Comparative neuroprotective effects of royal jelly and its unique compound 10-hydroxy-2-decenoic acid on ischemia-induced inflammatory, apoptotic, epigenetic and genotoxic changes in a rat model of ischemic stroke.

OBJECTIVES: This study aimed to compare the efficacy of royal jelly (RJ) and its major fatty acid 10-hydroxy-2-decenoic acid (10-HDA) on ischemic stroke-related pathologies using histological and molecular approaches. METHODS: Male rats were subjected to middle cerebral artery occlusion (MCAo) to induce ischemic stroke and were supplemented daily with either vehicle (control group), RJ or 10-HDA for 7 days starting on the day of surgery. On the eighth day, rats were sacrificed and brain tissue and blood samples were obtained to analyze brain infarct volume, DNA damage as well as apoptotic, inflammatory and epigenetic parameters. RESULTS: Both RJ and 10-HDA supplementation significantly reduced brain infarction and decreased weight loss when compared to control animals. These effects were associated with reduced levels of active caspase-3 and PARP-1 and increased levels of acetyl-histone H3 and H4. Although both RJ and 10-HDA treatments significantly increased acetyl-histone H3 levels, the effect of RJ was more potent than that of 10-HDA. RJ and 10-HDA supplementation also alleviated DNA damage by significantly reducing tail length, tail intensity and tail moment in brain tissue and peripheral lymphocytes, except for the RJ treatment which tended to reduce tail moment in lymphocytes without statistical significance. CONCLUSIONS: Our findings suggest that neuroprotective effects of RJ in experimental stroke can mostly be attributed to 10-HDA.

The effect of anastrozole therapy on final height and sex hormone levels in pubertal boys receiving growth hormone therapy

ABSTRACT Objective: This research aimed to evaluate retrospectively the effect of anastrozole on height gain and sex hormone levels in pubertal boys receiving growth hormone (GH). Materials and methods: Pubertal boys who received both GH and anastrozole (GH+A) were one-to-one matched with boys who received only GH (GH-Only) for chronological and bone age, pubertal stage and height before the GH initiation, treatment duration and midparental height. Anthropometric measurements throughout treatment and adult heights were compared between the groups. Sex hormone levels were evaluated longitudinally in the GH+A group. Results: Forty-eight cases (24 in each group) were included. There was no statistical difference in adult height between the GH+A and GH-Only (p = 0.071). However, when the analysis was limited to those receiving anastrozole for at least 2 years, mean adult height was higher in the GH+A than in the GH-Only group (173.1 ± 6.2/169.8 ± 5.6 cm, p = 0.044). Despite similar growth rates between the two groups, bone age advancement was slower in the GH+A than in the GH-Only in a mean anastrozole treatment period of 1.59 years (1.37 ± 0.80/1.81 ± 0.98 years, p = 0.001). The greatest increase for FSH, LH, total and free testosterone and decrease for estradiol levels were observed in the third month after anastrozole was started, albeit remaining within the normal ranges according to the actual pubertal stages. Conclusions: Using anastrozole with GH for at least 2 years decelerates the bone age advancement resulting in adult height gain with no abnormality in sex hormone levels. These results suggest anastrozole can be used as an additional treatment to GH for further height gain in pubertal boys.

The effect of anastrozole therapy on final height and sex hormone levels in pubertal boys receiving growth hormone therapy.

Objective: This research aimed to evaluate retrospectively the effect of anastrozole on height gain and sex hormone levels in pubertal boys receiving growth hormone (GH). Materials and methods: Pubertal boys who received both GH and anastrozole (GH+A) were one-to-one matched with boys who received only GH (GH-Only) for chronological and bone age, pubertal stage and height before the GH initiation, treatment duration and midparental height. Anthropometric measurements throughout treatment and adult heights were compared between the groups. Sex hormone levels were evaluated longitudinally in the GH+A group. Results: Forty-eight cases (24 in each group) were included. There was no statistical difference in adult height between the GH+A and GH-Only (p = 0.071). However, when the analysis was limited to those receiving anastrozole for at least 2 years, mean adult height was higher in the GH+A than in the GH-Only group (173.1 ± 6.2/169.8 ± 5.6 cm, p = 0.044). Despite similar growth rates between the two groups, bone age advancement was slower in the GH+A than in the GH-Only in a mean anastrozole treatment period of 1.59 years (1.37 ± 0.80/1.81 ± 0.98 years, p = 0.001). The greatest increase for FSH, LH, total and free testosterone and decrease for estradiol levels were observed in the third month after anastrozole was started, albeit remaining within the normal ranges according to the actual pubertal stages. Conclusion: Using anastrozole with GH for at least 2 years decelerates the bone age advancement resulting in adult height gain with no abnormality in sex hormone levels. These results suggest anastrozole can be used as an additional treatment to GH for further height gain in pubertal boys.

Inulinase and fructooligosaccharide production from carob using Aspergillus niger A42 (ATCC 204447) under solid-state fermentation conditions.

This study aimed to the production of inulinase and fructooligosaccharides (FOSs) from carob under the solid-state fermentation (SSF) conditions by using Plackett-Burman Design (PBD). Based on the results the maximum inulinase and specific inulinase activities were 249.98 U/mL and 318.29 U/mg protein, respectively. When the fructooligosaccharide (FOS) results were evaluated, the maximum values of 1,1,1-Kestopentaose, 1,1-Kestotetraose, and 1-Kestose were 182.01, 506.16, 132.16 ppm while the lowest and highest total FOS values were 179.35 and 516.66 ppm, respectively. On the other hand, it was observed that the maximum inulinase activity was found at the center points of the design. Therefore, validation fermentations were carried out at center point conditions. Subsequently, the yielded bulk enzyme extracts were partially purified using Spin-X UF membranes with 10, 30, and 50 kDa cut-off values. After purification, the maximum inulinase activity was 247.30 U/mg using a 50 kDa cut-off value. Followed by this process, the purified enzyme was used to produce FOSs and the results indicated that the maximum total FOS amount was 28,712.70 ppm. Consequently, this study successfully demonstrates that Aspergillus niger A42 inulinase produced from carob under the SSF conditions can be used in FOSs production.

Preventive effects of antenatal CDP-choline in a rat model of neonatal hyperoxia-induced lung injury.

Antenatal steroid administration to pregnant women at risk of prematurity provides pulmonary maturation in infants, while it has limited effects on incidence of bronchopulmonary dysplasia (BPD), the clinical expression of hyperoxia-induced lung injury (HILI). Cytidine-5'-diphosphate choline (CDP-choline) was shown to alleviate HILI when administered to newborn rats. Therefore, we investigated effects of maternal administration of CDP-choline, alone or in combination with betamethasone, on lung maturation in neonatal rats subjected to HILI immediately after birth. Pregnant rats were randomly assigned to one of the four treatments: saline (1 mL/kg), CDP-choline (300 mg/kg), betamethasone (0.4 mg/kg), or CDP-choline plus betamethasone (combination therapy). From postnatal day 1 to 11, pups born to mothers in the same treatment group were pooled and randomly assigned to either normoxia or hyperoxia group. Biochemical an d histopathological effects of CDP-choline on neonatal lung tissue were evaluated. Antenatal CDP-choline treatment increased levels of phosphatidylcholine and total lung phospholipids, decreased apoptosis, and improved alveolarization. The outcomes were further improved with combination therapy compared to the administration of CDP-choline or betamethasone alone. These results demonstrate that antenatal CDP-choline treatment provides benefit in experimental HILI either alone or more intensively when administered along with a steroid, suggesting a possible utility for CDP-choline against BPD.

Effects of uridine administration on hippocampal matrix metalloproteinases and their endogenous inhibitors in REM sleep-deprived rats.

Rapid eye movement (REM) sleep is associated with synaptic plasticity which is considered essential for long-term potentiation (LTP). The composition of extracellular matrix (ECM), in part, plays a role in REM sleep-associated synaptic functioning. The objective of this study was to investigate the effects of uridine administration on levels of matrix metalloproteinases (MMPs) and their endogenous inhibitors (TIMPs) in rats subjected to REM sleep deprivation (REMSD). REMSD was induced by modified multiple platform method for 96-hour. Rats were randomized to receive either saline or uridine (1 mmol/kg) intraperitoneally twice a day for four days. Rats were then decapitated and their hippocampi were dissected for analyzing the levels of MMP-2, MMP-3, MMP-9, TIMP-1, TIMP-2 and TIMP-3 by Western-blotting and the activities of MMP-2 and MMP-9 by Gelatin zymography. REMSD resulted in reduced levels of MMP-3, MMP-9, TIMP-3 and activity of MMP-9 in saline-treated rats, while uridine treatment significantly enhanced their impairment. TIMP-1 was enhanced following REMSD but uridine treatment had no significant effect on TIMP-1 levels. MMP-2, TIMP-2 levels and MMP-2 activity were not affected by either REMSD or uridine administration. These data show that REMSD significantly affects ECM composition which is ameliorated by uridine administration suggesting a possible use of uridine in sleep disorders.

Anti-Apoptotic and Anti-Oxidant Effects of Systemic Uridine Treatment in an Experimental Model of Sciatic Nerve Injury.

AIM: To investigate the anti-apoptotic and anti-oxidant effects of systemic uridine treatment in a rat model of sciatic nerve injury. MATERIAL AND METHODS: Thirty-two adult male rats were equally randomized to Sham, Control, U100, and U500 groups. Sham rats received a sham operation by exposing the right sciatic nerve without transection, while those in the Control, U100, and U500 groups underwent right sciatic nerve transection followed by immediate primary anostomosis. Sham and Control groups received saline (0.9% NaCl) injections intraperitoneally (i.p.), while U100 and U500 groups received 100 mg/kg and 500 mg/kg uridine injections (i.p.), respectively, once a day for 7 days after the surgery. Rats in all the groups were sacrificed on the eighth day; sciatic nerve samples were analyzed for apoptosis by Western Blotting and for oxidation parameters including myeloperoxidase (MPO), malondialdehyde (MDA), superoxide dismutase (SOD), glutathione peroxidase (GPx) and catalase (CAT) by Enzyme-Linked Immunosorbent Assay (ELISA). RESULTS: Uridine treatment at the dose of 500 mg/kg significantly decreased as apoptosis determined by Caspase-3/Actin ratio and exhibited significant anti-oxidant effects as determined by decreased levels of MPO and MDA as well as increased levels of SOD, GPx, and CAT compared to controls. Uridine at 100 mg/kg was only found to decrease the Caspase-3/Actin ratio, although it significantly decreased MDA and increased CAT levels compared to controls. CONCLUSION: Treatment with uridine reduces apoptosis and oxidation in a rat model of sciatic nerve injury dose-dependently. Thus, uridine may be beneficial in peripheral nerve regeneration by exhibiting anti-apoptotic and anti-oxidant effects.

Uridine treatment improves nerve regeneration and functional recovery in a rat model of sciatic nerve injury.

AIM: Peripheral nerve regeneration remains an issue, and novel therapeutic approaches are required for functional recovery. This study investigated the regenerative potential and long-term functional effects of Uridine treatment in a rat model of sciatic nerve injury. MATERIAL AND METHODS: Male Sprague-Dawley rats were randomized to receive sham surgery plus saline (Sham group), right sciatic nerve transection and primary repair plus saline (Control group), right sciatic nerve transection, and primary repair plus 500 mg/kg Uridine (Uridine group). Saline or Uridine was injected intraperitoneally (i.p.) for seven days, and the rats were monitored for 12 weeks after surgery. We evaluated electrophysiological and functional recovery using electromyography (EMG) and sciatic functional index (SFI) at six and 12 weeks, respectively. At 12 weeks, rats were decapitated and their right sciatic nerves were examined in macroscopic and histomorphologic manners. RESULTS: Functional evaluation by SFI and sciatic nerve conduction velocity analyzed by EMG both decreased in the Control group but recovered in the Uridine group 12 weeks after surgery. Additionally, upon experiment completion, Uridine treatment was observed to enhance nerve adherence, separability scores, and the number of myelinated axons. CONCLUSION: These results reveal that short-term Uridine treatment provides morphological and electrophysiological benefits, which are represented by long-term functional improvement in a rat model of sciatic nerve injury. These findings validate and extend our knowledge on Uridine's regenerative effects in peripheral nerve injuries.

Antioxidative effects of uridine in a neonatal rat model of hyperoxic brain injury

Background/aim: Premature birth is a major problem that results in an increased risk of mortality and morbidity. The management of such infants consists of supraphysiological oxygen therapy, which affects brain development due, in part, to the deterioration caused by reactive oxygen species (ROS). We showed previously that exogenously administered uridine provides neuroprotection in a neonatal rat model of hyperoxic brain injury. Hence, the aim of the present study was to investigate the effects of uridine on ROS in the same setting. Materials and methods: Hyperoxic brain injury was induced by subjecting a total of 53 six-day-old rat pups to 80% oxygen (the hyperoxia group) for a period of 48 h. The pups in the normoxia group continued breathing room air (21% oxygen). Normoxia + saline or hyperoxia + saline or hyperoxia + uridine 100 mg/kg or hyperoxia + uridine 300 mg/kg or hyperoxia + uridine 500 mg/kg was injected intraperitoneally (i. p.) 15 min prior to the hyperoxia procedure. The pups were decapitated and the brains were homogenized to analyze superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), myeloperoxidase (MPO), and malondialdehyde (MDA) enzymes as well as DJ-1 (protein deglycase DJ-1) ­ an oxidative stress-sensitive protein. Results: Hyperoxia-induced may cause overproduction of oxygen radicals and the oxidant/antioxidant balance may be disturbed in the brain. Brain MPO and MDA levels were significantly increased in saline-receiving pups exposed to hyperoxia. Brain SOD and GSH-Px levels were significantly decreased in saline-receiving pups exposed to hyperoxia. Our results showed that uridine administration prevented the hyperoxia-induced decrease in SOD and GSH-Px while counteracting the hyperoxia-induced increase in MPO and MDA in a dose-dependent manner. Uridine also increased the DJ-1 levels in brains of rat pups subjected to hyperoxia. Conclusion: These data suggest that uridine exhibits antioxidative properties which may mediate the protective effects of uridine in a neonatal rat model of hyperoxic brain injury.

Effects of CDP-choline administration on learning and memory in REM sleep-deprived rats.

Cytidine 5-diphosphocholine (CDP-choline) administration has been shown to improve learning and memory deficits in different models of brain disorders. In this study, effects of CDP-choline on the well known negative effects of Rapid Eye Movements (REM) sleep deprivation on learning and memory were investigated. Sleep deprivation was induced by placing adult male Wistar albino rats on 6.5 cm diameter platforms individually for 96 h according to flower pot method. Learning and memory performances were evaluated using Morris Water Maze (MWM) test during the same period of time. Saline or CDP-choline (100 µmol/kg, 300 µmol/kg or 600 µmol/kg) was administered intraperitoneally 30 min prior to the onset of MWM experiments. On completion of behavioral tests, rats were decapitated and hippocampi were assayed for total and phosphorylated Ca2+/calmodulin-dependent protein kinase II (tCaMKII and pCaMKII, respectively) and total antioxidant capacity. We observed that while REM sleep deprivation had no effect on learning, it diminished the memory function, which was associated with decreased levels of pCaMKII and total antioxidant capacity in the hippocampus. CDP-choline treatment blocked the impairment in memory function of sleep-deprived rats and, increased pCaMKII levels and total antioxidant capacity. These data suggest that CDP-choline reduces REM sleep deprivation-induced impairment in memory, at least in part, by counteracting the disturbances in biochemical and molecular biological parameters.

Proteomics Analysis of CA1 Region of the Hippocampus in Pre-, Progression and Pathological Stages in a Mouse Model of the Alzheimer's Disease.

BACKGROUND: CA1 subregion of the hippocampal formation is one of the primarily affected structures in AD, yet not much is known about proteome alterations in the extracellular milieu of this region. OBJECTIVE: In this study, we aimed to identify the protein expression alterations throughout the pre-pathological, progression and pathological stages of AD mouse model. METHODS: The CA1 region perfusates were collected by in-vivo intracerebral push-pull perfusion from transgenic 5XFAD mice and their non-transgenic littermates at 3, 6 and 12 wereßmonths of age. Morris water maze test and immunohistochemistry staining of A performed to determine the stages of the disease in this mouse model. The protein expression differences were analyzed by label-free shotgun proteomics analysis. RESULTS: A total of 251, 213 and 238 proteins were identified in samples obtained from CA1 regions of mice at 3, 6 and 12 months of age, respectively. Of these, 68, 41 and 33 proteins showed statistical significance. Pathway analysis based on the unique and common proteins within the groups revealed that several pathways are dysregulated during different stages of AD. The alterations in glucose and lipid metabolisms respectively in pre-pathologic and progression stages of the disease, lead to imbalances in ROS production via diminished SOD level and impairment of neuronal integrity. CONCLUSION: We conclude that CA1 region-specific proteomic analysis of hippocampal degeneration may be useful in identifying the earliest as well as progressional changes that are associated with Alzheimer's disease.

Uridine treatment prevents REM sleep deprivation-induced learning and memory impairment.

Previous studies have shown that sleep plays an important role in cognitive functions and sleep deprivation impairs learning and memory. Uridine is the main pyrimidine nucleoside found in human blood circulation and has beneficial effects on cognitive functions. The aim of the present study was to investigate the effects of uridine administration on learning and memory impairment in sleep-deprived rats. Flower pot method was used to induce REM sleep deprivation. Uridine-treated groups received 1 mmol/kg uridine and control groups received 1 ml/kg saline (0.9% NaCl) twice a day for four days and once a day on the 5th day intraperitoneally. Learning and memory performances were measured using Morris water maze (MWM) test. We also measured the ratios of total calcium-calmodulin dependent kinase II (tCaMKII)/ß-tubulin and phosphorylated cyclic adenosine monophosphate (cAMP) response element binding protein (pCREB)/ß-tubulin, long-term potentiation (LTP) related molecules, using western blot analysis on the hippocampus. The results showed that REM sleep deprivation impaired learning and memory and also decreased the ratios of tCaMKII and pCREB. Uridine treatment enhanced learning and memory parameters in REM sleep-deprived rats. Additionally, decreases in tCaMKII and pCREB were prevented by uridine treatment. These data suggest that administration of uridine for five consecutive days prevents REM sleep deprivation-induced deficits in learning and memory associated with enhanced tCaMKII and pCREB ratios in the hippocampus.