Investigation of the effects of rapamycin on the mTOR pathway and apoptosis in metastatic and non-metastatic human breast cancer cell lines.

AIM: The aim of this study was to analyze the effects of rapamycin treatment on apoptosis via mTOR pathway in metastatic and non-metastatic human breast cancer cell lines by immunohistochemical and TUNEL analysis. METHOD: MCF-7 and MDA-MB 231 cell lines were incubated under standard conditions forming Rapamycin and control groups. In immunohistochemical evaluation; mTOR pathway was evaluated with anti-IGF1, anti-PI3K, anti-pAKT1/2/3, anti-mTORC1, anti-mTORC2 and anti-ERK1 antibodies. The effect of apoptosis was also confirmed by TUNEL method. RESULTS: In this study, activation of PI3K/AKT/mTOR and related molecular pathways in the MDA-MB 231 and MCF-7 breast cancer cell line was evaluated and it was observed that these pathways could play a key role in cancer development. Increased apoptotic cells were observed in mTORC1 inhibition by Rapamycin administration. CONCLUSION: Targeting the mTOR pathway in breast cancer treatment may be a treatment option. In addition, the demonstration and confirmation of increased apoptosis in Rapamycin treated groups suggested that Rapamycin, an inhibitor of mTOR, is promising in the treatment of breast cancer (Tab. 2, Fig. 3, Ref. 66).

A randomized study to evaluate post-dural puncture headache after cesarean section: Comparison with median and paramedian approaches.

BACKGROUND: Although the most popular anesthesia technique for cesarean is spinal anesthesia, its most common complication is post-dural puncture headache (PDPH). AIM: We aimed to determine the effect of median and paramedian approaches during spinal anesthesia on PDPH in patients undergoing cesarean section. SUBJECTS AND METHODS: 200 pregnant women between the ages of 19-45 years, ASA physical status II, scheduled to undergo elective cesarean section under spinal anesthesia, were studied. The patients were randomized into two groups: Group M; (n = 100) spinal anesthesia with the median approach, Group PM; (n = 100) spinal anesthesia with paramedian approach. The patients were questioned for the possible occurrence of PDPH on the first, third and seventh postoperative days. A telephone follow-up call was used if the hospital stay was shorter than seven days. Post-dural puncture headache was evaluated according to the International Classification of Headache Disorders (ICHD-III) diagnostic criteria. Normally distributed data were summarized using mean and standard deviation. Skewed data were summarized using median (range). RESULTS: A total of 200 patients completed the study. There were no statistically different between the groups by comparing the incidence and characteristics of PDPH (32% vs. 28%, P = 0.548). Most patients rated their pain intensity during PDPH as mild to moderate in both groups (p = 0.721). PDPH onset time was 2 (1-4) days in Group PM versus 3 (1-7) days in Group M (p = 0.173). No patient needed for epidural blood patch in both groups. CONCLUSIONS: Spinal anesthesia with a median or paramedian approach at cesarean section has no effect on the incidence of PDPH, but we believe that there has been a need for further studies with larger or different patient populations.

The role of stem/progenitor cells and Wnt/ß-catenin signaling pathway in the patients with prostate cancer.

AIM: The aim of this paper was to investigate the possible effect of cancer stem cells (CSCs) and relationship with Wnt/ß-catenin signaling pathway progressing of prostate cancer. METHODS: Thirty men with a pathological diagnosis of benign prostate hyperplasia (BPH) (group 1, N.=10), prostate cancer with a gleason score of ≤6 (group 2, N.=10), and prostate cancer with a gleason score of >6 (group 3, N.=10) were included in the study. The patients' groups were compared in terms of immunoreactivity strength of prostatic stem/progenitor cell surface markers including CD133 and CD117. We also compared the immunoreactivity of Wnt7a, a part of Wnt signaling pathway which has a potential role in the progression of several cancers including prostate cancer. The immunoreactivity of Frizzled 6 (Fzd 6) which is the receptor of Wnt family was also evaluated in all groups. RESULTS: Immunohistochemical analyses demonstrated that although CD133 immunoreactivity was positive in all groups, immunoreactivity was significantly stronger in group 3 when compared to other groups. While CD117 immunoreactivity was negative in group 1 and 2, it was positive in group 3. Wnt7a immunoreactivity was weak in all groups and Fzd 6 immunoreactivity was stronger in group 1 and 3 when compared to group 2. CONCLUSION: Our findings demonstrated that CSCs and Wnt signaling pathway have a potential role in the development and progression of prostate cancer.

Continuous and intermittent parathyroid hormone administration promotes osteogenic differentiation and activity of programmable cells of monocytic origin.

Bone healing deficiencies are challenging for orthopedic practice. The use of stem cells with scaffolds to treat bone tissue losses currently is popular for promoting regeneration of tissue. Programmable cells of monocytic origin (PCMO) may differentiate into three germ layers and may be a promising alternative treatment due to their stem cell-like properties. Parathyroid hormone (PTH) participates in bone metabolism. Intermittent administration of PTH promotes osteogenic activity of mesenchymal stem cdells (MSC). We investigated the osteogenic effects of continuous and intermittent administration of PTH on PCMO. Mononuclear cells were harvested from the peripheral blood of healthy donors. Isolated cells were cultured for six days in a de-differentiation medium. Indirect immunocytochemistry using anti-CD14, anti-CD45 and anti-CD90 primary antibodies, as well as electron microscopy were used to detect PCMO. PCMO then were cultured in an osteogenic differentiation medium supplemented with continuous or intermittent 50 ng/ml PTH. The PTH-free control group (CG), intermittent PTH treated group (IPG) and continuous PTH treated group (CPG) were cultured and assessed for their differentiation into osteogenic lineage cells by indirect immunocytochemistry using anti-collagen I, anti-osteonectin and anti-osteocalcin primary antibodies. Osteoblast-like cells obtained by continuous or intermittent PTH administration exhibited increased levels of collagen I, osteonectin and osteocalcin immunoreactivity. We found that continuous and intermittent PTH administration to PCMO enhanced their differentiation to osteogenic lineage cells and increased osteoblastic activity.

Effects of caffeic acid phenethyl ester (CAPE) on angiogenesis, apoptosis and oxidative stress in various cancer cell lines.

Cancer is a common cause of death worldwide. Approximately 80% of cancer patients use complementary or alternative medicines for treatment. Caffeic acid phenethyl ester (CAPE), the main active component of propolis, exhibits cytotoxic, antiproliferative and anti-cancer effects. Despite its anticancer effects CAPE exhibits no known harmful effects toward normal cells. We investigated the effects of CAPE on angiogenesis, apoptosis and oxidative stress using MDA MB-231, N2a and COLO 320 cell lines and CAPE treatments at 24 and 48 h. A two dimensional cell culture system was used and the findings were evaluated by an indirect immunohistochemical method and H-scores were calculated. CAPE was effective for all three cancer cell lines. After 24 and 48 h, we found a significant decrease in live cells and increased stress in the cells based on e-NOS and i-NOS levels.

Keratinocytes derived from embryonic stem cells induce wound healing in mice.

The skin plays an important role in defending the body against the environment. Treatments for burns and skin injuries that use autologous or allogenic skin grafts derived from adult or embryonic stem cells are promising. Embryonic stem cells are candidates for regenerative and reparative medicine. We investigated the utility of keratinocyte-like cells, which are differentiated from mouse embryonic stem cells, for wound healing using a mouse surgical wound model. Mice were allocated to the following groups: experimental, in which dressing and differentiated cells were applied after the surgical wound was created; control, in which only the surgical wound was created; sham, in which only the dressing was applied after the surgical wound was created; and untreated animal controls with healthy skin. Biopsies were taken from each group on days 3, 5 and 7 after cell transfer. Samples were fixed in formalin, then stained with Masson's trichrome and primary antibodies to interleukin-8 (IL-8), fibroblast growth factor-2 (FGF-2), monocyte chemoattractant protein-1 (MCP-1), collagen-1 and epidermal growth factor (EGF) using the indirect immunoperoxidase technique for light microscopy. Wound healing was faster in the experimental group compared to the sham and control groups. The experimental group exhibited increased expression of IL-8, FGF-2 and MCP-1 during early stages of wound healing (inflammation) and collagen-1 and EGF expression during late stages of wound healing (proliferation and remodeling). Keratinocytes derived from embryonic stem cells improved wound healing and influenced the wound healing stages.

Effects of high doses of dexamethasone on hemodynamic and immunohistochemical characteristics of acute paraquat intoxication in rat kidneys.

Paraquat (1,1'-dimethyl-4,4'-bipyridinium) (PQ), is a nonselective contact herbicide that is highly toxic to humans. The kidney is affected during PQ intoxication. Dexamethasone (Dexa) has anti-inflammatory effects and is used to treat cases of PQ poisoning. We investigated in rat kidney hemodynamic effects and immunohistochemical characteristics of Dexa treatment in acute PQ poisoning. Adult male rats were divided into four groups: 1, untreated control; 2, treated with 100 mg/kg Dexa; 3, treated with 25 mg/kg PQ; 4, treated with PQ + Dexa. Mean arterial pressure (MAP) and heart rate (HR) were recorded during the experimental period (2 h). Tissues were removed after 2 h and immunohistochemistry was performed after 24 h. Paraffin sections of kidney were prepared and anti-cyclo-oxygenase-1 (COX-1), anti-cyclo-oxygenase-2 (COX-2), anti-angiotensin converting enzyme (ACE), anti-aquaporin-1 (AQU-1), anti-vascular cell adhesion molecule (VCAM) primary antibodies were used for immunohistochemical examination. Immunoreactivities were scored as: (1) minimal, (2) weak, (3) mild, (4) moderate, (5) strong and (6) very strong. MAP and HR were measured at 10 min, 20 min, 1 h and 2 h. MAP at 10 and 20 min and 1 h was increased in the Dexa group. HR also was increased in all groups compared to controls at 2 h. Compared to groups 2 and 4, MAP values decreased significantly in group 3 at 1 h. The intensity of all of immunoreactivities was decreased in group 2. In group 3, immunoreactivities of COX-1, COX-2 and ACE were decreased compared to the control and the other groups, whereas AQU-1 and VCAM immunoreactivities were the same as the control group. ACE and VCAM immunoreactivities were decreased in group 4 compared to the control group, while COX-1, COX-2 and AQU-1 immunoreactivities were close to those of the control group. Dexa appears to be useful for treating PQ intoxication.

The role of hypoxia related angiogenesis in uterine smooth muscle tumors.

Mechanisms of hypoxia-related angiogenesis are important for uterine smooth muscle tumors. Factors that are related to angiogenesis during hypoxia include vascular endothelial growth factor (VEGF), hypoxia inducible factor 1α (HIF1α), T-cell intracellular antigen1 (TIA1), eukaryotic translation initiation factor 2α (eIF2α) and thrombospondin 1 (TSP1). We investigated immunoreactivities of VEGF, HIF1α, TIA1, eIF2α and TSP1 using an indirect immunoperoxidase method for formalin fixed, paraffin embedded tumors that had been diagnosed as leiomyoma (LMY), cellular leiomyoma (CLM) or leiomyosarcoma (LMS). TSP1 immunoreactivity was scored as moderate, mild or minimal, while VEGF, eIF2α and TIA1 immunoreactivities were scored as mild, moderate and strong in LMY, CLM and LMS samples, respectively. HIF1α immunoreactivity was scored as mild to minimal in LMY, CLM and LMS samples, but showed no statistically significant differences among samples. Although angiogenic factors showed strong immunohistochemical staining intensity in LMS, anti-angiogenic factors showed minimal immunohistochemical intensity. There was no difference in HIF-1α immunoreactivity compared to LMY, CLM and LMS samples. We suggest that HIF1α protein synthesis could be suppressed by eIF2α and TIA1. Furthermore, VEGF could be activated by pathways such as COX2, Ras, NF-ĸB or c-myc instead of HIF1α. Angiogenesis could trigger and accelerate tumor development; therefore, anti-angiogenic therapy could be useful for treatment of tumors.