Detection of circulating tumor cells in patients with localized and metastatic prostatic carcinoma: clinical implications.

1995 ◽  
Vol 13 (5) ◽  
pp. 1195-1200 ◽  
Author(s):  
R A Ghossein ◽  
H I Scher ◽  
W L Gerald ◽  
W K Kelly ◽  
T Curley ◽  
...  
Keyword(s):  
Hormonal Therapy ◽  
Peripheral Blood ◽  
Prostatic Cancer ◽  
Mononuclear Cells ◽  
Limit Of Detection ◽  
Specific Antigen ◽  
Distant Metastases ◽  
Tumor Stage ◽  
Rt Pcr ◽  
Circulating Cells

PURPOSE To determine the frequency with which prostate-specific antigen (PSA)-positive cells can be detected in the peripheral blood of patients with prostatic cancer in different stages and with different sensitivities to hormonal therapy. PATIENTS AND METHODS Peripheral blood from 107 men with prostatic cancer and 27 non-prostate cancer controls was analyzed for PSA mRNA using reverse-transcriptase polymerase chain reaction (RT-PCR) and Southern blotting. RESULTS The lower limit of detection was one PSA-producing cell diluted into 1 x 10(6) blood mononuclear cells. The test detected PSA mRNA in four of 25 patients (16%) with clinically organ-confined (T1-2) disease, three of 10 (30%) with T3-4 or N+ tumors, and 25 of 72 (35%) with distant metastases. None of the control samples were positive. An increase in positivity was observed with increasing PSA levels. Within the subgroup of patients with distant metastases, positivity was observed in six of 16 patients (38%) with normal or undetectable PSA levels after hormonal therapy and, overall, in 37% of patients (21 of 57) with androgen-independent disease. CONCLUSION An RT-PCR-based assay for PSA mRNA can detect circulating cells in the peripheral blood of patients with prostatic cancer. The frequency of positivity increases with tumor stage. A unique observation was the detection of cells in patients with no measurable PSA on hormonal therapy. This suggests that continued seeding of distant sites may still be occurring in these patients, despite seemingly successful therapy. The relationship between continued seeding, disease progression, and survival will require further study.

Peripheral Blood Cells from Multiple Myeloma Patients Show Increased Osteoclast and Decreased Osteoblast Gene Expression.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 5098-5098
Author(s):  
Melinda S. Gordon ◽  
Ariana M. Berenson ◽  
Charles B. Drucker ◽  
Matthew Katz ◽  
Hee Jin Lee ◽  
...  
Keyword(s):  
Gene Expression ◽  
Multiple Myeloma ◽  
Alkaline Phosphatase ◽  
Bone Disease ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Bisphosphonate Treatment ◽  
Rt Pcr ◽  
Circulating Cells ◽  
Osteolytic Bone Disease

Abstract Bone resorption leading to osteolytic bone disease is characteristic of multiple myeloma (MM). Recent studies show the presence of bone-resorbing osteoclasts and bone-forming osteoclasts in the circulation, and these cells may correlate with bone disease and change with anti-bone resorptive therapies. We have investigated whether there is an imbalance in the expression of osteoblast and osteoclast genes in the peripheral blood mononuclear cells (PBMCs) from MM patients relative to normal age-matched controls and the effect of bisphosphonate treatment on the expression of these genes. We analyzed the expression of a panel of osteoblast-related (bone alkaline phosphatase [bone AP], bone morphogenic protein 2 [BMP2], collagen I and osteocalcin) and osteoclast-related (b3 integrin, calcitonin, receptor for activation of nuclear factor kappa B [RANK] and tartrate-resistant alkaline phosphatase [TRAP]) genes by semi-quantitative RT-PCR on total RNA isolated from PBMCs obtained following density gradient separation. We demonstrated that the expression of the osteoblast-related gene BMP2 was reduced in eight of nine MM patients when compared with normal donors. In marked contrast, three osteoclast-related genes, b3 integrin, RANK and TRAP, were more highly expressed in all nine MM patients compared to the normal donors; only calcitonin expression was similar to the control subjects. Interestingly, patients receiving bisphosphonate treatment appeared to show increased osteoblast gene expression with higher amounts of bone AP, BMP2 and osteocalcin RNA compared to the patients not receiving anti-bone resorptive therapy. However, there was no alteration in the level of the RNA in any of the four osteoclast genes compared to patients not receiving anti-bone resorptive therapy. We are extending our analysis to a larger panel of MM patients in order to determine the relationship between these circulating cells and bone disease, overall clinical status and change in their levels with anti-bone resorptive therapy. In addition, we are also investigating whether there exist larger and smaller numbers of circulating osteoclasts and osteoblasts, respectively, in MM patients, or whether these circulating cells show alteration of their expression of these genes. Our semi-quantitative RT-PCR results are being correlated with immunohistochemical staining results from osteoblast and osteoclast markers obtained on PBMCs from MM and normal subjects. These studies provide evidence that the number of circulating osteoblasts and osteoclasts is altered in patients with MM, and also may suggest that bisphosphonate therapy may also be associated with changes in these cell populations.

scholarly journals Molecular staging of prostatic cancer with RT-PCR assay for prostate-specific antigen in peripheral blood and lymph nodes: 5 Year follow-up

2007 ◽  
Vol 60 (10) ◽  
Author(s):  
Luis Martínez-Piñeiro ◽  
Montserrat Martínez-Gomariz ◽  
Emilio Rios ◽  
María L. Picazo ◽  
Hugo R. Arriaga ◽  
...  
Keyword(s):  
Lymph Nodes ◽  
Prostate Specific Antigen ◽  
Peripheral Blood ◽  
Prostatic Cancer ◽  
Specific Antigen ◽  
Rt Pcr ◽  
Pcr Assay ◽  
Molecular Staging

scholarly journals The Effect of Sulforaphane on Glyoxalase I Expression and Activity in Peripheral Blood Mononuclear Cells

Nutrients ◽  
2018 ◽  
Vol 10 (11) ◽  
pp. 1773 ◽  
Author(s):  
Michela Alfarano ◽  
Donato Pastore ◽  
Vincenzo Fogliano ◽  
Casper Schalkwijk ◽  
Teresa Oliviero
Keyword(s):  
Gene Expression ◽  
Peripheral Blood Mononuclear Cells ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Limit Of Detection ◽  
Peripheral Blood Mononuclear ◽  
Potential Health ◽  
Age Related ◽  
Blood Mononuclear Cells ◽  
Expression Activity

Studies demonstrate that the potential health-beneficial effect of sulforaphane (SR), a compound formed in broccoli, is the result of a number of mechanisms including upregulation of phase two detoxification enzymes. Recent studies suggest that SR increases expression/activity of glyoxalase 1 (Glo1), an enzyme involved in the degradation of methylglyoxal, is major precursor of advanced glycation end products. Those compounds are associated with diabetes complications and other age-related diseases. In this study, the effect of SR on the expression/activity of Glo1 in peripheral blood mononuclear cells (PBMCs) from 8 healthy volunteers was investigated. PBMCs were isolated and incubated with SR (2.5 μM-concentration achievable by consuming a broccoli portion) for 24 h and 48 h. Glo1 activity/expression, reduced glutathione (GSH), and glutathione-S-transferase gene expression were measured. Glo1 activity was not affected while after 48 h a slight but significant increase of its gene expression (1.03-fold) was observed. GSTP1 expression slightly increased after 24 h incubation (1.08-fold) while the expressions of isoform GSTT2 and GSTM2 were below the limit of detection. GSH sharply decreased, suggesting the formation of GSH-SR adducts that may have an impact SR availability. Those results suggest that a regular exposure to SR by broccoli consumption or SR supplements may enhance Glo1.

Downregulating the Fibromodulin Gene by Short Interfering RNA Causes B-CLL Cell Death.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 176-176 ◽  
Author(s):  
Aniruddha Choudhury ◽  
Katja Derkow ◽  
Eva Mikaelsson ◽  
Parviz Kokhaei ◽  
Anders Österborg ◽  
...  
Keyword(s):  
Peripheral Blood Mononuclear Cells ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Rt Pcr ◽  
Sirna Transfection ◽  
Peripheral Blood Mononuclear ◽  
Normal Peripheral Blood ◽  
Extracellular Matrix Molecule ◽  
Rnai Technology ◽  
Blood Mononuclear Cells

Abstract Development of targeted therapies against B-CLL is dependent on the identification of molecules that are essential for the proliferation and survival of the leukemic cells. One such molecule investigated by us as a putative leukemia-associated target is Fibromodulin (FIM). FIM is an extracellular matrix molecule belonging to the leucin-rich proteoglycan family. Our laboratory studies have verified that expression of FIM is upregulated in B-CLL cells. Moreover, this molecule is specifically overexpressed in B-CLL cells and not on normal peripheral blood mononuclear cells. Analysis of FIM expression on various other hematological tumor cells have revealed that with the exception of mantle cell lymphoma, FIM is not expressed in other hematological malignancies. RNAi technology has recently emerged as a powerful method to specifically silence the expression of a gene. We have generated three siRNA against various segments of FIM and tested the effects of these siRNA on B-CLL cells. Transfection of B-CLL cells with these siRNA significantly diminished, or completely abrogated the expression of FIM mRNA as detected by RT-PCR. The figure below shows silencing of the FIM gene following siRNA transfection, as assayed by RT-PCR. “Untrans” and “ctrl” represents untransfected and control siRNA-transfected B-CLL cells respectively. As noted, transfection with each of the three siRNA against FIM completely abrogated the expression of FIM siRNA. 24–48 hours after siRNA transfection, a substantial fraction (30–70%) of the B-CLL cells, compared to cells transfected with control non-silencing siRNA, went into apoptosis as assayed by Annexin-V-propidium iodide staining. Time kinetic studies revealed that siRNA mediated silencing and apoptosis occurred between 18 and 48 hours. No such effect was noted when peripheral blood mononuclear cells of healthy donors or FIM-positive fibroblast cell lines were transfected with siRNA. In reconstitution experiments, siRNA treated B-CLL cells were co-cultured with various numbers of FIM-positive fibroblasts. The presence of these fibroblasts greatly diminished the apoptosis of B-CLL cells induced after siRNA treatment. Our results indicate that overexpression of FIM in B-CLL cells are critical for their survival and FIM may serve as a leukemia-specific target for B-CLL therapy. Figure Figure

scholarly journals Thyroglobulin mRNA quantification in the peripheral blood is not a reliable marker for the follow-up of patients with differentiated thyroid cancer

2002 ◽  
pp. 575-582 ◽  
Author(s):  
M Eszlinger ◽  
S Neumann ◽  
L Otto ◽  
R Paschke
Keyword(s):  
Thyroid Cancer ◽  
Real Time ◽  
Peripheral Blood ◽  
Differentiated Thyroid Cancer ◽  
Mononuclear Cells ◽  
Radioiodine Therapy ◽  
Rt Pcr ◽  
Beta Actin ◽  
Citrate Blood

BACKGROUND: The detection of serum thyroglobulin (Tg) by immunoassay is widely used to detect residual, recurring or metastatic thyroid carcinoma tIssue in patients with differentiated thyroid cancer (DTC) after total thyroidectomy and radioiodine therapy. However, this method requires thyroid hormone withdrawal to increase sensitivity and is limited by the interference of anti-Tg antibodies. To solve these problems, the detection of Tg mRNA from circulating thyroid cells by reverse transcription (RT)-PCR has been suggested as an alternative method. However, different previous reports show discrepant conclusions as to the clinical usefulness of Tg mRNA quantification. METHODS: We compared three methods of blood collection and RNA extraction, and quantified Tg mRNA (by real time RT-PCR) in the peripheral blood of a) probands without thyroid disease (n=42), patients with b) thyroid autonomy (n=15), c) Graves' disease (n=22), d) euthyroid goiter (n=6), and in DTC-patients after thyroidectomy and radioiodine therapy e) with (n=16) and f) without (n=37) metastasis. As the use of citrate blood in combination with a subsequent separation of mononuclear cells showed a significantly better RNA yield than the extraction of RNA from EDTA or citrate blood without the separation of mononuclear cells, this was the method used. Total RNA was reverse transcribed with random hexamer primers and Tg mRNA was amplified by real time RT-PCR using specific primers and hybridization probes. The Tg mRNA concentrations were normalized to beta-actin mRNA concentrations. RESULTS: Mean circulating Tg mRNA for each group detailed above, expressed as the ratio of Tg to beta-actin concentrations x 1000, were: a) 2.3 (range 0.03-70.89), b) 0.25 (range 0.02-0.55), c) 0.31 (range 0.05-1.36), d) 0.18 (range 0.08-0.35), e) 0.57 (range 0.03-3.03) and f) 0.17 (range 0.02-0.60). Furthermore, we found no correlation between serum Tg and Tg mRNA. CONCLUSIONS: In summary, our data do not show significant differences in Tg mRNA expression between the investigated groups. Therefore, the detection and quantification of Tg mRNA in peripheral blood is unlikely to be suitable for the follow-up of DTC.

scholarly journals Differential Expression of Neurokinin-1 Receptor by Human Mucosal and Peripheral Lymphoid Cells

2000 ◽  
Vol 7 (3) ◽  
pp. 371-376 ◽  
Author(s):  
Triona Goode ◽  
Joe O'Connell ◽  
Wen-Zhe Ho ◽  
Gerald C. O'Sullivan ◽  
J. Kevin Collins ◽  
...  
Keyword(s):  
Mrna Expression ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Peripheral Blood Lymphocytes ◽  
Lymphoid Cells ◽  
Receptor Expression ◽  
Rt Pcr ◽  
Peripheral Blood Mononuclear ◽  
Neurokinin 1 Receptor ◽  
Neurokinin 1

ABSTRACT Substance P (SP) has been implicated in peripheral and mucosal neuroimmunoregulation. However, confusion remains regarding immunocyte expression of the receptor for SP, neurokinin-1 receptor (NK-1R), and whether there is differential NK-1R expression in the mucosal versus the peripheral immune system. In the same assay systems, we examined the expression of NK-1R in human lamina propria mononuclear cells (LPMC), peripheral blood mononuclear cells (PBMC), peripheral blood lymphocytes (PBL), monocytes, and monocyte-derived macrophages (MDM). Using standard reverse transcription (RT)-PCR, mRNA expression of both the long and the short isoforms of the NK-1R was evident in LPMC but not in PBMC, PBL, monocytes, or MDM. However, by using nested RT-PCR NK-1R mRNA expression was detected in PBMC, PBL, monocytes, and MDM. This level of expression was found to represent one NK-1R mRNA transcript in >1,000 cells. In contrast, by using competitive RT-PCR we demonstrate that LPMC express a more biologically significant level of eight NK-1R mRNA transcripts per cell. Flow cytometric detection of NK-1R expression at the protein level was evident in LPMC but not in PBMC. These findings illustrate the extreme sensitivity of nested RT-PCR and the advantages of competitive RT-PCR in comparative studies of receptor expression in different cell populations. This study suggests that, under normal conditions, readily detectable expression of NK-1R in human mononuclear cells occurs at the mucosal level rather than in the peripheral circulation.

Livin Expression in Normal Hematopoietic Cells and in Hematologic Malignancies.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 4301-4301
Author(s):  
Ihab Abd-Elrahman ◽  
Vered Bucholtz ◽  
Klilah Hershko ◽  
Gail Amir ◽  
Riki Perlman ◽  
...  
Keyword(s):  
Bone Marrow ◽  
B Cells ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Hematopoietic Cells ◽  
Hematologic Malignancies ◽  
Inhibitor Of Apoptosis ◽  
Mrna Levels ◽  
Rt Pcr ◽  
Immunohistochemistry Staining

Abstract Livin is a member of the Inhibitor of Apoptosis Proteins (IAP) family, a novel family of intracellular anti-apoptotic proteins that act by binding and inhibiting caspases. We found that Livin is unique among the IAP members as upon strong apoptotic stimuli, it is specifically cleaved by caspases to produce a large C-terminal subunit. This subunit has a paradoxical pro-apoptotic activity. Thus, Livin is not merely an inhibitor of apoptosis. Rather, it is a regulator of apoptosis that can protect against apoptosis but upon continuous apoptotic signals it helps to assure cell death. We showed that Livin plays a major role in melanoma. The level of the Livin protein is directly correlated to the resistance of melanoma cells to chemotherapy and to the survival rate of melanoma patients. Livin was also shown to be over expressed in other solid tumors such as nasopharyngeal, neuroblastoma, colorectal and lung cancers. In this work we studied Livin expression in normal hematopoietic cells as well as hematologic malignancies. Using immunohistochemistry staining for Livin we evaluated its expression in reactive lymph nodes (LN) and showed that in contrast to Bcl2, Livin was detected in highly proliferating germinal centers. In normal bone marrow Livin was detected in Megakaryocytes and immature myeloid precursors. In peripheral blood mononuclear cells, using quantitative RT-PCR, we found that Livin expression was down regulated in activated monocytes and T cells while in B cells, Livin was upregulated upon activation. We studied bone marrow and LNs from 84 patients (pts) with hematologic malignancies. Positive immunohistochemistry staining was found in the malignant cells of all pts with DLBC NHL (31 pts), follicular lymphoma (12 pts) and multiple myeloma (15 pts). Peripheral blood samples from 28 B-CLL pts were compared with healthy controls’ B cells. High mRNA levels were detected in 43% of the pts, in correlation with older age (p<0.05). On the other hand, Livin was not expressed in Hodgkin’s disease (4 pts and 4 cell lines) and only 6/29 pts with AML had high levels of Livin in RT-PCR without any clinical correlation. Our data demonstrate that Livin is over expressed in activated normal B cells both in peripheral blood and LN as well as in most B cell lymphoproliferative diseases. Further investigation will establish the role of Livin over expression in hematologic malignancies.

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