Apoptosis Induction with Combined Use of Cisplatin and Fisetin in Cisplatin-Resistant Ovarian Cancer Cells (A2780).

BACKGROUND: Ovarian cancer is the leading cause of death caused by genital cancers. One of the most common treatments for this type of cancer is chemotherapy by cisplatin, which induces apoptosis in cancer cells. Apoptosis is a type of physiological cell death. Cisplatin chemotherapy usually has several side effects and cellular resistance to cisplatin is a common incidence. In order to overcome these problems, the use of combination therapies using natural substances has been considered. Fisetin is a flavonoid with anti-cancer activity which induces apoptosis. In this study, the apoptosis induced by cisplatin along with Fisetin in cisplatin-resistant ovarian cancer cell line (A2780) was investigated. METHODS: In the present experimental study, the effect of combined use of Fisetin and cisplatin on ovarian cancer cell lines (A2780) was investigated by using MTT assay. Cell death was also determined by DAPI, acridine orange/propidium iodide, and Annexin/PI assay. Apoptotic gene expression of Bax, BCL-2, caspase 3, and caspase 9 was also assessed by real time PCR. RESULTS: The results of MTT assay indicated that the combined treatment of Fisetin and cisplatin effectively inhibits proliferation of A2780 cells. The results of DAPI staining showed that fragmentation of chromatin in cells occurred in the combined treatment. Acridine orange-propidium iodide staining and Annexin/PI staining showed an increase in the rate of apoptotic cells in cells under combined treatment. The results of the study regarding changes in gene expression also indicated that Bax pro-apoptotic gene expression and BCL-2 anti-apoptotic gene expression increased in cells under treatment; moreover, gene expression of caspases 3 and 9 significantly increased as well. CONCLUSION: According to the findings of this study, the combined use of cisplatin and Fisetin increases the induction of apoptosis in cisplatin-resistant ovarian cancer cells (A2780); therefore, the combined use of cisplatin and Fisetin can be considered a promising strategy in the treatment of ovarian cancer.

The effect of hydroalcoholic extract of Ziziphora clinopodioides L. on spatial memory and neuronal density of hippocampal CA1 region in rats with sporadic Alzheimer's disease.

OBJECTIVE: Alzheimer's disease is a neurodegenerative disorder associated with gradual loss of cognitive and memory abilities. It was shown that the hippocampus is one of the first structures in the brain that is affected by the disease. Ziziphora clinopodioides (Z. clinopodioides) is a member of Lamiaceae family and contains various substances. MATERIALS AND METHODS: In this experimental study, 72 adult male Wistar rats were used for behavioral and histopathologic studies. They were divided into nine groups included: control, negative control (Alzheimer), positive control (Alzheimer's treated with rivastigmine), aCSF (artificial cerebrospinal fluid) + ziziphora extract with doses of 200,400, and 600 mg/kg, and STZ (stereptozotocine)+ziziphora extract in 200,400,600 mg/kg doses. The injury was created with bilaterally intraventricular injection. The spatial memory was studied by passive avoidance test and neuronal density was evaluated by dissector method. To examine the histopathological lesions, Congo red and toluidine blue staining were done. Data were analyzed by ANOVA Minitab software. RESULTS: The memory index (neuronal density and passive avoidance test results) showed a significant decrease in negative control group compared to control (p≤0.001). Treatment with the hydroalcoholic extract at the doses of 400 and 600 mg/kg showed a significant increase in memory index in rats with Alzheimer's disease (p≤0.001). The effect of 200 mg/kg extract was not significantly different from that of the negative control group. The results of histological analysis indicated beta-amyloid plaques formation in the control group as compared to the negative control group while treatment with the extract at the doses of 400 and 600 mg/kg, significantly reduced beta-amyloid plaques formation. CONCLUSION: These findings suggest that the extract of Z. clinopodioides can improve Alzheimer's condition and alleviate memory and histopathologic damages; also, it decreases beta-amyloid plaques and apoptosis in CA1 region of the hippocampus.

Preventive effects of the aqueous extract of Cichorium intybus L. flower on ethylene glycol-induced renal calculi in rats.

OBJECTIVE: Urolithiasis remains a global problem. Despite the availability of numerous methods, no definite therapeutic agent has been yet introduced for the prevention or treatment of kidney stones. In this study, we evaluated the possible preventive effects of aqueous extract of Cichorium intybus L. (chicory) flowers on ethylene glycol-induced renal calculi in rats. MATERIALS AND METHODS: A total of 24 Wistar rats were randomly divided into four groups and were treated for 30 days. Group A received drinking tap water, while groups B, C, and D were administered with 1% ethylene glycol for induction of calcium oxalate stone formation. Rats in groups C and D received intraperitoneal injections of the aqueous extract of chicory flowers (50 and 200 mg/kg, respectively) since the first day of the experiment. The urine volume, urine pH, and urinary levels of oxalate, citrate, calcium, uric acid, and creatinine as well as serum levels of calcium, uric acid, and creatinine were measured. After 30 days, the rats' kidneys were removed and prepared for histological evaluation of calcium oxalate deposits. One-way analysis of variance (ANOVA), followed by Tukey's test, was performed, using SPSS version 20. RESULTS: The number of calcium oxalate crystals was significantly higher in group B (ethylene glycol-only treated animals), compared to group A (control), group C (50 mg/kg of aqueous extract), and group D (200 mg/kg of aqueous extract) (p<0.05). On day 30, the urine level of citrate, oxalate (p>0.05), and creatinine (p<0.05), as well as urine pH (p<0.05) decreased in groups C and D, compared to group B. Also, urine calcium level, urine uric acid (p>0.05), and urine volume (p<0.05) were higher in group D, compared to group B. In addition, the serum level of calcium, creatinine (p<0.05), and uric acid (p<0.001) decreased in groups C and D. CONCLUSION: The aqueous extract of chicory flower (50 mg/kg) could reduce the number of calcium oxalate deposits in the urine and reduce the level of serum parameters.

Different responses of nitric oxide synthase inhibition on morphine-induced antinociception in male and female rats.

The roles of gonadal hormones and nitric oxide on pain perception and their interaction have been widely investigated. In the present study the chronic effect of l-NAME (NOS inhibitor) on morphine-induced antinociception in male and female rats was investigated. Forty rats were divided into four groups: (1) female (2) female-LN (3) male (4) and male-LN. The animals of groups 2 and 4 received daily injection of l-NAME (10mg/kg) during 3 weeks. The animals of control groups 1 and 3 received 2ml/kg saline instead of l-NAME. Finally, all animals were tested on the hot plate test (52±0.2°C; Cut-off 80s) to evaluate the antinociceptive effects of morphine. The hot plate test was performed for base record 15min before the injection of morphine (10mg/kg; s.c.) and consequently it was repeated every 15min after the injection. There were no significant differences in baseline latencies among all groups. Reaction times after injection of morphine in female-LN were higher than in the female control group (p<0.01). There was, however, no significant difference between male control and male-LN groups. Reaction times in the female-LN group were significantly higher than in the male-LN group. Reaction times after injection of morphine in the male group was longer than in the female group (p<0.01). It is concluded that sex hormones such as testosterone and estrogen have a role in pain perception and analgesia. NO has a modulatory effects on functions of sex hormones in pain perception and analgesic effects of opioids.

The preventive effect of N-butanol fraction of Nigella sativa on ethylene glycol-induced kidney calculi in rats.

BACKGROUND: The current study was carried out to determine whether the aqueous-ethanolic extract or the butanolic fraction of Nigella sativa (NS) seeds could prevent or reduce calculi aggregation in experimental calcium oxalate nephrolithiasis in Wistar rats. MATERIALS AND METHODS: Male Wistar rats were randomly divided into 5 groups: group A received tap drinking water for 28 days. Groups B, C, D and E received 1% ethylene glycol for induction of calcium oxalate (CaOx) calculus formation for 28 days. Rats in groups C, D and E also received aqueous-ethanolic extract of NS, N-butanol fraction and N-butanol phase remnant of NS, respectively, in drinking water at a dose of 250 mg/kg for 28 days. Urine concentration of oxalate, citrate, and calcium on days 0, 14, and 28, and also serum concentration of magnesium and calcium on days 0 and 28, were measured. On day 29, kidneys were removed for histopathologic study and examined for counting the calcium oxalate deposits in 10 microscopic fields. RESULT: Treatment of rats with N-butanol fraction and N-butanol phase remnant of NS significantly reduced the number and size of kidney calcium oxalate deposits compared with ethylene glycol group. Urinary concentration of oxalate in all experimental groups increased compared with control group on days 14 and 28, whereas the urine citrate concentration was lower in all experimental groups compared with control group on days 14 and 28. CONCLUSION: N-butanol fraction and N-butanol phase remnant of NS showed a beneficial effect on calcium oxalate deposition in the rat kidney. Therefore, the butanolic fraction of NS may be suggested for prevention of calcium oxalate calculi in humans.

Neuroprotective effects of testosterone on regenerating spinal cord motoneurons in rats.

Degeneration in the CNS and peripheral nervous system consists of degradation and phagocytosis of axons and their myelin sheath distal to the site of injury. Testosterone is a gonadal sex steroid hormone that plays an important role in CNS development. One of the lesser-known testosterone actions is neuroprotection. In the present study, the authors investigated the neuroprotectective effect of intracerebral ventricular injection of testosterone on the number of spinal motoneurons after sciatic nerve crush. In all, 32 male Wistar rats were divided to 4 groups (control, compression, compression + castration, compression + testosterone injections; n = 8). Four weeks after compression the lumber segments of spinal cord were sampled, processed, sectioned serially, and stained with toluidine blue (pH = 4.65) by using steriological quantitative technique (physical dissector), the number of alpha motoneurons in the right ventral horns of spinal cord were counted and compared between groups. Statistical analyses showed that testosterone injections (1 microl icv, 4 times, 1 week interval between injections) significantly (p < .05) reduced neuronal damage. These results indicated that testosterone has an obvious neuroprotective effect on lumbar spinal motoneurons.