scholarly journals Sustained AMPK Activation and Proline Metabolism Play Critical Roles in the Survival of Matrix-Deprived Transformed Cells

2021 ◽  
Vol 9 ◽  
Author(s):  
Manipa Saha ◽  
Neha Deshpande ◽  
Abhinav Dubey ◽  
Debnath Pal ◽  
Hanudatta S. Atreya ◽  
...  
Keyword(s):  
Cancer Progression ◽  
Simian Virus 40 ◽  
Transformation Model ◽  
Transformed Cells ◽  
Ampk Activation ◽  
Proline Metabolism ◽  
Functional Link ◽  
Atp Levels ◽  
The Matrix

Attachment to the matrix is critical for the survival of adherent cells, whereas detachment triggers death by apoptosis. Therefore, solid tumors must acquire the ability to survive the stress of matrix-detachment to transit through circulation and seed metastases. Although a central role for energy metabolism in cancer progression is well established, what distinguishes its role in the cellular state of the matrix-deprived form compared to the matrix-attached form is not fully understood yet. Using an in vitro transformation model dependent on simian virus 40 (SV40) small t (ST) antigen for cellular survival and proliferation in matrix-deprived conditions, we demonstrate that 5′-adenosine monophosphate-activated protein kinase (AMPK) activity is elevated and sustained under matrix-deprived conditions in ST-expressing fibroblasts. Additionally, these cells display elevated energy (ATP) levels under matrix-deprived conditions in contrast to cells lacking ST expression. The elevated ATP levels are coupled to increased levels of proline in ST-expressing cells, as revealed by metabolomics studies. The AMPK-dependent upregulation of proline oxidase, an enzyme of proline degradation, is a key link for elevated ATP levels. This functional link is further established by proline supplementation concomitant with AMPK activation in matrix-deprived cells lacking ST antigen, yielding ATP and enhancing survival. Thus, our data establishes a key role for AMPK-dependent regulation of proline metabolism in mediating energy homeostasis and promoting survival of matrix-deprived cells. These findings identify key markers that distinguish the metabolic states of matrix-detached and matrix-attached transformed cells and have implications in developing novel therapeutic strategies for specifically targeting matrix-detached metastasizing cancer cells.

scholarly journals Signaling Pathways Regulating TC21-induced Tumorigenesis

2007 ◽  
Vol 282 (38) ◽  
pp. 27713-27720 ◽  
Author(s):  
Mete Erdogan ◽  
Ambra Pozzi ◽  
Neil Bhowmick ◽  
Harold L Moses ◽  
Roy Zent
Keyword(s):  
P38 Mapk ◽  
Cancer Progression ◽  
Transforming Growth Factor ◽  
Cellular Transformation ◽  
Tumor Formation ◽  
Transformed Cells ◽  
Growth Inhibitory ◽  
Phosphoinositide 3 Kinase

TC21(R-Ras2), a Ras-related GTPase with transforming potential similar to H-, K- and N-Ras, is implicated in the pathogenesis of human cancers. Transforming growth factor β (TGF-β), a cytokine that plays a significant role in modulating tumorigenesis, normally prevents uncontrolled cell proliferation but paradoxically induces proliferation in H-Ras-transformed cancer cells. Although TC21 activates some pathways that mediate cellular transformation by the classical Ras proteins, the mechanisms through which TC21 induces tumor formation and how TGF-β regulates TC21 transformed cells is not known. To better understand the role of TC21 in cancer progression, we overexpressed an activated G23V mutant of TC21 in a nontumorigenic murine mammary epithelial (EpH4) cell line. Mutant TC21-expressing cells were significantly more oncogenic than cells expressing activated G12V H-Ras both in vivo and in vitro. TC21-induced transformation and proliferation required activation of p38 MAPK, mTOR (the mammalian target of rapamycin), and phosphoinositide 3-kinase but not Akt/PKB. Transformation by TC21 rendered EpH4 cells insensitive to the growth inhibitory effects of TGF-β, and the soft agar growth of these cells was increased upon TGF-β stimulation. Despite losing responsiveness to TGF-β-mediated growth inhibition, both Smad-dependent and independent pathways remained intact in TC21-transformed cells. Thus, overexpression of active TC21 in EpH4 cells induces tumorigenicity through the phosphoinositide 3-kinase, p38 MAPK, and mTOR pathways, and these cells lose their sensitivity to the normal growth inhibitory role of TGF-β.

scholarly journals Long noncoding RNA GAS5 inhibits progression of colorectal cancer by interacting with and triggering YAP phosphorylation and degradation and is negatively regulated by the m6A reader YTHDF3

2019 ◽  
Vol 18 (1) ◽  
Author(s):  
Wen Ni ◽  
Su Yao ◽  
Yunxia Zhou ◽  
Yuanyuan Liu ◽  
Piao Huang ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Cancer Progression ◽  
Noncoding Rna ◽  
Function Analysis ◽  
Loss Of Function ◽  
Functional Link ◽  
Protein Levels ◽  
Lncrna Gas5

Abstract Background YAP activation is crucial for cancer development including colorectal cancer (CRC). Nevertheless, it remains unclear whether N6-Methyladenosine (m6A) modified transcripts of long noncoding RNAs (lncRNAs) can regulate YAP activation in cancer progression. We investigated the functional link between lncRNAs and the m6A modification in YAP signaling and CRC progression. Methods YAP interacting lncRNAs were screened by RIP-sequencing, RNA FISH and immunofluorescence co-staining assays. Interaction between YAP and lncRNA GAS5 was studied by biochemical methods. MeRIP-sequencing combined with lncRNA-sequencing were used to identify the m6A modified targets of YTHDF3 in CRC. Gain-of-function and Loss-of-function analysis were performed to measure the function of GAS5-YAP-YTHDF3 axis in CRC progression in vitro and in vivo. Results GAS5 directly interacts with WW domain of YAP to facilitate translocation of endogenous YAP from the nucleus to the cytoplasm and promotes phosphorylation and subsequently ubiquitin-mediated degradation of YAP to inhibit CRC progression in vitro and in vivo. Notably, we demonstrate the m6A reader YTHDF3 not only a novel target of YAP but also a key player in YAP signaling by facilitating m6A-modified lncRNA GAS5 degradation, which profile a new insight into CRC progression. Clinically, lncRNA GAS5 expressions is negatively correlated with YAP and YTHDF3 protein levels in tumors from CRC patients. Conclusions Our study uncovers a negative functional loop of lncRNA GAS5-YAP-YTHDF3 axis, and identifies a new mechanism for m6A-induced decay of GAS5 on YAP signaling in progression of CRC which may offer a promising approach for CRC treatment.

Post-translational regulation of the 54K cellular tumor antigen in normal and transformed cells

1981 ◽  
Vol 1 (2) ◽  
pp. 101-110
Author(s):  
M Oren ◽  
W Maltzman ◽  
A J Levine
Keyword(s):  
Tumor Antigen ◽  
Ribonucleic Acid ◽  
Translational Regulation ◽  
Simian Virus 40 ◽  
T Antigen ◽  
Transformed Cells ◽  
Permissive Temperature ◽  
3T3 Cells ◽  
Messenger Ribonucleic Acid

The 54K cellular tumor antigen has been translated in vitro, using messenger ribonucleic acids from simian virus 40 (SV40)-transformed cells or 3T3 cells. The in vitro 54K product could be immunoprecipitated with SV40 tumor serum and had a peptide map that was similar, but not identical, to the in vivo product. The levels of this 54K protein in SV3T3 cells were significantly higher than those detected in 3T3 cells (D. I. H. Linzer, W. Maltzman, and A. J. Levine, Virology 98:308-318, 1979). In spite of this, the levels of translatable 54K messenger ribonucleic acid from 3T3 and SV3T3 cells were roughly equivalent or often greater in 3T3 cells. Pulse-chase experiments with the 54K protein from 3T3 or SV3T3 cells demonstrated that this protein, once synthesized, was rapidly degraded in 3T3 cells but was extremely stable in SV3T3 cells. Similarly, in an SV40 tsA-transformed cell line, temperature sensitive for the SV40 T-antigen, the 54K protein was rapidly turned over at the nonpermissive temperature and stable at the permissive temperature, whereas the levels of translatable 54K messenger ribonucleic acid at each temperature were roughly equal. These results demonstrate a post-translational regulation of the 54K cellular tumor antigen and suggest that this control is mediated by the SV40 large T-antigen.

Tumorigenicity of cells transformed by Simian virus 40 and of hybrids between such cells and normal diploid cells

1979 ◽  
Vol 36 (1) ◽  
pp. 223-240
Author(s):  
C.J. Gee ◽  
H. Harris
Keyword(s):  
Mouse Embryo ◽  
Simian Virus 40 ◽  
T Antigen ◽  
Transformed Cells ◽  
Sv40 T Antigen ◽  
Malignant Phenotype ◽  
Embryo Cells ◽  
Diploid Cells

A number of newly isolated clonal cell lines derived from diploid mouse embryo cells transformed by SV40 were examined in vitro and in vivo. Although these lines showed the properties that define transformation in vitro, they were not tumorigenic for many passages after their initial isolation. Cells from tumours eventually produced by the SV40-transformed cells were fused with diploid mouse embryo cells. The hybrids formed were initially non-tumorigenic. This indicates that a normal diploid cell can suppress the malignant phenotype of a tumorigenic SV40-transformed cell. The hybrid cells did, however, express the SV40 T antigen and they nad a clearly transformed phenotype in vitro. It thus appears that neither the transformed phenotype nor the expression of the SV40 T antigen are enough to endow a cell with the ability to grow progressively in vivo. The relationship between the transformed phenotype and tumorigenicity was further studied by fusing malignant mouse melanoma cells with non-tumorigenic SV40-transformed cells. The hybrids expressed the transformed phenotype in vitro but unable to form tumours in vivo. The changes that occur in cells after transformation by SV40 do not apparently affect the ability of these cells to suppress the malignant phenotype of tumour cells.

scholarly journals Post-translational regulation of the 54K cellular tumor antigen in normal and transformed cells.

1981 ◽  
Vol 1 (2) ◽  
pp. 101-110 ◽  
Author(s):  
M Oren ◽  
W Maltzman ◽  
A J Levine
Keyword(s):  
Tumor Antigen ◽  
Ribonucleic Acid ◽  
Translational Regulation ◽  
Simian Virus 40 ◽  
T Antigen ◽  
Transformed Cells ◽  
Permissive Temperature ◽  
3T3 Cells ◽  
Messenger Ribonucleic Acid

The 54K cellular tumor antigen has been translated in vitro, using messenger ribonucleic acids from simian virus 40 (SV40)-transformed cells or 3T3 cells. The in vitro 54K product could be immunoprecipitated with SV40 tumor serum and had a peptide map that was similar, but not identical, to the in vivo product. The levels of this 54K protein in SV3T3 cells were significantly higher than those detected in 3T3 cells (D. I. H. Linzer, W. Maltzman, and A. J. Levine, Virology 98:308-318, 1979). In spite of this, the levels of translatable 54K messenger ribonucleic acid from 3T3 and SV3T3 cells were roughly equivalent or often greater in 3T3 cells. Pulse-chase experiments with the 54K protein from 3T3 or SV3T3 cells demonstrated that this protein, once synthesized, was rapidly degraded in 3T3 cells but was extremely stable in SV3T3 cells. Similarly, in an SV40 tsA-transformed cell line, temperature sensitive for the SV40 T-antigen, the 54K protein was rapidly turned over at the nonpermissive temperature and stable at the permissive temperature, whereas the levels of translatable 54K messenger ribonucleic acid at each temperature were roughly equal. These results demonstrate a post-translational regulation of the 54K cellular tumor antigen and suggest that this control is mediated by the SV40 large T-antigen.

scholarly journals β-Elemene Restrains PTEN mRNA Degradation to Restrain the Growth of Lung Cancer Cells via METTL3-Mediated N6 Methyladenosine Modification

2022 ◽  
Vol 2022 ◽  
pp. 1-11
Author(s):  
Yuxu Feng ◽  
Chenchen Li ◽  
Siwen Liu ◽  
Fei Yan ◽  
Yue Teng ◽  
...  
Keyword(s):  
Lung Cancer ◽  
Cancer Progression ◽  
Vital Role ◽  
Pten Expression ◽  
Rna Modification ◽  
Functional Link ◽  
Side Impacts ◽  
Pten Mrna

Lung cancer is one of the most fatal malignancies and the leading cause of cancer death worldwide. β-Elemene, a well-known anticancer drug, has drawn a great deal of attention from researchers attributed to its limited side impacts. N6-Methyladenosine (m6A) modification is the most common RNA modification and plays a vital role in the pathogenesis of multiple tumors. However, the functional link between β-elemene and the m6A modification in lung cancer development remains unexplored. In this study, we investigated whether m6A modification was responsible for the impacts of β-elemene on lung cancer. Firstly, outcomes suggested that β-elemene restrained the malignant behaviors of A549 together with H1299 cells. Thereafter, we observed that β-elemene markedly regulated METTL3, YTHDF1, and YTHDC1 among various m6A modulators. METTL3 was selected for further study because of its oncogenic function in lung cancer. RT-qRCR and western blot assays exhibited that the mRNA and protein expression levels of METTL3 were lessened by the administration of β-elemene. Mechanistically, β-elemene exerted the restrictive impacts on the cell growth of lung cancer in vivo and in vitro through targeting METTL3. More importantly, β-elemene contributed to the augmented PTEN expression via suppressing its m6A modification. To sum up, we provided strong clues that β-elemene promoted PTEN expression to retard lung cancer progression by the regulation of METTL3-mediated m6A modification.

scholarly journals An Organotypic Mammary Duct Model Capturing Distinct Events of DCIS Progression

2020 ◽  
Author(s):  
Jonathan Kulwatno ◽  
Xiangyu Gong ◽  
Rebecca DeVaux ◽  
Jason I. Herschkowitz ◽  
Kristen Lynn Mills
Keyword(s):  
Breast Cancer ◽  
Cell Lines ◽  
Cancer Progression ◽  
Ductal Carcinoma ◽  
Biomechanical Properties ◽  
Breast Cancer Progression ◽  
Surrounding Tissue ◽  
Mammary Duct ◽  
The Matrix

ABSTRACTDuctal carcinoma in situ (DCIS) is a pre-cancerous stage breast cancer, where abnormal cells are contained within the duct, but have not invaded into the surrounding tissue. However, only 30-40% of DCIS cases are likely to progress into an invasive ductal carcinoma (IDC), while the remainder are innocuous. Since little is known about what contributes to the transition from DCIS to IDC, clinicians and patients tend to opt for treatment, leading to concerns of overdiagnosis and overtreatment. In vitro models are currently being used to probe how DCIS transitions into IDC, but many models do not take into consideration the macroscopic tissue architecture and the biomechanical properties of the microenvironment. Here, we developed an organotypic mammary duct model by molding a channel within a collagen matrix and lining it with a basement membrane. By adjusting the concentration of collagen, we effectively modulated the stiffness and morphological properties of the matrix and examined how an assortment of breast cells responded to changing density and stiffness of the matrix. We first validated the model using two established, phenotypically divergent breast cancer cell lines by demonstrating the ability of the cells to either invade (MDA-MB-231) or cluster (MCF7). We then examined how cells of the isogenic MCF10 series—spanning the range from healthy to aggressive—behaved within our model and observed distinct characteristics of breast cancer progression such as hyperplasia and invasion, in response to collagen concentration. Our results show that the model can recapitulate different stages of breast cancer progression and that the MCF10 series is adaptable to physiologically relevant in vitro studies, demonstrating the potential of both the model and cell lines to elucidate key factors that may contribute to understanding the transition from DCIS to IDC.IMPACT STATEMENTThe success of early preventative measures for breast cancer has left patients susceptible to overdiagnosis and overtreatment. Limited knowledge of factors driving an invasive transition has inspired the development of in vitro models that accurately capture this phenomenon. However, current models tend to neglect the macroscopic architecture and biomechanical properties of the mammary duct. Here, we introduce an organotypic model that recapitulates the cylindrical geometry of the tissue and the altered stroma seen in tumor microenvironments. Our model was able to capture distinct features associated with breast cancer progression, demonstrating its potential to uncover novel insights into disease progression.

Radiolabeled r-Hirudin as a Measure of Thrombin Activity at, or within, the Rabbit Aorta Wall In Vitro and In Vivo

1994 ◽  
Vol 71 (04) ◽  
pp. 499-506 ◽  
Author(s):  
Mark W C Hatton ◽  
Bonnie Ross-Ouellet
Keyword(s):  
Rabbit Aorta ◽  
Balloon Injury ◽  
Recombinant Hirudin ◽  
Aorta Wall ◽  
Thrombin Activity ◽  
Before And After ◽  
The Matrix

SummaryThe behavior of 125I-labeled recombinant hirudin towards the uninjured and de-endothelialized rabbit aorta wall has been studied in vitro and in vivo to determine its usefulness as an indicator of thrombin activity associated with the aorta wall. Thrombin adsorbed to either sulfopropyl-Sephadex or heparin-Sepharose bound >95% of 125I-r-hirudin and the complex remained bound to the matrix. Binding of 125I-r-hirudin to the exposed aorta subendothelium (intima-media) in vitro was increased substantially if the tissue was pre-treated with thrombin; the quantity of l25I-r-hirudin bound to the de-endothelialized intima-media (i.e. balloon-injured in vitro) correlated positively with the quantity of bound 131I-thrombin (p <0.01). Aortas balloon-injured in vivo were measured for thrombin release from, and binding of 125I-r-hirudin to, the de-endothelialized intimal surface in vitro; 125I-r-hirudin binding correlated with the amount of active thrombin released (p <0.001). Uptake of 125I-r-hirudin by the aorta wall in vivo was proportional to the uptake of 131I-fibrinogen (as an indicator of thrombin activity) before and after balloon injury. After 30 min in the circulation, specific 125I-r-hirudin binding to the uninjured and de-endo- thelialized (at 1.5 h after injury) aorta wall was equivalent to 3.4 (± 2.5) and 25.6 (±18.1) fmol of thrombin/cm2 of intima-media, respectively. Possibly, only hirudin-accessible, glycosaminoglycan-bound thrombin is measured in this way.

Formulation and Evaluation of Galantamine Hydrobromide Floating Matrix Tablet

2018 ◽  
Vol 8 (2) ◽  
Author(s):  
Poreddy Srikanth Reddy ◽  
Penjuri Subhash Chandra Bose ◽  
Vuppula Sruthi ◽  
Damineni Saritha
Keyword(s):  
Sodium Alginate ◽  
Xanthan Gum ◽  
Lag Time ◽  
Matrix Tablets ◽  
In Vitro Dissolution ◽  
Matrix Tablet ◽  
Compression Technique ◽  
Weight Variation ◽  
The Matrix

The aim of the present work was to prepare floating tablets of galantamine HBr using sodium alginate and xanthan gum as matrix forming carriers. Galantamine HBr is used for the treatment of mild to moderate Alzheimer's disease and various other memory impairments, in particular those of vascular origin. The matrix tablet formulations were prepared by varying the concentrations of sodium alginate and xanthan gum. The tablets were prepared by direct compression technique using PVP K-30 as a binder and sodium bicarbonate for development of CO2. The prepared matrix tablets were evaluated for properties such as hardness, thickness, friability, weight variation, floating lag time, compatibility using DSC and FTIR. In vitro dissolution was carried out for 12 hrs in 0.1N HCl at 37±0.5 ºC using USP paddle type dissolution apparatus. It was noted that, all the prepared formulations had desired floating lag time and constantly floated on dissolution medium by maintaining the matrix integrity. The drug release from prepared tablets was found to vary with varying concentration of the polymers, sodium alginate and xanthan gum. From the study it was concluded that floating drug delivery system for galantamine HBr can be prepared by using sodium alginate and xanthan gum as a carrier.

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