scholarly journals Expression of the E5 Oncoprotein of HPV16 Impacts on the Molecular Profiles of EMT-Related and Differentiation Genes in Ectocervical Low-Grade Lesions

2021 ◽  
Vol 22 (12) ◽  
pp. 6534
Author(s):  
Danilo Ranieri ◽  
Deborah French ◽  
Salvatore Raffa ◽  
Luisa Guttieri ◽  
Maria Rosaria Torrisi ◽  
...  
Keyword(s):  
Significant Positive Correlation ◽  
Expression Profiles ◽  
Human Keratinocytes ◽  
Specific Gene ◽  
Low Grade ◽  
Neoplastic Progression ◽  
Differentiation Markers ◽  
Viral Oncoprotein ◽  
Positive Correlation

Infection with human papillomavirus type 16 (HPV16) is one of the major risk factors for the development of cervical cancer. Our previous studies have demonstrated the involvement of the early oncoprotein E5 of HPV16 (16E5) in the altered isoform switch of fibroblast growth factor receptor 2 (FGFR2) and the consequent expression in human keratinocytes of the mesenchymal FGFR2c isoform, whose aberrant signaling leads to EMT, invasiveness, and dysregulated differentiation. Here, we aimed to establish the possible direct link between these pathological features or the appearance of FGFR2c and the expression of 16E5 in low-grade squamous intraepithelial lesions (LSILs). Molecular analysis showed that the FGFR2c expression displayed a statistically significant positive correlation with that of the viral oncoprotein, whereas the expression values of the epithelial FGR2b variant, as well as those of the differentiation markers keratin 10 (K10), loricrin (LOR) and involucrin (INV), were inversely linked to the 16E5 expression. In contrast, the expression of EMT-related transcription factors Snail1 and ZEB1 overlapped with that of 16E5, becoming a statistically significant positive correlation in the case of Snail2. Parallel analysis performed in human cervical LSIL-derived W12 cells, containing episomal HPV16, revealed that the depletion of 16E5 by siRNA was able to counteract these molecular events, proving to represent an effective strategy to identify the specific role of this viral oncoprotein in determining LSIL oncogenic and more aggressive profiles. Overall, coupling in vitro approaches to the molecular transcript analysis in ectocervical early lesions could significantly contribute to the characterization of specific gene expression profiles prognostic for those LSILs with a greater probability of direct neoplastic progression.

scholarly journals TMOD-13. MODELING THE GENETIC, TRANSCRIPTOMIC, AND CELLULAR HETEROGENEITY OF GLIOBLASTOMA USING TUMOR ORGANOIDS

2019 ◽  
Vol 21 (Supplement_6) ◽  
pp. vi265-vi265
Author(s):  
Daniel Zhang ◽  
Fadi Jacob ◽  
Ryan Salinas ◽  
Phuong Nguyen ◽  
Guo-li Ming ◽  
...  
Keyword(s):  
Gene Expression ◽  
Expression Profiles ◽  
Gene Expression Profiles ◽  
Cellular Heterogeneity ◽  
Definitive Treatment ◽  
Functional Study ◽  
Specific Gene ◽  
Fresh Tissue ◽  
Mechanistic Investigation

Abstract Glioblastoma exhibits enormous genetic, transcriptional, and cellular heterogeneity at the macroscopic level across regions of the tumor as well as at the microscopic level between neighboring cells, all of which present significant challenges towards creating a definitive treatment for this devastating disease. We have developed a method of generating glioblastoma organoids (GBOs) from fresh tissue obtained directly from surgical resection and maintaining them in a defined medium without bFGF/EGF. Whole exome sequencing revealed that GBOs maintain the genomic landscape of their parent tumors. Somatic and copy number variants are present in the GBOs at similar allele frequencies or copy ratios as in the parent tumor, suggesting that the relative proportions of clonal populations are largely maintained in the organoids. Bulk transcriptomic analysis demonstrated strong gene expression correlations between the parent tumor and corresponding GBOs through 12 weeks of culture. Some tumors were sampled at multiple different anatomic regions, and the corresponding GBOs maintained region-specific gene expression signatures and genomic variants. EGFRvIII, a tumor-specific variant targeted in a number of emerging therapies, also remains present in the GBOs at similar transcript frequencies, reflecting the native heterogeneity of the parent tumor. Finally, we used single cell transcriptomics to examine cellular heterogeneity and find that GBOs contain many different cell types that exhibit similar gene expression profiles as the matching cell type in the corresponding parent tumor. Notably, these GBOs retain neoplastic as well as non-neoplastic cells, such as tumor associated macrophages / microglia, T-cells, endothelial cells, stromal cells, and oligodendrocytes. These GBOs preserve complex tumor heterogeneity an in vitro environment, creating opportunities for extended manipulation, characterization, and functional study for mechanistic investigation and therapeutic testing.

In Vitro Radiation-Induced Neoplastic Progression of Low-Grade Uroepithelial Tumors

1994 ◽  
Vol 138 (1) ◽  
pp. 86 ◽  
Author(s):  
Simonetta Pazzaglia ◽  
Xiao-Rong Chen ◽  
Carla B. Aamodt ◽  
Shi-Qi Wu ◽  
Chinghai Kao ◽  
...  
Keyword(s):  
Low Grade ◽  
Neoplastic Progression ◽  
Radiation Induced

scholarly journals EST analysis of cDNA libraries from the entomopathogenic fungus Beauveria (Cordyceps) bassiana. I. Evidence for stage-specific gene expression in aerial conidia, in vitro blastospores and submerged conidia

Microbiology ◽  
2006 ◽  
Vol 152 (9) ◽  
pp. 2843-2854 ◽  
Author(s):  
Eun-Min Cho ◽  
Li Liu ◽  
William Farmerie ◽  
Nemat O. Keyhani
Keyword(s):  
Gene Expression ◽  
Entomopathogenic Fungus ◽  
Expressed Sequence Tag ◽  
Biological Control Agent ◽  
Expression Profiles ◽  
Cdna Libraries ◽  
Specific Gene ◽  
Cell Type ◽  
Aerial Conidia

The entomopathogenic fungus Beauveria (Cordyceps) bassiana holds much promise as a pest biological control agent. B. bassiana produces at least three in vitro single cell infectious propagules, including aerial conidia, vegetative cells termed blastospores and submerged conidia, that display different morphological, biochemical and virulence properties. Populations of aerial conidia, blastospores and submerged conidia were produced on agar plates, rich liquid broth cultures and under conditions of nutrient limitation in submerged cultures, respectively. cDNA libraries were generated from mRNA isolated from each B. bassiana cell type and ∼2500 5′ end sequences were determined from each library. Sequences derived from aerial conidia clustered into 284 contigs and 963 singlets, with those derived from blastospores and submerged conidia forming 327 contigs with 788 singlets, and 303 contigs and 1079 contigs, respectively. Almost half (40–45 %) of the sequences in each library displayed either no significant similarity (e value >10−4) or similarity to hypothetical proteins found in the NCBI database. The expressed sequence tag dataset also included sequences representing a significant portion of proteins in cellular metabolism, information storage and processing, transport and cell processes, including cell division and posttranslational modifications. Transcripts encoding a diverse array of pathogenicity-related genes, including proteases, lipases, esterases, phosphatases and enzymes producing toxic secondary metabolites, were also identified. Comparative analysis between the libraries identified 2416 unique sequences, of which 20–30 % were unique to each library, and only ∼6 % of the sequences were shared between all three libraries. The unique and divergent representation of the B. bassiana transcriptome in the cDNA libraries from each cell type suggests robust differential gene expression profiles in response to environmental conditions.

scholarly journals Phosphoprotein Phosphatase 1 Is Required for Extracellular Calcium-Induced Keratinocyte Differentiation

2016 ◽  
Vol 2016 ◽  
pp. 1-11 ◽  
Author(s):  
Chandrama Shrestha ◽  
Yuanyuan Tang ◽  
Hong Fan ◽  
Lusha Li ◽  
Qin Zeng ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Second Messengers ◽  
Human Keratinocytes ◽  
Extracellular Calcium ◽  
Keratinocyte Differentiation ◽  
Differentiation Markers ◽  
Calcium Stimulation ◽  
E Cadherin

Extracellular calcium is a major regulator of keratinocyte differentiation in vitro and appears to play that role in vivo, but the mechanism is unclear. We have previously demonstrated that, following calcium stimulation, PIP5K1αis recruited by the E-cadherin-β-catenin complex to the plasma membrane where it provides the substrate PIP2 for both PI3K and PLC-γ1. This signaling pathway is critical for calcium-induced generation of second messengers including IP3 and intracellular calcium and keratinocyte differentiation. In this study, we explored the upstream regulatory mechanism by which calcium activates PIP5K1αand the role of this activation in calcium-induced keratinocyte differentiation. We found that treatment of human keratinocytes in culture with calcium resulted in an increase in serine dephosphorylation and PIP5K1αactivation. PP1 knockdown blocked extracellular calcium-induced increase in serine dephosphorylation and activity of PIP5K1αand induction of keratinocyte differentiation markers. Knockdown of PLC-γ1, the downstream effector of PIP5K1α, blocked upstream dephosphorylation and PIP5K1αactivation induced by calcium. Coimmunoprecipitation revealed calcium induced recruitment of PP1 to the E-cadherin-catenin-PIP5K1αcomplex in the plasma membrane. These results indicate that PP1 is recruited to the extracellular calcium-dependent E-cadherin-catenin-PIP5K1αcomplex in the plasma membrane to activate PIP5K1α, which is required for PLC-γ1 activation leading to keratinocyte differentiation.

scholarly journals Type 1 Chemokine Receptor Expression in Chagas' Disease Correlates with Morbidity in Cardiac Patients

2005 ◽  
Vol 73 (12) ◽  
pp. 7960-7966 ◽  
Author(s):  
Juliana A. S. Gomes ◽  
Lilian M. G. Bahia-Oliveira ◽  
Manoel Otávio C. Rocha ◽  
Solange C. U. Busek ◽  
Mauro M. Teixeira ◽  
...  
Keyword(s):  
Chagas Disease ◽  
Significant Positive Correlation ◽  
Receptor Expression ◽  
Necrosis Factor Alpha ◽  
Positive Correlation ◽  
Intracellular Cytokines ◽  
Tnf Α ◽  
Ifn Γ

ABSTRACT Chemokines and chemokine receptors (CKRs) control the migration of leukocytes during the inflammatory process and are important immunological markers of type 1 (CCR5 and CXCR3) and type 2 (CCR3 and CCR4) responses. The coexpression of CKRs (CCR2, CCR3, CCR5, CXCR3, and CXCR4) and intracellular cytokines (interleukin-10 [IL-10], IL-4, tumor necrosis factor alpha [TNF-α], and gamma interferon [IFN-γ]) on T CD4+ and CD8+ peripheral cells from individuals with indeterminate (IND) or cardiac (CARD) clinical forms of Chagas' disease after in vitro stimulation with Trypanosoma cruzi antigens, were evaluated in this study. The percentage of T CD4+ and CD8+ cells coexpressing CCR5 and IFN-γ, CXCR3 and IFN-γ, and CXCR3 and TNF-α were higher in CARD than in IND individuals; on the other hand, the percentage of T CD4+ or CD8+ cells coexpressing CCR3 and IL-10 or coexpressing CCR3 and IL-4 were lower in CARD individuals than in IND individuals. In addition, a significant positive correlation between the expression of CCR5 or CXCR3 and IFN-γ was observed in CARD individuals contrasting with a significant positive correlation between the expression of CCR3 and IL-4 and of CCR3 and IL-10 in IND patients. These results reinforce the hypothesis that a T. cruzi-exacerbated specific type 1 immune response developed by CARD chagasic patients is associated with the development of heart pathology.

Mycophenolic acid affects basic functions of human keratinocytes in the IMPDH-dependent manner

2013 ◽  
Vol 91 (5) ◽  
pp. 333-340 ◽  
Author(s):  
J. Borowczyk ◽  
E. Laczna ◽  
K. Sporniak-Tutak ◽  
Z. Madeja ◽  
J. Drukala
Keyword(s):  
Wound Healing ◽  
Mycophenolic Acid ◽  
De Novo ◽  
Human Keratinocytes ◽  
Pcr Analysis ◽  
Dependent Manner ◽  
Differentiation Markers ◽  
Impaired Wound Healing ◽  
Basic Functions

Clinical studies suggest that the immunosuppressant MPA is associated with impaired wound healing. It is believed that the main cause of impairment is the inhibition of inflammatory response. However, it is unknown whether MPA may directly affect epidermal cells. The aim of our study was to examine the direct influence of mycophenolic acid, the selective blocker of de novo purine synthesis, on human epidermal keratinocyte morphology, proliferation, motile activity, and differentiation in in vitro culture. The number of keratinocytes cultured in the presence of MPA was counted and cell motility was measured by a time-lapse computer-aided method. Cell morphology was determined by flow and image cytometry methods. Real-time RT-PCR analysis was employed to investigate the expression of markers of differentiation. We showed that MPA induces irreversible inhibition of cell proliferation, causes cell enlargement and impairs cell locomotion in a time-dependent manner. The level of expression of differentiation markers was significantly reduced by MPA treatment. All these effects were reversed by the addition of guanine. Our results indicated that MPA impairs basic functions of human skin keratinocytes via intracellular guanosine nucleotide depletion, which may be directly reflected in wound healing problems in patients treated with this immunosuppressant.

scholarly journals YY1 directly interacts with myocardin to repress the triad myocardin/SRF/CArG box-mediated smooth muscle gene transcription during smooth muscle phenotypic modulation

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Jian-Pu Zheng ◽  
Xiangqin He ◽  
Fang Liu ◽  
Shuping Yin ◽  
Shichao Wu ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Gene Transcription ◽  
Molecular Mechanisms ◽  
Serum Response Factor ◽  
Specific Gene ◽  
Phenotypic Modulation ◽  
Differentiation Markers ◽  
Carg Box

AbstractYin Yang 1 (YY1) regulates gene transcription in a variety of biological processes. In this study, we aim to determine the role of YY1 in vascular smooth muscle cell (VSMC) phenotypic modulation both in vivo and in vitro. Here we show that vascular injury in rodent carotid arteries induces YY1 expression along with reduced expression of smooth muscle differentiation markers in the carotids. Consistent with this finding, YY1 expression is induced in differentiated VSMCs in response to serum stimulation. To determine the underlying molecular mechanisms, we found that YY1 suppresses the transcription of CArG box-dependent SMC-specific genes including SM22α, SMα-actin and SMMHC. Interestingly, YY1 suppresses the transcriptional activity of the SM22α promoter by hindering the binding of serum response factor (SRF) to the proximal CArG box. YY1 also suppresses the transcription and the transactivation of myocardin (MYOCD), a master regulator for SMC-specific gene transcription by binding to SRF to form the MYOCD/SRF/CArG box triad (known as the ternary complex). Mechanistically, YY1 directly interacts with MYOCD to competitively displace MYOCD from SRF. This is the first evidence showing that YY1 inhibits SMC differentiation by directly targeting MYOCD. These findings provide new mechanistic insights into the regulatory mechanisms that govern SMC phenotypic modulation in the pathogenesis of vascular diseases.

scholarly journals Viral E6-E7 Transcription in the Basal Layer of Organotypic Cultures without Apparent p21cip1 Protein Precedes Immortalization of Human Papillomavirus Type 16- and 18-Transfected Human Keratinocytes

1998 ◽  
Vol 72 (1) ◽  
pp. 749-757 ◽  
Author(s):  
Renske D. M. Steenbergen ◽  
Jacqueline N. Parker ◽  
Sharon Isern ◽  
Peter J. F. Snijders ◽  
Jan M. M. Walboomers ◽  
...  
Keyword(s):  
Human Papillomavirus ◽  
Cellular Proliferation ◽  
Basal Layer ◽  
Human Keratinocytes ◽  
Model System ◽  
Basal Cells ◽  
Organotypic Cultures ◽  
Differentiation Markers ◽  
Ki 67

ABSTRACT Organotypic cultures of human keratinocytes provide a useful model system to study human papillomavirus (HPV)-host cell interactions. In this study, we analyzed organotypic cultures of two HPV type 16 (HPV16) (FK16A and FK16B)- and two HPV18 (FK18A and FK18B)-transfected keratinocyte cell lines through the process of immortalization in vitro. For FK16A and FK18B cells, passages of both mortal cells in their extended life span and subsequent immortal stages were studied. Mortal cells of FK16A and FK18B showed a morphology reminiscent of mild to moderate dysplasia, whereas in their immortal descendants, severely dysplastic features were observed. Immortal FK18A cells were mildly to moderately dysplastic, while FK16B cells were severely dysplastic. The increasing degrees of dysplasia were associated with a decreasing expression of differentiation markers cytokeratin 10 and profilaggrin. All raft cultures expressed E6-E7 mRNAs in the basal layer, while the amount of viral transcripts in the suprabasal cells was in general proportional to the degree of dysplasia. In all cases, E6-E7 transcription and dysplastic features were highly correlated with cellular proliferation, as assessed by Ki-67 (MIB-1) antigen expression. Moreover, high levels of E6-E7 transcription and expression of p21cip1 protein in the basal layer seemed to be mutually exclusive. We conclude that expression of E6-E7 in the basal cells associated with increased proliferation in the absence of detectable p21cip1 protein is apparently necessary but not sufficient for immortalization, or for the loss of terminal differentiation, for which yet to be discovered additional events are required. The model system described in this study provides a valuable tool to analyze alterations in viral transcription regulation during HPV-mediated cell transformation.

Blood bisphenol A concentration in patient with infertility affects in vitro fertilization outcomes

GYNECOLOGY ◽  
2021 ◽  
Vol 23 (2) ◽  
pp. 161-166
Author(s):  
Anastasia G. Syrkasheva ◽  
Svetlana V. Kindysheva ◽  
Nataliya L. Starodubtseva ◽  
Vladimir E. Frankevich ◽  
Nataliya V. Dolgushina
Keyword(s):  
In Vitro Fertilization ◽  
Bisphenol A ◽  
Significant Positive Correlation ◽  
Threshold Level ◽  
Early Embryogenesis ◽  
Miscarriage Rate ◽  
Blood Samples ◽  
Vitro Fertilization ◽  
Positive Correlation

Aim. To analyze the relationship between bisphenol A levels in the blood of patients with infertility and in vitro fertilization (IVF) outcomes. Materials and methods. Infertility treatment of 301 married couples was performed using IVF. The level of bisphenol A in the blood of patients was determined by gas chromatography-mass spectrometry. The influence of bisphenol A levels on parameters of early embryogenesis and outcomes of IVF programs was assessed. Results. Bisphenol A was detected in 92.9% (277/298) blood samples of women and in 92.7% (141/154) blood samples of men. Bisphenol A levels in women had a statistically significant positive correlation with the level of bisphenol A in their spouses (r=0.533, p0.0001). No differences were found in the main parameters of early embryogenesis and pregnancy rates in quartile bisphenol A subgroups in women and men. There was a trend towards an increase in the miscarriage rate with an increase in the level of bisphenol A. The threshold level of bisphenol A that increased the miscarriage rate was 0.9 ng/ml for women and 0.4 ng/ml for men. With an increase in the threshold level of bisphenol A in both spouses (men 0.4 ng/ml and women 0.9 ng/ml), the odds ratio for miscarriage after IVF 8.8 (95% confidence interval 0.88113.08). Conclusions. Bisphenol A was found in the majority of infertile patients. A significant positive correlation was noted between the level of bisphenol A in patients and their spouses. An increase in the level of bisphenol A in the blood is associated with an increased risk of early reproductive losses after IVF, which requires further research.

The influence of growth hormone/insulin-like growth factor deficiency on prostatic dysplasia in pbARR2-cre PTEN knockout mice.

2012 ◽  
Vol 30 (5_suppl) ◽  
pp. 66-66
Author(s):  
Naokazu Ibuki ◽  
Kiyoshi Takahara ◽  
Mazyar Gaffari ◽  
Howard Tearle ◽  
Christopher J. Ong ◽  
...  
Keyword(s):  
Growth Factor ◽  
Germ Line ◽  
Growth Factor Signaling ◽  
In Vitro Growth ◽  
Neoplastic Progression ◽  
Differentiation Markers ◽  
Prostate Carcinogenesis ◽  
Igf I

66 Background: Signaling through GH/IGF-I axis has been linked to PCa risk. The GH/IGF-I axis is an important regulator of growth, survival, and metastatic potential in a variety of malignancies and is strongly implicated in PCa etiology. A range of therapeutic approaches including reducing ligand availability by GH antagonist, IGF-I antibodies, and recombinant IGFBPs, reducing IGF-1R expression by antisense and RNA interference, or inhibiting of IGF-1R signaling by IGF-1R antibodies and small molecule tyrosine kinase inhibitors are under investigation in PCa. Methods: To investigate the role of GH/IGF-I axis on in vivo prostate carcinogenesis and neoplastic progression, we crossed pbARR2-Cre, PTEN(fl/fl) mice (PTEN-/-) with IGF-I deficient lit mice, and produced lit/lit and lit/+ PTEN-/- mice. To complement the in vivo experiments, in vitro growth and growth factor signaling of murine PTEN-/- cells derived from the prostate (MPPK) using serum from lit/lit or lit/+ mice was examined. Results: No obvious differences in prostatic dysplasia were observed in lit/lit mice at 15 and 20 weeks of age when compared to lit/+ littermates measured as normalized prostatic wet weight or for expression of the murine prostatic differentiation markers, probasin and PSP94. However the rate of decreased expression of E-cadherin and increased expression of N-cadherin was slightly delayed in PTEN-/- prostates from lit/lit mice as compare to lit/+ mice. In vitro, growth of MPPK cells was decreased when cultured in serum from lit/lit mice as compared with serum from lit/+ mice. Suppressed growth of MPPK cells in lit/lit serum could be restored by addition of IGF-I, and to a lesser extent, GH. Addition of GH or IGF-I to lit/lit serum increased steady-state activation of AKT without affecting ERK1/2 activation. Conclusions: Our data suggest that initiation of prostate carcinogenesis by loss of PTEN is not influenced by germ line variation of genes encoding signaling molecules in the GH/IGF-I axis, but suggests that such factors may affect the progression of dysplasic phenotype.

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