Investigation of the effects of bisphenol-A exposure on lymphoid system in prenatal stage.

Bisphenol-A (BPA) used in the production of plastic materials is a temperature-soluble agent. It also has a steroid hormone-like activity; therefore, it poses a danger to human health. In our study, we aimed to investigate the effects of BPA on lymph node and spleen in male rats exposed to this agent during prenatal stage. The pregnant female rats were divided into four groups: control, sham, low dose (300 µg/kg BPA), and high dose (900 µg/kg BPA). BPA was dissolved in 1 mL of corn oil and administered to the pregnant rats every day during pregnancy. On the 21st and 45th day after the birth, male rats' lymph node and spleen samples were taken and histopathological examination was performed. Samples were stained with hematoxylin and eosin to determine the general histological appearance, and with CD3 and CD20 immunohistochemically. The results of staining were evaluated by H-score, and statistical analysis was performed. In the samples, BPA applications were not found to cause significant tissue damage. But there was a significant decrease in the immunoreactivities of CD3 and CD20 after BPA applications in both 21st and 45th day samples. After high dose BPA administration, decreased CD3 immunoreactivity was statistically significant. It is thought that BPA does not cause histologically significant tissue damage, but it may impair organ function at cellular level. The investigation of molecules involved in organ function will be useful in revealing the mechanisms that will cause dysfunction.

Acute and Chronic Pretreatment With Atenolol Attenuates Intestinal Ischemia and Reperfusion Injury in Hypercholesterolemic Rats.

OBJECTIVE: To evaluate the protective effects of preinjury atenolol (acute v chronic) on apoptosis, contractility, oxidative stress, and inflammatory markers in hypercholesterolemic rats undergoing intestinal ischemia-reperfusion (I/R) injury. DESIGN: Prospective, experimental animal study. SETTING: University laboratory. PARTICIPANTS: Male Wistar rats (n = 32). INTERVENTIONS: Rats were divided into the following 4 groups: 1 group was fed a normal diet (ND) (group ND+NoAT [no atenolol]), and the other 3 groups were fed a high-cholesterol diet (HCD)-group HCD+NoAT, group HCD+ChAT (chronic atenolol, 3 mg/kg/day for 8 weeks), and group HCD+AcAT (acute atenolol, 1.5 mg/kg, given 5 minutes before intestinal clamping). All rats underwent I/R injury. The superior mesenteric artery was clamped for 60 minutes, then opened for 120 minutes (reperfusion). Apoptotic cells and stimulated contractions of ileal segments were examined. Tissue markers of intestinal I/R injury were examined. Intestinal malondialdehyde, superoxide dismutase, and nitrate/nitrite levels were measured. MEASUREMENTS AND MAIN RESULTS: The chronic atenolol group had fewer apoptotic cells and higher superoxide dismutase activity compared with the other groups. Intestinal contraction was higher in both atenolol pretreatment groups compared with the NoAT groups. Chronic and acute atenolol resulted in lower ileal levels of malondialdehyde and immunolabeling-positive cells (intestinal inducible nitric oxide synthase, endothelial nitric oxide synthase, interleukin-1, and interleukin-8) after I/R injury compared with the no atenolol groups. CONCLUSIONS: Both chronic and acute pre-I/R injury treatment with atenolol attenuated I/R injury in this hypercholesterolemic rat model. These findings should encourage future studies of atenolol in hypercholesterolemic patients undergoing procedures with a high risk of intestinal ischemia.

Investigation of distribution of GSK-3ß signal pathway by age groups in cases of ulcerative colitis.

Ulcerative colitis (UC), one of the inflammatory bowel diseases, has been reported to increase in recent years. Although the exact cause is unknown, disruptions in the molecular pathways are thought to trigger UC. We aimed to examine the distributions of glycogen synthase kinase-3beta (GSK-3ß), nuclear factor-kappa B (NF-κB) and wingless∕int-1 (Wnt-1) in different age groups diagnosed with UC. Patients diagnosed with UC were divided into four groups according to their ages: Group 1, aged 18-30 (n=20); Group 2, aged 31-45 (n=20); Group 3, aged 46-60 (n=20); Group 4, aged 61-75 (n=20). Tissue sections were histochemically stained to examine the parameters of epithelial cell height, length of crypt, thickness of muscularis mucosa and extent of submucosal fibrosis. The immunohistochemistry assay was performed using cell survival and for GSK-3ß, NF-κB and Wnt-1 cell growth markers. Immunoreactivities were evaluated using H-score and analyzed using the one-way analysis of variance (ANOVA) test for statistics. It was detected a decrease in the histopathological parameters whereas the immunoreactivities of GSK-3ß, NF-κB and Wnt-1 were increased with increasing age. The levels of GSK-3ß immunoreactivity were similar in both epithelium and submucosa in all groups. NF-κB immunoreactivity was higher in submucosa of Groups 1, 2 and 3, while Wnt-1 was enhanced in Groups 1 and 3. The results of histopathology showed that the integrity of the epithelial tissue in the colon deteriorated with increasing age. The expressions of GSK-3ß, NF-κB and Wnt-1 were detected in all age groups. We thought that there was a synergistic activation between these three markers. Nevertheless, studies are needed to investigate this molecular pathway.

The Condylar Effects of Mesenchymal Stem Cells, Low-Level Laser Therapy and Grape Seed Extract on Functional Mandibular Advancement of the Rat Mandible.

Objective: Functional treatment of Class II malocclusion is expected to lead to adaptation in the condyle. This study aimed to evaluate the effects of adipose tissue-derived mesenchymal stem cells (ADMSCs), low-level laser therapy (LLLT), and grape-seed extract (GSE) on condylar growth after functional mandibular advancement. Methods: Forty-five rats were randomly divided into 8 groups. Functional appliances were applied to all groups (n=6) except the control group (n=3). One group was treated with appliances only; the other six groups received various combinations of ADMSCs, LLLT, and GSE. Analyses for new osteoblasts and new bone formation, vascular endothelial growth factor, and Type II collagen were performed on condylar tissues, after an experimental period of four weeks. The quantitative data obtained from the results of the experiments were evaluated by H-score and analyzed using One-Way ANOVA by Tukey-Kramer multiple comparisons test (p≤0.05). Results: Levels of all investigated parameters increased in all groups (p≤0.05). The highest increases were achieved by a combined application of functional appliance, ADMSCs, LLLT and GSE (p≤0.05). Single LLLT administrations or single GSE applications did not create a statistical difference from appliance alone (p>0.05). A positive effect of ADMSCs or LLLT on osteoblast formation, neovascularization, and Type II collagen level was apparent (p≤0.05), however, neither affected new bone formation (p>0.05). Conclusion: This study shows that ADMSCs with LLLT and GSE applications provide differing levels of new osteoblast and bone formation, new vascular formation, and Type II collagen formation in rat condyles after functional mandibular advancement.

The Inhibitory Effect of Capparis Ovata Polysaccharides on Cultured Pterygium Fibroblasts.

OBJECTIVES: Pterygium recurrence after removal surgery is an important problem. Polysaccharides obtained from Capparis species have been shown to possess various biological properties, including anti-tumor activity. This study was an investigation of the effect of Capparis ovata polysaccharides on cultured pterygium fibroblasts and a comparison with the effects of mitomycin C (MMC). METHODS: Pterygium tissue samples were obtained during excision surgery from 3 patients with primary pterygium, and fibroblasts were isolated. Pterygium fibroblast cultures and L929 cell cultures were treated with different concentrations of MMC and Capparis ovata polysaccharides. Cell proliferation and migration were evaluated. RESULTS: An MTT assay revealed that cell viability decreased with increasing concentrations of Capparis ovata polysaccharides in both cell types. MMC also inhibited the proliferation of both cell types. A scratch-wound assay indicated that both Capparis ovata polysaccharides and MMC molecules reduced proliferation and migration in the pterygium fibroblasts and L929 cells. CONCLUSION: The in vitro Capparis ovata polysaccharide inhibition of proliferation and migration of pterygium fibroblasts was similar to that of MMC. The results of this study suggested that Capparis ovata polysaccharides may be a valuable candidate drug to treat pterygium.

Effects of oxidative stress and apoptosis on vascularity and viability of perforator flaps.

We investigated lateral thoracic and posterior thigh perforator flaps for viability, vascularization, perfusion and apoptosis in a rat model. Wistar albino rats were divided into six groups: lateral thoracic artery perforator flap (LTPF) sham, 3 × 2 cm2 LTPF, 3 × 6 cm2 LTPF, posterior thigh perforator flap (PTPF) sham, 3 × 2 cm2 PTPF, and 3 × 6 cm2 PTPF. Flap viability was determined on postoperative days 1 and 7. On day 7, flaps were photographed and their viability was measured using two-dimensional planimeter paper. Tissue samples were harvested for examination by histology and immunohistochemistry. Viability differences were statistically significant. Epithelial thickness, vascularity and number of fibroblasts were reduced in the 3 × 6 cm2 groups. Neovascularization and apoptosis based on molecular tests were not significantly different among groups. Flap size and location are important factors for closure of surgical or traumatic defects. We suggest that for clinical application, wound complications will occur less frequently with perforators that nourish large areas of flaps.

Therapeutic effects of Lacosamide in a rat model of traumatic brain injury: A histological, biochemical and electroencephalography monitoring study.

OBJECTIVE: Traumatic Brain Injury (TBI) is a major cause of death and disability worldwide, especially in children and young adults. TBI can be classified based on severity, mechanism or other features. Inflammation, apoptosis, oxidative stress, and ischemia are some of the important pathophys-iological mechanisms underlying neuronal loss after TBI. Lacosamide (LCM) is an anticonvulsant compound approved for the adjunctive treatment of partial-onset seizures and neuropathic pain. This study aimed to investigate possible neuroprotective effects of LCM in a rat model of TBI. MATERIAL AND METHODS: Twenty-eight adult male, Wistar albino rats were used. The rats were divided into 4 groups. Group 1 was the control group (n=7). Group 2 was the trauma group (n=7) where rats were treated with 100 mg/kg saline intraperitoneally (IP) twice a day. Groups 3 and 4, rats were treated with 6 (group 3, n=7) or 20 (group 4, n=7) mg/kg Lacosamide IP twice a day. For each group, brain samples were collected 72 hours after injury. Brain samples and blood were evaluated with histopathological and biochemical methods. In addition, electroencephalograpy monitoring results were compared. RESULTS: The immunoreactivity of both iNOS and eNOS (oxidative stress markers) were decreased with LCM treatment compared to trauma group. The results were statistically significant (***P<0.001). The treatments of low (56,17±9,69) and high-dose LCM (43,91±9,09) were decreased the distribution of HIF-1α compared to trauma group (P<0.01). The number of apoptotic cells were decreased with LCM treatment the difference between the trauma group and 20mg/kg LCM treated group (9,55±1,02) was statistically significant (***P<0.001). Malondialdehyde level was reduced with LCM treatment. MDA level was significantly higher in trauma group compared to LCM treated groups (***P<0.001). The level of Superoxide dismutase in the trauma group was 1,86 U/ml, whereas it was 36,85 U/ml in 20mg/kg LCM treated group (***P<0.001). Delta strength of EEG in 20mg/kg LCM treated group were similar to control group values after LCM treatment. CONCLUSION: No existing study has produced results suggesting that different doses of LCM has therapeutic effect against TBI, using EEG recording in addition to histological and biochemical evaluations in rats.

Inhibitory Effect of Valsartan on Pterygium Fibroblasts.

PURPOSE: Angiotensin receptor blockers (ARBs) were shown to have antifibrotic properties in ocular and systemic diseases. In this study, our aim was to investigate the effect of an angiotensin receptor blocker, valsartan, on pterygium fibroblasts and compare this effect with that of mitomycin C (MMC). METHODS: Pterygium tissue samples were obtained from 3 patients during surgical excision. Primary cultured pterygium fibroblasts and L929 cell cultures were treated with different concentrations of MMC and valsartan. RESULTS: The cell viability decreased with increasing concentrations of valsartan at 48 hours for both cell types. MMC inhibited the proliferation of both cell types at 48 hours. Both agents significantly decreased the cell migration of the 2 cell types, although it was more prominent in the MMC-treated group. CONCLUSIONS: Valsartan inhibited the proliferation and migration of pterygium fibroblasts. The known favorable safety profile of these drugs and the results of this study showing inhibitory effect on pterygium fibroblasts make valsartan a potential therapeutic agent for pterygium treatment.

Adjuvant effects of chemotherapeutics and Metformin on MFE-319 endometrial carcinoma cell line.

We aimed to investigate the cytotoxicity of Metformin, Cisplatin, and Paclitaxel on MFE-319 endometrial carcinoma cell line using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and immunocytochemistry assays. Half maximal inhibitory concentration (IC50) doses of three drugs alone and in the dual combinations were applied to the cells. Immunocytochemical method was performed for the cell survival and for phosphatidylinositol 3-kinase (PI3K), phosphorylated extracellular regulated kinases (pErk)-1∕2, Akt-1, phosphorylated Akt (pAkt)-1∕2∕3 cell growth markers and angiogenic vascular endothelial growth factor (VEGF). Immunoreactivities were evaluated using H-score and analyzed using the one-way analysis of variance (ANOVA) test for statistics. It was found that these drugs caused a decrease in the immunoreactivities of these markers. Particularly, dual combination of Paclitaxel and Cisplatin decreased the immunoreactivities of PI3K, pErk-1∕2, Akt-1, and pAkt-1∕2∕3. Cisplatin and Paclitaxel were more effective than Metformin; on the other hand, Metformin has been shown to enhance the efficacy of these two drugs. In vitro or in vivo further studies are needed to investigate the efficacy of these three drugs via PI3K∕Akt signal pathway.

Inhibiting effect of oleocanthal on neuroblastoma cancer cell proliferation in culture.

We investigated the potential anticancer effects of oleocanthal (OC) on neuroblastoma cells. Cells were divided into four groups: group 1, neuroblastoma cells were treated with OC; group 2, neurons that differentiated from neuroblastoma cells were treated with phosphate-buffered saline(PBS); group 3, bone marrow derived neuronal (BMDN) cells that were differentiated from bone marrow derived mesenchymal stem cells (BMSCs) were treated with OC; group 4, BMDN cells that were differentiated from BMSCs were treated with PBS. Groups 2 and 4 were control groups. The effects of OC on cell viability, oxidative stress, neurite inhibition and apoptosis at IC50 dose were investigated using MTT analysis, i-NOS and e-NOS measurement, neurotoxicity screening test (NST) and TUNEL staining, respectively. MTT analysis demonstrated that cells were significantly less viable in group 1 than in group 3. i-NOS and e-NOS staining intensity was significantly greater in group 1 than in group 3. NST revealed that OC inhibited neurite growth in both neuroblastoma and BMND cells; inhibition was significantly less in group 3 than in group 1. Significantly more TUNEL labeled cells were found in group 1 than in group 3. We found that OC prevented growth and proliferation of neuroblastoma cells in culture by increasing oxidative stress and apoptosis. We also found that the cytotoxicity of OC is negligible in BMDN cells.

Histological and immunohistochemical studies of the proximal caecum and caecal tonsils of quail (Coturnix coturnix japonica).

The proximal caecum in quails consists of lymphoid and non-lymphoid structures. The caecal tonsils in the proximal part of the caecum are units of gut-associated lymphoid tissue in poultry. This study aimed to examine the histological characteristics of the proximal caecum, as well as compositions of dendritic cells (DCs) and antigen-presenting cells (APCs) in the caecal tonsil of quails. Tissue sections were stained with Crossman's triple, periodic acid-Schiff, Gordon and Sweet's silver, Congo red and methyl green-pyronin dyes, as well as immunohistochemically by the streptavidin-biotin-peroxidase complex method. Caecal lymphoid tissue was located in the lamina propria and submucosa. Germinative centres were observed within the lymphoid tissue. Reticular fibres were mainly distributed in the border area of the germinal centre with only a few fibres scattered in the centre. Plasma cells were observed in the subepithelial region and germinal centres. Eosinophil granulocytes were prevalent in the lymphoid tissue. Additionally, CD83-immunoreactive DCs and MHC class II immunoreactive APCs were present in the subepithelial area and diffuse lymphoid tissue. While DCs were seen in the germinal centres of tonsillar units, APCs were rarely present in the germinal centres, but they were noticed around the germinal centres. In conclusion, the histological structure of the proximal caecum in quails and the distributions of some immunological cells in the caecal tonsils were revealed. Therefore, the defensive role of the caecal tonsils in the digestive system may be better understood, and comparative studies may be carried out.

Punicic Acid Inhibits Glioblastoma Migration and Proliferation via the PI3K/AKT1/mTOR Signaling Pathway.

BACKGROUND: Punicic Acid (PA) is a polyunsaturated fatty acid that accounts for approximately 70%- 80% of Pomegranate Seed Oil (PSO). PA possesses strong antioxidant, anti-inflammatory, anti-atherogenic effects, and anti-tumorigenic properties. Pomegranate extracts have been shown to have anticancer activity in many studies. However, there is no evidence for the effect of PSO on T98 glioblastoma cells. Therefore, the present study was the first to investigate the mechanisms induced by PA on T98 cells, which is one of the major compounds extracted from PSO. METHODS: The effects of PA on cell viability; oxidative stress; and migration, proliferation, and apoptosis at the IC50 dose were studied. RESULTS: The proliferation and migration were inhibited in the treated group compared to the non-treated group by 9.85µl/ml PA. The difference was statistically significant (***p<0.001). Furthermore, PA-induced apoptosis in the T98 glioblastoma cells compared to non-treated group and the difference was statistically significant (***p<0.001). Apoptosis was determined via immunocytochemistry staining of caspase-3, caspase-9 and TUNEL methods. Apoptosis was checked by flow cytometry (using caspase 3 methods) and Scanning Electron Microscopy Analysis. We also investigated the potential signaling pathway underlying this apoptotic effect. The immunocytochemical stainings of PI3K/ Akt-1/ mTOR-1 demonstrated that Akt-1 staining was increased with PA treatment similar to mTOR-1 and PI3K staining (***p<0.001). These increases were statistically significant compared to the non-treated group. CONCLUSION: PA exhibited exceptional abilities as an anticancer agent against GBM cells. The use of punicic acid in combination with other drugs used in the treatment of glioblastoma may increase the efficacy of the treatment. This study provided a basis for future investigation of its use in preclinical and clinical studies.

Histological investigations on thymus of male rats prenatally exposed to bisphenol A.

Bisphenol A is called as a endocrine-distrupting chemical because of the its steroid-like activity and it used in the construction of plastic containing materials. It is indicated that bisphenol A can pass the human serum, urine, follicular fluid, placenta and umblical cord as a result of the use of substances containing this agent. In this study, we aimed to investigate the effects of bisphenol A on the development of the thymus, a primary lymphoid organ which plays an important role in the specific immunity. The adult pregnant female rats were administered orally with bisphenol A (for 21 days) and postnatal thymus samples were obtained on day 21, 45 and 90 and were performed for histochemical and immunohistochemical staining for CD3, CD4, CD8 and CD79a and TUNEL assay for the apoptotic cells. Evaluation of all groups, CD3, CD4, CD8 and CD79a stainings were decreased in the experimental groups compared with control group. The apoptotic cells were determined in the all groups on day 90 as a result of the thymus involution. It is noted that there was not any histological and morphological damages in the rats prenatally exposed the bisphenol A. The effect of the bisphenol A is unknown in the future, but there is no problem in the adult rats.

Neuroprotective Effects of Oleocanthal, a Compound in Virgin Olive Oil, in a Rat Model of Traumatic Brain Injury.

AIM: To evaluate the neuroprotective effects of deocanthal OC in a rat model of traumatic brain injury (TBI). MATERIAL AND METHODS: Twenty-six adult male, Wistar albino rats were used. The rats were divided into 4 groups. Group 1 was the sham group (n=5). Group 2 was the trauma group (n=5) where rats were treated with 10 mg/kg saline intraperitoneally (IP) twice a day. Groups 3 and 4, rats were treated with 10 (group 3, n=8) or 30 (group 4, n=8) mg/kg OC IP twice a day. For each group, brain samples were collected 72 hours after injury. Brain samples and blood were evaluated with histopathological and biochemical methods. RESULTS: Histopathological evaluation revealed a significant difference between Group 2 and Group 4. Biochemical findings demonstrated that the oxidative stress index was highest in Group 2 and lowest in Group 4. CONCLUSION: OC has a protective effect on neural cells after TBI. This effect is achieved by reducing oxidative stress and apoptosis.

Comparison of intraperitoneal and intratesticular ozone therapy for the treatment of testicular ischemia-reperfusion injury in rats.

We compare the efficacy of intratesticular ozone therapy with intraperitoneal ozone therapy in an experimental rat model. For this purpose, 24 rats were divided into four groups including sham-operated, torsion/detorsion, torsion/detorsion plus intraperitoneal ozone (O-IP), and torsion/detorsion plus intratesticular ozone (O-IT). The O-IP ozone group received a 4 mg kg-1 intraperitoneal injection of ozone, and the O-IT group received the same injection epididymally. At 4 h after detorsion, the rats were sacrificed and orchiectomy materials were assessed histopathologically. Spermatogenesis in the seminiferous tubules and damage to the Sertoli cells were histopathologically evaluated in the testes using the Johnsen scoring system. i-NOS and e-NOS activities in the testis tissue were also evaluated. Torsion-detorsion caused a decreased Johnsen score and increased apoptosis of spermatogonial and Sertoli cells. Ozone injection prevented increases in Johnsen score and i-NOS level. e-NOS level of the O-IP group was significantly lower than that of the O-IP group, and i-NOS level of the O-IT group was significantly lower than that of the O-IP group. Local ozone therapy is more effective than systemic ozone therapy at improving IRI-related testicular torsion. Our study is the first to show that the efficacy of intratesticular implementation of ozone therapy is higher than that of intraperitoneal ozone therapy.

Neuroprotective effects of bone marrow-derived mesenchymal stem cellsand conditioned medium in mechanically injured neuroblastoma cells.

BACKGROUND/AIM: Bone marrow-derived mesenchymal stem cells (BMSCs) possess self-renewal characteristics that distinguish them from other cell types. Recent studies have focused on the effects of conditioned medium (CM) that includes the extracellular matrix. Here we examined the neuroprotective effects of BMSCs and CM on damaged neuroblastoma cells. MATERIALS AND METHODS: The cells were divided into five groups: 1) healthy controls, 2) damaged cells alone, 3) damaged cells treated with BMSCs, 4) damaged cells treated with CM, and 5) damaged cells treated with both BMSCs and CM. Neuroprotective effects were then evaluated based upon the levels of oxidative stress, antitransforming growth factor ß1 (anti-TGFß1) production, and apoptosis. RESULTS: Significant differences were observed between healthy controls and damaged cells (P < 0.001), as well as between damaged cells and those treated with BMSCs alone (P < 0.05), CM alone (P < 0.05), and both BMSCs and CM in combination (P < 0.01). Among the treated groups, the strongest neuroprotective effects were seen in cells treated with both BMSCs and CM. CONCLUSION: These results show that both BMSCs and CM exhibit neuroprotective effects in damaged neuroblastoma cells. The strongest benefits were seen following treatment with both BMSCs and CM.

Effects of 5-fluorouracil and gemcitabine on a breast cancer cell line (MCF-7) via the JAK/STAT pathway.

Aberrant activation of the JAK/STAT pathway may predispose to malignancy as a consequence of the deregulation of cell proliferation, differentiation or apoptosis such as in cancer of the blood, head and neck, and breast. In our study we aimed to investigate the effects of 5-fluorouracil (5-FU) and gemcitabine on a breast cancer cell line (MCF-7 cells) via the JAK/STAT pathway. Distribution of JAK1, JAK2, JAK3 and STAT2, STAT3, STAT4, STAT5 were evaluated on MCF-7 cells following gemcitabine and 5-FU treatment and in the absence of drug treatment by an indirect immunohistochemical method. It was observed that JAK1, JAK3, STAT5 and particularly STAT2 activation were more effective than the other JAK/STATs in breast cancer progression. Following treatment with 5-FU, JAK1 and STAT5 immunoreactivities were decreased in MCF-7 cells in comparison with both gemcitabine-treated and non-treated groups. These results suggest that the JAK/STAT pathway plays an important role in breast cancer pathogenesis and may be more affected after 5-FU treatment rather than gemcitabine. Drugs which block STAT5 may provide a novel therapeutic approach for the treatment of breast cancer.