scholarly journals Exploring the Crosstalk between Hydrostatic Pressure and Adipokines: An In Vitro Study on Human Osteoarthritic Chondrocytes

2021 ◽  
Vol 22 (5) ◽  
pp. 2745
Author(s):  
Sara Cheleschi ◽  
Sara Tenti ◽  
Marcella Barbarino ◽  
Stefano Giannotti ◽  
Francesca Bellisai ◽  
...  
Keyword(s):  
Gene Expression ◽  
Antioxidant Enzymes ◽  
Hydrostatic Pressure ◽  
Cyclin D1 ◽  
Mirna Gene ◽  
In Vitro Study ◽  
B Cell Lymphoma ◽  
Superoxide Anion Production

Obesity is a risk factor for osteoarthritis (OA) development and progression due to an altered biomechanical stress on cartilage and an increased release of inflammatory adipokines from adipose tissue. Evidence suggests an interplay between loading and adipokines in chondrocytes metabolism modulation. We investigated the role of loading, as hydrostatic pressure (HP), in regulating visfatin-induced effects in human OA chondrocytes. Chondrocytes were stimulated with visfatin (24 h) and exposed to high continuous HP (24 MPa, 3 h) in the presence of visfatin inhibitor (FK866, 4 h pre-incubation). Apoptosis and oxidative stress were detected by cytometry, B-cell lymphoma (BCL)2, metalloproteinases (MMPs), type II collagen (Col2a1), antioxidant enzymes, miRNA, cyclin D1 expressions by real-time PCR, and β-catenin protein by western blot. HP exposure or visfatin stimulus significantly induced apoptosis, superoxide anion production, and MMP-3, -13, antioxidant enzymes, and miRNA gene expression, while reducing Col2a1 and BCL2 mRNA. Both stimuli significantly reduced β-catenin protein and increased cyclin D1 gene expression. HP exposure exacerbated visfatin-induced effects, which were counteracted by FK866 pre-treatment. Our data underline the complex interplay between loading and visfatin in controlling chondrocytes’ metabolism, contributing to explaining the role of obesity in OA etiopathogenesis, and confirming the importance of controlling body weight for disease treatment.

The Role of PIM1 in the Ibrutinib-Resistant ABC Subtype of Diffuse Large B-Cell Lymphoma

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 699-699 ◽  
Author(s):  
Hsu-Ping Kuo ◽  
Sidney Hsieh ◽  
Karl J. Schweighofer ◽  
Leo WK Cheung ◽  
Shiquan Wu ◽  
...  
Keyword(s):  
Gene Expression ◽  
Tumor Growth ◽  
B Cell ◽  
Repeated Measures ◽  
Cell Lymphoma ◽  
Xenograft Model ◽  
B Cell Lymphoma ◽  
Bcr Signaling

Abstract Introduction: Diffuse large B-cell lymphoma (DLBCL) is the most common subtype of non-Hodgkin lymphoma (NHL), accounting for roughly 30% of newly diagnosed cases in the United States (US). DLBCL is a heterogeneous lymphoma, including the activated B cell-like (ABC) and germinal center B cell-like (GCB) subtypes, which have different gene expression profiles, oncogenic aberrations, and clinical outcomes (Alizadeh, Nature 2000; Staudt, Adv Immunol 2005). ABC-DLBCL is characterized by chronic active B-cell receptor (BCR) signaling (Davis, Nature 2010), which is required for cell survival. Thus, the BCR signaling pathway is an attractive therapeutic target in this type of B-cell malignancy. Bruton's tyrosine kinase (BTK), which plays a pivotal role in BCR signaling, is covalently bound with high affinity by ibrutinib, a first-in-class BTK inhibitor approved in the US for mantle cell lymphoma and chronic lymphocytic leukemia (CLL) patients (pts) who have received at least one prior treatment, CLL with del17p, and WaldenstršmÕs macroglobulinemia. A recent phase 2 clinical trial of single-agent ibrutinib in DLBCL pts revealed an overall response rate of 40% for ABC-DLBCL (Wilson, Nat. Med 2015); however, responses to single kinase-targeted cancer therapies are often limited by the cellÕs ability to bypass the target via alternative pathways or acquired mutations in the target or its pathway (Nardi, Curr Opin Hematol 2004; Gazdar, Oncogene 2009). The serine/threonine-protein kinase PIM1 is one of several genes exhibiting differential expression in ibrutinib-resistant ABC-DLBCL cells compared with wild-type (WT) cells. We identified and report herein the role of PIM1 in ABC-DLBCL ibrutinib-resistant cells. Methods: PIM1 gene expression was analyzed by RT-qPCR. In vitro, cell viability was assessed in the human ABC-DLBCL cell line HBL-1 after treatment with ibrutinib and/or a pan-PIM inhibitor for 3 days, and the effect on colony formation was determined 7 days post-treatment. PIM1 mutational analysis was performed with clinical tumor biopsy samples from 2 studies, PCYC-04753 (NCT00849654) and PCYC-1106-CA (NCT01325701). PIM1 protein stability was analyzed by treating cells with cycloheximide and examining protein levels at different time points up to 8 hours. Results: Gene expression profiling of ibrutinib-resistant ABC-DLBCL cells revealed an upregulation of PIM1 (15-fold increase compared with WT cells) as well as PIM2 and PIM3. We also found that, compared with single-drug treatment, in vitro cell growth could be synergistically suppressed with a combination of ibrutinib and a pan-PIM inhibitor. This effect was observed in both WT (combination index (C.I.) = 0.25; synergy score = 3.18) and ibrutinib-resistant HBL-1 cells (C.I. = 0.18; synergy score = 4.98). In HBL-1 cells, this drug combination reduced colony formation and suppressed tumor growth in a xenograft model (Figure 1). In 48 DLBCL patient samples with available genomic profiling, PIM1 mutations appeared more frequently in pts diagnosed with ABC-DLBCL compared with GCB-DLBCL (5 out of 6 DLBCL pts with PIM1 mutations were ABC-subtype). 4 of these 5 pts exhibited a poor clinical response to ibrutinib, ie, 80% of ABC-DLBCL pts with PIM1 mutations had progressive disease, compared with only 13 of 26 (ie, 50%) ABC-DLBCL pts without PIM1 mutations. Subsequent characterization of the mutant PIM1 proteins (L2V, P81S, and S97N) confirmed that they were more stable than WT PIM1, suggesting increased protein levels by 2 potential mechanisms (WT PIM1 gene up-regulation or increased mutant PIM1 protein half-life). The impact of these mutations on PIM1 function and ibrutinib sensitivity is under investigation. Conclusions: Ibrutinib-resistant ABC-DLBCL cells have increased PIM1 expression, and synergistic growth suppression was observed when ibrutinib was combined with a pan-PIM inhibitor. PIM1 mutations identified in ABC-DLBCL pts with poor responses to ibrutinib contributed to increased PIM1 protein stability. A better understanding of the role of PIM1 in ibrutinib-resistant ABC-DLBCL tumors could provide a rationale for the design of combination therapies. Figure 1. Combination of ibrutinib and a pan-PIM inhibitor in the HBL-1 xenograft model. Ibrutinib and PIM inhibitor treatment suppressed tumor growth by 62% compared with the vehicle-treated group (*p < 0.01, repeated measures MANOVA adjusted univariate F-test). Figure 1. Combination of ibrutinib and a pan-PIM inhibitor in the HBL-1 xenograft model. Ibrutinib and PIM inhibitor treatment suppressed tumor growth by 62% compared with the vehicle-treated group (*p < 0.01, repeated measures MANOVA adjusted univariate F-test). Disclosures Kuo: Pharmacyclics LLC, an AbbVie Company: Employment. Hsieh:pharmacyclics LLC, an AbbVie Company: Employment. Schweighofer:Pharmacyclics LLC, an AbbVie Company: Employment. Cheung:Pharmacyclics LLC, an AbbVie Company: Employment. Wu:Pharmacyclics LLC, an AbbVie Company: Employment. Apatira:Pharmacyclics LLC, an AbbVie Company: Employment. Sirisawad:Pharmacyclics LLC, an AbbVie Company: Employment. Eckert:Pharmacyclics LLC, an AbbVie Company: Employment. Liang:Pharmacyclics LLC, an AbbVie Company: Employment. Hsu:Pharmacyclics LLC, an AbbVie Company: Employment. Chang:Pharmacyclics LLC, an AbbVie Company: Employment.

scholarly journals Hydrostatic Pressure Regulates Oxidative Stress through microRNA in Human Osteoarthritic Chondrocytes

2020 ◽  
Vol 21 (10) ◽  
pp. 3653
Author(s):  
Sara Cheleschi ◽  
Marcella Barbarino ◽  
Ines Gallo ◽  
Sara Tenti ◽  
Maria Bottaro ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Antioxidant Enzymes ◽  
Hydrostatic Pressure ◽  
B Cell Lymphoma ◽  
Type Ii Collagen ◽  
Mrna Levels ◽  
Osteoarthritic Chondrocytes ◽  
The Relationship ◽  
And Oxidative Stress

Hydrostatic pressure (HP) modulates chondrocytes metabolism, however, its ability to regulate oxidative stress and microRNAs (miRNA) has not been clarified. The aim of this study was to investigate the role of miR-34a, miR-146a, and miR-181a as possible mediators of HP effects on oxidative stress in human osteoarthritis (OA) chondrocytes. Chondrocytes were exposed to cyclic low HP (1–5 MPa) and continuous static HP (10 MPa) for 3~h. Metalloproteinases (MMPs), disintegrin and metalloproteinase with thrombospondin motif (ADAMTS)-5, type II collagen (Col2a1), miR-34a, miR-146a, miR-181a, antioxidant enzymes, and B-cell lymphoma 2 (BCL2) were evaluated by quantitative real-time polymerase chain reaction qRT-PCR, apoptosis and reactive oxygen species ROS production by cytometry, and β-catenin by immunofluorescence. The relationship among HP, the studied miRNA, and oxidative stress was assessed by transfection with miRNA specific inhibitors. Low cyclical HP significantly reduced apoptosis, the gene expression of MMP-13, ADAMTS5, miRNA, the production of superoxide anion, and mRNA levels of antioxidant enzymes. Conversely, an increased Col2a1 and BCL2 genes was observed. β-catenin protein expression was reduced in cells exposed to HP 1–5 MPa. Opposite results were obtained following continuous static HP application. Finally, miRNA silencing enhanced low HP and suppressed continuous HP-induced effects. Our data suggest miRNA as one of the mechanisms by which HP regulates chondrocyte metabolism and oxidative stress, via Wnt/β-catenin pathway.

scholarly journals A Combination of Celecoxib and Glucosamine Sulfate Has Anti-Inflammatory and Chondroprotective Effects: Results from an In Vitro Study on Human Osteoarthritic Chondrocytes

2021 ◽  
Vol 22 (16) ◽  
pp. 8980
Author(s):  
Sara Cheleschi ◽  
Sara Tenti ◽  
Stefano Giannotti ◽  
Nicola Veronese ◽  
Jean-Yves Reginster ◽  
...  
Keyword(s):  
In Vitro Study ◽  
B Cell Lymphoma ◽  
Type Ii Collagen ◽  
Inhibitory Effect ◽  
Glucosamine Sulfate ◽  
Nuclear Factor ◽  
Superoxide Anion Production ◽  
Qrt Pcr ◽  
Anti Inflammatory

This study investigated the possible anti-inflammatory and chondroprotective effects of a combination of celecoxib and prescription-grade glucosamine sulfate (GS) in human osteoarthritic (OA) chondrocytes and their possible mechanism of action. Chondrocytes were treated with celecoxib (1.85 µM) and GS (9 µM), alone or in combination with IL-1β (10 ng/mL) and a specific nuclear factor (NF)-κB inhibitor (BAY-11-7082, 1 µM). Gene expression and release of some pro-inflammatory mediators, metalloproteinases (MMPs), and type II collagen (Col2a1) were evaluated by qRT-PCR and ELISA; apoptosis and mitochondrial superoxide anion production were assessed by cytometry; B-cell lymphoma (BCL)2, antioxidant enzymes, and p50 and p65 NF-κB subunits were analyzed by qRT-PCR. Celecoxib and GS alone or co-incubated with IL-1β significantly reduced expression and release of cyclooxygenase (COX)-2, prostaglandin (PG)E2, IL-1β, IL-6, tumor necrosis factor (TNF)-α, and MMPs, while it increased Col2a1, compared to baseline or IL-1β. Both drugs reduced apoptosis and superoxide production; reduced the expression of superoxide dismutase, catalase, and nuclear factor erythroid; increased BCL2; and limited p50 and p65. Celecoxib and GS combination demonstrated an increased inhibitory effect on IL-1β than that observed by each single treatment. Drugs effects were potentiated by pre-incubation with BAY-11-7082. Our results demonstrated the synergistic effect of celecoxib and GS on OA chondrocyte metabolism, apoptosis, and oxidative stress through the modulation of the NF-κB pathway, supporting their combined use for the treatment of OA.

scholarly journals Role of Stat3 Activation in Cell-Cell Interaction between B-Cell Lymphoma and Macrophages : The in vitro Study

2013 ◽  
Vol 53 (2) ◽  
pp. 127-133 ◽  
Author(s):  
Bing Bai ◽  
Hasita Horlad ◽  
Yoichi Saito ◽  
Koji Ohnishi ◽  
Yukio Fujiwara ◽  
...  
Keyword(s):  
B Cell ◽  
Cell Lymphoma ◽  
In Vitro Study ◽  
B Cell Lymphoma ◽  
Cell Interaction ◽  
Vitro Study ◽  
Cell Cell

163 Effect of melatonin and its receptors on bovine oocyte maturation and cumulus cell gene expression after heat shock in vitro: preliminary results

2019 ◽  
Vol 31 (1) ◽  
pp. 206 ◽  
Author(s):  
H. Fernandes ◽  
F. C. Castro ◽  
L. Schefer ◽  
D. M. Paschoal ◽  
C. L. V. Leal
Keyword(s):  
Gene Expression ◽  
Antioxidant Enzymes ◽  
Heat Stress ◽  
Heat Shock ◽  
Geometric Mean ◽  
Stress Conditions ◽  
Melatonin Receptors ◽  
Nuclear Maturation

The aim of this study was to assess the effect of melatonin (MLT) during in vitro maturation (IVM) of bovine cumulus-oocyte complexes (COC) under normal and heat stress conditions, on oocyte nuclear maturation and expression of genes related to antioxidant, apoptotic, and expansion activities in cumulus cells (CC), and whether effects are mediated by melatonin receptors (MT1, MT2, or both). The COC were submitted to IVM (25-30/group) in TCM-199 supplemented with 3mg mL−1 BSA, 0.5µg mL−1 FSH, 11µg mL−1 sodium pyruvate and 10µg mL−1 gentamicin. Melatonin (10−6 M) was used alone or associated with luzindole (10−6 M; LUZ) or 4-phenyl-2-propionamidotetralin (10−6 M; 4-P-PDOT). Groups were matured under normal conditions for 24h (38.5°C, control, and 38.5°C+MLT) and heat shock (cultured for 14h at 41°C, 41°C+MLT, 41°C+MLT+LUZ, and 41°C+MLT+4-P-PDOT), and then incubated for the remaining 10h at 38.5°C. The CC were collected and evaluated by real-time quantitative PCR for GPX4, SOD1, and SOD2 (antioxidant enzymes); BAX, CASP3, and P53 (apoptosis-related); and HAS1, HAS2, and PTX3 (expansion-related) transcripts. Gene expression data were normalized by the geometric mean of housekeeping genes ACTB, GAPDH, and PPIA. Relative expression levels were calculated by the comparative method of 2−ΔΔCt (Livak and Schmittgen 2001 Methods, 25, 402-408). Maturation rates (metaphase II; MII) were determined by staining oocytes with Hoechst 33342. Data were analysed by ANOVA followed by Tukey’s test to compare effects of treatments on maturation rate (3 replicates/treatment). Significance level was 5% (GraphPrism software, GraphPad Inc., San Diego, CA, USA). After 24h of IVM, the MII rate decreased (P&lt;0.05) for the 41°C group (30±14.1%, 21/70) compared with the control at 38.5°C (76.1±14.7%, 51/67; P&lt;0.05) and addition of MLT to the 41°C group reversed the decrease (41°C+MLT, 55.3±7.1%, 26/47) and was similar to that of the control (P&gt;0.05). When the heat stress with MLT group was associated with MT inhibitors, MII rates decreased relative to control (P&lt;0.05) and were similar to 41°C without MLT and to each other (41°C+MLT+LUZ, 31.6±8.9%, 24/76; and 41°C+MLT+4-P-PDOT, 30±9.0%, 21/70; P&gt;0.05). Addition of MLT did not differ from control without the hormone (38.5°C+MLT, 90.5±6.7%, 38/42; P&gt;0.05). For gene expression in CC, only the 41°C+MLT+LUZ group increased (P&lt;0.05) transcripts for CASP3 compared with control. There was no difference for antioxidants (GPX4, SOD1, and SOD2), expansion (HAS1, HAS2, and PTX3), or apoptotic (BAX and P53) transcripts for any of the groups (P&gt;0.05). In conclusion, MLT reversed the negative effects on nuclear maturation caused by heat stress conditions during IVM; this effect was probably mediated by the MT2 receptor. However, MLT did not influence the antioxidant enzymes, apoptosis, or expansion-related transcripts in CC, but inhibition of MT1/MT2 receptors increased CASP3 transcripts, suggesting a possible role of the receptors on apoptosis in these cells. Further studies are necessary to improve our knowledge on the role of MLT in heat stress during IVM. The authors acknowledge FAPESP for funding (HF-2016/24884-3; CLVL-2015/20379-0).

scholarly journals Role of glyoxalases in wound healing process: an in vitro study

2021 ◽  
Vol 165 ◽  
pp. 39
Author(s):  
Francesca Lombardi ◽  
Silvano Santini ◽  
Paola Palumbo ◽  
Valeria Cordone ◽  
Virginio Bignotti ◽  
...  
Keyword(s):  
Wound Healing ◽  
Healing Process ◽  
In Vitro Study ◽  
Vitro Study ◽  
Wound Healing Process

scholarly journals Validation of epigenetic mechanisms regulating gene expression in canine B-cell lymphoma: An in vitro and in vivo approach

PLoS ONE ◽  
2018 ◽  
Vol 13 (12) ◽  
pp. e0208709 ◽  
Author(s):  
Silvia Da Ros ◽  
Luca Aresu ◽  
Serena Ferraresso ◽  
Eleonora Zorzan ◽  
Eugenio Gaudio ◽  
...  
Keyword(s):  
Gene Expression ◽  
B Cell ◽  
Cell Lymphoma ◽  
B Cell Lymphoma ◽  
Epigenetic Mechanisms

scholarly journals Antiproliferative Effect of Androgen Receptor Inhibition in Mesenchymal Stem-Like Triple-Negative Breast Cancer

2016 ◽  
Vol 38 (3) ◽  
pp. 1003-1014 ◽  
Author(s):  
Aiyu Zhu ◽  
Yan Li ◽  
Wei Song ◽  
Yumei Xu ◽  
Fang Yang ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Androgen Receptor ◽  
Cyclin D1 ◽  
Cancer Cells ◽  
Triple Negative Breast Cancer ◽  
Breast Cancer Cells ◽  
Triple Negative

Background/Aims: Androgen receptor (AR), a steroid hormone receptor, has recently emerged as prognostic and treatment-predictive marker in breast cancer. Previous studies have shown that AR is widely expressed in up to one-third of triple-negative breast cancer (TNBC). However, the role of AR in TNBC is still not fully understood, especially in mesenchymal stem-like (MSL) TNBC cells. Methods: MSL TNBC MDA-MB-231 and Hs578T breast cancer cells were exposed to various concentration of agonist 5-α-dihydrotestosterone (DHT) or nonsteroidal antagonist bicalutamide or untreated. The effects of AR on cell viability and apoptosis were determined by MTT assay, cell counting, flow cytometry analysis and protein expression of p53, p73, p21 and Cyclin D1 were analyzed by western blotting. The bindings of AR to p73 and p21 promoter were detected by ChIP assay. MDA-MB-231 cells were transplanted into nude mice and the tumor growth curves were determined and expression of AR, p73 and p21 were detected by Immunohistochemistry (IHC) staining after treatment of DHT or bicalutamide. Results: We demonstrate that AR agonist DHT induces MSL TNBC breast cancer cells proliferation and inhibits apoptosis in vitro. Similarly, activated AR significantly increases viability of MDA-MB-231 xenografts in vivo. On the contrary, AR antagonist, bicalutamide, causes apoptosis and exerts inhibitory effects on the growth of breast cancer. Moreover, DHT-dependent activation of AR involves regulation in the cell cycle related genes, including p73, p21 and Cyclin D1. Further investigations indicate the modulation of AR on p73 and p21 mediated by direct binding of AR to their promoters, and DHT could make these binding more effectively. Conclusions: Our study demonstrates the tumorigenesis role of AR and the inhibitory effect of bicalutamide in AR-positive MSL TNBC both in vitro and in vivo, suggesting that AR inhibition could be a potential therapeutic approach for AR-positive TNBC patients.

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