Ferit Avcu

Low-Level Laser Irradiation Affects the Release of Basic Fibroblast Growth Factor (bFGF), Insulin-Like Growth Factor-I (IGF-I), and Receptor of IGF-I (IGFBP3) from Osteoblasts

OBJECTIVE It was the aim of the present study to evaluate whether the laser irradiation of osteoblasts could enhance the release of growth factors including basic fibroblast growth factor (bFGF), insulin-like growth factor-I (IGF-I), and receptor of IGF-I (IGFBP3). BACKGROUND DATA Low-level laser therapy (LLLT) has been shown to have biostimulatory effects on various cell types by enhancing production of some cytokines and growth factors. MATERIALS AND METHODS Human mesenchymal stem cells (MSCs) were seeded in osteogenic medium and differentiated into osteoblasts. Three groups were formed: in the first group (single dose group), osteoblasts were irradiated with laser (685 nm, 25 mW, 14.3 mW/cm(2), 140 sec, 2 J/cm(2)) for one time; and in the second group, energy at the same dose was applied for 2 consecutive days (double dose group). The third group was not irradiated with laser and served as the control group. Proliferation, viability, bFGF, IGF-I, and IGFBP3 levels were compared between groups. RESULTS Both of the irradiated groups revealed higher proliferation, viability, bFGF, IGF-I, and IGFBP3 expressions than did the nonirradiated control group. There was increase in bFGF and IGF-I expressions and decrease in IGFBP3 in the double dose group compared to single dose group. CONCLUSIONS The results of the present study indicate that LLLT increases the proliferation of osteoblast cells and stimulates the release of bFGF, IGF-I, and IGFBP3 from these cells. The biostimulatory effect of LLLT may be related to the enhanced production of the growth factors.

In vitro synergistic cytoreductive effects of zoledronic acid and radiation on breast cancer cells

IntroductionBisphosphonates are mostly used in the treatment of bone metastases. They have been shown to act synergistically with other chemotherapeutic agents. It is not known, however, whether similar synergistic effects exist with radiation on breast cancer cells.MethodsHuman MCF-7 breast cancer cells were treated with up to 100 μM zoledronic acid, were irradiated with up to 800 cGy or were exposed to combinations of both treatments to determine the antiproliferative effects of zoledronic acid and radiation.ResultsZoledronic acid and radiation caused a dose-dependent and time-dependent decrease in cell viability (approximate 50% growth inhibition values were 48 μM and 20 μM for 24 hours and 72 hours, respectively, for zoledronic acid and 500 cGy for radiation). A synergistic cytotoxic effect of the combination of zoledronic acid and radiation was confirmed by isobologram analysis.ConclusionThese data constitute the first in vitro evidence for synergistic effects between zoledronic acid and radiation. This combination therapy might thus be expected to be more effective than either treatment alone in patients with metastatic breast carcinoma.

The bisphosphonate zoledronic acid inhibits the development of plasmacytoma induced in BALB/c mice by intraperitoneal injection of pristane

Abstract:  Objectives: Bisphosphonates (BPs) are mostly used in the palliative care of myeloma‐associated osteolytic lesions. Recent studies have suggested that BPs may also exert direct antitumor effects on myeloma cells. We have investigated the effect of the potent bisphosphonate, zoledronic acid (ZOL), on the development of pristane (2,6,10,14‐tetramethylpentadecane)‐induced plasmacytoma (PCT) in six‐week‐old BALB/c mice. Methods: Different groups of pristane‐treated mice also received ZOL (100 μg/kg) commencing after the development of PCT or ZOL (20 μg/kg) from the first day. Control groups received pristane alone, ZOL alone (20 μg/kg), or phosphate‐buffered saline. The study was terminated on day 300, and the remaining mice were autopsied and abdominal tissues were examined histologically for PCT. Results and conclusions: Statistical analysis revealed a significant delay in PCT development in the group receiving pristane plus ZOL (20 μg/kg) from the first day compared to the groups receiving pristane alone and pristane combined with ZOL (100 μg/kg) after the appearance of PCT (Log‐rank, P = 0.0001 and 0.0001; respectively). Kaplan–Meier analysis revealed a significant difference in survival between the group treated with pristane alone and the groups receiving pristane plus ZOL (20 μg/kg) from the first day or ZOL (100 μg/kg) after the appearance of PCT (Log‐rank, P = 0.016 and 0.023; respectively). These results indicate a direct anti‐tumor effect of ZOL in pristane‐induced PCT development BALB/c mice, which may contribute to their significantly increased survival. This hypothesis should now be further investigated in clinical trials.

Upregulation of multi drug resistance genes in doxorubicin resistant human acute myelogeneous leukemia cells and reversal of the resistance

Abstract The major problem in the treatment of acute myeloid leukemia (AML) patients results from multidrug resistance to administered anticancer agents. Drug resistance proteins, MDR1 and MRP1, which work as drug efflux pumps, can mediate the multidrug resistance of human leukemia cells. In this study, the mechanisms of resistance to doxorubicin-induced cell death in human HL60 AML cells were examined. Continuous exposure of cells to step-wise increasing concentrations of doxorubicin resulted in the selection of HL60/DOX cells, which expressed about 10.7-fold resistance as compared to parental sensitive cells. The expression analyses of MRP1 and MDR1 drug efflux proteins in doxorubicin-sensitive and -resistant HL60 cells revealed that there was an upregulation of MRP1 gene in HL60/DOX cells as compared to parental sensitive cells. On the other hand, while there was no expression of MDR1 gene in parental cells, the expression of MDR1 gene was upregulated in HL60/DOX cells. HL60/DOX cells also showed cross-resistance to cytosine arabinoside (Ara-c). This resistance was reversed by a combination therapy of Ara-c and cyclosporine A. However, the expression levels of CD15 and CD16 surface markers were significantly decreased in HL60/DOX cells.

A novel mechanism of dasatinib-induced apoptosis in chronic myeloid leukemia; ceramide synthase and ceramide clearance genes.

Sphingolipids are bioeffector molecules that control various aspects of cell growth, proliferation, apoptosis, and drug resistance. Ceramides, the central molecule of sphingolipid metabolism, are inducer of apoptosis and inhibitors of proliferation. Sphingosine-1-phosphate (S1P) and glucosyleceramide, converted from ceramides by sphingosine kinase-1 (SK-1) and glucosyleceramide synthase (GCS) enzymes, respectively, inhibit apoptosis and develop resistance to chemotherapeutic drugs. In this study, we examined the therapeutic potentials of bioactive sphingolipids in chronic myeloid leukemia (CML) alone and in combination with dasatinib in addition to investigate the roles of ceramide-metabolizing genes in dasatinib-induced apoptosis. Cytotoxic effects of dasatinib, C8:ceramide, PDMP, and SK-1 inhibitor were determined by XTT cell proliferation assay. Changes in caspase-3 enzyme activity and mitochondrial membrane potential (MMP) were measured using caspase-3 colorimetric assay and JC-1 MMP detection kit. Expression levels of ceramide-metabolizing genes were examined by qRT-PCR. Application of ceramide analogs and inhibitors of ceramide clearance genes decreased cell proliferation and induced apoptosis. Targeting bioactive sphingolipids towards generation/accumulation of ceramides increased apoptotic effects of dasatinib, synergistically. It was shown for the first time that dasatinib induces apoptosis through downregulating expression levels of antiapoptotic SK-1 but not GCS, and upregulating expression levels of ceramide synthase (CerS) genes, especially CerS1, in K562 cells. On the other hand, dasatinib downregulates expression levels of both GCS and SK-1 and upregulate apoptotic CerS2, −5 and −6 genes in Meg-01 cells. Increasing endogenous ceramide levels and decreasing prosurvival lipids, S1P, and GC, can open the way of more effective treatment of CML.

Roles of ceramide synthase and ceramide clearence genes in nilotinib- induced cell death in chronic myeloidleukemia cells

In this study, we aimed to increase the sensitivity of human K562 and Meg-01 chronic myeloid leukemia (CML) cells to nilotinib by targeting bioactive sphingolipids, in addition to investigating the roles of ceramide metabolizing genes in nilotinib induced apoptosis. Cytotoxic effects of nilotinib, C8:ceramide, glucosyle ceramide synthase (GCS) and sphingosine kinase-1 (SK-1) inhibitors were determined by XTT cell proliferation assay and synergism between the agents was determined by isobologram analysis. Also, quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) results demonstrated that expression levels of longevity assurance (LASS) genes in response to nilotinib were correlated with sensitivity to nilotinib. For the first time, The results of this study showed for the first time that nilotinib induces apoptosis through upregulating ceramide synthase genes and downregulating SK-1 in CML cells in addition to inhibition of BCR/ABL. On the other hand, manipulating bioactive sphingolipids toward generation/accumulation of ceramides increased the apoptotic effects of nilotinib in CML cells.

Proteasome inhibitor bortezomib increases radiation sensitivity in androgen independent human prostate cancer cells.

OBJECTIVES To investigate the effects of a strong proteasome inhibitor, bortezomib alone or in combination with radiotherapy on androgen-independent DU145 human prostate cancer cells. Proteasomes play important roles in cell cycle, proliferation, apoptosis, angiogenesis, and cellular resistance to chemotherapy and radiotherapy. METHODS Increasing concentrations of bortezomib alone or in combination with radiation were applied to DU145 cells and IC(50) values that inhibited cell growth by 50% were determined by 3-(4,5-dimethylthiazol-2-yl)-2,5 diphenyltetrazolium-bromide assay. Apoptosis was determined using annexin V staining by flow cytometry. mRNA levels of proapoptotic caspase-3 and antiapoptotic Bcl-2 genes were examined by reverse transcriptase polymerase chain reaction. RESULTS The IC(50) value of bortezomib was found to be 28 microm although 400- and 800-cGy radiation decreased the cell proliferation by 14% and 28%, respectively. In 400- and 800-cGy radiation applied DU145 cells, IC(50) value of bortezomib decreased to 23- and 12 microm, respectively. Exposure to 5 microm bortezomib for 48 hours caused apoptosis in 35% of the population whereas 800-cGy radiation resulted apoptosis in 14% of cells. However, 42% of DU145 cells that were exposed to 800 cGy and 5 microm bortezomib underwent apoptosis. Reverse transcriptase polymerase chain reaction results showed a significant decrease in mRNA levels of antiapoptotic Bcl-2 gene and an increase in proapoptotic caspase-3 gene expression in the combination group compared to control group. CONCLUSIONS Bortezomib increases radiation sensitivity in androgen-independent human DU145 prostate cancer cells through inhibition of Bcl-2 and induction of caspase-3 genes.

Aberrant expressions of leptin and adiponectin receptor isoforms in chronic myeloid leukemia patients

BACKGROUND Leptin and adiponectin receptors mediate the role of leptin in stimulating the growth of leukemic cells and the protective function of adiponectin undertaken in several malignancies such as leukemia. In this study, we investigated the involvement of the expression of leptin and adiponectin receptors in chronic myeloid leukemia (CML) pathogenesis. METHODS The expression of leptin receptor isoforms, OB-Rt, OB-Ra, and OB-Rb, and the expression of adiponectin receptors, AdipoR1 and AdipoR2, were measured as mRNA levels in two CML cell lines (K562 and Meg-01) and 20 CML patients and 24 healthy controls by using RT-PCR. RESULTS OB-Rt and OB-Ra isoforms expression of the leptin receptors were found to be significantly lower in Meg-01 cell lines than K562 cells. All leptin receptors were downregulated in CML patients and more particularly OB-Rb level was found to be undetectably low in normal PBMC as well as in CML patients. AdipoR1 expression level was higher in Meg-01 than in K562, whereas AdipoR2 level was found to be unchanged in both cell lines. Interestingly, while AdipoR1 expression increased in CML patients, AdipoR2 decreased. Moreover, imatinib therapy did not affect both leptin and adiponectin isoform expressions. CONCLUSION While the decrease in leptin receptor levels in CML patients was confirmed, the increase in AdipoR1 levels and relevant decrease in AdipoR2 levels depicted their possible involvement in CML pathogenesis. This suggests different functions of adiponectin receptors in CML development.

Effect of cobalt-60 (γ radiation) on multidrug-resistant multiple myeloma cell lines

Emergence of resistance to chemotherapy and radiotherapy is a major obstacle for the successful treatment of MM (multiple myeloma). Prednisone, vincristine and melphalan are commonly used chemotherapeutic agents for the treatment of MM. In the current study, we examined the presence of possible cross‐resistance between these drugs and gamma (γ) radiation. Prednisone, vincristine and melphalan resistant RPMI‐8226 and U‐266 MM cells were generated by stepwise increasing concentrations of the drugs. The sensitive and resistant cells were exposed to 200‐ and 800 cGy γ radiation, and proliferation was examined by XTT {2,3‐bis(2‐methoxy‐4‐nitro‐5‐sulfophenyl)‐5‐[(phenylamino)carbonyl]‐2H‐tetrazolium hydroxide} assay. The results showed that Prednisone‐ and melphalan‐resistant RPMI‐8226 cells were also cross‐resistant to 200 and 800 cGy γ radiation application, while vincristine‐resistant cells did not show resistance. On the other hand, Prednisone‐, vincristine‐ and melphalan‐resistant U‐266 cells showed cross‐resistance to 200‐ and 800 cGy γ radiation application. These results demonstrated that MM cells resistant to anticancer agents respond to radiation in different levels. These findings may be important in the clinical applications of radiation therapy in the treatment of vincristine resistant MM.

Soluble CD40 ligand, soluble P-selectin and von Willebrand factor levels in subjects with prediabetes: the impact of metabolic syndrome.

OBJECTIVES The data regarding circulating levels of markers of platelet activation and endothelial function in people with prediabetes are scant. The aim of the present study was to search blood levels of soluble CD40 ligand (sCD40L), soluble P-selectin (sP-sel) and von Willebrand Factor (vWF) in subjects with prediabetes, along with the effects of the metabolic syndrome (MetS) on these markers. DESIGN AND METHODS A total of 77 prediabetic individuals and 81 age, sex and body mass index matched healthy subjects with normal glucose tolerance (NGT) were prospectively analyzed. Anthropometric parameters, fasting plasma glucose, blood d lipid profiles and insulin resistance indexes were determined. Plasma sCD40L, sP-sel and vWF levels were measured by ELISA. RESULTS sCD40L, sP-sel and vWF levels in the prediabetic group were similar to those in the controls. However, prediabetic subjects with the MetS had significantly higher level of sCD40L compared to those without MetS. Moreover, sCD40L level correlated significantly with waist circumference, systolic blood pressure and HDL-cholesterol level in the patient group. CONCLUSION These data imply that MetS may contribute, at least in part, to the mechanism of platelet activation and endothelial dysfunction in people with prediabetes.