Melamine exposure during the weaning period negatively affects ovarian reserve.

In this study, it was aimed to investigate the effects of melamine exposure since the weaning period on ovarian tissue and ovarian reserve. Melamine is illegally added to milk and formula to provide high false protein positivity. Female rats (the weaning period = 21 days old, n = 18) were divided into 3 groups. 0.1 mL saline was applied to the control group by gavage for 21 days. Fifty mg/kg and seventy-five mg/kg melamine was dissolved in 0.1 mL of saline and applied by gavage for 21 days, respectively. At the end of the experiment, plasma anti-Mullerian hormone (AMH) was measured, follicle count and ovarian diameter measurement were performed in the right ovaries, and flow cytometric analysis for apoptosis was performed in the left ovaries. While a statistically significant decrease was not observed in the number of the follicle and ovarian diameter between the control and melamine-treated groups (p > 0.05), a significant decrease in the corpus luteum and a significant increase in the number of atretic follicles were observed (p < 0.05). Apoptosis (Annexin V) increased in both melamine groups and AMH plasma level decreased significantly in the 75 mg/kg group (p < 0.05). Melamine exposure from the weaning (early postnatal) period may cause a decrease in ovarian reserve in parallel with a dose increase.

Use of hyaluronic acid matrix in dorsal augmentation rhinoplasty.

Augmentation rhinoplasty sometimes is required for patients with saddle nose deformity caused by failed rhinoplasty or facial trauma; finding appropriate grafting material remains a significant problem for this procedure. We investigated hyaluronic acid matrix as an allograft for dorsal augmentation rhinoplasty in a rabbit model. We performed an osteotomy on the nasal bones of eight rabbits. Four animals were sham operated as the control group and four were administered a mixture of saline-gelled hyaluronic acid matrix and sliced cartilage. Ultrasonography and three-dimensional reconstruction tomography were performed at the end of the experimental period. After sacrifice of the animals, nasal tissues were examined for histopathology, and both collagen scores and number of capillaries were compared between the two groups. Increased collagen and capillaries were apparent in the hyaluronic acid matrix group compared to controls. The median collagen score was significantly greater for the hyaluronic acid matrix group than for the control group. Although the number of capillaries for the hyaluronic acid matrix group was greater than for the control group, the difference was not statistically significant. Three weeks is sufficient for adhesion of ends of fractures in clinical practice; however, we found no ossification at this time in either group. A hyaluronic acid matrix may be a useful alternative supplement for dorsal augmentation rhinoplasty. Development of collagen was commensurate with membranous ossification; however, assessment of complete ossification requires a longer experimental period.

Comparison of Protective Effect of Aqueous and Ethanolic Extracts of Propolis against Gastric Ulceration in Rats.

This study focused on the protective effects of different types of propolis extracts on gastric mucosa in indomethacin-induced rats. The animals were divided into nine groups: control, negative control (ulcer), positive control (omeprazole), and experimental groups, which were summarized by 200, 400, and 600 mg/kg, bw for aqueous-based and ethanol, respectively. According to the histopathological evaluation, more than others, the doses of 200 and 400 mg/kg of aqueous-based propolis extracts had different degrees of positive effects on the gastric mucosa. Generally, the biochemical analyses of the gastric tissue showed a correlation with microscopic evaluations. According to the phenolic profile analysis, while pinocembrin (684.34±1.70 µg/ml) and chrysin (540.54±9.06 µg/ml) were the most abundant phenolics in the ethanolic extract, ferulic acid (53.77±0.07 µg/ml) and p-coumaric acid (52.61±0.42 µg/ml) dominated the aqueous-based extract. Also, the total phenolic content (TPC), total flavonoid content (TFC), and DPPH radical scavenging activity of the ethanolic extract showed almost nine-fold superiority compared to the aqueous-based extracts. Based on data from preclinical data, it was decided that the best doses for the main goal of the study were 200 mg and 400 mg/kg, bw for aqueous-based propolis extract.

Methotrexate-induced hepatotoxicity in rats and the therapeutic properties of vitamin E: a histopathologic and flowcytometric research.

Aim of the study: Methotrexate (MTX) causes oxidative stress-related liver damage. Our objective was to investigate the protective effects of vitamin E against MTX-induced hepatotoxicity through histopathological methods and flow cytometry. Material and methods: The rats were assigned to four groups: Control (2 ml saline for 5 days), MTX (20 mg/kg intraperitoneally (i.p.) only on the initial day of the study), MTX + vitamin E (20 mg/kg MTX (i.p.) only on the first day, and 100 mg/kg vitamin E (i.p.) was applied for 5 days during the study), Vitamin E (100 mg/kg of vitamin E (i.p.) was given for five days). Histopathologic changes and the flow cytometric apoptotic index were evaluated for liver tissue. The Kruskal-Wallis test was used for comparisons between groups. The statistical significance level was accepted as p < 0.05. Results: In the histopathological analysis, hepatocyte degeneration, dilatation of sinusoids, mononuclear cell infiltration, hydropic degeneration in hepatocytes, vacuolization, and pycnotic nucleus were observed in the MTX group. In the MTX + vitamin E group, hepatocyte degeneration, pycnotic nuclei, and dilatation in sinusoids were significantly lower compared to the MTX group. In the MTX group, glycogen accumulation in hepatocytes was lower compared to the control group. In the MTX + vitamin E group, glycogen accumulation in hepatocy-tes was higher compared to the MTX group. The flowcytometric apoptotic index (AI) percentage in the MTX group was 34.4% and in the MTX + vitamin E group the value was 9.4%. Conclusions: Our results demonstrated that vitamin E ameliorates MTX-induced liver damage. Co-using vitamin E and MTX drugs will be beneficial for the treatment of various diseases.

Effects of hyaluronic acid on the bone healing after lateral osteotomy: an experimental animal model.

OBJECTIVE: Postoperative comfort of the patients undergoing rhinoplasty might be poor because of edema and ecchymosis caused by lateral osteotomy. In this animal experiment, we aimed at performing a quantitative assessment of effects of hyaluronic acid usage on healing process of lateral osteotomy. METHODS: Fourteen New Zealand rabbits with a weight of 2000-2500 kg and an age of 8-12 weeks were included. Under anesthesia, nasal dorsums were exposed with midline incision and lateral osteotomies on both sides were performed using a 2 mm chisel. A hyaluronic acid-based mesh (Hyalonect®) (1 × 1 cm) was embedded on the left osteotomy areas of all rabbits. Right osteotomy areas were left blank as control group. Collagen density and capillary development were quantitatively compared. RESULTS: Convergence of fracture lines was observed in 6 (60%) of 10 samples from Hyalonect® group, while was observed in 4 (40%) of 10 samples from control group. Although a higher rate of convergence was seen in the Hyalonect® group (60% vs 40%), the difference was not statistically significant (p = 0.5). Median collagen score was 2 (1-3) in the Hyalonect® group and 1 (1-2) in the control group. Median capillary count value was 4 (1-23) in the Hyalonect® group and 3 (1-17) in the control group. Both collagen score and capillary count values were significantly greater in the in the Hyalonect® group compared with the control group (p = 0.023 and p = 0.019, respectively). CONCLUSION: The effects of hyaluronic acid-based meshes on the bone healing process of the lateral osteotomy area might be investigated furthermore, on more comprehensive studies, as a material facilitating collagen organization and capillary development.

Hepatotoxic effects of melamine exposure from the weaning period in rats: a flow cytometric, electron microscopic, and histopathologic study.

This study aims to investigate the effects of melamine exposure from the weaning period (21st postnatal days in rats) on liver tissue. Female Wistar albino rats (n = 18) were divided into three groups. About 0.1-ml saline was applied to the control group by gavage for 21 days from the postnatal 21st day. The second group was taken 50-mg/kg melamine (in 0.1-ml saline) and the third group was taken 75-mg/kg melamine (in 0.1-ml saline) p.o. On the postnatal 45th day, all rats were sacrificed under anesthesia. Then, liver tissues were cut into three parts and two of them placed in neutral formalin for histopathological and flow cytometric analysis, and one of them placed in 2.5% glutaraldehyde. Histopathological analysis was performed with hematoxylin & eosin, Masson trichrome, periodic acid Schiff stained sections, and also with transmission electron microscopy. Apoptosis (Annexin V positivity) was analyzed by flow cytometry. According to histopathological analysis, hepatocyte damage, sinusoidal dilatation, and inflammatory cell infiltration significantly increased in both melamine groups compared with the control group. Apoptosis significantly increased in the 50 and 75-mg melamine groups compared with the control group. In the results of transmission electron microscopy analysis, there was abnormal chromatin distribution in the hepatocyte nuclei, loss in the cristae of the mitochondria, and organelle loss in large areas in the cytoplasm in both melamine exposure groups. As result, melamine exposure from the weaning period causes liver damage with increasing doses.

Melamine exposure from the weaning period causes apoptosis, inflammation, and damage to the blood-brain barrier.

The aim of this study to investigate the effects of melamine exposure from the weaning period on the developing brain in rats. Female Wistar albino rats (21 days old, n = 18) were divided into 3 groups, all animals were weighed daily and dose-adjusted. For 3 weeks, 0.1 mL of saline was administered by oral gavage to the control group, 50 mg/kg of melamine to the second group, and 75 mg/kg of melamine to the third group were administered by oral gavage by dissolving in 0.1 mL of saline. On the postnatal 45th day, the rats were sacrificed under anesthesia and brain tissues placed in neutral formalin. After routine tissue processing, brain sections were stained with hematoxylin&eosin(H&E) and Terminal deoxynucleotidyl transferase(TdT) dUTP Nick-End Labeling(TUNEL), IBA-1, Glial Fibrillar Acidic Protein(GFAP), Tumor necrosis factor-α (TNF-α), and SMI-70 antibodies as immunohistochemically. In the results, according to apoptotic index(AI) results, there was a significant increase in the 75 mg/kg and 50 mg/kg melamine groups compared to the control groups (p < 005). There was a significant increase in the number of anti- TNF-α positive neurons and the number of anti-GFAP positive astrocytes in both melamine groups compared to the control group (p < 001). In terms of SMI-71, an increase was found in the 75 mg/kg group compared to other groups (p < 001), while no significant difference was found between the groups in terms of IBA-1 (p > 0.05). It has been observed an increase in dilatation of blood vessels, inflammatory cell infiltration, and endothelial cell degeneration in the 50 mg/kg and 75 mg/kg melamine groups compared to the control group (p < 0.01). there was no statistically significant difference in the body and brain weight between both melamine treatment groups (75 mg/kg and 50 mg/kg) and the control group (p > 005). Melamine exposure (50 mg/kg and 75 mg/kg) from the weaning period causes apoptosis and inflammation in the developing brain, and the disruptions in the blood-brain barrier (BBB) significantly increase exposure to 75 mg/kg.

Does melamine exposure during infancy cause rhabdomyolysis?

We investigated the possible toxic effects of melamine on muscle tissue in rats using biochemical, hematological and histopathological methods. We used three groups of female albino Wistar rats. The first group was given 0.1 ml saline. The second and third groups were given 50 and 75 mg/kg melamine dissolved in 0.1 ml saline, respectively, daily for 21 days. On day 45, all rats were sacrificed, and whole blood and plasma were analyzed for hematologic and biochemical characteristics. Muscle samples were stained with hematoxylin and eosin for histopathological investigation. Other sections were immunostained for matrix metalloproteinase-9 (MMP-9) and type IV collagen. We found a significant increase in the lymphocytosis-compliant leukocyte number in the 75 mg/kg melamine group compared to the other groups. We also found significant decreases in the hemoglobin levels and hematocrit values in the 75 mg/kg compared to the other groups. We found that the 75 mg/kg melamine group exhibited a significant increase in plasma aspartate aminotransferase (AST) activity compared to the other groups. Changes in plasma creatine kinase and lactate dehydrogenase activity were not statistically significant. Plasma AST activity and mean corpuscular hemoglobin levels were correlated with the lymphocyte:neutrophil ratio. We found mononuclear cell infiltration at the periphery of muscle bundles and in the connective tissue bundles in the melamine treated group. We found MMP-9 expression in muscle cell membranes and type IV collagen expression in degenerative connective tissue fibers. Whole blood, plasma and muscle tissue analysis indicated that the 75 mg/kg melamine group exhibited rhabdomyolysis that was associated with lymphocytosis and anemia. The underlying mechanisms by which melamine causes rhabdomyolytic effects remain unclear.

The effects of intravitreal H2 S application on apoptosis in the retina and cornea in experimental glaucoma model.

One of the most important causes of visual loss (blindness) is glaucoma, which occurs due to the degeneration of the ganglion cells in retina. It has been shown that hydrogen sulphide (H2 S) acts an antioxidant, neuroprotective and neuromodulator and provides protection against oxidative stress and apoptosis. This study aims to examine through which apoptotic pathway H2 S acts in experimental glaucoma model. Twenty-two male wistar albino rats were used in this study. Group 1 (n = 6, control group): Intravitreal saline was given in the third week without inducing ocular hypertension (OHT) with laser photocoagulation. Group 2 (n = 8): After the induction of OHT with laser photocoagulation, intravitreal saline was given in the third week. Group 3 (n = 8): After the induction of OHT with laser photocoagulation, intravitreal H2 S's donor sodium hydrosulphide (NaSH) 100 nmol/L was given in the third week. At the end of the 6th week, the eyes of the rats were sacrified under anaesthesia and extracted and then routine tissue follow-up was undertaken. Besides haematoxylin & eosin (H&E) staining, Bax, Bcl-2, p53 and caspase-3 activation were examined immunohistochemically in the retina and the cornea. This showed that ocular hypertension caused apoptosis through the intrinsic pathway, due to Bax and caspase-3 activation, in both retina and cornea, and that this led to DNA damage due to p53 activation. Also, we found that H2 S exposure in glaucoma distinctly suppressed Bax, caspase-3 and p53 activations in retina but that it has a limited effect on the cornea. According to these results, glaucoma caused apoptosis in the retina through intrinsic pathway, and the damage to the retina could be compensated partially by H2 S but would have limited on the cornea.

Protective effects of aqueous extracts of some honeys against HCl/ethanol-induced gastric ulceration in rats.

This study focused on the concept of the possible protective effect of some honey types against HCl/ethanol-induced gastric ulcers in male Wistar albino rats. Rats were pretreated with rhododendron, chestnut, and oak honey orally with doses of 1.25 and 2.5 g/kg, bw (body weight) for three consecutive days. On fourth day, nothing was applied, and after the administration of anesthesia on the fifth day, their stomachs were surgically removed to investigate the histopathological examinations. Besides analyses of some blood serum profiles and antioxidant parameters of gastric tissue, some biochemical properties of honeys were investigated to support the histopathological results. The degrees of ulcer lesions in all groups revealed a statistically significant difference (p = .011). Although this difference originated from the additional ulcerative inducing effect of some honeys, the lower concentration rhododendron honey indicated more promising data than the positive control group (pantoprazole) in consequence of the microscopic and macroscopic evaluations. PRACTICAL APPLICATIONS: As being a member of natural products, honey has acquired fame among the studies in recent years due to its versatility as a source of food and complementary medicine. For contributing to this argument, this comprehensive study was performed and results were focused on the lower concentration of rhododendron honey thanks to its clinical potential with protecting the gastric mucosa. According to the obtained results, our suggestion came into prominence that this honey might be protecting the mucosa, better than the different concentrations of chestnut and oak honeys, by being better-absorbed through the gastric mucosa.

The neurotoxic effects of prenatal gabapentin and oxcarbazepine exposure on newborn rats.

AIM: Teratogenicity is a problematic issue for pregnant women because of X-ray radiation, drugs, and genetic and unknown variables. First-generation antiepileptic drugs (AED) like valproic acid are well-known teratogens for developing fetuses. However, their usage is necessary in order to prevent maternal seizures. The underlying mechanism of birth defects associated with AED exposure remains unclear and information about the neurotoxic effects of prenatal exposure to AED is still limited. Oxcarbazepine (OXC) and gabapentin (GBP) are second-generation AED. It still remains unclear how much these drugs are safe during pregnancy. This study aimed to investigate whether any neurotoxic effect of OXC and GBP in utero exposure on the developing brain. METHODS: Eighteen pregnant Wistar albino rats were divided into six groups. The first group was exposed to OXC at 100 mg/kg/day, the second to GBP at 50 mg/kg/day, and third to saline (0.9% NaCl) at 1.5 ml/day between the first and the fifth days of gestation. The same procedure was applied at the same dosages between the 6th and the 15th days of gestation for the 2nd three groups. Five female offspring (total n = 30, 45 days old) were taken from each group and stereological methods were applied in order to analyze the total and dopaminergic neuron number of the substantia nigra pars compacta (SNc). CONCLUSION: The result is that the OXC and GBP exposure at different gestational periods may not give rise to congenital malformation and it appears that the GBP exposure during the organogenesis period proliferatively affects the total number of neurons.

Intra-articular injection of hydrogen sulfide decreased the progression of gonarthrosis.

Hydrogen sulfide (H2S) is found in both the plasma and synovial fluid of patients with gonarthrosis. In the present study, we investigated whether intra-articular injection of sodium hydrosulfide (NaSH) (1 mM, 30 µL), a H2S donor, might affect gonarthrosis in rats. Gonarthrosis was induced surgically in the left knees of rats and left for 6 weeks for the development of disease. Then, intra-articular injections of NaSH or methylprednisolone (1 mg/kg, 30 µL) were administered to rats. Half of each group was sacrificed at the end of the first day and the other half was sacrificed at the end of 4 weeks to evaluate early and later effects of injections on gonarthrosis. The injury induced by anterior cruciate ligament resection and medial meniscectomy in rats caused the development of gonarthrosis. As the duration lengthened after gonarthrosis induction, the progression of the disease continued. According to the modified Mankin Scoring System, intra-articular injection of NaSH histopathologically slowed the progression of gonarthrosis, whereas methylprednisolone was ineffective. In addition, NaSH decreased apoptosis in rat knees with gonarthrosis. Each treatment did not cause injury to healthy knees. Our results lead to the consideration that intra-articular NaSH administration may be effective in the progression of gonarthrosis.

Protective effects of Nigella sativa oil against carboplatin-induced liver damage in rats.

OBJECTIVE: This study aimed to investigate the protective effects of nigella sativa oil (NSO) against liver damage due to intraperitoneal (i.p.) usage of carboplatin which is commonly used as a chemotherapeutic agent. MATERIAL AND METHOD: Twenty four female Wistar-albino rats (about 200-350 grams each) were divided into 4 groups. Group 1 (n = 6) was administered 4 ml/kg intraperitoneal (i.p.) saline 48 and 24 h before. Group 2 (n = 6) was i.p. administered 4 ml/kg NSO 48 h before and 4 ml/kg saline 24 h before. Group 3 (n = 6) was i.p. administered 4 ml/kg saline 48 h before and 80 mg/kg carboplatin 24 h before. Group 4 (n = 6) was i.p. administered 4 ml/kg NSO 48 h before and 80 mg/kg carboplatin 24 h before. At the end of 48 h, all rats were sacrificed, and liver tissues were put into 10% neutral formalin. After the routine tissue follow-up, histopathological changes and collagen fiber density were evaluated with Hematoxylin-Eosin and Masson's Trichrome staining. Apoptotic index was determined with TUNEL staining. RESULTS: The degeneration in hepatocytes, fiber distribution and density around central vein and portal space was observed in the carboplatin group compared to the control and NSO groups, hepatocyte cords preserved integrity, partial degeneration in hepatocytes and decreased collagen fiber distribution around central vein was noted in the NSO-carboplatin group compared to the carboplatin group. The apoptosis was lower in the NSO-carboplatin group compare with the carboplatin group, but no statistically significant difference was found between the two groups (p = 0.449). CONCLUSION: When used NSO before carboplatin exposure, it may protect against liver damage.

Effects of grape seed extract on periodontal disease: an experimental study in rats.

OBJECTIVE: This study aimed to demonstrate the effect of grape seed extract (GSE) on periodontitis. MATERIAL AND METHODS: Ligature induced periodontitis was created in 40 rats and they were assigned to four equal groups. One group was fed laboratory diet (group A) while three groups received GSE additionally. Silk ligatures were placed around the cervical area of the mandibular first molars for four weeks to induce periodontitis. The GSE groups were reallocated regarding GSE consumption as: for two weeks before ligation (group B; totally eight weeks), from ligation to two weeks after removal of the ligature (group C; totally six weeks), and for two weeks from ligature removal (group D; totally two weeks). Sections were assessed histologically and immunohistochemically. Inflammatory cell number (ICN), connective tissue attachment level (CAL), osteoclast density (OD), IL-10 and TGF-ß stainings in gingival epithelium (GE), connective tissue (GC), and periodontal ligament (PL) were used as the study parameters. RESULTS: Lower ICN, higher CAL, and lower OD were observed in the GSE groups (p<0.05). IL-10 was more intensive in the GSE groups and in the GEs (p<0.05). Group B showed the highest IL-10 for PL (p<0.05). TGF-ß was higher in the GEs of all groups (p<0.017). CONCLUSIONS: The results suggest anti-inflammatory activities of GSE, but further investigations are needed for clarification of these activities.

Assessment of MMP-1, MMP-8 and TIMP-2 in experimental periodontitis treated with kaempferol.

PURPOSE: The objective of this study was to investigate the effect of a dietary flavonoid, kaempferol, which has been shown to possess antiallergic, anti-inflammatory, anticarcinogenic, and antioxidant activities on the periodontium by histomorphometric analysis and on gingival tissue matrix metalloproteinase-1 (MMP-1), MMP-8, and tissue inhibitor of metalloproteinase-2 (TIMP-2) by biochemical analysis of rats after experimental periodontitis induction. METHODS: Sixty Wistar rats were randomly divided into six groups of ten rats each, and silk ligatures were placed around the cervical area of the mandibular first molars for 15 days, except in the healthy control rats. In the experimental periodontitis groups, systemic kaempferol (10 mg/kg/2d) and saline were administered by oral gavage at two different periods (with and without the presence of dental biofilm) to all rats except for the ten non-medicated rats. Alveolar bone area, alveolar bone level, and attachment level were determined by histomorphometric analysis, and gingival tissue levels of MMP-1, MMP-8, and TIMP-2 were detected by biochemical analysis. RESULTS: Significantly greater bone area and significantly less alveolar bone and attachment loss were observed in the kaempferol application groups compared to the control groups (P<0.05). In addition, gingival tissue MMP-1 and -8 levels were significantly lower in the kaempferol application groups compared to the control groups and the periodontitis group (P<0.001). There were no statistically significant differences in TIMP-2 levels between the kaempferol and saline application groups (P>0.05). CONCLUSIONS: Kaempferol application may be useful in decreasing alveolar bone resorption, attachment loss, and MMP-1 and -8 production in experimental periodontitis.

Effects of tacrolimus and nifedipine, alone or in combination, on gingival tissues.

BACKGROUND: The aim of this study is to compare gingival changes induced by short- and long-term tacrolimus and nifedipine administration, alone or in combination, and evaluate the expression levels of tumor suppressor phosphatase and tensin homolog (PTEN) in drug-induced gingival overgrowth. METHODS: Eighty rats were equally divided into eight groups: 1) tacrolimus for 8 weeks; 2) nifedipine for 8 weeks; 3) tacrolimus and nifedipine for 8 weeks; 4) 8-week control; 5) tacrolimus for 24 weeks; 6) nifedipine for 24 weeks; 7) tacrolimus and nifedipine for 24 weeks; and 8) 24-week control. Histomorphometric analyses included measurements of epithelial thickness, connective tissue thickness, and height. Stereologic analyses included measurements of volumetric densities of fibroblasts (Vf), collagen fibers (Vcf), and blood vessels (Vbv). In addition, PTEN expression was analyzed using immunohistochemistry. RESULTS: Epithelial thickness and connective tissue thickness were significantly increased in groups 5, 6, and 7 compared to group 8 (P <0.05), whereas connective tissue height was significantly increased in groups 5 and 7 (P <0.001). Vf and Vcf were significantly increased in group 7 compared to group 8 (P <0.001). PTEN immunoreactivity was significantly decreased in all experimental groups compared to the control groups (P <0.05). CONCLUSIONS: Results suggest that duration of drug administration is a more important risk factor than drug combination. The results include a potentially new insight about PTEN's role in the etiology of drug-induced gingival overgrowth.