Metformin represses the carcinogenesis potentially induced by 50 Hz magnetic fields in aged mouse fibroblasts via inhibition of NF-kB.

Aging is a risk factor for various human disorders, including cancer. Current literature advocates that the primary principles of aging depend on the endogenous stress-induced DNA damage caused by reactive oxygen species 50 Hz low-frequency magnetic field was suggested to induce DNA damage and chromosomal instability. NF-kB, activated by DNA damage, is upregulated in age-related cancers and inhibition of NF-kB results in aging-related delayed pathologies. Metformin (Met), an NF-kB inhibitor, significantly reduces both NF-kB activation and expression in aging and cancer. This in vitro study, therefore, was set out to assess the effects of 5mT MF in 50 Hz frequency and Met treatment on the viability and proliferation of aged mouse NIH/3T3 fibroblasts and expression of RELA/p65, matrix metalloproteinases MMP2 and MMP9, and E-cadherin (CDH1) genes. The trypan blue exclusion assay was used to determine cell viability and the BrdU incorporation assay to determine cell proliferation. The MMP-2/9 protein analysis was carried out by immunocytochemistry, NF-kB activity by ELISA and the expressions of targeted genes by qRT-PCR methods. Four doses of Met (500 uM, 1 mM, 2 mM and 10 mM) suppressed both the proliferation and viability of fibroblasts exposed to the MF in a dose-dependent pattern, and the peak inhibition was recorded at the 10 mM dose. Met reduced the expression of NF-kB, and MMP2/9, elevated CDH1 expression and suppressed NF-kB activity. These findings suggest that Met treatment suppresses the carcinogenic potential of 50 Hz MFs in aged mouse fibroblasts, possibly through modulation of NF-kB activation and epithelial-mesenchymal transition modulation.

Midkine silencing enhances the anti-prostate cancer stem cell activity of the flavone apigenin: cooperation on signaling pathways regulated by ERK, p38, PTEN, PARP, and NF-κB.

Prostate cancer (PCa) is the most common cancer in men worldwide. Midkine (MK) is overexpressed in PCa, as well as in tumor-initiating cells termed cancer stem cells (CSCs). Apigenin is a dietary flavone with considerable anti-tumor activities. In this study, we explored the possible therapeutic use of MK silencing, apigenin treatment, and a combination of both on human PCa and prostate cancer stem cells (PCSCs). CD44+CD133+ PC3 and CD44+ LNCaP CSCs were isolated from their parent cell lines. Both MK knockdown and apigenin treatment resulted in loss of cell viability in PCSCs, and these effects were significantly elevated when apigenin was applied with MK silencing. Combined treatment of CD44+CD133+ PC3 cells with apigenin and MK siRNA was also more effective in inducing apoptotic and non-apoptotic cell death when compared with individual applications. Treatment of CD44+ LNCaP cells with apigenin significantly decreased viability, although the combination treatment did not markedly alter the individual therapy. Molecular events underlying cell cycle arrest and inhibition of the survival, proliferation, and migration of CD44+CD133+ PC3 cells were found to be associated with upregulated p21, p27, Bax, Bid, caspase-3, and caspase-8 expression, as well as downregulated p-p38, p-ERK, NF-κB, and PARP. In addition, the combination of apigenin treatment and MK silencing showed better outcomes on the anticancer efficacy of docetaxel in CD44+CD133+ PC3 cells. In conclusion, MK-regulated events are different between PCSCs, and when combined with apigenin plus MK silencing, docetaxel treatment may be a valuable approach for the eradication of PCSCs.

Involvement of midkine in autoimmune and autoinflammatory diseases.

Midkine (MK) is a heparin-binding growth factor that markedly expressed during embryogenesis but downregulated to inconsiderable levels in healthy adults. However, MK is upregulated during tissue repair and in many pathologic conditions, mostly malignancies and inflammatory diseases. MK promotes a number of functions in target cells such as migration, proliferation, survival, growth, reproduction and repair, angiogenesis, and gene expression. It acts as a pro-inflammatory cytokine and contributes to chronic inflammation via promoting chemotaxis and tissue infiltration of neutrophils and macrophages. Furthermore, MK upregulated the production of various inflammatory mediators (i.e. interleukin (IL) 6 and IL8). Recent studies have demonstrated strong evidence that MK is involved in the onset and progression of autoimmune rheumatic diseases, including rheumatoid arthritis (RA), systemic lupus erythematosus (SLE), and Sjögren's syndrome (SS) and other autoimmune conditions such as multiple sclerosis (MS). Additionally, it has been shown that MK is overexpressed in two major clinically defined forms of inflammatory bowel diseases (IBD), Crohn's disease (CD) and ulcerative colitis (UC), which are classified as autoinflammatory diseases. Taken together, MK is involved in the pathogenesis of autoimmune and autoinflammatory diseases and may serve as an indicator and biomarker in these conditions. Furthermore, MK inhibitors are expected to contribute in the management of these diseases.

1,25-Dihydroxyvitamin D3 stimulates odontoblastic differentiation of human dental pulp-stem cells in vitro.

BACKGROUND: 1,25-Dihydroxyvitamin D3 (1,25-OH D3) plays an important role in mineralized tissue metabolism, including teeth. However, few studies have addressed its role in odontoblastic differentiation of human dental pulp-stem cells (hDPSCs). AIM: This study aimed to understand the influence of various concentrations of 1,25-OH D3 on the proliferation capacity and early dentinogenesis responses of hDPSCs. MATERIALS AND METHODS: hDPSCs were obtained from the impacted third molar teeth. Monolayer cultured cells were incubated with a differentiation medium containing different concentrations of 1,25-OH D3 (0.001, 0.01, and 0.1 µM). All groups were evaluated by S-phase rate [immunohistochemical (IHC) bromodeoxyuridine (BrdU) staining], STRO-1 and dentin sialoprotein (DSP)+ levels (IHC), and alkaline phosphatase (ALP, enzyme-linked immunosorbent assay (ELISA)) levels. RESULTS: The number of cells that entered the S-phase was determined to be the highest and lowest in the control and 0.001 µM 1,25-OH D3 groups, respectively. The 0.1 µM vitamin D3 group had the highest increase in DSP+ levels. The highest Stro-1 levels were detected in the control and 0.1 µM 1,25-OH D3 groups, respectively. The 0.1 µM 1,25-OH D3 induced a mild increase in ALP activity. CONCLUSIONS: This study demonstrated that 1,25-OH D3 stimulated odontoblastic differentiation of hDPSCs in vitro in a dose-dependent manner. The high DSP + cell number and a mild increase in ALP activity suggest that DPSCs treated with 0.1 µM 1,25-OH D3 are in the later stage of odontoblastic differentiation. The results confirm that 1,25-OH D3-added cocktail medium provides a sufficient microenvironment for the odontoblastic differentiation of hDPSCs in vitro.

Ficus carica latex prevents invasion through induction of let-7d expression in GBM cell lines.

Glioblastoma multiforme (GBM) is one of the deadliest human malignancies. A cure for GBM remains elusive, and the overall survival time is less than 1 year. Thus, the development of more efficient therapeutic approaches for the treatment of these patients is required. Induction of tumor cell death by certain phytochemicals derived from medicinal herbs and dietary plants has become a new frontier for cancer therapy research. Although the cancer suppressive effect of Ficus carica (fig) latex (FCL) has been determined in a few cancer types, the effect of this latex on GBM tumors has not been investigated. Therefore, in the current study, the anti-proliferative activity of FCL and the effect of the FCL-temozolomide (TMZ) combination were tested in the T98G, U-138 MG, and U-87 MG GBM cell lines using the WST-1 assay. The mechanism of cell death was analyzed using Annexin-V/FITC and TUNEL assays, and the effect of FCL on invasion was tested using the chick chorioallantoic membrane assay. To determine the effect of FCL on GBM progression, the expression levels of 40 GBM associated miRNAs were analyzed in T98G cells using RT-qPCR. According to the obtained data, FCL causes cell death in GBM cells with different responses to TMZ, and this effect is synergistically increased in combination with TMZ. In addition, the current study is the first to demonstrate the effect of FCL on modulation of let-7d expression, which may be an important underlying mechanism of the anti-invasive effect of this extract.

Antibacterial effect of Kenger gum on mutans streptococci and its cytotoxic effect on the 3T3 fibroblast cell line.

PURPOSE: To evaluate the antibacterial effect of Kenger gum on mutans streptococci (in vivo) and Streptococcus mutans (in vitro) and its cytotoxic effect on the 3T3 fibroblast cell line. MATERIALS AND METHODS: In vitro antibacterial activity of Kenger gum extracts against S.mutans was determined by the disk-diffusion method. The broth dilution method was used to determine the minimum inhibitory concentration (MIC). The cytotoxic effect on 3T3 fibroblast cells at different time intervals was determined using cell culture and viability assays. Clinical studies were then performed on 20 healthy adult subjects, where a sugar-free chewing gum was used as a control. To determine the MS counts, oral rinse samples were taken before chewing as well as 30 and 60 min after 15 min of chewing. Repeated-measures ANOVA was used to compare the bacteria level in the oral rinse samples between the two chewing gums. The Least Significant Difference test was used for adjustment for multiple comparisons. RESULTS: The MIC of the acetone extract of Kenger gum was 30 µg/ml. The acetone extract of Kenger gum possessed moderate antiproliferative properties against the non-tumorigenic cell line 3T3. A statistically significant decrease was observed for both chewing gums at 30 and 60 min. The decrease continued at 60 min after chewing Kenger gum, while the values for control gum tended to approach the baseline after 60 min. CONCLUSION: This preliminary study showed that Kenger gum had particular and prolonged antibacterial activity against S. mutans and salivary mutans streptococci.

Notch signaling-related therapeutic strategies with novel drugs in neuroblastoma spheroids.

Neuroblastoma is a severe pediatric tumor characterized by poor prognosis. Identification of novel molecular targets and diversion of investigations on new drug trials is mandatory for cancer therapy. In this study, vinorelbine tartrate, lithium chloride, clomipramine, and medroxyprogesterone acetate are used for the possible new treatment modalities in neuroblastoma cells. Notch and c-kit are novel molecules in cancer research, and Notch pathway is one of the emerging molecules in the neuroblastoma pathogenesis. Cytotoxic effects of these drugs at different time points, with different doses were studied in the SH-SY5Y human neuroblastoma cell line. Analysis of Notch and c-kit signaling with immunohistochemistry were constituted in multicellular tumor spheroids, and morphologic investigation was performed for digital imaging of cancer stem cells (CSCs) with electron microscopy. Size kinetics of spheroids was also determined after drug treatment. Results showed that all drugs were cytotoxic for neuroblastoma cells. Yet, this cytotoxic action did not correlate with the inhibitory effects in cell signaling. Neuroblastoma spheroids showed increased immunoreactivity of Notch signaling and c-kit. Altered ultrastructural CSCs morphology was observed after clomipramine and medroxyprogesterone acetate treatment compared with other drugs. Lithium chloride showed cellular membrane destruction for both CSCs and the remaining population. In this study, independent effects of cytotoxicity in tumor cells with respect to CSCs were determined. Redundant cells, which are the bulk population in tumor a compound, destroyed with therapy, were neither a target for treatment nor a remarkable investigation of cancer.

Effects of hemostatic agents on fibroblast cells.

PURPOSE: Hemostatic agents may be used topically to control hemorrhage, especially in patients with bleeding disorders. The agent used may have a negative effect on the tissue prolonging the healing time. The aim of this study was to compare the effects of 3 different hemostatic agents on fibroblast cells on a rat primary fibroblast cell culture model. MATERIALS AND METHODS: Ankaferd Blood Stopper (ABD) (Ankaferd Pharmaceuticals Cosmetics Production and Marketing Co.), fibrin glue, and tranexamic acid were the agents to be evaluated for their effects on cell proliferation, cell numbers, cell viability, and cell morphology. Also lactate dehydrogenase, basic fibroblast growth factor, and vascular endothelial growth factor C levels were measured. RESULTS: It was found that all of the agents used in the study have negative effects on fibroblasts, with ABD having the lowest values of cell proliferation, cell number, and cell viability. CONCLUSION: The results of this study indicate that ABD, fibrin glue, and tranexamic acid may negatively affect tissue healing.

WNT1 gene expression alters in heterogeneous population of prostate cancer cells; decreased expression pattern observed in CD133+/CD44+ prostate cancer stem cell spheroids.

PURPOSE: Established cancer cell lines contain cancer stem cells (CSCs) which can propagate to form three dimensional (3D) tumor spheroids in vitro. Aberrant activation of WNT signaling is strongly implicated in the progression of cancer and controls CSCs properties. In this study we hypothesized that when cells were maintained as spheroids, the structure of CSCs could show differentiation between CSCs and non- CSCs. METHODS: CD133+/CD44+ cancer-initiating cells were isolated from DU-145 human prostate cancer cell line monolayer cultures, propagated as tumor spheroids and compared with the remaining heterogeneous cancer cells bulk population. The expression levels of WNT1, FZD1, ADAR, APC, AXIN, BTRC, FRAT1 and PPARD genes were measured by polymerase chain reaction (PCR) array assay and the protein expression levels of WNT1, FZD and AXIN by immunohistochemistry. RESULTS: The expression levels of WNT pathway-related molecules were found to increase in both CSCs and non- CSCs when CSCs were maintained as spheroids. However, different expression profiles were observed when CSCs and non-CSCs were compared. In spheroids, the expression levels of FZD1, APC, ADAR, WNT1, PPARD genes in CSCs decreased when compared to non-CSCs. Interestingly, when CSCs from spheroids were compared with CSCs from monolayers the most significant decrease was observed in FZD1 and increase in APC genes. CONCLUSION: It is possible to assume that intracellular signaling of WNT-related molecules in the nucleus and/or cytoplasm might play an important role but it is independent from increased ligand expression and this expression strongly differentiate CSCs and non-CSCs population. This unexpected expression could be important for CSCs behavior and targeting this pathway could have therapeutic implications in cancer.

Midkine: A Cancer Biomarker Candidate and Innovative Therapeutic Approaches.

Midkine (MDK) is a protein that contributes to both physiological and pathological processes. Several studies provide insight into the different roles of MDK in development, tissue repair, neural plasticity, and health and disease processes. This research further examined how MDK contributed to conditions, including neurological diseases, inflammation, and ischaemia. Furthermore, MDK overexpression has been reported in many kinds of cancer and MDK is recognized as a malignancy marker. MDK stimulates pro-tumor activity by regulating a number of signaling pathways, which increase cancer cell proliferation, survival, metastasis, and treatment resistance. However, studies have shown that MDK also functions as a molecule that regulates drug resistance. Several cancer therapy techniques have been suggested to modify MDK function, including antibody-based therapies, oligonucleotides, oncolytic viruses, and small compounds. Further research and experimentation will be required to establish the therapeutic relevance and efficacy of these treatments. This review focuses on the role of MDK in cancer biology, as well as its multiple different roles in health and disease processes.

Notch signaling pathway in cumulus cells can be a novel marker to identify poor and normal responder IVF patients.

PURPOSE: To identify expression of Notch signaling proteins and its ligands in human cumulus cells which were obtained by follicle aspiration and to compare the differences of this protein expression between the normal and poor responder patients. METHODS: 47 patients who applied to the assisted reproductive treatments with various infertility problems were included to the study. Controlled ovarian hyperstimulation was performed by using GnRH agonist and gonadotropins. Serum hormon levels were measured by using Chemilluminescent Microparticle Immunoassay method for each patient. After ultrasonographic ovarian follicle screening, oocytes were retrievaled. Cumulus cells obtained from the follicles were cultured for 72 h and immunuhistochemistry were performed for Notch1, Notch2, Notch3, Notch4, Jagged1 and Jagged2 proteins. Histological score (HSCORE) were applied to all of the samples. The association between Notch and its ligands protein expressions and the oocyte-embryo quality and fertilization rates were investigated. RESULTS: Significant differences were observed between the mean values of age, AMH and FSH in the 2 groups, respectively (p < 0.05). However, the mean female infertility duration and total gonadotropin dose did not differ significantly between normal and poor responder groups. All the patients cumulus cells expressed Notch1, Notch2, Notch3, Notch4, Jagged1 and Jagged2. There was a significant difference (p < 0.05) only for Notch2 between the 2 groups and a positive correlation between Notch2 and Notch3 (r = 547, p = 0.00) expressions were noted. Furthermore, no correlations were observed between the following: Notch1, Notch2, Notch3, Notch4, Jagged1, and Jagged2 expression; mature oocyte number; fertilization rates, and embryo quality percentage in both of the groups. CONCLUSION: Notch signalling proteins can be an indicator for understanding the ovarian response in ovulation induction.

Low-level laser therapy vs. pulsed electromagnetic field on neonatal rat calvarial osteoblast-like cells.

To compare the effects of pulsed electromagnetic field (PEMF) and low-level laser therapy (LLLT) on osteoblast cells in a cell culture model. Fifty thousand neonatal rat calvarial osteoblast-like cells per milliliter were seeded and 0.06 mT PEMF, 0.2 mT PEMF, and LLLT at 808 nm were applied for 24 and 96 h on the cells. To evaluate cellular proliferation and differentiation, specimens were examined for DNA synthesis, alkaline phosphatase (ALP) activity, cell numbers, and viability of the cells. Morphological appearances of the cells were observed using scanning electron microcopy after 24 and 96 h of incubation. At 24 and 96 h, the control group had a higher cell proliferation than 0.06 and 0.2 mT PEMF groups (p=0.001). At 96 h, 0.2 mT PEMF group had higher cell proliferation rate than 0.06 mT PEMF and LLLT groups (p=0.001). The cell count and cell viability in 0.2 mT PEMF group were higher than the 0.06-mT PEMF and LLLT groups, although these differences were not statistically significant at 96 h (p>0.05). At 24 and 96 h, cell viability in the control group was higher than the test groups. Alkaline phosphatase levels of the groups were comparable in both time intervals (p>0.05). 0.2 mT PEMF application on osteoblast-like cells led to cell proliferation and differentiation better than 0.06 mT PEMF and LLLT at 808 nm, although a remarkable effect of both PEMF and LLLT could not be detected. The ALP activity of 0.2 and 0.06 mT PEMF and LLLT were comparable.

The effects of pulsed electromagnetic field (PEMF) on osteoblast-like cells cultured on titanium and titanium-zirconium surfaces.

BACKGROUND: Commercially pure Ti, together with Ti Ni, Ti-6Al-4V, and Ti-6Al-7Nb alloys, are among the materials currently being used for this purpose. Titanium-zirconium (TiZr) has been developed that allows SLActive surface modification and that has comparable or better mechanical strength and improved biocompatibility compared with existing Ti alloys. Furthermore, approaches have targeted making the implant surface more hydrophilic, as with the Straumann SLActive surface, a modification of the SLA surface. PURPOSE: The aim of this study is to evaluate the effects of pulsed electromagnetic field (PEMF) to the behavior of neonatal rat calvarial osteoblast-like cells cultured on commercially pure titanium (cpTi) and titanium-zirconium alloy (TiZr) discs with hydrophilic surface properties. MATERIALS AND METHODS: Osteoblast cells were cultured on titanium and TiZr discs, and PEMF was applied. Cell proliferation rates, cell numbers, cell viability rates, alkaline phosphatase, and midkine (MK) levels were measured at 24 and 72 hours. RESULTS: At 24 hours, the number of cells was significantly higher in the TiZr group. At 72 hours, TiZr had a significantly higher number of cells when compared to SLActive, SLActive + PEMF, and machine surface + PEMF groups. At 24 hours, cell proliferation was significantly higher in the TiZr group than SLActive and TiZr + PEMF group. At 72 hours, TiZr group had higher proliferation rate than machine surface and TiZr + PEMF. Cell proliferation in the machine surface group was lower than both SLActive + PEMF and machine surface + PEMF. MK levels of PEMF-treated groups were lower than untreated groups for 72 hours. CONCLUSIONS: Our findings conclude that TiZr surfaces are similar to cpTi surfaces in terms of biocompatibility. However, PEMF application has a higher stimulative effect on cells cultured on cpTi surfaces when compared to TiZr.

In vitro effects of heparin-binding epidermal growth factor on adhesion stage of implantation.

A member of the epidermal growth factor (EGF) family, the heparin-binding EGF (HB-EGF) is expressed in the uteri of both humans and mice during the implantation process. To study the effects of HB-EGF on adhesion stage, we developed an in vitro implantation model employing Ishikawa cell line and JAR cell line, which may attach to Ishikawa cells. For 1, 6, 12, and 24 hours, co-cultures of JAR spheroids grown on Ishikawa monolayers were treated with 1, 10, and 100 ng∕mL doses of HB-EGF. Using immunocytochemistry and Western blot analysis, the effects of HB-EGF on the protein expressions of E-cadherin, Erb-B2 receptor tyrosine kinase 4 (ErbB4), and integrin ανß3 in Ishikawa and JAR cells were examined semi-quantitatively and quantitatively. Ultrastructural changes of in vitro implantation model were investigated by transmission electron microscopy. We revealed that HB-EGF influenced trophoblast cell adhesion to endometrial cells by upregulating the expression of the proteins ErbB4 and trophoblastic integrin ανß3. Decrease in trophoblastic E-cadherin expression and increase in endometrial E-cadherin expression were demonstrated accompanying morphological variations in cells required for the invasion. We discovered ultrastructurally that Ishikawa cells acquired uterodome-like appearance, including the organelles, when 10 and 100 ng∕mL dosages of HB-EGF were administered for 12 and 24 hours. However, following additional hours of adhesion and invasion, their intercellular spaces enlarged. The trafficking of vesicular transport was enhanced by JAR spheroids. We therefore discovered that in this implantation paradigm, HB-EGF may enhance the receptivity of Ishikawa cells and the adherence of JAR cells.

Enhanced nerve growth factor expression by mast cells does not differ significantly between idiopathic and allergic rhinitis.

BACKGROUND: The role of neurotrophins in allergic rhinitis (AR) has been well studied, but it has not been evaluated in idiopathic rhinitis (IR). OBJECTIVE: We aimed to evaluate the nasal ß-nerve growth factor (ß-NGF) expressions of mast cells in patients with AR and IR. METHODS: Seventeen patients with house dust mites-induced persistent moderate/severe allergic rhinitis (mean age: 29.7 ± 11.96), 14 patients with idiopathic rhinitis (mean age, 29.3 ± 10.62), and 16 healthy controls (29.9 ± 11.57) were included in the study. Nasal biopsy specimens were taken from the posterior part of the inferior turbinate from all of the study subjects. Nasal ß-nerve growth factor and its receptors, pan-neurotrophin receptor p75, and tyrosine kinase A (trkA) were assessed with an immunofluorescence assay. Mast cells were determined by both an immunofluorescence assay and immunohistochemistry as tryptase-positive cells. RESULTS: The ß-NGF, trkA, and p75 receptor counts were significantly higher in AR and IR patients than in the control group (P < .001, for each), but they were not different between AR and IR patients. Similarly, the ratio of ß-NGF+ mast cells/total mast cells and the ratio of ß-NGF+ mast cells/total ß-NGF+ cells in AR and IR patients was found to be elevated when compared with the control group (P < .001, P < .001, P < .001, and P = .046, respectively); furthermore, the 2 ratios were not statistically different between the 2 patient groups. CONCLUSION: The increase in ß-NGF-expressing mast cells does not differ between idiopathic and allergic rhinitis. Therefore, we propose that mast cells do play a role in the pathogenesis of IR as important as in that of AR.

Cancer stem cell and embryonic development-associated molecules contribute to prognostic significance in ovarian cancer.

OBJECTIVES: Embryonic molecules and cancer stem cell signaling resemble each other, and they organize cancer modality. We hypothesized that similar immunohistochemical expressions between tumor spheroids and patients' samples compared with clinical relevance would give an important clue in patients' prognosis. METHODS: Immunohistochemical expression of c-kit, Notch1, Jagged1, and Delta1 in 50 cases of primary ovarian tumors (10 endometrioid, 10 serous, 10 mucinous adenocarcinoma, 10 borderline serous, and 10 borderline mucinous tumors) and MDAH-2774 spheroids were investigated. Results were compared in both spheroids and tumor samples with morphologic parameters (histological grade) and clinical data (age, stage, tumor size, and metastasis). RESULTS: High c-kit and Notch1 immunoreactivity was shown in spheroids, but interestingly immunoreactivity of these molecules in tumor samples was different from patients' clinicopathological characteristics. In serous carcinoma, metastasis correlated with Notch1 immunoexpression; in mucinous carcinoma, Jagged1 immunohistochemistry correlated with grade, stage, and metastasis of tumor; in borderline serous and mucinous tumors, Jagged1 correlated with high grade. Moreover, Jagged1 correlated with stage and Notch1 with size in borderline mucinous tumor. Endometrioid carcinoma statistics showed that there was a correlation between age and Notch1 expression. CONCLUSION: Notch1, Jagged1, and Delta1 expressions might be useful markers for clinical prognosis of ovarian carcinomas; and Notch pathway, one of the most intensively studied putative therapeutic targets, may be a useful marker for cancer. Consequently, Jagged1 could be a marker for tumor grades and Notch1 as a marker for metastases.

Inhibition of cell survival, viability and proliferation by dentin adhesives after direct and indirect exposure in vitro.

OBJECTIVES: The influence of dentin adhesive systems (Scotchbond Multi-Purpose, XP Bond, Xeno V, Clearfil Protect Bond, AdheSE) on cell survival, viability and proliferation was characterized after direct and indirect exposure using different cell culture techniques. MATERIALS AND METHODS: The primers and cured bonding parts were directly exposed to cells using cell culture inserts, and complete materials were analyzed in a dentin barrier test. Cell responses were examined in 3T3 mouse fibroblasts after 24- and 72-h exposure periods by the estimation of total cell numbers (survival), apoptosis (viability) and cell proliferation. RESULTS: Cell numbers were effectively reduced by the primers of AdheSE, Protect Bond, and Scotchbond Multi-Purpose as well as XP bond after direct exposure in a cell culture insert test device. Likewise, Scotchbond Multi-Purpose primer induced a rate of apoptosis (93.9%) even higher than detected with Protect Bond primer (91.6%). Cell proliferation was entirely inhibited by primers and by Xp Bond as well. The Scotchbond Multi-Purpose was most cytotoxic in a dentin barrier test device after a 24-h indirect exposure. It also increased the percentage of cells in apoptosis to 15.4% compared to untreated controls. CONCLUSION: Unpolymerized primers of dentin adhesives were more cytotoxic than polymerized bonding counterparts. Moreover, total etch dentin adhesives were more cytotoxic than self-etch adhesives. CLINICAL RELEVANCE: When dentin adhesives are used in deep cavities without a protective dentin barrier the leachable hydrophobic and hydrophilic component of dentin adhesive systems can penetrate to the pulp and may induce cytotoxic responses in pulp tissues.

Delayed cutaneous wound healing in aged rats compared to younger ones.

Delayed wound healing in elderly males is a complex process in which the factors responsible are not fully understood. This study investigated the hormonal, oxidative and angiogenic factors affecting wound healing in aged rats. Two groups consisting of eight healthy male Wistar Albino rats [young (30 ± 7 days) and aged (360 ± 30 days)], and a cutaneous incision wound healing model were used. Scar tissue samples from wounds on the 7th, 14th and 21st days of healing were evaluated for hydroxyproline and vascular endothelial growth factor content. Macrophage, lymphocyte, fibroblast and polymorphonuclear cell infiltration; collagen formation and vascularization were assessed by light and electron microscopy. The free oxygen radical content of the wounds was measured by a chemiluminescence method. Blood sample analysis showed that the hydroxyproline and total testosterone levels were significantly higher, and the oxygen radical content was significantly lower in young rats. Histopathological, immunohistochemical and ultrastructural evaluations revealed higher amounts of fibroblasts and collagen fibers, and more vascularization in young rats. These results are indicative of the delayed wound healing in aged rats. A combination of multiple factors including hormonal regulation, free oxygen radicals and impaired angiogenesis appears to be the cause of delayed cutaneous healing.

Decreased therapeutic effects of noscapine combined with imatinib mesylate on human glioblastoma in vitro and the effect of midkine.

BACKGROUND: Glioblastoma (GBM) develops resistance to the advances in chemotherapy leading to poor prognosis and life quality. Consequently, new treatment modalities are needed. Our aims were to investigate the effects of combined noscapine (NOS) and imatinib mesylate (IM) on human GBM in vitro and the role of midkine (MK) in this new combination treatment. METHODS: Monolayer and spheroid cultures of T98G human GBM cell line were used to evaluate the effects of IM (10 µM), Nos (10 µM) and their combination on cell proliferation and apoptotic indexes, cell cycle, the levels of antiapoptotic MK, MRP-1, p170, PFGFR-α, EGFR, bcl-2 proteins, apoptotic caspase-3 levels, morphology (SEM) and ultrastructure (TEM) for 72 hrs. Results were statistically analyzed using the Student's t-test. RESULTS: The combination group induced highest decrease in cell proliferation and apoptotic indexes, caspase-3 levels, MRP-1 and PDGFR-α levels. The decrease in p170 levels were lower than IM but higher that NOS. The highest increases were in EGFR, MK, bcl-2 and cAMP levels in the combination group. The G0+G1 cell cycle arrest at the end of 72nd hr was the lowest in the combination group. Apoptotic appearence was observed rarely both in the morphologic and ultrastructural evaluation of the combination group. In addition, autophagic vacuoles which were frequently observed in the IM group were observed rarely. CONCLUSIONS: The combination of Nos with IM showed antagonist effect in T98G human GBM cells in vitro. This antagonist effect was correlated highly with MK levels. The effects of NOS on MRP-1, MK and receptor tyrosine kinase levels were firstly demonstrated in our report. In addition, we proposed that MK is one of the modulator in the switch of autophagy to cell death or survival/resistance.

Evaluation of light-emitting diode (LED-660 nm) application over primary osteoblast-like cells on titanium surfaces: an in vitro study.

BACKGROUND: The goal of this study was to evaluate the behavior of neonatal rat calvarial osteoblast-like cells cultured on different implant surfaces and exposed once or three times to a 660-nm light-emitting diode (LED). METHODS: An LED with a 660-nm wavelength was applied once or three times to cultured cells on standard and modified sandblasted acid-etched surfaces (SLA and SLActive; Straumann, Basel, Switzerland). To analyze the effect of the LED on cell proliferation, numbers, and viability, cells were cultured on titanium discs, and measurements were taken after 72 h. Cell proliferation rates were assessed using a bromodeoxyuridine immunohistochemical technique. Cell morphologies were evaluated by scanning electron microscopy (SEM). RESULTS: Osteoblast-like cells proliferated on all tested surfaces, with differences among groups in cell counts and DNA synthesis values. The application of one LED treatment caused a significant increase in cell count in the SLActive group in comparison with the SLA group (p = 0.001), whereas the application of three LED treatments caused a significant decrease in cell count in the SLA group compared with the SLActive group (p < 0.001). After 72 h, the number of cells was highest in the SLActive group exposed once to the LED. CONCLUSIONS: One LED application in the SLActive group resulted in significantly increased cell numbers. However, these findings were not exactly compatible with the SEM findings, which demonstrated fewer cells and weak attachments between cells and to the surface. Thus, further studies using different LED application times are needed to clarify the reason for the increased number of cells that are apparently incapable of attaching to the titanium surfaces after 72 h.