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.

507-520

Authors: Daude, R. B.; Bhadane, Rajendra; Shah, J. S.

Objective: To investigate the renoprotective effects of luteolin on diabetes in rats. Methods: One week after administration of streptozotocin 55 mg/kg intraperitoneally, rats were given 25, 50, and 75 mg/kg/day of luteolin orally for another eight weeks. At the end of the experiment, body weight, blood glucose level, biochemical parameters for renal function (serum creatinine, blood urea nitrogen, uric acid, serum albumin, and total protein), kidney histology, matrix metalloproteinase (MMP)-2, MMP-9, and histone deacetylase 2 (HDAC-2) expression, and malondialdehyde, myeloperoxidase, and hydroxyproline content in renal tissue were evaluated. High glucose- induced damage using NRK-52E cell line was studied to evaluate cell viability and metalloenzyme expression. Additionally, in silico studies including docking and molecular dynamics simulations were conducted. Results: MMP-2, MMP-9, and HDAC-2 expressions were significantly increased in high glucose-induced NRK-52E cells and the renal tissue of diabetic rats. However, these changes were reversed by luteolin at the administered doses. Additionally, luteolin significantly reduced oxidative stress, inflammation, and fibrosis, as well as improved biochemical parameters in diabetic rats. Furthermore, luteolin at the examined doses markedly alleviated diabetes-induced histopathological changes in renal tissues. Conclusions: Luteolin effectively attenuates streptozotocin- induced diabetic nephropathy in rats by inhibiting MMP-2, MMP- 9, and HDAC-2 expression, and reducing oxidative stress and inflammation.

532-538

Authors: Liu, Yu; Wang, Chao-Qun; Zhu, Yong-Kang; Xu, Jia-Fang; Yin, Si-Qi; Hu, Qing-Jie; Yang, Rui-Qi

Objective: To investigate the role of macrophages in regulating breast cancer cell migration and its related mechanisms. Methods: Human leukemia monocytic cell line THP-1-secreted exosomes were isolated using multi-step ultracentrifugation and verified using nanoparticle tracking analysis. Differentially expressed miRNAs were identified using RNA sequencing. Overexpression of inhibitors of hsa-miR-101-3p in breast cancer MDA-MB-231 cells was performed by infecting their lentiviral constructs. The luciferase reporter assay was used to evaluate the interaction of DLG5 and miR-101. DGL5 expression was detected using qRT-PCR and Western blot analyses. Results: The migration of breast cancer cells was significantly inhibited after addition of exosomes. RNA sequencing results showed that miR-101-3p expression was significantly upregulated. Targetscan analysis predicted that miR-101-3p could target DLG5, and this prediction was verified using the luciferase assay. The addition of the miR-101-3p precursor significantly increased the expression of miR-101-3p, and the mRNA and protein levels of DLG5 were suppressed. In contrast, inhibiting the expression of miR-101-3p increased the mRNA and protein levels of DLG5. Furthermore, the scratch assay showed that inhibiting miR-101-3p could promote the migration of MDA-MB-231 cells. Conclusions: Macrophage exosomes can inhibit the migration of breast cancer cells, and increasing the expression of miR-101-3p to inhibit DLG5 expression may play an important role in this process, which needs further investigation.

460-468

Authors: Sethumathi, P. P.; Uddandrao, V. V. Sathibabu; Chandrasekaran, P.; Sengottuvelu, S.; Tamilmani, P.; Ponmurugan, P.; Vadivukkarasi, S.; Santhanakumar, M.; Begum, M. Shabana; Saravanan, G

Objective: To determine how biochanin-A (BCA) affects high-fat diet and streptozotocin-induced pathological changes in lipid metabolism and antioxidant status in diabetic rats. Methods: Diabetic rats were orally administered BCA (10 mg/kg body weight) for 30 days to investigate its effects on lipid profiles and oxidative stress markers in the liver and kidney. In addition, the mRNA expression of antioxidant and lipid metabolism enzymes in the liver was examined. Results: BCA attenuated hyperlipidemia by regulating mRNA expressions of HMG-CoA reductase, fatty acid synthase, carnitine palmitoyl transferase, and acetyl-CoA carboxylase. Additionally, BCA reduced high-fat diet and streptozotocin-induced oxidative stress by suppressing lipid peroxidation, improving superoxide dismutase, catalase, and glutathione peroxidase levels, and upregulating mRNA expressions of these enzymes. Conclusions: BCA may be a promising nutraceutical for the treatment of dyslipidemia and oxidative stress associated with diabetes.

479-487

Authors: Pattnaik, Soumya Sucharita; Sarangi, Sudhir Chandra; Sharma, Deeksha; Sinha, Surabhi; Nag, Tapas C; Nambirajan, Aruna; Tripathi, Manjari

Objective: To investigate the effect of Ocimum sanctum hydroalcoholic extract (OSHE) on seizure control and neuronal injury in rats with lithium-pilocarpine-induced status epilepticus (SE). Methods: SE was induced by administering lithium chloride followed by pilocarpine 24 h later. OSHE was administered either alone or in combination with valproate (VPA) 3 days before SE induction until 14 days post-SE induction. Seizure parameters were recorded on day 1 (0-3 h), day 1-3 and day 4-14 post-SE. On day 14 post-SE, neurobehavioural tests (elevated plus maze and passive avoidance) were done followed by total antioxidant capacity, neuron-specific enolase, immunohistochemistry, and electron microscopic assessment in the hippocampus and cortex tissue. Results: OSHE+VPA provided more significant seizure protection (75%) than VPA (62.5%), OSHE (62.5%), or SE control (12.5%) (overall P=0.003). The latency to stage-3/4 seizures was increased and the number of stage-3/4 seizures was reduced in all treatment groups compared to the SE control group (P=0.002 and <0.001, respectively). The OSHE+VPA group also had better memory retention than other treatment groups (P<0.001) in the passive avoidance test. Total antioxidant capacity level was significantly higher and neuron-specific enolase was lower in the OSHE and OSHE+VPA groups compared to the SE control group. Electron microscopic study showed significant myelin sheath damage (67.5%, P<0.05) and axonal degeneration (51.8%, P<0.001) in the hippocampus of the SE control group, which were alleviated by OSHE or OSHE+VPA treatment. In immunohistochemical analysis, the OSHE, OSHE+VPA, and VPA groups had a significantly higher number of viable neurons and less neuronal loss compared to the SE control in the hippocampus (P<0.001). Conclusions: OSHE either alone or in combination with VPA shows better seizure control by preservation of neuronal echotexture and reducing oxidative stress in the hippocampus.

421-430

Authors: Ravindran, Rajeswari; Manuel, Malathi; Mohan, Thangarajeswari; Jaganathan, Ravindran; Periandavan, Kalaiselvi

Objective: To explore the combinatorial effect of epigallocatechin-3-gallate (EGCG) and calorie restriction on activation of nuclear factor erythroid 2-related factor 2 (Nrf2), a transcription factor involved in the antioxidant defense system of aged rats. Methods: Aged male Wistar rats were calorie-restricted and treated with EGCG orally for 45 days. The initial body weight of aged rats was recorded, and the final body weight was measured at the end of the experimental period. Serum lipid and lipoprotein status, oxidative stress markers such as free radicals and malondialdehyde levels, and reduced glutathione were assessed. In addition, RT-PCR and Western blotting analyses were performed. Results: Calorie restriction potentiated the effect of EGCG on enhancing antioxidant status, improving the levels of serum lipid and lipoproteins, upregulating Nrf2 and Bcl2, and downregulating Keap1, cullin3, Bax and cytochrome c in aged rats. Conclusions: Calorie restriction can promote EGCG-mediated Nrf2 activation in aged rats. This preliminary finding paves the way for a combinatory approach to replenishing the antioxidant status during aging, thereby reducing the risk for age-associated degenerative diseases.

443-450

Authors: Eldourghamy, Ayman; Hossam, Toka; Hussein, Mohammed Abdalla; Abdel-Aziz, Amal; El-masry, Samir A

Objective: To investigate the cardioprotective effect of naringenin against isoproterenol (ISO)-induced cardiotoxicity in rats. Methods: Rats were divided into five groups: the normal group, the ISO group (85 mg/kg b.w.); the ISO+naringenin (50 mg/kg b.w.) group, the ISO+naringenin (100 mg/kg b.w.) group and the ISO+propranolol (10 mg/kg b.w.) group. Plasma creatine kinase-MB (CK-MB), cardiac troponin T, lactate dehydrogenase, brain natriuretic peptide (BNP), and IL-10, as well as cardiac transforming growth factor-β1 (TGF-β1), vascular endothelial growth factor (VEGF) and malondialdehyde (MDA) were examined. In addition, NLRP3 and mRNA-208a expressions were evaluated by RT-PCR analysis. Histopathological examination was also performed to assess cardiac damages. Results: Naringenin treatment significantly decreased plasma lactate dehydrogenase, CK-MB, cardiac troponin T, BNP, and IL-10, as well as cardiac TGF-β1, VEGF, and MDA while increasing p-Akt and superoxide dismutase in ISO-administered rats. It also reduced NLRP3 and mRNA-208a gene expression levels. Furthermore, naringenin improved ISO-induced cardiac damage. Conclusions: Naringenin attenuates myocardial dysfunction in ISO-treated rats by decreasing oxidative stress and increasing cardiac endogenous antioxidant system, which may be modulated partly by improvement of NLRP3 and mRNA-208a gene expression.

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.