Asian Pacific Journal of Tropical Biomedicine

60-69

Authors: Youssef Elouafy, Adil El Yadini, Salma Mortada, Mohamed Hnini, Hicham Harhar, Asaad Khalid, Ashraf N Abdalla, Abdelhakim Bouyahya, Khang Wen Goh, Long Chiau Ming, My El Abbes Faouzi, Mohamed Tabyaoui

Objective: To investigate the relationship between triterpenoid saponin content and antioxidant, antimicrobial, and α-glucosidase inhibitory activities of 70% ethanolic, butanolic, aqueous, supernate and precipitate extracts of Juglans regia leaves. Methods: Triterpenoid saponins of different Juglans regia leaf extracts were measured by the vanillin method. Antioxidant activity was evaluated against DPPH and ABTS free radicals. We also assessed α-glucosidase inhibitory and antimicrobial activities of the leaf extracts. Pearson's correlation coefficient was evaluated to determine the correlation between the saponin content and biological activities. Results: The butanolic extract was most effective against DPPH with an IC50 of 6.63 μg/mL, while the aqueous extract showed the highest scavenging activity against ABTS free radical with an IC50 of 42.27 μg/mL. Pearson's correlation analysis indicated a strong negative correlation (r = -0.956) between DPPH radical scavenging activity (IC50) and the saponin content in the samples examined. In addition, the aqueous extract showed the best α-glucosidase inhibitory activity compared with other extracts. All the extracts had fair antibacterial activity against Bacillus subtilis, Escherichia coli, and Klebsiella pneumoniae except for the aqueous extract. Conclusions: Juglans regia extracts show potent antioxidant, antimicrobial, and α-glucosidase inhibitory activities. There is a correlation between saponin levels in Juglans regia leaf extracts and the studied activities. However, additional research is required to establish these relationships by identifying the specific saponin molecules responsible for these activities and elucidating their mechanisms of action.

80-92

Authors: Srivarshini Sankar, Gothandam Kodiveri Muthukaliannan

Objective: To investigate the potential synergistic activity of diclofenac with piperine and D-limonene in inducing apoptosis and cell cycle arrest in breast cancer MCF-7 cells. Methods: Molecular docking study was conducted to evaluate the binding affinity of diclofenac with piperine and D-limonene against p53, Bax, and Bcl-2. The MTT assay was used to determine IC50, and the Chou-Talay method was used to determine the synergistic concentration of the combination treatment of diclofenac plus piperine and diclofenac plus D-limonene. Apoptosis detection, cell cycle arrest, reactive oxygen species production, and mitochondrial membrane potential were also investigated. Results: Diclofenac, piperine, and D-limonene showed potent binding affinity for p53, Bax, and Bcl-2. Diclofenac plus piperine and diclofenac plus D-limonene enhanced the formation of reactive oxygen species, which also had an effect on the mitochondrial membrane's integrity and caused DNA fragmentation. Diclofenac plus piperine and diclofenac plus D-limonene arrested the cells in the sub-G0 phase while drastically lowering the percentage of cells in the G2/M phase. Furthermore, the elevated apoptosis in the combined therapy was confirmed by annexin V/propidium iodide staining. Conclusions: The combined therapy prominently enhanced the anti-proliferative and apoptotic effects on MCF-7 cells compared with treatment with diclofenac, piperine, and D-limonene alone.

9-16

Authors: Asma Neisy, Farhad Koohpeyma, Majid Jafari Khorchani, Fatemeh Karimi, Fatemeh Zal

Objective: To examine the effect of quercetin on stereological parameters and autophagy-related genes in ovaries of polycystic ovary syndrome (PCOS) rats. Methods: Fifty female Sprague-Dawley rats were randomly divided into five groups: the control group, the ethanol group, the quercetin group (15 mg/kg/day), the PCOS group, as well as the PCOS + quercetin group. After the induction of PCOS, quercetin was administered orally for 30 days. Histological, stereological and real-time PCR analyses were carried out to evaluate the effect of quercetin on PCOS rats. Results: Stereological analysis revealed that quercetin significantly increased the number of ovarian follicles and the volume of corpus luteum and induced a significant decrease in atretic follicles in comparison to the PCOS group. In addition, quercetin markedly increased mTOR gene expression while decreasing Beclin-1 and LC3 gene expression. Conclusions: Quercetin strongly modulates the expression of ovarian autophagy-related genes and stereological parameters in PCOS rats. Therefore, it can be considered as an ameliorative component for ovarian follicular impairments.

26-34

Authors: Ahmad Oryan, Effat Bemani, Somayeh Bahrami

Objective: To evaluate the in vitro and in vivo efficacy of quercetin and its immunomodulatory and anti-oxidative activity against Leishmania major (L. major). Methods: L. major promastigotes and amastigotes were incubated with different concentrations of quercetin to estimate EC50. For in vivo study, the base of tails of mice was infected with L. major. After developing ulcers in the inoculation site, mice were treated with 50 mg/kg quercetin orally for 28 consecutive days. The wound-healing potential of quercetin was evaluated by histopathological analysis of tissue sections stained by hematoxylin and eosin as well as Masson's trichrome. In addition, the levels of tumor necrosis factor-α, interleukin-6, malondialdehyde, and adiponectin, the ferric reducing ability of plasma, as well as superoxide dismutase and glutathione peroxidase activities were measured. Results: The EC50 values of quercetin against L. major promastigotes and intracellular amastigotes were 0.27 and 0.85 μM, respectively. Histopathological analysis showed that fewer inflammatory cells, more fibroblasts, and more collagen deposition were observed in tissue sections of quercetin-treated mice. In addition, treatment with quercetin markedly increased glutathione peroxidase activity, the ferric reducing ability of plasma and adiponectin levels while decreasing malondialdehyde, interleukin-6, and tumor necrosis factor-α levels. Conclusions: Quercetin shows anti-leishmanial activity, immunomodulatory, anti-oxidative, and anti-inflammatory effects. Therefore, it may be further explored as an effective drug in treating leishmaniasis.

374-382

Authors: Dina F Elmaghraby, Fatma A.M. Salem, Esraa S.A. Ahmed

Objective: To explore the effect of Persea americana supplementation on inflammation, oxidative stress, and lipid profiles in ovariectomized rats fed with a high-fat diet and exposed to radiation. Methods: The control group was sham operated, while groups 2-5 were ovariectomized and fed a high-fat diet. Groups 4 and 5 were exposed to γ-radiation (1 Gy/week for 5 weeks) after ovariectomy. Groups 3 and 5 were treated with 1 mL/250 g/day of Persea americana for one month. Serum levels of estrogen, alanine aminotransferase, aspartate aminotransferase, cholesterol, triglycerides and lipoproteins were measured. Additionally, hepatic oxidative stress, inflammatory and fibrogenic markers were evaluated. Results: Persea americana treatment reduced the oxidative stress markers as well as the levels of triglyceride, total cholesterol, and low-density lipoprotein cholesterol, which in turn lowered hepatic fat accumulation. Moreover, it suppressed hepatic inflammatory mediators (interleukin-6, tumor necrosis factor-α, and C-reactive protein) and downregulated pro-fibrogenic markers (transforming growth factor-β and tissue inhibitor of metalloproteinase-1). Conclusions: Persea americana provides protection against ovariectomy, and gamma radiation-mediated hepatic inflammation not only through its antioxidant, anti-inflammatory, lipid-lowering effect but also by modulating the fibrogenic markers.

391-399

Authors: Regildo M. G. Silva, Gustavo R Martins, Laura M. B. Nucci, Filipe O Granero, Célia C. M. Figueiredo, Patrícia S Santiago, Luciana P Silva

Objective: To evaluate the photoprotective, antioxidant, antiglycation, and antiacne activities of crude extract (CESs) and triterpene saponin fraction (TSSs) of Sapindus saponaria. Methods: HPLC-MS purification was performed on a Symmetry TM C18 column. The saponins were identified by a UV detector. Antioxidant activity was evaluated by DPPH and O2- radicals scavenging, and FRAP and TBARS assays. Glycation activity was assessed by relative electrophoretic mobility and inhibition of advanced glycation end products (AGEs) formation. Additionally, antiacne activity was determined by inhibition of Cutibacterium acnes, and photoprotective effect was evaluated by Mansur’s method. Results: Most of the triterpene saponins detected in the fraction by HPLC-MS analysis were hederagenin as the aglycon. CESs and TSSs presented varying antioxidant activity in DPPH (CESs: 75.69% and TSSs: 83.65%), FRAP (CESs: 425.39 μM TE/g DW and TSSs: 649.36 μM TE/g DW), TBARS (CESs: 42.96% and TSSs: 52.16%) and O2- radicals scavenging (CESs: 61.33% and TSSs: 86.69%) tests. CESs and TSSs also exhibited antiglycation activity comparable to bovine serum albumin treated with aminoguanidine. In addition, CESs and TSSs showed inhibition of AGE formation (34.48% and 61.85%, respectively). Antiacne activity against Cutibacterium acnes was observed with a minimum inhibitory concentration equal to minimum bactericidal concentration (CESs: 36.11 μg/mL and TSSs: 18.34 μg/mL). In photoprotective assays, CESs and TSSs showed maximum absorbance of 1.42 to 0.20 and 2.80 to 1.30, respectively, in the wavelength range of 260 to 400 nm. Furthermore, CESs and TSSs showed sun protection factors of 8.89 and 14.89, respectively. Conclusions: Sapindus saponaria fruit extracts show strong antioxidant potential and antiglycation activity against bovine serum albumin glycation and AGE formation. Besides, they presented antibacterial activity against Cutibacterium acnes and photoprotective effect against UV-A and UV-B.

333-342

Authors: Qi Li, Bo Wang, Kai-Wen Lin, Tang Deng, Qi-Feng Huang, Shuang-Qin Xu, Hang-Fei Wang, Xin- Xin Wu, Nan Li, Yang Yi, Ji-Chao Peng, Yue Huang, Jin Qian, Xiao-Ran Liu

Objective: To explore the protective effects of anthrahydroquinone- 2,6-disulfonate (AH2QDS) on the kidneys of paraquat (PQ) poisoned rats via the apelin-APJ pathway. Methods: Male Sprague Dawley rats were divided into four experimental groups: control, PQ, PQ+sivelestat, and PQ+AH2QDS. The PQ+sivelestat group served as the positive control group. The model of poisoning was established via intragastric treatment with a 20% PQ pesticide solution at 200 mg/kg. Two hours after poisoning, the PQ+sivelestat group was treated with sivelestat, while the PQ+AH2QDS group was given AH2QDS. Six rats were selected from each group on the first, third, and seventh days after poisoning and dissected after anesthesia. The PQ content of the kidneys was measured using the sodium disulfite method. Hematoxylin-eosin staining of renal tissues was performed to detect pathological changes. Apelin expression in the renal tissues was detected using immunofluorescence. Western blotting was used to detect the expression levels of the following proteins in the kidney tissues: IL- 6, TNF-α, apelin-APJ (the apelin-angiotensin receptor), NF-κB p65, caspase-1, caspase-8, glucose-regulated protein 78 (GRP78), and the C/EBP homologous protein (CHOP). In in vitro study, a PQ toxicity model was established using human tubular epithelial cells treated with standard PQ. Twenty-four hours after poisoning, sivelestat and AH2QDS were administered. The levels of oxidative stress in human renal tubular epithelial cells were assessed using a reactive oxygen species fluorescence probe. Results: The PQ content in the kidney tissues of the PQ group was higher than that of the PQ+AH2QDS group. Hematoxylin-eosin staining showed extensive hemorrhage and congestion in the renal parenchyma of the PQ group. Vacuolar degeneration of the renal tubule epithelial cells, deposition of crescent-like red staining material in renal follicles, infiltration by a few inflammatory cells, and a small number of cast formation were also observed. However, these pathological changes were less severe in the PQ+sivelestat group and the PQ+AH2QDS group (P<0.05). On the third day after poisoning, immunofluorescence assay showed that the level of apelin in the renal tissues was significantly higher in the PQ+AH2QDS group than in the PQ group. Western blotting analysis results showed that IL-6, TNF-α, NF-κB p65, caspase-1, caspase-8, GRP78, and CHOP protein levels in the PQ group were higher than in the PQ+AH2QDS group (P<0.05). The expression of apelin-APJ proteins in the PQ+AH2QDS group was higher than in the PQ+sivelestat and PQ groups (P<0.05); this difference was significant on Day 3 and Day 7. The level of oxidative stress in the renal tubular epithelial cells of the PQ+AH2QDS group and the PQ+sivelestat group was significantly lower than in the PQ group (P<0.05). Conclusions: This study confirms that AH2QDS has a protective effect on PQ-poisoned kidneys and its positive effect is superior to that of sivelestat. The mechanism of the protective effects of AH2QDS may be linked to reduction in cellular oxidative stress, PQ content of renal tissue, inflammatory injury, endoplasmic reticulum stress, and apoptosis. AH2QDS may play a role in the treatment of PQ poisoning by upregulating the expression of the apelin-APJ.

351-356

Authors: Masud Eneji Sadiq, Chibuzo Egwuenu, Rabiu Saidu Umar Wasagu, Usman Zayyanu Umar, Bello Usman

Objective: To investigate the role of Mirabilis jalapa root extracts in restoration of glucose homeostasis in alloxan-induced hyperglycemic Wistar albino rats. Methods: Experimental hyperglycemic rats were treated daily with 200 and 400 mg/kg of Mirabilis jalapa extracts after initial fasting for 6 h. Two-hour postprandial glucose and changes in body weight were monitored during treatment. After 14 d, the rats were sacrificed and blood was collected for biochemical assessment of serum glucose and insulin levels, lipid profile, and oxidative stress markers. Histopathological examinations of harvested pancreas were also carried out. Results: Mirabilis jalapa root extracts at 200 and 400 mg/kg increased the body weight of hyperglycemic rats. Postprandial glucose levels of the extract-treated hyperglycemic groups progressively declined during treatment compared with the untreated hyperglycemic control group (P<0.05). The lipid profile indices of the untreated negative control group were significantly elevated (P<0.05), which were reversed by treatment with Mirabilis jalapa extracts. The remarkable increases in antioxidant enzyme activities and a significant decrease in malondialdehyde levels were observed in the hyperglycemic group treated with Mirabilis jalapa extracts. Mirabilis jalapa extracts also significantly increased serum insulin levels (P<0.05). In addition, histopathological examinations of the pancreas revealed a significant cell population within the islet nests of the extract-treated hyperglycemic groups. Conclusions: Mirabilis jalapa extract can restore glucose homeostasis and show hypoglycemic and hypolipidemic effects in hyperglycemic rats. Further studies are needed to verify the active components of the plant and the underlying mechanism of action in the future.

290-299

Authors: Ruttiya Thongrung, Laddawan Senggunprai, Wiphawi Hipkaeo, Panot Tangsucharit, Patchareewan Pannangpetch

Objective: To investigate the effect of Moringa oleifera leaf extract on angiogenesis and inflammatory process in a rat model of streptozotocin-induced diabetic nephropathy. Methods: Four weeks after a single injection of 50 mg/kg streptozotocin, rats were treated with 100 or 200 mg/kg/day Moringa oleifera leaf extract, 1 mg/kg/day dapagliflozin, or a combination of Moringa oleifera leaf extract and dapagliflozin for further eight weeks. Renal function, kidney histology, and gene expression were evaluated at the end of the experiment. Results: Renal function of diabetic rats was significantly impaired as evidenced by increased blood urea nitrogen, albuminuria, 24-h proteinuria, and high creatinine clearance which indicated glomerular hyperfiltration. In addition, diabetic rats showed an increase in gene expressions of vascular endothelial growth factor-A (VEGF-A), angiopoietin-2 (Ang2), the Ang2/Ang1 ratio, tumor necrosis factor-α, interleukin-1β and monocyte chemoattractant protein-1. Immunohistochemical staining demonstrated a significant increase in the density of glycoprotein CD34. Moringa oleifera leaf extract markedly improved all renal dysfunction markers and modulated the upregulated expression of angiogenic factors and inflammatory genes. Conclusions: Moringa oleifera leaf extract could suppress abnormal angiogenesis and inflammatory processes possibly by downregulating gene expression of angiogenesis factors and proinflammatory cytokines.

312-322

Authors: Md Jamir Anwar, Sattam Khulaif Alenezi, Faizul Azam, Danish Mahmood, Faisal Imam, Khalid Saad Alharbi

Objective: To investigate the effect of Nigella sativa oil on cardiomyopathy and neurobehavioral changes induced by doxorubicin in mice. Methods: Swiss strain of albino female mice were divided into 6 groups of 5 animals in each: GroupⅠ(control group), group Ⅱ (doxorubicin, 10 mg/kg, i.v.), group Ⅲ, Ⅳ, and Ⅴ (Nigella sativa oil; 1.5, 3, and 6 mL/kg, respectively), group Ⅵ (Nigella sativa oil per se; 6 mL/kg, p.o.). The duration of treatment was 15 d (10 days’ pre-treatment and 5 days’ post-treatment) and doxorubicin was administered on day 11th of the treatment schedule. Following Nigella sativa oil treatment, neurobehavioral tests, cardiac hypertrophy tests, and biochemical tests in serum and tissues were performed. Neurological tests included assessment of anxiety-like behavior in the elevated plus maze, spontaneous alternation behavior in the cross maze, and depression-like behavior in modified forced swim tests. Biochemical tests included serum lactate dehydrogenase and creatinine kinase-MB, malondialdehyde and reduced glutathione in tissues. Lastly, molecular docking was used to estimate the affinity of the phytoconstituents of Nigella sativa oil with histone deacetylases. Results: Nigella sativa oil treatment significantly (P<0.001) restored doxorubicin-induced neurobehavioral changes, decreased lactate dehydrogenase and creatinine kinase-MB in the plasma, malondialdehyde contents in tissues, and increased reduced glutathione level. Besides, no significant alteration was observed in Nigella sativa oil per se group as compared to the control. Molecular docking showed that Nigella sativa oil components had appreciable binding affinitiy with the protein cavities of HDAC1 and HDAC6. Conclusions: The result shows that Nigella sativa oil exerts anxiolytic, antidepressant, and memory-enhancing effects in addition to cardioprotective effect against doxorubicin-induced cardiomyopathy in mice. The modulatory effect of Nigella sativa oil on oxidative stress could contribute to the cardioprotective effect and associated neurobehavioral changes in mice.