In Vitro Model of Metastasis to Bone Marrow Mediates Prostate Cancer Castration Resistant Growth through Paracrine and Extracellular Matrix Factors

AbstractThe gut microbiota-derived metabolites of ellagitannins and green tea catechins, urolithin A (uroA) and 5-(3′,4′,5′-trihydroxyphenyl)-γ-valerolactone (M4), respectively, are among the main compounds absorbed into human system after ingestion of these polyphenols. The aim of this study was to establish the effects of M4, uroA, and their combinations on LNCaP cells, an androgen dependent prostate cancer in vitro model.. The LNCaP cells were incubated with increasing concentrations of tested metabolites. The cell proliferation was determined by measurement of DNA-bisbenzimide H 33 258 complexes fluorescence. The isobolographic analysis was used to establish the type of interaction between metabolites. The apoptosis, androgen receptor (AR) localization, and phosphorylation of Akt kinase were measured by flow cytometry. Prostate-specific antigen (PSA) secretion was determined by ELISA. M4 showed modest antiproliferative activity in LNCaP cells (IC50 = 117 µM; CI: 81 – 154). UroA decreased proliferation (IC50 = 32.7 µM; CI: 24.3 – 41.1) and induced apoptosis of LNCaP cells. The mixture of M4 with uroA had synergistic antiproliferative effect. Moreover, M4 potentiated inhibition of PSA secretion and enhanced retention of AR in cytoplasm caused by uroA. Interestingly, uroA increased levels of pSer473 Akt in LNCaP cells. These results show that colonic metabolites may contribute to chemoprevention of prostate cancer by varied polyphenol-rich diet or composite polyphenol preparations.

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An in vitro model of erythroid egress in bone marrow

Blood ◽

10.1182/blood.v68.1.250.250 ◽

1986 ◽

Vol 68 (1) ◽

pp. 250-257 ◽

Cited By ~ 7

Author(s):

RE Waugh ◽

M Sassi

Keyword(s):

Bone Marrow ◽

In Vitro Model ◽

Physical Parameters ◽

In Vitro System ◽

Wafer Thickness ◽

Pore Cells ◽

A Cell ◽

Vitro Model ◽

Passage Times

Abstract An in vitro system has been developed that mimics the passage of erythrocytes from the bone marrow to the circulation. Bone marrow egress and its proper regulation are vital physiologic processes. However, because of the inaccessibility of the marrow, it is difficult to evaluate the various factors important in controlling these processes or even to define the precise mechanism by which egress occurs. The in vitro system has been designed to evaluate the importance of different physical parameters in regulating egress. It consists of a thin silicon wafer (thickness approximately equal to 1.0 micron) cemented over the tip of a large (15.0 micron ID) micropipette. The wafer contains a single circular pore. Cells were observed under the microscope as they passed through the pore under controlled pressures. The rate and duration of passage were obtained from videorecordings of the experiment. The measured passage times agreed well with the predictions of a simple analytical model of a cell passing through a thin aperture. The experimental results confirm the conclusion reached from the analysis that the pressures needed to drive a cell through the pore are well within the physiologic range, and the time needed to complete egress is typically less than 1.0 seconds. These results support the hypothesis that erythrocyte egress may be driven by a hydrostatic pressure difference across the pore.

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Amniotic Membrane as an in vitro Model for Endometrium- Extracellular Matrix Interactions

Gynecologic and Obstetric Investigation ◽

10.1159/000009914 ◽

1998 ◽

Vol 45 (1) ◽

pp. 7-11 ◽

Cited By ~ 10

Author(s):

Paul J.Q. van der Linden ◽

Anton F.P.M. de Goeij ◽

Gerard A.J. Dunselman ◽

Heleen W.H. Erkens ◽

Johannes L.H. Evers

Keyword(s):

Extracellular Matrix ◽

Amniotic Membrane ◽

In Vitro Model ◽

Matrix Interactions ◽

Vitro Model ◽

Extracellular Matrix Interactions

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An In Vitro Model of Proliferating CLL Cells for Drug Sensitivity Analysis.

Blood ◽

10.1182/blood.v106.11.2956.2956 ◽

2005 ◽

Vol 106 (11) ◽

pp. 2956-2956

Author(s):

K. Ganeshaguru ◽

N. I. Folarin ◽

R. J. Baker ◽

A. M. Casanova ◽

A. Bhimjiyani ◽

...

Keyword(s):

Bone Marrow ◽

Lymph Nodes ◽

Peripheral Blood ◽

Drug Sensitivity ◽

In Vitro Model ◽

Survivin Expression ◽

Lymphoid Cells ◽

Malignant Cells ◽

Vitro Model

Abstract B-cell chronic lymphocytic leukaemia (CLL) is a heterogeneous disease with a variable clinical course. The disease is characterised by the proliferation in the bone marrow and lymph node of a clonal population of CD5+ve cells that accumulates in the peripheral blood. Therefore, the characteristics of the proliferative compartment are important in determining the kinetics of disease progression in CLL and the sensitivity of the malignant cells to cytotoxic drugs. However, laboratory studies on drug sensitivity of CLL have been performed exclusively on resting circulating peripheral blood cells since it is not feasible to obtain cells from the proliferating pool in sufficient numbers for in vitro analysis. CLL cells can be stimulated to proliferate in vitro using CpG oligonucleotides (ODN) and other factors. The aim of the present study was to generate and validate an in vitro model using malignant cells from the peripheral blood of patients with CLL. The expression pattern of proteins eg., survivin in this model should mimic that in proliferating CLL cells in the bone marrow and lymph nodes. Survivin is a member of the family of inhibitor of apoptosis (IAP) proteins with an additional role in cell cycle progression. Survivin has been shown to be expressed in proliferating bone marrow and lymphoid cells. Cells from patients with CLL were activated for 72h with a combination of ODN (1μM), IL-2 (100u/ml) and CD40L (0.5μg/ml) (ODN*). Activated cells retained their characteristic CLL immunophenotype as determined by the continued expression of CD5, CD19, CD23 and CD25 (n=5). Cell proliferation was confirmed by increased incorporation of 3H-thymidine into DNA in activated cells (n=12). Novel findings in the ODN* activated CLL cells were significant increases in expression of CD38 (n=7, p=0.0001) and of T-cell zeta associated protein (ZAP-70) tyrosine kinase (n=14, p=0.0005). The increased expression of both these proteins in circulating peripheral blood CLL cells has been associated with poor prognosis. All six ODN* activated CLL isolates analysed by western blotting showed increased survivin expression with no constitutive expression in the controls. Drug sensitivity was studied in cells from eight patients using the MTT assay. Activated cells showed significantly greater resistance to chlorambucil (median IC50=164.4±28.18μM) compared to control cells (median IC50=93.63±14.96μM, p=0.044). Figure 1 shows representative IC50 curves. The increased resistance of the activated cells to chlorambucil may be a consequence of the upregulation of survivin. In summary, the in vitro model replicates several key features of authentic proliferating CLL cells found in bone marrow and lymph nodes. It also shows increased resistance to the conventional drug chlorambucil. This model may be of value in evaluating novel drugs and drug combinations which may be more effective in killing the proliferating population that maintain the malignant cell population in CLL. Figure Figure

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