Asian Pacific Journal of Tropical Biomedicine

231-243

Authors: Arnica F Lal, Shaminder Singh, Francisco C Franco, Sonam Bhatia

Polyphenols are the secondary metabolic products of plants and are considered as active constituents to possess therapeutic effects. To date, a vast number of scientific literature addressed the potential of polyphenols as bio-efficient compounds owing to their structural diversity. Due to the presence of several hydroxyl groups, they are metabolized quickly due to conjugation reaction and thus, readily produce toxic metabolites as a defense material against many pathogens, reflecting their safety strategy. This review focuses on the anti-quorum sensing and biofilm inhibition activity of polyphenols, which display their potential to treat bacterial infections by combating the virulence caused by pathogenic agents. Thus, for mitigating quorum sensing-controlled pathogenesis, the use of polyphenol-based phytochemicals holds immense potential to cure infections. The application of polyphenol as sensitizing agent/ adjuvant therapeutics which act in synergism with antibiotics is highly remarkable.

254-262

Authors: Raghda A El-Sayed, Fatma M El-Demerdash, Mohammed A El-Magd

Objective: To investigate the protective and therapeutic role of ginseng against silicon dioxide nanoparticles (SiO2NPs)-induced toxicity in the lungs. Methods: Sixty male rats were divided into five groups (n = 12/ group); group 1 was used as a control, group 2 received ginseng, group 3 was treated with SiO2NPs, and group 4 was pretreated with ginseng one week before SiO2NPs, while group 5 was given SiO2NPs one week before supplementation with ginseng. Animals were treated with both ginseng and SiO2NPs orally for five weeks. Real-time PCR was used to measure gene expression. Besides, DNA damage and cell cycle changes were determined by comet assay and flow cytometry, respectively. Histological study was also done to assess the effect of ginseng on SiO2NPs-induced toxicity. Results: SiO2NPs increased lipid peroxidation and decreased the activities of antioxidant enzymes. SiO2NPs induced apoptosis in lung tissues as revealed by upregulation of Bax and caspase 3 and downregulation of Bcl-2 as well as the induction of DNA damage. SiO2NPs also caused inflammation as indicated by upregulation of the inflammation-related genes [interleukin 1 beta (IL-1β), tumor necrosis factor-alpha (TNF-α), nuclear factor kappa B (NF-κB), cyclooxygenase 2 (COX2), and transforming growth factor-beta 1 (TGFβ1)] as well as cell cycle arrest in the G0/G1 phase of lung cells. Moreover, histopathological examination proved the biochemical and molecular perturbations that occurred due to SiO2NPs toxicity. However, ginseng alleviated SiO2NPs-induced toxicity in rat lung. Conclusions: Ginseng has a potent preventive and therapeutic effect and could be used in the treatment of SiO2NPs-induced pulmonary toxicity.

273-284

Authors: Li-Shan Yan, Li Wang, Brian Chi-Yan Cheng, Yu Ding, Jing Kong, Qing Gao Wang, Xiu-Qiong Fu, Shuo-Feng Zhang, Gan Luo, Yi Zhang

Objective: To investigate the anti-inflammatory effects of the total flavonoids from Saussurea involucrata on lipopolysaccharides (LPS)-stimulated murine RAW264.7 macrophages and explore its underlying mechanism of action. Methods: Total flavonoids from Saussurea involucrata were extracted using chromatographic column method. Cell viability was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay. The production of nitric oxide was detected by Griess assay and the release of cytokines (IL-10 and TNF-α) and chemokines (MCP-1, MIP-1a, and CCL5/RANTES) was determined by ELISA to evaluate the anti-inflammatory activity of total flavonoids from Saussurea involucrata. Moreover, nuclear translocation of p65, c-Jun, and IRF3 was detected by immunofluorescence microscopy and Western blotting analysis was performed to determine the expression of related proteins. Results: Total flavonoids extracted from Saussurea involucrata were 751.5 mg/g and the content of rutin was 506.5 mg/g. The production of inflammatory mediators including nitric oxide, cytokines, and chemokines was effectively inhibited by total flavonoids from Saussurea involucrata. Meanwhile, total flavonoids also suppressed the nuclear translocation of p65, c-Jun, and IRF3 in LPS-stimulated RAW264.7 cells. The LPS-induced expression of iNOS and COX-2 was remarkably reduced by treatment with total flavonoids from Saussurea involucrata. Moreover, total flavonoids decreased the expression levels of p-IKKa/β, p-TBK1, p-p38, p-ERK, p-JNK, p-p65, p-c-Jun, and p-IRF3 in LPS-exposed RAW264.7 macrophages. Conclusions: Total flavonoids from Saussurea involucrata potentially inhibit the secretion of pro-inflammatory mediators, which may be related to inhibition of p65, c-Jun, and IRF3 signaling pathways in LPS-stimulated RAW264.7 cells.

183-193

Authors: Ketmanee Senaphan, Upa Kukongviriyapan, Pisit Suwannachot, Geerasak Thiratanaboon, Weerapon Sangartit, Supawan Thawornchinsombut, Akkasit Jongjareonrak

Objective: To examine the ameliorative effect of rice bran hydrolysates (RBH) on metabolic disorders, cardiac oxidative stress, heart rate variability (HRV), and cardiac structural changes in high fat and high fructose (HFHF)-fed rats. Methods: Male Sprague-Dawley rats were daily fed either standard chow diet with tap water or an HFHF diet with 10% fructose in drinking water over 16 weeks. RBH (500 and 1 000 mg/kg/day) was orally administered to the HFHF-diet-fed rats during the last 6 weeks of the study period. At the end of the treatment, metabolic parameters, oxidative stress, HRV, and cardiac structural changes were examined. Results: RBH administration significantly ameliorated metabolic disorders by improving lipid profiles, insulin sensitivity, and hemodynamic parameters. Moreover, RBH restored HRV, as evidenced by decreasing the ratio of low-frequency to high-frequency power of HRV, a marker of autonomic imbalance. Cardiac oxidative stress was also mitigated after RBH supplementation by decreasing cardiac malondialdehyde and protein carbonyl, upregulating eNOS expression, and increasing catalase activity in the heart. Furthermore, RBH mitigated cardiac structural changes by reducing cardiac hypertrophy and myocardial fibrosis in HFHF-diet-fed rats. Conclusions: The present findings suggest that consumption of RBH may exert cardioprotective effects against autonomic imbalances, cardiac oxidative stress, and structural changes in metabolic syndrome.

205-213

Authors: Aarti S Kale, Avinash R Wadkar, Umesh B Mahajan, Lalit A Birari, Sateesh Belemkar, Sameer N Goyal, Shreesh Ojha, Sanjay J Surana, Chandragouda R Patil, Kalpesh R Patil

Objective: To investigate the effect of aloin against chronic constriction injury (CCI)-induced neuropathic pain in rats. Methods: Rats were randomly divided into 7 groups: Group I (normal control), Group II (sham-operated), Group III (CCI control) and Group IV, V, VI, and VII, which underwent CCI surgery and then were administered with aloin (5 mg/kg, p.o.; 25 mg/kg, p.o.; 125 mg/kg, p.o.) and gabapentin (50 mg/kg, p.o.), respectively for 14 days. Peripheral neuropathy was induced by silk ligatures (4-0) loosely placed around the sciatic nerve. Nociceptive thresholds against mechanical stimuli (Von-Frey filaments) and thermal stimuli (12 °C and 40 °C) were measured at mid-plantar paw region ipsilateral to the compressed nerve on day-3, 7, 11, and 14. The concentration of cytokines including tumor necrosis factor-α (TNF-α), interleukin-6, and interleukin-1β was estimated at day-7. At day 14, motor nerve conduction velocity was determined under urethane anesthesia (1.25 g/kg). Oxidative stress parameters (malondiadehyde, glutathione, catalase, and superoxide dismutase) were estimated in sciatic nerve homogenates at day 14. Representative nerve samples were processed for histological investigations. Results: Aloin significantly reduced CCI-induced mechanical and thermal allodynia. It also improved motor nerve conduction velocity and decreased oxidative stress in nerve tissues. In addition, it decreased pro-inflammatory cytokine levels and restored the histoarchitecture of compressed sciatic nerve. Conclusions: Aloin mitigates CCI-induced neuropathic pain in rats by inhibiting oxidative stress and pro-inflammatory cytokines in the afflicted sciatic nerve.

222-230

Authors: Yung Hyun Choi

Objective: To investigate the effect of honokiol on oxidative damage in HaCaT human keratinocytes. Methods: HaCaT cells were exposed to hydrogen peroxide (H2O2), following pretreatment with various concentrations of honokiol. The alleviating effects of honokiol on HaCaT cell viability and cell death, reactive oxygen species (ROS) production, DNA damage, mitochondrial dynamics, and inhibition of adenosine triphoaphate production against H2O2 were investigated. Western blotting analysis was used to analyze the expression levels of specific proteins. Results: Honokiol suppressed H2O2-induced cytotoxicity and DNA damage by blocking abnormal ROS accumulation. Honokiol also prevented apoptosis by inhibiting loss of mitochondrial membrane potential and release of cytochrome c from the mitochondria into the cytosol, decreasing the Bax/Bcl-2 ratio, and reducing the activity of caspase-3 in H2O2-stimulated HaCaT cells. In addition, honokiol attenuated H2O2-induced reduction of adenosine triphosphate content, and activation of AMP-activated protein kinase (AMPK) was markedly promoted by honokiol in H2O2-stimulated cells. Importantly, the anti-apoptosis and anti-proliferative activity of honokiol against H2O2 was further enhanced by adding an activator of AMPK, indicating that honokiol activated AMPK in HaCaT keratinocytes to protect against oxidative damage. Conclusions: The present results indicate that honokiol may be useful as a potential therapeutic agent against various oxidative stress-related skin diseases.

141-147

Authors: Clarice C.V. Moura, Rafaela S dos Santos, Lucienir P Duarte, Giovane Galdino

Objective: To investigate the antinociceptive effect of tingenone on inflammatory pain, as well as and the involvement of the cannabinoid receptors type 2 (CB2) and spinal microglia in this process. Methods: Male Swiss mice were subjected to inflammatory pain induced by intraplantar injection of carrageenan. The nociceptive threshold was measured by von Frey filaments test. Tingenone was administered orally 60 min before carrageenan injection. To evaluate the involvement of CB2 receptor, endocannabinoids, and microglia, AM630 (a CB2 receptor antagonist), MAFP (an inhibitor of an enzyme that hydrolyses endocannabinoids), and minocycline (a microglial inhibitor) were given intrathecally 20 min before tingenone administration. In addition, an immunofluorescence assay was used to evaluate CB2 receptor and CD11B (a microglial marker) expression in the spinal cord dorsal horn. Results: Tingenone significantly reduced carrageenan-induced hyperalgesia, which was reversed by pretreatment with AM630. MAFP and minocycline potentiated and prolonged the tingenone- induced antinociception. CD11B expression was increased in the spinal cord dorsal horn of mice with inflammatory pain pretreated with tingenone, which was reduced by AM630, MAFP, and minocycline. Conclusions: CB2 receptors and endocannabinoids participate in the tingenone-induced antinociception which may involve the inhibition of microglia at spinal level.

155-163

Authors: Amaresh Mishra, Yamini Pathak, Anuj Kumar, Surabhi Kirti Mishra, Vishwas Tripathi

Objective: To explore natural compounds as potential inhibitors against main protease (Mpro) of SARS-CoV-2. Methods: In the current study, systematic molecular docking analysis was conducted using AutoDock 4.2 to determine the binding affinities and interactions between natural compounds and Mpro. Selected natural compounds were further validated using a combination of molecular dynamic (MD) simulations and molecular mechanic Poisson-Boltzmann surface area (MM/PBSA) free energy calculations. Results: Out of twenty natural compounds, four natural metabolites namely, amentoflavone, guggulsterone, puerarin, and piperine were found to have strong interaction with Mpro of SARS-CoV-2 based on docking analysis. During MD simulations, all four natural compounds bound to Mpro at 50 ns and MM/G/P/BSA free energy calculations showed that all four shortlisted ligands had stable and favorable energies with strong binding to Mpro protein. Conclusions: Guggulsterone is a potential inhibitor of COVID- 19 main protease Mpro. Further in vitro and pre-clinical studies are needed.

174-182

Authors: Ummi Maryam Zulfin, Ave Rahman, Mila Hanifa, Rohmad Yudi Utomo, Sari Haryanti, Edy Meiyanto

Objective: To determine the effect of rice bran extract (RBE) in combination with doxorubicin on 4T1 triple-negative breast cancer cells and NIH-3T3 cells. Methods: RBE was obtained by maceration with n-hexane. The phytochemical profile of RBE was observed using highperformance liquid chromatography. Cytotoxic effect of RBE was evaluated through MTT assay. In addition, flow cytometry was used for cell cycle and apoptosis analysis. Cellular senescence was observed using SA-β-Gal assay and intracellular reactive oxygen species (ROS) levels were evaluated using DCFDA staining. The pro-oxidant property of RBE was also evaluated through 1-chloro- 2,4-dinitrobenzene spectrophotometry and molecular docking. Results: RBE was obtained with a yield of 18.42% w/w and contained tocotrienols as the major compound. RBE exerted no cytotoxic effect on 4T1 and NIH-3T3 cells. However, RBE in combination with doxorubicin decreased 4T1 cell viability synergistically (combination index<0.9) and induced apoptosis and senescence on 4T1 cells. RBE significantly decreased senescence in doxorubicin-treated NIH-3T3 cells. Additionally, RBE did not increase ROS levels in doxorubicin-treated 4T1 cells. Meanwhile, the combination of RBE and doxorubicin reduced ROS levels in NIH-3T3 cells. RBE significantly reduced glutathione-S-transferase activity and alpha-tocotrienol interacted with glutathione-S- transferase in the glutathione binding site. Conclusions: Rice bran may be used as a co-chemotherapeutic agent to improve the therapeutic effectiveness of doxorubicin while protecting against the cellular senescence effects of doxorubicin on healthy cells.

97-104

Authors: Jarinyaporn Naowaboot, Urarat Nanna, Linda Chularojmontri, Pholawat Tingpej, Patchareewan Pannangpetch

Objective: To examine the effect of water extract of Thunbergia laurifolia on hepatic insulin resistance in high-fat diet-induced obese mice. Methods: High-fat diet with 45 kcal% lard fat was used for obesity induction in ICR mice. The mice were fed with high-fat diet for 16 weeks, and during the last 8 weeks, they were treated with 200 mg/ kg/day of water extracts from Thunbergia laurifolia leaf, stem and flower. Serum biochemistry, liver histology, and protein expression were examined after the treatment. Results: Extracts from all of the three parts of Thunbergia laurifolia significantly alleviated hyperglycemia, hyperlipidemia, hyperinsulinemia, and hyperleptinemia. The stem and flower extracts improved glucose tolerance. All of the extracts significantly reduced serum TNFα and monocyte chemoattractant protein-1 levels. Liver weight, triglyceride levels, and lipid accumulation were also decreased. Moreover, hepatic glucose-6-phosphatase level was significantly decreased, while the levels of PPARα, phosphorylated AMPK, and phosphorylated Akt were significantly increased with treatment of Thunbergia laurifolia extracts. Conclusions: Thunbergia laurifolia extracts can ameliorate hepatic insulin resistance in high-fat diet-induced obese mice by improving glucose and lipid homeostasis, which may be associated with stimulating phosphorylation of AMPK and Akt pathways.

115-121

Authors: Zeinab Rafiee, Maasoumeh Zare Moaiedi, Armita Valizadeh Gorji, Esrafil Mansouri

Objective: To evaluate the effect of p-coumaric acid against adriamycin-induced hepatotoxicity in rats. Methods: The rats were divided into 4 groups. The control group received solvent; the p-coumaric acid group was treated with 100 mg/kg of p-coumaric acid orally for five consecutive days; the adriamycin group was administered with a single dose of adriamycin (15 mg/kg, i.p.), and the p-coumaric acid + adriamycin group was given p-coumaric acid five days before adriamycin administration. Twenty-four hours after the last administration, blood samples were collected for biochemical analysis, and liver tissues were removed for histopathological and immunohistochemistrical studies. Moreover, the levels of tissue lipid peroxidation and enzyme activities of glutathione peroxidase, superoxide dismutase, and catalase in liver tissue were measured. Results: Treatment with p-coumaric acid protected the liver from the toxicity of adriamycin by attenuating the increase in alkaline phosphatase, alanine transaminase, aspartate transaminase, total bilirubin, total cholesterol, triglyceride, and low-density lipoprotein cholesterol and lessening the decrease in high-density lipoprotein cholesterol and albumin. p-Coumaric acid also raised the levels of glutathione peroxidase, superoxide dismutase, and catalase, as well as decreased lipid peroxidation in liver tissue and hepatic IL- 1β expression. Additionally, histopathological study confirmed the protective effect of p-coumaric acid against liver damage. Conclusions: p-Coumaric acid can alleviate adriamycin-induced hepatotoxicity.

132-140

Authors: Koloko Brice Landry, Shafiq Azam, Sidra Rehman, Somayya Tariq, Bushra Iqbal, Mateen Abbas, Dieudonné Massoma Lembè, Bushra Ijaz

Objective: To evaluate the antiviral activity and phytochemicals of selected plant extracts and their effect on the mitogen-activated protein kinase (MAPK) signaling pathway modulated by hepatitis C virus (HCV) nonstructural protein 5A (NS5A). Methods: A total of ten plant extracts were initially screened for their toxicities against HepG2 cells. The non-toxic plants were tested for their inhibitory effect on the expression of HCV NS5A at both mRNA and protein levels using real-time PCR and Western blotting assays, respectively. The differential expression of the genes associated with MAPK pathway in the presence of NS5A gene and plant extract was measured through real-time PCR. Subsequently, the identification of secondary metabolites was carried out by phytochemical and HPLC analysis. Results: The phytochemical profiling of Berberis lyceum revealed the presence of alkaloids, phenols, saponins, tannins, flavonoids, carbohydrates, terpenoids, steroids, and glycosides. Similarly, quercetin, myricetin, gallic acid, caffeic acid, and ferulic acid were identified through HPLC analysis. The methanolic extract of Berberis lyceum strongly inhibited HCV RNA replication with an IC50 of 11.44 µg/mL. RT-PCR and Western blotting assays showed that the extract reduced the expression of HCV NS5A in a dose- dependent manner. Berberis lyceum extract also attenuated NS5A- induced dysregulation of the MAPK signaling pathway. Conclusions: Our findings suggest that Berberis lyceum extract strongly inhibits HCV propagation by reducing HCV NS5A- induced perturbation of MAPK signaling.

47-58

Authors: Mohsen Akbaribazm, Nader Goodarzi, Mohsen Rahimi, Leila Naseri, Mozafar Khazaei

Objective: To evaluate the effect of Heracleum persicum L. against gentamicin-induced nephrotoxicity in rats. Methods: Thirty-six Wistar rats were divided into 6 groups including control (normal saline), gentamicin (80 mg/kg/d for 10 d), Heracleum persicum (750 mg/kg/d), and gentamicin (10 d) + Heracleum persicum extract at three different doses (250, 500, and 750 mg/kg/d for 40 d). Urine creatinine, urea, protein, and albumin levels were determined. In addition, serum urea, creatinine, sodium, potassium, cytokines (TNF-α, IL-1β, IL-6, and IL-10), glutathione peroxidase activity, total antioxidant capacity, kidney malondialdehyde, stereological parameters, and expressions of apoptosis-related genes (p53, Bax, Bcl-2, and caspase-3) were measured. The LD50 of Heracleum persicum extract was determined based on Lorke’s method. Histopathological evaluation was also performed. Results: In addition to decreased urine protein and albumin, and increased creatinine and urea, co-treatment with gentamicin and Heracleum persicum significantly reduced levels of creatinine and urea, and increased sodium and potassium in serum. Heracleum persicum treatment also improved stereological parameters and serum inflammatory cytokines. There was a significant increase in serum glutathione peroxidase activity and total antioxidant capacity as well as a reduction in malondialdehyde level. Furthermore, treatment with Heracleum persicum extracts downregulated p53, caspase-3, and Bax and upregulated Bcl-2 expressions. In histopathological evaluation, Heracleum persicum extracts showed protection against gentamicin-induced renal damages. Conclusions: Heracleum persicum exhibits protective effects against gentamicin-induced structural and functional renal impairments.

66-73

Authors: Hanan M Al-Yousef, Perwez Alam, Zakia Khanam, Musarat Amina, Wafaa H. B. Hassan

Objective: To investigate the effect of Corchorus olitorius aqueous fraction (COAF) on quorum sensing (QS)-regulated virulence factors and biofilm formation in Pseudomonas aeruginosa (PAO1). Methods: The preliminary screening of the anti-QS effect of COAF was performed by evaluating the anti-pathogenic activity against Chromobacterium violaceum CV026 biosensor strain. Next, the inhibitory effects of COAF on QS-regulated pyocyanin production, proteolytic and elastolytic activities, swarming motility, and biofilm formation were evaluated in PAO1. Results: The results showed that the treatment of COAF significantly decreased the biofilm biomass, attenuated virulence factors, and inhibited swarming motility of PAO1 without affecting the growth of the bacteria in a dose-dependent manner. COAF at 2 000 μg/mL significantly decreased Las B elastase activity in PAO1 culture, exopolysaccharide production, swarming motility, pyocyanin level, and biomass of PAO1 by 55% (P<0.05), 60% (P<0.01), 61% (P<0.01), 65%(P<0.01) and 73% (P<0.01), respectively. In addition, the production of violacein was decreased by 62% (P<0.01) with the treatment of a high dose of COAF. Conclusions: These findings indicate that COAF can be a potential source of anti-QS agents.

81-88

Authors: Ornanong Tusskorn, Kanoktip Pansuksan, Kwanchayanawish Machana

Objective: To investigate the effects of Borassus flabellifer L. extracts on antioxidant activity, maintenance of cellular redox, and mitochondrial function in cisplatin-induced kidney injury. Methods: The extracts of Borassus flabellifer were obtained from crude male flowers using ethyl acetate and methanol. The antioxidant potential was evaluated by 2,2-azino-bis-(3-ethylbenzothiazoline- 6-sulfonic acid), 2,2-diphenyl-1-picrylhydrazyl, and ferric reducing antioxidant power, and total phenolic content was also determined. Cytoprotective activity of ethyl acetate and methanolic extracts was assessed after kidney cells were treated with cisplatin. Oxidative stress was determined by glutathione (GSH) assay, and formation of reactive oxygen species (ROS) and changes in mitochondrial transmembrane potential (ΔΨm) using 2’,7’-dichlorofluorescin diacetate and JC-10 assays, respectively. Results: Borassus flabellifer methanolic extract exhibited greater antioxidant activity than the ethyl acetate extract. Cytoprotective effect was demonstrated in both extracts, particularly in the ethyl acetate extract. The extracts showed protection against the cytotoxic effect of cisplatin by prevention of the increased GSSG and declined GSH/GSSG ratio. Both extracts also prevented the increase in ROS formation, and loss of ΔΨm. Conclusions: Both Borassus flabellifer extracts show antioxidant activity and cytoprotective effect against cisplatin-induced cytotoxicity of NRK-52E cells by preventing oxidative stress and maintenance of GSH redox status. Borassus flabellifer extracts may possess beneficial effects on the prevention of oxidative stress- induced cell injury.