Protective effect of alpha lipoic acid against oxidative damage caused by valproic acid on lung tissue in rats

To determine whether the possible oxidative effect of methotrexate (Mtx) on ovary and to evaluate the effectiveness of alpha lipoic acid (ALA), which may be useful in many oxidative stress models.Thirty-two female Wistar-albino rats were randomly divided into four groups; control group, alpha lipoic acid group (ALA 100 mg/kg, 10 days), multiple dose Mtx group (Mtx 1 mg/kg 1, 3, 5, 7 days) and Mtx and ALA group (Mtx 1 mg/kg 1, 3, 5, 7 days and ALA 100 mg/kg, 10 days). Serum total antioxidant status (TAS), total oxidant status (TOS) and oxidative stress index (OSI), tumor necrosis factor-alpha (TNF-α), tissue malondialdehyde (MDA) and activities of glutathione peroxidase (GSH-Px) and catalase (CAT) and anti-Mullerian hormone (AMH) and total ovarian follicle count were evaluated.Mtx administration caused a significant decrease in TAS, a significant increase in TOS and OSI, a significant increase in MDA levels and a decrease in GSH-Px and CAT activity. Moreover the proinflammatory cytokine (TNF-α) was increased in the Mtx group. And AMH values and total follicle count were significantly decreased in Mtx group. However, ALA treatment reversed biochemical results and AMH levels and total follicle count.Alpha lipoic acid ameliorates methotrexate induced oxidative damage of ovarian in rats.

We investigated the effects of alpha lipoic acid (ALA) on blood and lung tissue exposed chronically to cigarette smoke (CS). Female Sprague-Dawley rats were divided into three groups. Group 1 was the control group (CON): fresh air was supplied twice daily and 0.1 ml physiological saline was given orally for 8 weeks. Group 2 was exposed to CS: 12 cigarettes were smoked daily at two sessions for 1 h and 0.1 ml saline was given orally for 8 weeks. Group 3 (CS + ALA) was exposed to 12 cigarettes daily in two sessions for 1 h and 100 mg/kg/day ALA was given orally for 8 weeks. DNA damage was assessed using comet analysis; oxidative damage was assessed using ischemia-modified albumin (IMA) from blood; and total oxidant status (TOS), total antioxidant status (TAS) and oxidative stress index (OSI) were measured in blood and lung tissue. Histopathological and immunohistochemical evaluation of hypoxia-inducible factor (HIF)-1α, and −2α, caspase-3, vascular endothelial growth factor (VEGF) and fibroblast growth factor (FGF2) were conducted using lung tissue. The oxidative markers, TOS, OSI and IMA, and the comet analysis score were increased and the TAS level was decreased in the blood of the CS group compared to the CON group. IMA levels in blood, and TOS and OSI levels in the lung were decreased significantly in the CS + ALA group compared to the CS group. We observed increased septal wall thickness, marked and diffuse inflammatory reaction, emphysema, and necrotic cell debris in bronchial and bronchiolar lumens in the CS group. HIF-1α, HIF-2α, caspase-3 and FGF2 expressions were increased, while VEGF expression decreased in the lung tissues of the CS group compared to the CON group. ALA slightly ameliorated the damage caused by chronic exposure to CS in the lungs, but further investigation is needed to determine its possible protective effects at different dosages.

The protective effects of alpha lipoic acid on methotrexate induced testis injury in rats

Alpha lipoic acid is a potent antioxidant that plays numerous roles in human health. This study examined the effect of ALA on rat sciatic nerve ischemia reperfusion damage. Protective effect of alpha lipoic acid (ALA) on sciatic nerve following ischemia-reperfusion in rats was investigated by using light microscopy and biochemical methods. Provided that the protective effect of ALA on sciatic nerve is proven, we think the damage to the sciatic nerve that has already occurred or might occur in patients for various reasons maybe prevented or stopped by giving ALA in convenient doses. Animal experiment. Forty-two adult male Sprague-Dawley rats (250-300 grams) were used in this study. Rats were randomly divided into six groups including one control (Group 1), one sham (Group 2), two ischemia-reperfusion (Groups 3 and 4) and two treatment groups (Groups5 and 6). Doses of 60 and 100 mg/kg ALA were given (Group 5 and 6) intra peritoneally twice, 1 and 24 hours before the ischemia to each treatment group. Ischemia was carried out the abdominal aorta starting from the distal part of the renal vein for two hours followed by reperfusion for three hours. In immunohistochemical methods, fibronectin immunoreactivity was analyzed. For biochemical analyses, the tissues were taken in eppendorf microtubes and superoxide dismutase (SOD) and glutathione peroxidase (GSHPx) enzyme activities as well as malondialdehyde (MDA) and nitricoxide (NO) levels were measured. Fibronectin was observed to have increased significantly in the ischemia group; on the other hand, it was observed to have decreased in parallel to the doses in the ALA groups. Biochemical studies showed that SOD and GSHPx declined with ischemia-reperfusion, but the activities of these enzymes were increased in the treatment groups in parallel with the dose. It was found that increased MDA levels with ischemia-reperfusion were decreased in parallel with ALA dose. There were no statistically significant changes in NO. Increased fibronectin observed after ischemia/reperfusion of rat sciatic nerve is reduced after the administration of ALA. This indicates that the function of fibronectin, to reconnect cut nerve segments and regenerate nerves, is more prominent than its function in tissue healing after ischemia. ALA administered before ischemia decreases MDA and increases SOD and GSHPx. We think that ALA may protect against the pathological changes in ischemic nerve and may be used to devise more efficient treatments.

Radioiodine (RI) therapy is known to cause salivary gland (SG) dysfunction. The effects of antioxidants on RI-induced SG damage have not been well described. This study was performed to investigate the radioprotective effects of alpha lipoic acid (ALA) administered prior to RI therapy in a mouse model of RI-induced sialadenitis. Four-week-old female C57BL/6 mice were divided into four groups (n = 10 per group): group I, normal control; group II, ALA alone (100 mg/kg); group III, RI alone (0.01 mCi/g body weight, orally); and group IV, ALA + RI (ALA at 100 mg/kg, 24 h and 30 min before RI exposure at 0.01 mCi/g body weight). The animals in these groups were divided into two subgroups and euthanized at 30 or 90 days post-RI treatment. Changes in salivary 99mTc pertechnetate uptake and excretion were tracked by single-photon emission computed tomography. Salivary histological examinations and TUNEL assays were performed. The 99mTc pertechnetate excretion level recovered in the ALA treatment group. Salivary epithelial (aquaporin 5) cells of the ALA + RI group were protected from RI damage. The ALA + RI group exhibited more mucin-containing parenchyma and less fibrotic tissues than the RI only group. Fewer apoptotic cells were observed in the ALA + RI group compared to the RI only group. Pretreatment with ALA before RI therapy is potentially beneficial in protecting against RI-induced salivary dysfunction.

P8 – 2228: The effects of anti-epileptic drugs on the growth plates of infant rats

A rodent autism spectrum disorder (ASD) model based on prenatal exposure to valproic acid (VPA) is widely recognized as a prominent model. Social behavior in rodent ASD models has primarily been evaluated through a three-chamber approach test. However, in this study, we focused on social attention in the VPA model of ASD. In male C57BL/6 J mice, attentional behaviors toward conspecifics were examined through reaching tasks around 9-11 weeks of age. On embryonic day 12.5, pregnant mice underwent a subcutaneous injection of 600 mg/kg VPA sodium salt dissolved in 0.9% saline solution (VPA group) or saline solution alone (Sal group) into their neck fat. Thirty-six mice-nine each in the VPA and saline groups, and 18 partners-underwent training in reaching behavior. Subsequently, we examined whether the VPA or Sal group demonstrated focused attention toward their partners during reaching tasks. A two-way analysis of variance (ANOVA) (condition [VPA/Sal] × situation [face-to-face (attention)/not paying attention (not attention)]) was conducted on the average success rate of the situation. Additionally, we measured the duration of sniffing behavior between pairs of mice in an open field twice in total at 4 and 8 weeks of age before reaching task. The pairs were constructed by pairing a VPA or Sal group mouse with its partner, with the objective of facilitating initial encounters between the mice. A one-way ANOVA was conducted on the average duration of sniffing behavior data from 4 weeks and a second one-way ANOVA on data from 8 weeks. The analysis revealed a significant interaction between condition and situation in the reaching task [F (1, 28) = 6.75, p = 0.015, ηp 2 = 0.19]. The simple main effect test exhibited that the "not paying attention" rate was significantly higher than that of the "face-to-face" in the VPA group (p < 0.01). The results revealed a not significant difference in the average duration of sniffing behavior at 4 weeks [F (3, 32) = 2.71, p = 0.06, n.s., ηp 2 = 0.20], but significant difference at 8 weeks [F (3, 32) = 4.12, p < 0.05, ηp 2 = 0.28]. Multiple comparisons using the Bonferroni method revealed significant differences in the sniffing duration at 8 weeks between from the partner toward the VPA mouse and from the partner toward the Sal mouse (p < 0.05). The VPA rodent model of ASD exhibited differences in social attention compared to the saline group. By focusing on social attention and exploring various ASD models, insights can be gained from the neural mechanisms underlying gaze abnormalities during social interaction in individuals with ASD.

230 Background: Histone deacetylase (HDAC) is strongly associated with epigenetic regulation and carcinogenesis, and its inhibitors induce the differentiation or apoptosis of cancer cells. Valproic acid (VPA) is one of the clinically available HDAC inhibitors. We previously showed that VPA augmented antitumor effect of GEM in choalngiocarcinoma cell line (2010 GI Symposium); this time, we performed microarray analysis and Ingenuity Pathways Analysis (IPA) to identify the systematic mechanism of the augmentative effect of VPA. Methods: Human cholangiocarcinoma cell line (HuCCT1) was used. The anticancer effects of VPA or gemcitabine (GEM), and the effects of VPA combined with GEM were studied by MTT assay. We divided the following four groups: control group, VPA group, GEM group, VPA plus GEM combination group. The gene expressions of p21, HDAC, VEGF, and HIF-1 were evaluated by RT-PCR. And, the microarray analysis was performed, the genes were picked up using Gene Spring GX10, and then IPA was performed. Results: In GEM alone group, no effect of GEM was observed in dose of 5 mm, and 16% of proliferation-inhibitory effects were observed in dose of 10 nm. In VPA alone group, no effect of VPA was observed in dose of 0.5 mm, and 12%, 35%, and 67% of proliferation-inhibitory effects were observed in dose of 1.0, 5.0, and 10mm, respectively. GEM (5 nm) and VPA (0.5 mm) reduced by 23%, which significantly augmented the anticancer effect of GEM alone or VPA alone (p&lt;0.01). Furthermore, GEM combined with VPA upregulated the p21 expression compared with single agent (p&lt;0.05). And, in regard to microarray analysis, we analyzed in 28,869 genes. The 24 genes were picked up with the comparison between VPA group and VPA plus GEM combination group using Gene Spring GX10, and the gene network of the cellular development containing the gene relevant to the differentiation of cancer cell, HLA-DR, was formed with IPA. Conclusions: VPA augmented the effects of anticancer agents in a cholangiocarcinoma cell line. Such effects may be owing to the gene network of the cellular development. HDAC inhibitor may have the effect of the differentiation of cancer cell. No significant financial relationships to disclose.

Adrenomedullin expression in aortic artery wall of diabetic rats given alpha lipoic acid.

Valproic Acid Increased Autophagic Flux in human Multiple Myeloma Cells in Vitro

Objective To explore the effects of histone acetylation and deacetylation on C2C12 myoblasts differentiation. Methods Based on the differentiation of C2C12 myoblasts into myotubes using high glucose dulbecco's modified eagle medium (DMEM) containing 2% horse serum in vitro, valproic acid(VPA) was given to C2C12 myoblasts during differentiation with different concentrations, which contained 1 mmol/L VPA, 2 mmol/L VPA, 4 mmol/L VPA and 8 mmol/L VPA in the final concentrations with 2% horse serum and high glucose DMEM. So that this experiment was scheduled into 6 groups as control group(contain 10% fetal bovine serum in growth medium), horse serum induced differentiation group, 1 mmol/L VPA group, 2 mmol/L VPA group, 4 mmol/L VPA group and 8 mmol/L VPA group according to different growth medium. There were 6 samples in each group. The myotube differentiation rate were compared in different concentration VPA groups and horse serum induced differentiation group. Besides, mRNA and protein expression levels of muscle-related proteins(including Myosin, Troponin I-SS, myogenic differentiation 1) and histone deacetylases (HDAC, including HDAC1, 2, 3) were also evaluated with real time polymerase chain reaction (RT-PCR) and Western blotting. The mRNA and protein expression levels of them were compared and analyzed. Results ①The mRNA and protein expression levels of muscle-related proteins of horse serum induced differentiation group were significantly higher than those of control group, and the differences were statistically significant (P 0.05). ②The myotube differentiation rate in every concentration VPA group compared with horse serum induced differentiation group were significantly lower, and the differences were statistically significant (P<0.05). ③ The mRNA and protein expression levels of muscle-related proteins and HDAC in 4 mmol/L VPA group or 8 mmol/L VPA group compared with horse serum induced differentiation group were significantly decreased, and the differences were statistically significant (P<0.05). ④The impacts of VPA on the expression of muscle-related proteins and HDAC in mRNA level and protein level during C2C12 differentiation were identified as a concentration dependent pattern, indicated a negative linear correlation(r=-0.92, -0.89, -0.93, -0.84, -0.93, -0.97; r=-0.99, -0.86, -0.92, -0.95, -0.91, -0.96; P<0.05). Conclusions Histone acetylation and deacetylation played an important role in skeletal muscle cell differentiation. The inhibition of HDAC could result in the reduction of transcription and expression of muscle-related proteins and suppressed the differentiation of myoblasts. Key words: Histone acetyltransferases; Histone deacetylases; Cell differentiation; Valproic acid

Objective To investigate the relationship between uridine diphosphate glucuronosyltransferases (UGT) 1A6 rs2070959 polymorphism and serum concentration of valproic acid (VPA) in patients with epilepsy. Methods We selected 200 cases of epilepsy patients treated in our hospital from June 2014-January 2017. All the patients were treated with VPA monotherapy > 3months. When the VPA of patients reached steady state, we detected the VPA blood level. The genotypes and allele frequencies of UGT1A6 rs2070959 in 200 epilepsy patients were determined. The average standard deviation of 1-fold VPA was used as high VPA group and vice versa as low VPA group. The genotype and allele frequencies of UGT1A6 rs2070959 were compared between the two groups, and the influencing factors of VPA concentration in epileptic patients were analyzed. Results The frequencies of A genotype, AG genotype and GG genotype were 77.01%, 18.39% and 4.60% in the high VPA group, 67.26%, 19.47% and 13.27% in the low VPA group, with no significant difference between the two groups (P>0.05); the allele G frequency in the high VPA group was significantly lower than that in the low VPA group (13.79%). The age and weight of the high VPA group were significantly higher than those of the low VPA group (P 0.05). Logistic regression analysis showed that age and weight gain were positively correlated with VPA concentration (P<0.05), allele G expression was negatively correlated with VPA concentration (P<0.05). Conclusions Allele G expression at UGT1A6 rs2070959 is associated with decreased VPA concentration in epileptic patients, which may require a higher dose of VPA. Key words: Epilepsy/DT; Valproic acid/TU; Glucuronosyltransferase/GE; Polymorphism, single nucleotide; Pharmacokinetics

Valproic acid (VPA) is widely used antiepileptic agent which is associated with reproductive toxicity via impairment in oxidative redox. Zinc (Zn) and selenium (Se) are trace element with antioxidant effect that known to be essential for spermatogenesis. In the current study, the protective effect of co-administration of Zn and Se on VPA-induced reproductive toxicity in male rats was evaluated. Forty-eight male rats were divided into 8 groups of six (n=6): Control group (treated with normal saline); VPA only (250, 500, 1,000 mg/kg) group; VPA (500 mg/kg) plus Zn (2 mg/kg) group; VPA (500 mg/kg) plus Se (1.5 mg/kg) group; VPA (500 mg/kg) plus a combination of Zn and Se group; and VPA+vitamin E (20 mg/kg) group. The Animals were sacrificed after 28 days of treatment and sperm analysis was taken. Also, evaluation of oxidative stress markers including malondialdehyde (MDA), protein carbonyl (PC), glutathione (GSH) and histopathological changes were done on testis tissue. Morphological changes and a significant decrease in motility and sperm count in rats treated with VPA were observed. Also, an increase in oxidative stress marker, including MDA and PC and a decrease in GSH level was evident in VPA group. Zn and Se administration was able to protect against sperm abnormality, ameliorate the histological change in testis tissue, and suppressed the increase in oxidative stress markers induced by VPA. These results indicated that combination therapy with Zn and Se showed better an ameliorative effect than each one alone. Therefore, it can be suggested as an effective supplement for reproductive impairment in VPA-treated patient.

Background: Paracetamol, is the most widely used over-the-counter analgesic and antipyretic medication in the world, which has minimal adverse effects at therapeutic dosages. But in high doses causes hepatic damage and oxidative stress. Objectives: The current study was designed to investigate paracetamol toxic effects upon the liver and oxidative stress after repeated oral dose and evaluate possible protective effect of alpha lipoic acid when co-administered with and after paracetamol. Methods: forty eight white albino rats were divided equally into four groups. Each group was subdivided into two sub groups A & B. Group I received gum acacia suspension. Group II received Alpha lipoic acid (50mg/kg) orally. Group III received paracetamol (1 gm/ kg orally) for 4 weeks. Group IV received paracetamol and alpha lipoic acid at the same doses. Sub groups A were euthanized after 4 weeks, while sub groups B were euthanized after 8 weeks. Blood was collected for evaluation of liver functions and oxidative stress marker. The livers were preserved for histopathological examinations. Results: The study proved that repeated administration of paracetamol induced disturbed liver functions and oxidative stress. But this toxic effects decline markedly when alpha lipoic acid (ALA) was coadministered with paracetamol. And more improvement occurs when ALA was administered for another 4 weeks after stoppage of paracetamol. Conclusions: The present study concluded that repeated paracetamol administration has hepatotoxic and oxidative stress effect and alpha lipoic acid has a protective effect against such harmful effects especially when ALA was administered for another 4 weeks after stoppage of paracetamol.

Antioxidant and anti-inflammatory activities of alpha lipoic acid protect against indomethacin-induced gastric ulcer in rats