 36 (2) ◽  
pp. 318-319
Author(s):  
N. P. Dmitrieva
Keyword(s):  
Cancer Cells ◽  
Human Thyroid ◽  
Adjacent Cell ◽  
Main Role ◽  
Cellular Interactions ◽  
Electron Microscopic ◽  
Cancer Tumor ◽  
Normal Human ◽  
Characteristic Features

One of the most characteristic features of cancer cells is their ability to metastasia. It is suggested that the modifications of the structure and properties of cancer cells surfaces play the main role in this process. The present work was aimed at finding out what ultrastructural features apear in tumor in vivo which removal of individual cancer cells from the cell population can provide. For this purpose the cellular interactions in the normal human thyroid and cancer tumor of this gland electron microscopic were studied. The tissues were fixed in osmium tetroxide and were embedded in Araldite-Epon.In normal human thyroid the most common type of intercellular contacts was represented by simple junction formed by the parallelalignment of adjacent cell membranees leaving in between an intermembranes space 15-20 nm filled with electronlucid material (Fig. 1a). Sometimes in the basal part of cells dilatations of the intercellular space 40-50 nm wide were found (Fig. 1a). Here the cell surfaces may form single short microvilli.

Scanning Microscopy of Human Intestinal Explants in Organ Culture

1972 ◽  
Vol 30 ◽  
pp. 396-397
Author(s):  
Bruce Wetzel ◽  
Robert Buscho ◽  
Raphael Dolin
Keyword(s):  
Electron Microscopy ◽  
Room Temperature ◽  
Culture Conditions ◽  
Human Fetuses ◽  
Enteric Disease ◽  
Organ Cultures ◽  
Growth Of A Variety ◽  
Normal Human ◽  
Fetal Intestine ◽  
Scanning Electron

It has been reported that explants of human fetal intestine can be maintained in culture for up to 21 days in a viable condition and that these organ cultures support the growth of a variety of known viral agents responsible for enteric disease. Scanning electron microscopy (SEM) has been undertaken on several series of these explants to determine their appearance under routine culture conditions.Fresh specimens of jejunum obtained from normal human fetuses were washed, dissected into l-4mm pieces, and cultured in modified Leibowitz L-15 medium at 34° C as previously described. Serial specimens were fixed each day in 3% glutaraldehyde for 90 minutes at room temperature, rinsed, dehydrated, and dried by the CO2 critical point method in a Denton DCP-1 device. Specimens were attached to aluminum stubs with 3M transfer tape No. 465, and one sample on each stub was carefully rolled along the adhesive such that villi were broken off to expose their interiors.

Sign in / Sign up

Export Citation Format

Share Document