scholarly journals Inhibition of the Ras oncoprotein reduces proliferation of hepatocytes in vitro and in vivo in rats

2007 ◽  
Vol 114 (1) ◽  
pp. 73-83 ◽  
Author(s):  
Alain da Silva Morais ◽  
Alain Saliez ◽  
Isabelle Leclercq ◽  
Yves Horsmans ◽  
Peter Stärkel
Keyword(s):  
Cell Proliferation ◽  
Hepg2 Cells ◽  
Cyclin E ◽  
Hepatocyte Proliferation ◽  
Brdu Incorporation ◽  
Cyclin D ◽  
Ras Inhibition ◽  
The Impact

Ras oncoproteins are probably implicated in normal and malignant cell growth in various organs. Inhibition of Ras interferes with cell proliferation of non-hepatic cells in vitro and in vivo. A potential role for Ras in normal and malignant hepatocyte proliferation prompted us to evaluate the impact of Ras inhibition by FTS (S-farnesylthiosalicylic acid) on hepatocyte proliferation in vitro in the human hepatic tumour cell line HepG2 and in vivo after PH (partial hepatectomy) in rats. Rats were administered with FTS intraperitoneally (1, 8 and 16 h after PH) and killed 12, 24 and 48 h after PH. Cell proliferation, phosphorlyation of members of the MAPK (mitogen-activated protein kinase) pathway and levels and activity of cell cycle effectors (cyclin D, cyclin E, Cdk2 and Cdk4) were assessed in FTS-treated rats compared with controls. FTS significantly decreased overall cell count, PCNA (proliferating-cell nuclear antigen) expression and BrdU (bromodeoxyuridine) incorporation into HepG2 cells after 7 days of culture. FTS treatment significantly reduced BrdU incorporation and PCNA expression in hepatocytes after PH. Unlike control rats, cell-membrane expression of Ras was decreased in FTS-treated animals after PH, resulting in decreased Raf membrane recruitment and phosphorylation and in reduced phosphorylation of ERK1/2 (extracellular-signal-regulated kinase 1/2). The antiproliferative effect of FTS was linked to a decrease in expression and activity of the cyclin E/Cdk2 complex, without affecting cyclin D and Cdk4. Ras inhibition by FTS significantly decreased proliferation of HepG2 cells and normal hepatocytes after a strong and highly synchronized proliferation stimulus elicited by PH. The inhibitory effect was at least partially mediated by inhibition of Ras/Raf/MAPK signalling. It appears worthwhile to evaluate the impact of Ras inhibition on the development of hepatocarcinomas in vivo in adequate animal models.

Amplified LncRNA PVT1 promotes lung cancer proliferation and metastasis by facilitating VEGFC expression

2020 ◽  
Vol 98 (6) ◽  
pp. 676-682
Author(s):  
Yanming Pan ◽  
Lantao Liu ◽  
Yongxia Cheng ◽  
Jianbo Yu ◽  
Yukuan Feng
Keyword(s):  
Lung Cancer ◽  
Cell Proliferation ◽  
Tumor Model ◽  
Cancer Proliferation ◽  
Survival Prognosis ◽  
Proliferation And Metastasis ◽  
Factor C ◽  
The Impact

Although the abundance of long non-coding RNA (lncRNA) plasmacytoma variant translocation 1 (PVT1) in lung cancer has been well researched, the underlying mechanisms behind its effects were unknown. Here we investigated the molecular events regulating PVT1 in lung cancer. The pro-proliferative property of PVT1 was examined using a xenograft tumor model. Transwell chambers were used to analyze the impact of PVT1 expression on cell invasiveness and migration. In vivo metastasis was examined by tail-vein-injection in mice. Direct binding of miR-128 to PVT1 was investigated using a probe pulldown assay. The relative expression levels of miR-128 and PVT1 were quantified by real-time polymerase chain reaction and Western blotting. We show here that when PVT1 is amplified, there is a poor survival prognosis for patients with lung cancer. Elevated levels of PVT1 promoted lung cancer cell proliferation and metastasis, both in vitro and in vivo. Mechanistically, we found that PVT1 competes endogenously with miR-128 in the regulation of vascular endothelial growth factor C (VEGFC) expression, which is significantly associated with an unfavorable prognosis in lung cancer. We identified that copy number amplification significantly contributes to the high level of PVT1 transcripts in lung cancer, which promotes cell proliferation and metastatic behavior via modulating VEGFC expression by endogenous competition with miR-128.

scholarly journals Sigma 1 Receptor is Overexpressed in Hepatocellular Adenoma: Involvement of ERα and HNF1α

Cancers ◽  
2020 ◽  
Vol 12 (8) ◽  
pp. 2213
Author(s):  
Laure Villemain ◽  
Sylvie Prigent ◽  
Aurélie Abou-Lovergne ◽  
Laura Pelletier ◽  
Magali Chiral ◽  
...  
Keyword(s):  
Knockout Mice ◽  
Hepatocellular Adenoma ◽  
Integral Membrane Protein ◽  
Hepatocyte Proliferation ◽  
Sigma 1 Receptor ◽  
Cellular Models ◽  
Huh7 Cells ◽  
The Impact

Sigma receptor 1 (SigR1) is an endoplasmic reticulum resident integral membrane protein whose functions remain unclear. Although the liver shows the highest expression of SigR1, its role in this organ is unknown. SigR1 is overexpressed in many cancers and its expression is correlated to hormonal status in hormone-dependent cancers. To better understand the role of SigR1 in hepatocytes we focused our work on the regulation of its expression in tumoral liver. In this context, hepatocellular adenomas, benign hepatic tumors associated with estrogen intake are of particular interest. The expression of SigR1 mRNA was assessed in hepatocellular adenoma (HCA) patients using qPCR. The impact of estrogen on the expression of SigR1 was studied in vivo (mice) and in vitro (HepG2 and Huh7 cells). The effect of HNF1α on the expression of SigR1 was studied in vivo by comparing wild type mice to HNF1 knockout mice. Estrogen enhanced SigR1 expression through its nuclear receptor ERα. HNF1α mutated HCA (H-HCA) significantly overexpressed SigR1 compared to all other HCA subtypes. HNF1 knockout mice showed an increase in SigR1 expression. Overexpressing SigR1 in cellular models increases proliferation rate and storage of lipid droplets, which phenocopies the H-HCA phenotype. SigR1 is involved in hepatocyte proliferation and steatosis and may play an important role in the control of the H-HCA phenotype.

scholarly journals Trilobatin, a Novel SGLT1/2 Inhibitor, Selectively Induces the Proliferation of Human Hepatoblastoma Cells

Molecules ◽  
2019 ◽  
Vol 24 (18) ◽  
pp. 3390 ◽  
Author(s):  
Lujing Wang ◽  
Min Liu ◽  
Fei Yin ◽  
Yuanqiang Wang ◽  
Xingan Li ◽  
...  
Keyword(s):  
Hepg2 Cells ◽  
S Phase ◽  
Hepatocyte Proliferation ◽  
Hepatocyte Nuclear Factor ◽  
Interacting Protein ◽  
Glucose Lowering ◽  
Anti Cancer ◽  
High Concentrations

Studies have indicated that Na+-d-glucose co-transporter (SGLT) inhibitors had anti-proliferative activity by attenuating the uptake of glucose in several tumor cell lines. In this study, the molecular docking showed that, trilobatin, one of the dihydrochalcones from leaves of Lithocarpus polystachyus Rehd., might be a novel inhibitor of SGLT1 and SGLT2, which evidently attenuated the uptake of glucose in vitro and in vivo. To our surprise, we observed that trilobatin did not inhibit, but promoted the proliferation of human hepatoblastoma HepG2 and Huh 7 cells when it was present at high concentrations. At the same time, incubation with high concentrations of trilobatin arrested the cell cycle at S phase in HepG2 cells. We also found that treatment with trilobatin had no significant effect on the expression of hepatitis B x-interacting protein (HBXIP) and hepatocyte nuclear factor (HNF)-4α, the two key regulators of hepatocyte proliferation. Taken together, although trilobatin worked as a novel inhibitor of SGLTs to attenuate the uptake of glucose, it also selectively induced the cell proliferation of HepG2 cells, suggesting that not all the SGLT inhibitors inhibited the proliferation of tumor cells, and further studies are needed to assess the anti-cancer potentials of new glucose-lowering agents.

Deterministic Trigger of Self-Renewal in Expanded HSCs Expressing Hoxb4 + Antisense Pbx1.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 800-800
Author(s):  
Sonia Cellot ◽  
Jana Krosl ◽  
Keith Humphries ◽  
Guy Sauvageau
Keyword(s):  
Cell Proliferation ◽  
Stem Cell ◽  
Cell Fate ◽  
Time Course ◽  
Cell Cycle Distribution ◽  
Self Renewal ◽  
Primary Mouse ◽  
The Impact

Abstract We previously reported the generation of pluripotent and ultracompetitive HSCs through modulation of Hoxb4 and Pbx1 levels. These Hoxb4hiPbx1lo HSCs display a tremendous regenerative potential, yet they are still fully responsive to in vivo regulatory signals that control stem cell pool size (20 000 HSCmouse) and differentiation pathways. Further work in our laboratory attempted to circumvent these physiological constraints by expanding Hoxb4hiPbx1lo transduced HSCs in vitro, and hence revealing their intrinsic expansion potential. Independent experiments were performed where primary mouse BM cells were co-infected with retroviruses encoding antisense Pbx1 cDNA plus YFP, and Hoxb4 plus GFP (double gene transfer ranged between 20–50%). Hoxb4hiPbx1lo HSCs measured using the CRU assay expanded by 105-fold during a 12 day in vitro culture. Following serial transplantations, these cells displayed an additional 4–5 log expansion in vivo. Total stem cell content per animal remained within normal limits. Southern blot analyses of proviral integrations showed that the expansion was polyclonal, and analyses of individually expanded clones provided a molecular proof of in vitro self-renewal (SR). This unprecedented level of HSC expansion in such a short time course (105-fold in 12 days) implies an absolute HSC doubling time of approximately 17 hours in our culture, raising the possibility that virtually all dividing HSCs undergo self-renewal. This analysis prompted us to dissect the impact of Hoxb4 on cell proliferation versus cell fate (SR?). When analyzed during the period of maximal HSC expansion, the cell cycle distribution of Sca+ or Sca+Lin− cells were comparable between the cultures initiated with neo control versus Hoxb4 BM cells (CTL vs Hoxb4: G0/G1: 66% vs 83%; S: 15% vs 9%; G2/M: 18% vs 7%). Correspondingly, CFSE tracking studies confirmed the identical, or even lower, number of cellular divisions in Sca+ cells isolated from cultures initiated with Hoxb4 versus neo transduced cells. Annexin V studies precluded protection from apoptosis as the major mechanism to increase HSC numbers since similar results (3–10% positive cells) were observed in the Hoxb4 versus neo-transduced cells. In summary, our studies support the emerging concept that distinct molecular pathways regulate cell proliferation and self-renewal, suggesting that Hoxb4 + antisense Pbx1 predominantly triggers self-renewal over HSC proliferation.

Cellular Senescence Contributes to the Outcome of Lymphoma Chemotherapy by Cell-Autonomous and Paracrine Immune Mechanisms

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 3617-3617
Author(s):  
Jan Dörr ◽  
Yong Yu ◽  
Bernd Dörken ◽  
Clemens A. Schmitt
Keyword(s):  
Dna Damage ◽  
Immune Cell ◽  
Genetic Alterations ◽  
Cell Interactions ◽  
Brdu Incorporation ◽  
Tumor Site ◽  
Immune Cell Subsets ◽  
The Impact

Abstract Introduction: Premature senescence reflects an acutely inducible, irreversible growth arrest as a cellular response to stresses such as oncogenic activation and DNA damage, including chemotherapeutic anticancer agents. Senescence complements apoptosis as a tumor suppressive and therapeutic effector principle, but whether a selective disruption of the senescence machinery impairs treatment outcome is unknown. Moreover, function and fate of senescent tumor cells within the tumor site remain unclear. Here, we analyze the impact of defined genetic alterations, i.e. Bcl2 overexpression (blocking apoptosis), deletion of the histone H3 lysine 9 methyltransferase Suv39h1 (controlling senescence), and conditional expression of p53 (mediating both apoptosis and senescence), on therapy-induced senescence (TIS) in the Eμ-myc mouse lymphoma model with specific emphasis on immunological tumor-host and growth-modulating senescent/non-senescent cell interactions as a consequence of TIS in vitro and in vivo. Methods: Lymphoma cells (LCs) of various genetic backgrounds were retrovirally transduced with the bcl2 gene to study TIS in the absence of drug-induced apoptosis. Bcl2-protected LCs were treated with the DNA damaging anticancer agent adriamycin in vitro, or were exposed to the alkylating agent cyclophosphamide upon lymphoma formation in normal immunocompetent mice in vivo. TIS was detected by staining for senescence-associated β-galactosidase activity (SA-β-gal) and other senescence-related markers, including Ki67 and BrdU incorporation. To study tumor-host cell interactions, isolated normal splenocytes were co-incubated with proliferating or senescent LCs in vitro. Immunophenotyping was carried out with antibodies specific for macrophages, granulocytes, natural killer cells and T-lymphocytes. Cytokine production was measured by protein arrays. Results: Senescent LCs engage in cell-cell interactions with different immune cell subsets, in particular macrophages, granulocytes and T-cells in vitro. Fluorescence microscopy reveals that macrophages engulf LCs after they entered TIS. In vivo, TIS correlates with the quantitative attraction of immune cell populations to the tumor site and subsequent clearing of senescent cells. Ongoing mechanistic studies on underlying ligand/receptor interactions will be reported at the meeting. TIS cells exhibit a specific pro-inflammatory secretory profile whose functional impact on tumor and bystander cells is currently being investigated. Importantly, this profile is distinguishable from cytokine profiles of senescence-compromised Suv39h1- or p53-deficient lymphomas, and, thus, reflects a senescence - rather than a DNA damage-associated secretory response. Discussion: The study unveils a functional interaction of senescent LCs with different immune cell subsets in vitro and in vivo. The cytokine arrays show that senescent cells produce a specific secretory profile, which might stimulate immune cell attraction. Therefore, immune cells could be recruited to lymphomas in vivo specifically after TIS with the potential to clear senescent – and possibly non-senescent – cells from the tumor site. The data demonstrate genetically that senescence is a beneficial effector principle of DNA damaging chemotherapy and encourage further exploration of this program to limit cancer expansion in vivo.

scholarly journals The impact of genistein supplementation on tendon functional properties and gene expression in estrogen deficient rats

2019 ◽  
Author(s):  
Chad C. Carroll ◽  
Shivam H. Patel ◽  
Jessica Simmons ◽  
Ben DH. Gordon ◽  
Jay F. Olson ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Altered Expression ◽  
Collagen Remodeling ◽  
Ovx Rats ◽  
Genistein Treatment ◽  
Droplet Pcr ◽  
Old Rats ◽  
The Impact

ABSTRACTPurposeTendinopathy risk increases with menopause. The phytoestrogen genistein prevents collagen loss during estrogen deficiency [ovariectomy (OVX)]. The influence of genistein on tendon function and extracellular matrix (ECM) regulation are not well known. We determined the impact of genistein on tendon function and examined potential mechanisms by which genistein alters tendon ECM.Materials and MethodsEight-week-old rats (n=42) were divided into three groups: intact, OVX, or OVX-genistein (6mg/kg/day) for 6-weeks. Tail fascicles were assessed with a Deben tensile stage. Achilles tendon mRNA expression was determined with digital droplet PCR. Tendon-derived fibroblasts were also treated with genistein in the presence of estrogen receptor (ER) antagonists.ResultsCompared to intact, stress tended to be lower in untreated OVX rats (p=0.022). Further, modulus and energy density were greater in genistein-treated rats (p<0.05) compared to intact. Neither OVX nor genistein altered expression of Col1a1, Col3a1, Casp3, Casp8, Mmp1a, Mmp2, or Mmp9 (p>0.05). Compared to intact, Tnmd and Esr1 expression was greater and Pcna and Timp1 expression lower in OVX rats (p<0.05). Genistein treatment returned Tnmd, Pcna, and Timp1 to levels of Intact-Vehicle (p<0.05), but did not alter Scx or Esr1 (p>0.05). Several β-catenin/Wnt signaling related molecules were not altered by OVX or genistein (p>0.05). In vitro, genistein blunted cell proliferation but not via ERs.ConclusionsOur findings demonstrate that genistein improves tendon function. Genistein inhibits cell proliferation in vitro but not via ER. The effect of genistein in vivo was predominately on genes related to cell proliferation rather than collagen remodeling.

scholarly journals 253.Follicle stimulating-hormone (FSH) withdrawal induces germ cell apoptosis in the mature rat testis in vitro

2004 ◽  
Vol 16 (9) ◽  
pp. 253
Author(s):  
S. M. Degen ◽  
P. G. Stanton ◽  
K. L. Loveland ◽  
S. J. Meachem
Keyword(s):  
Cell Proliferation ◽  
Germ Cell ◽  
Cell Development ◽  
Brdu Incorporation ◽  
Proliferating Cells ◽  
Germ Cell Development ◽  
Sperm Output ◽  
The Impact ◽  
Germ Cell Proliferation

FSH is a key determinant of adult sperm output influencing both Sertoli and germ cell development. The aim of this study was to assess the impact of FSH action on Sertoli and germ cell proliferation and survival in vitro, and to identify FSH-regulated genes that may underpin these responses. Testis fragments from 17-day-old rats were cultured with recombinant human FSH for 2 or 24 h and then labelled with bromodeoxyuridine (BrdU) to identify proliferating cells. The testis fragments were then processed for analysis of cell numbers by stereology, BrdU incorporation by immunohistochemistry, and apoptosis by TUNEL. The TUNEL assay revealed that without FSH, spermatogonial apoptosis was induced to 195% and 179% (P�<�0.05) compared to fragments with FSH after 2 and 24 h, respectively. No difference in apoptosis was observed in spermatocyte or Sertoli cell populations at these time points. No differences in Sertoli or germ cell proliferation were observed with or without FSH. To understand how FSH mediates spermatogonial apoptosis the response of 5 testicular genes of interest was examined. Expression of cyclin D2 (cell cycle, G1-S), N-cadherin (N-Cad; adhesion molecule), Bax (pro-apoptotic), Bcl-w (anti-apoptotic), and stem cell factor (SCF; pro-apoptotic and other functions) was elevated to 151%, 348%, 209%, 258%, and 198%, respectively (all P�<�0.001), in fragments cultured without FSH for 24 h, compared to fragments with FSH. No gene expression differences were observed at 2 h, except for SCF, which was elevated to 135% (P�<�0.01). In conclusion, these studies have examined apoptosis and proliferation activities simultaneously in testis fragments in vitro, and demonstrated that FSH withdrawal induces both spermatogonial apoptosis and expression of testicular genes known to be involved in cell survival. This model will now be used to further investigate FSH-mediation of Sertoli and germ cell development.

scholarly journals Identification of human and mouse p19, a novel CDK4 and CDK6 inhibitor with homology to p16ink4.

1995 ◽  
Vol 15 (5) ◽  
pp. 2682-2688 ◽  
Author(s):  
F K Chan ◽  
J Zhang ◽  
L Cheng ◽  
D N Shapiro ◽  
A Winoto
Keyword(s):  
Cell Cycle ◽  
T Cell ◽  
Mammalian Cells ◽  
Cyclin E ◽  
G1 Arrest ◽  
Cdk Inhibitor ◽  
Cyclin D ◽  
Human And Mouse

The cell cycle in mammalian cells is regulated by a series of cyclins and cyclin-dependent kinases (CDKs). The G1/S checkpoint is mainly dictated by the kinase activities of the cyclin D-CDK4 and/or cyclin D-CDK6 complex and the cyclin E-CDK2 complex. These G1 kinases can in turn be regulated by cell cycle inhibitors, which may cause the cells to arrest at the G1 phase. In T-cell hybridomas, addition of anti-T-cell receptor antibody results not only in G1 arrest but also in apoptosis. In searching for a protein(s) which might interact with Nur77, an orphan steroid receptor required for activation-induced apoptosis of T-cell hybridomas, we have cloned a novel human and mouse CDK inhibitor, p19. The deduced p19 amino acid sequence consists of four ankyrin repeats with 48% identity to p16. The human p19 gene is located on chromosome 19p13, distinct from the positions of p18, p16, and p15. Its mRNA is expressed in all cell types examined. The p19 fusion protein can associate in vitro with CDK4 but not with CDK2, CDC2, or cyclin A, B, E, or D1 to D3. Addition of p19 protein can lead to inhibition of the in vitro kinase activity of cyclin D-CDK4 but not that of cyclin E-CDK2. In T-cell hybridoma DO11.10, p19 was found in association with CDK4 and CDK6 in vivo, although its association with Nur77 is not clear at this point. Thus, p19 is a novel CDK inhibitor which may play a role in the cell cycle regulation of T cells.

The two major glucokinase isoforms show conserved functionality in β-cells despite different subcellular distribution

2018 ◽  
Vol 399 (6) ◽  
pp. 565-576 ◽  
Author(s):  
Brian Lu ◽  
Miguel Munoz-Gomez ◽  
Yasuhiro Ikeda
Keyword(s):  
Cell Proliferation ◽  
Subcellular Distribution ◽  
Normal Diet ◽  
Β Cells ◽  
Β Cell ◽  
Functional Conservation ◽  
Exon 1 ◽  
The Impact

Abstract Glucokinase (GCK) is crucial to regulating glucose metabolism in the liver and in pancreatic β-cells. There are two major GCK isoforms, hepatic and pancreatic GCKs, which differ only in exon 1. However, the functional differences between the two GCK isoforms remain poorly understood. Here, we used a β-cell-targeted gene transfer vector to determine the impact of isoform-specific GCK overexpression on β-cells in vitro and in vivo. We showed that pancreatic GCK had a nuclear localization signal unique to the pancreatic isoform, facilitating its nuclear distribution in β-cells. Despite the difference in subcellular distribution, overexpression of GCK isoforms similarly enhanced glucose uptake and β-cell proliferation in vitro. Overexpression of hepatic or pancreatic GCK also similarly enhanced β-cell proliferation in normal diet mice without affecting fasting glucose and intraperitoneal glucose tolerance tests (IPGTT). Our further study on human GCK sequences identified disproportional GCK amino acid variants in exon 1, while mutations linked to maturity onset diabetes of the young type 2 (MODY2) were disproportionally found in exons 2 through 10. Our results therefore indicate functional conservation between the two major GCK isoforms despite their distinct subcellular distribution.

scholarly journals M2 macrophage-induced lncRNA PCAT6 facilitates tumorigenesis and angiogenesis of triple-negative breast cancer through modulation of VEGFR2

2020 ◽  
Vol 11 (9) ◽  
Author(s):  
Fang Dong ◽  
Shengnan Ruan ◽  
Jinlong Wang ◽  
Yun Xia ◽  
Kehao Le ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Cell Proliferation ◽  
Triple Negative Breast Cancer ◽  
Noncoding Rna ◽  
Triple Negative ◽  
M2 Macrophage ◽  
Breast Cancers ◽  
The Impact

Abstract As a common female malignancy, triple-negative breast cancer (TNBC) is the most malignant subtype of breast cancers (BC). This study further studied the role of long noncoding RNA (lncRNA) prostate cancer-associated transcript 6 (PCAT6) in TNBC. Functional assays, including EdU, wound healing, transwell, and immunofluorescence staining, revealed the effect of PCAT6 on cell proliferation, migration, and EMT process. The tube-formation assay disclosed the function of PCAT6 on angiogenesis. In vivo assays were also established to explore the impact of PCAT6 on tumor growth and microangiogenesis. The results revealed that PCAT6 boosted TNBC cell proliferation, migration, and angiogenesis both in vitro and in vivo. Then, this study unveiled that M2 macrophage secreted VEGF to stimulate the upregulation of PCAT6, thus promoting angiogenesis in TNBC. Next, through bioinformatics analysis and mechanism assays, we identified that PCAT6 positively regulated VEGFR2 expression via ceRNA pattern and then participated in VEGFR/AKT/mTOR signaling pathway to accelerate angiogenesis. Moreover, PCAT6 bound USP14, a deubiquitinase, to induce the deubiquitination of VEGFR2. On the whole, M2 macrophage-induced upregulation of PCAT6 facilitates TNBC tumorigenesis through modulation of VEGFR2 expression via ceRNA and deubiquitination patterns.

Sign in / Sign up

Export Citation Format

Share Document