Asian Pacific Journal of Tropical Biomedicine
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    2021,11(2):47-58, DOI: 10.4103/2221-1691.298628
    Abstract:
    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.
    2021,11(2):59-65, DOI: 10.4103/2221-1691.303604
    Abstract:
    Objective: To evaluate the anti-inflammatory potential of aqueous extract of Pterocarpus santalinus L.f. heartwood using molecular docking and in vivo experiment. Methods: An aqueous extract of Pterocarpus santalinus heartwood was prepared using a Soxhlet apparatus. Phytocompounds in the extract were tentatively identified using high-resolution mass spectrometry. Molecular docking experiments were carried out to evaluate the binding affinity of selected compounds, phloridzin to cyclooxygenase-1 (COX-1), cyclooxygenase-2 (COX-2), prostaglandin E synthase-1 (PGES-1) and 5-lipoxygenase (5-LOX). Anti-inflammatory potential was evaluated by carageenan induced paw edema model in rats. Results: The presence of major component phloridzin along with quercetin, parthenin, ginkgolide B, picrotoxinin, usnic acid, octopine, and epigallocatechin was detected in the extract. Molecular docking study showed that phloridzin inhibited COX-1, COX-2, PGES-1 and 5-LOX with more affinity than ibuprofen and paracetamol. Pterocarpus santalinus heartwood extract at 200 and 400 mg/kg BW showed significant reduction in carageenan-induced hind paw edema in a dose-dependent manner, but the effect was slow when compared with the standard ibuprofen (30 mg/kg p.o.). Conclusions: The study indicated that after clinical trials, the aqueous extract of Pterocarpus santalinus heartwood can be effectively used in phytotherapy to treat inflammation.
    2021,11(2):66-73, DOI: 10.4103/2221-1691.303605
    Abstract:
    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.
    2021(2):74-80, DOI: 10.4103/2221-1691.303606
    Abstract:
    Objective: To investigate the anti-senescence effect of 3-bromo-4,5- dihydroxybenzaldehyde (BDB) from Polysiphonia morrowii Harvey in human dermal fibroblasts (HDF). Methods: HDF were subjected to treatment of BDB and then treated with hydrogen peroxide (H2O2) to induce premature senescence. Senescence-associated β-galactosidase (SA-β-gal) activity in HDF was determined using the SA-β-gal staining method. Intracellular reactive oxygen species (ROS) production was measured using the 2’,7’-dichlorodihydrofluorescein diacetate assay. Western blotting assay was performed to assess the level of antioxidant enzyme glutathione peroxidase 1 (GPX1). In addition, intracellular collagen and collagenase contents were analyzed using the respective ELISA kits. Elastase activity in HDF supernatants was measured from p-nitroaniline release and normalized using total protein content. Results: Treatment of HDF with H2O2 increased the activity of SA-β-gal, but BDB pre-treatment resulted in the reduction of SA-β-gal activity. Moreover, BDB significantly reduced H2O2-induced intracellular ROS production. BDB also markedly increased the level of GPX1, which was inhibited by 400 μM of H2O2. Furthermore, in in vitro study, BDB significantly increased intracellular collagen content and decreased matrix metalloproteinase-1 and elastase activities in HDF. Conclusions: Our results demonstrate that BDB shows antisenescence and anti-wrinkle activities in vitro.
    2021(2):81-88, DOI: 10.4103/2221-1691.303607
    Abstract:
    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.
    2021(2):89-96, DOI: 10.4103/2221-1691.301058
    Abstract:
    Objective: To investigate the potential anti-aging mechanism of 9-hydroxy-6,7-dimethoxydalbergiquinol (HDDQ) on hydrogen peroxide (H2O2)-induced oxidative stress in human dermal fibroblasts (HDFs). Methods: The effect of HDDQ on cell viability was assessed by MTT assay, and the effects of HDDQ on senescence-like phenotypes were determined by senescence-associated β-galactosidase (SA- β-gal) staining, Western blotting analysis, and a cell proliferation assay. The expression level and activity of sirtuin-1 (SIRT1) induced by HDDQ were also measured. Results: HDDQ reversed senescence-like phenotypes in the oxidant-challenged model, through reducing SA-β-gal activity and promoting cell growth. Meanwhile, decreases in ac-p53, p21Cip1/WAF1, and p16Ink4a and an increase in pRb were observed. HDDQ induced the expression of SIRT1 in a concentration- and time-dependent manner. Moreover, HDDQ inhibited H2O2-induced phosphorylation of Akt by SIRT1 up-regulation and reduced SA-β-gal staining. Conclusions: HDDQ inhibits H2O2-induced premature senescence and upregulation of SIRT1 expression plays a vital role in the inhibition of the senescence phenotype in HDFs.
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    Available online:January 05, 2021, DOI: 10.4103/2221-1691.306133
    Abstract:
    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 methanolic extract of Berberis lyceum strongly inhibited HCV RNA replication with an IC50 of 11.44 μg/mL. RTPCR 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. 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. Conclusions: Our findings suggest that Berberis lyceum extract strongly inhibits HCV propagation by reducing HCV NS5Ainduced perturbation of MAPK signaling.
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    2019(7):299-306, DOI: 10.4103/2221-1691.261810
    [Abstract] (150) [HTML] (0) [PDF 1.67 M] (1020)
    Abstract:
    Objective: To investigate the effects of atranorin, a lichen secondary metabolite, on SPC212 malignant mesothelioma cells in vitro. Methods: SPC212 malignant mesothelioma cell line was used. 3-(4,5-dimethylthiazol-2-yl)- 2,5-diphenyltetrazolium bromide assay was used to evaluate cytotoxic effects of atranorin and cisplatin at 24, 48 and 72 h. Hematoxylin-eosin staining and 4',6-diamidino-2-phenylindole, dihydrochloride staining were used for determining cell and nucleus morphology, respectively. Wound healing assay was used for investigating cell migration. The xCELLigence real-time cell analysis system was used for determining cell proliferation. Results: Atranorin at 5-450 μM decreased cell viability at 24, 48 and 72 h. IC50 values of atranorin were 300.94, 292.6 and 278.02 μM at 24, 48 and 72 h, respectively; meanwhile, the IC50 values of cisplatin were 128.00, 34.37 and 17.05 μM at 24, 48 and 72 h, respectively. Furthermore, atranorin disrupted cell and nuclear morphology with increasing concentrations. Atranorin significantly reduced cell migration by 38%, 37% and 35% at 300, 250 and 200 μM, respectively (P<0.000). Atranorin at 160-450 μM decreased cell proliferation at 72 h (P<0.000). Conclusions: Atranorin has cytotoxic, antiproliferative, apoptotic and cell migration inhibitory effects on SPC212 malignant mesothelioma cancer cells.
    2019(7):271-277, DOI: 10.4103/2221-1691.261742
    Abstract:
    Objective: To determine the effects of toll-like receptor 2 (TLR-2) agonist, Pam3CSK4, on cellular and humoral immune response against recombinant Mycobacterium bovis bacilli Calmette-Guérin (rBCG) expressing the C-terminus of merozoite surface protein-1 of Plasmodium falciparum. Methods: Six groups of mice (n=6 per group) received intraperitoneal phosphate buffered saline T80 (PBS-T80), BCG or rBCG in the presence or absence of Pam3CSK4. Enzymelinked immunosorbent assay was carried out to measure serum total IgG, IgG1, IgG2a, and IgG2b production. Spleens were also harvested and splenocytes were co-cultured with rBCG antigen for in vitro determination of IL-4 and IFN-γ via enzyme-linked immunosorbent assay. Results: The production of total IgG and the isotype IgG1, IgG2a and IgG2b was significantly higher in rBCG-immunised mice than in the BCG and PBS-T80-immunised mice, and Pam3CSK4 further enhanced their productions. A similar pattern was also observed in IFN-γ production. Moreover, there was no significant difference in IL-4 production in all groups either in the presence or absence of Pam3CSK4. Conclusions: We present evidence of the adjuvant effects of TLR-2 agonist in enhancing the production of total IgG, IgG1, IgG2a, IgG2b, as well as IFN-γ in response to rBCG. However, the presence or absence of Pam3CSK4 had no effect on IL-4 production.
    2019(7):284-291, DOI: 10.4103/2221-1691.261764
    Abstract:
    Objective: To assess the protective effect of galangin on membrane bound enzymes in rats with streptozotocin-induced diabetes. Methods: A single low dose of streptozotocin was injected to adult male albino rats to induce hyperglycemia. Galangin (8 mg/kg) or glibenclamide 600 μg/kg as a standard drug was given orally once daily for 45 days by gavage. Membrane-bound adenosine triphosphatases were determined including total ATPase, sodium-potassium-ATPase, calcium-ATPase and magnesium-ATPase in erythrocytes and tissues (kidney, liver, and heart). Results: The levels of total ATPases, sodium-potassium-ATPase, calcium-ATPase and magnesium-ATPase in erythrocytes and tissues were significantly altered in diabetic rats as compared to that in normal rats. After 45 days of treatment with galangin or glibenclamide, the levels of these enzymes were similar to that of normal control rats. Conclusions: Oral administration of galangin or glibenclamide can improve activities of these membrane-bound ATPases towards normal levels. Mechanism of galangin needs to be further explored in future.
    2019(7):307-314, DOI: 10.4103/2221-1691.261822
    Abstract:
    Objective: To evaluate the antimicrobial, antioxidant, cytotoxicity and anticancer activity of fractions from Jatropha zeyheri roots and to explore the phytochemical profile of the most biologically active fraction. Methods: Fractions from Jatropha zeyheri ethyl acetate extract were investigated for antimicrobial activity against a plethora of pathogenic microorganisms of different origins. The cytotoxicity studies of fractions were evaluated in vitro using tetrazolium-based calorimetric assay against human dermal fibroblast, colon adenocarcinoma (Caco-2), breast cancer (MCF-7) and lung cancer (A547) cell lines. The anti-oxidant activity of fractions was determined in vitro against 2,2-diphenyl-1-picrylhydrazyl (DPPH), 2,2′-azino-bis-(3-ethylbenzothiazoline- 6-sulfonic acid) diammonium salt (ABTS) and chelation of iron (Fe2+). Gas chromatography mass spectrometry analysis was performed to detect phytochemical constituents in fraction with potent biological activity. Results: Fraction 2 of Jatropha zeyheri roots exhibited the lowest minimum inhibitory concentration of 40 μg/mL against Klebsiella pneumoniae and 80 μg/mL against Candida albicans, Staphylococcus aureus and Mycoplasma hominis. The fractions revealed some varying degrees of cytotoxicity against human dermal fibroblasts yielding LC50 values ranging from 28.96 to 166.52 μg/mL. Fraction 3 exhibited the highest selectivity index value of 2.08 against Klebsiella pneumoniae. Moreover, fraction 2 selectively inhibited the growth of Caco-2 with LC50 of 8.83 μg/mL, compared to other cancerous cell lines. Fraction 2 of Jatropha zeyheri further exhibited IC50 of 19.66, 22.63 and 1.82 μg/mL against DPPH, ABTS and Fe2+, respectively. Gas chromatography mass spectrometry analysis revealed the presence of cyclotetracosane (10.08%), 9-hexacosene (9.40%), hexadecanoic acid (3.90%), (Z)-9-octadecenamide (3.63%), octacosane (2.27%), 11-n-decylheneicosane (2.23%), ethyl vallesiachotamate (2.17%), heneicosanoic acid (2.10%), and octadecanoic acid (2.08%) in fraction 2 of Jatropha zeyheri. Conclusions: These identified compounds, particularly cyclotetracosane (hydrocarbon), may well explain the biological activity of fraction 2 of Jatropha zeyheri, which possesses higher biological activity than other fractions. These compounds can be further investigated for use in treating various bacterial and fungal opportunistic infections associated with HIV-AIDS and related cancers.
    2019(7):278-283, DOI: 10.4103/2221-1691.261763
    Abstract:
    Objective: To explore the effect of hydroalcoholic extract of Cynodon dactylon on the whole body radiation-induced oxidative status of the cerebellum and cognitive impairments in mice. Methods: Swiss albino mice were randomly divided into the control group, radiation control group, low dose and high dose Cynodon dactylon extract treated groups and pre-treated with Cynodon dactylon extract before irradiation. Cynodon dactylon extract was administered for 7 d daily in low dose (0.25 g/kg) and high dose (1 g/kg). On day 7, mice were irradiated with a sublethal dose of 5 Gy gamma rays. Motor coordination was assessed by elevated rotarod test and spatial memory was studied by water maze test. Subsequently, biochemical markers (glutathione, lipid peroxidation and nitric oxide levels) in the cerebellum were evaluated. Results: The gamma irradiated group showed significant impairment in motor coordination and spatial memory compared to normal mice. Mice treated by Cynodon dactylon extract prior to gamma radiation showed good improvement in both paradigms compared to the radiation control group. Moreover, glutathione level was increased, while lipid peroxidation and nitric oxide levels were significantly reduced in mice receiving low dose and high dose of Cynodon dactylon extract compared to the radiation control group. Conclusions: The present study suggests the neuroprotective role of Cynodon dactylon against radiation-induced cognitive impairment and oxidative stress on the cerebellum of mice.
    2019(7):291-298, DOI: 10.4103/2221-1691.261809
    Abstract:
    Objective: To study the inhibitory effect of rice bran extracts of Thai black Kam Muang and red Hawm Dawk Mali Deang on oxidative stress factors including superoxide (O2 ?-), nitric oxide (NO?), and inducible nitric oxide synthase (iNOS). Methods: Bran extracts (40% ethanol) of Kam Muang and Hawm Dawk Mali Deang were obtained and evaluated for in vitro 2-2′-azino-di-(3-ethylbenzthiazoline sulfonate) (ABTS) and NO?scavenging activity. Their inhibitory effects on cellular O2 ?- and NO? were measured in phorbol 12-myristate 13-acetate-stimulated neutrophil-like HL-60 cells and lipopolysaccharidestimulated RAW264.7 macrophages, respectively, and their viability was monitored using the MTT assay. The effect on iNOS expression was also assessed by the Western blotting assay. Total contents of phenolics, flavonoids, and subtypes were also determined. Results: Hawm Dawk Mali Deang exhibited about 3.5-fold greater cellular O2 ?- inhibitory activity than Kam Muang [EC50 values of (23.57±4.54) and (81.98±1.45) μg/mL, respectively] in phorbol 12-myristate 13-acetate-stimulated HL-60 cells. Hawm Dawk Mali Deang exhibited about 2-fold higher in vitro ABTS?+ and NO? scavenging activity than Kam Muang, but it exerted cellular NO?inhibitory activity of only about 26% (undetermined EC50 value) in lipopolysaccharide-stimulated RAW264.7 cells. Conversely, Kam Muang exerted potent cellular NO? inhibitory activity [EC50 value: (281.13±59.18) μg/mL] and dose-dependently decreased iNOS levels. No cytotoxicity of both extracts was detected in both cell types. As for corresponding contents, Hawm Dawk Mali Deang contained higher contents of phenolics and flavonoids than Kam Muang. Moreover, Kam Muang and Hawm Dawk Mali Deang had a high content of total anthocyanins [(14.73±0.52) mg C3GE/g of extract] and total proanthocyanidins [(115.13±1.47) mg CE/g of extract], respectively. Conclusions: Based on these data, bran extracts of Thai black Kam Muang and red rice Hawm Dawk Mali Deang can help lower oxidative stress and inflammation attributed partly to O2 ?- and NO?.
    2019(5):209-216, DOI: 10.4103/2221-1691.259001
    Abstract:
    Objective: To evaluate the phytochemical constituents, antimicrobial and antioxidant activities of the extracts of Distemonanthus benthamianus (D. benthamianus) stem bark and Solanum torvum (S. torvum) fruit which have been used as traditional medicinal herbs in Gabon. Methods: Plant extracts were subjected to a qualitative study (phytochemical screening) and a quantitative (dosing) study of secondary metabolites. Antioxidant activity was tested by 1,1- diphenyl-2-picrylhydrazyl and 2,2'-azino-bis 3-ethylbenzothiazoline-6-sulfonic acid assay. Bacteria and fungi susceptibility tests were performed on Mueller Hinton medium and solid Sabouraud, respectively, using the diffusion method, while minimum inhibitory concentration, minimum fungicidal concentration and minimum bactericidal concentration were evaluated by microdilution method. Results: The total phenol and tannin contents were significantly higher in the water-ethanol extract compared to the other extracts of D. benthamianus and S. torvum. The water-ethanol and water-acetone extracts showed significantly higher antioxidant activity than the aqueous extracts of the two medicinal plants. However, the extracts presented weak antioxidant activities compared to standards (Vitamin C, BHA). The water-acetone and water-ethanol extracts of S. torvum showed the highest antimicrobial activity against Bacillus cereus LMG 13569 BHI, /i>Shigella dysenteriae 5451 CIP, Shigella dysenteriae and Neisseria gonorrhoeae. Conclusions: Our results show that D. benthamianus and S. torvum can be promising sources of natural products with potential antimicrobial and antioxidant activities.
    2019(5):188-195, DOI: 10.4103/2221-1691.258998
    Abstract:
    Objective: To examine the effects of p-coumaric acid on ethanol-induced kidney injury in Swiss Wistar rats. Methods: Ethanol (25% v/v) was used to induce nephrotoxicity in rats. p-Coumaric acid was orally administered at 50, 100, or 200 mg/kg body weight. The levels of oxidative parameters were determined; pro-inflammatory biomarkers were analyzed by Western blotting and apoptotic protein was analyzed by immunohistochemistry. Results: Ethanol treated rats showed decreased levels of antioxidants and aberrant production of pro-inflammatory cytokines (IL-6, IL1β, TNF-α), NF-κB activation and imbalance of proand anti-apoptotic proteins (Bcl-2, Bax, caspase 3). Meanwhile, p-coumaric acid restored antioxidant levels and decreased the levels of inflammatory cytokines, NF-κB, and proapoptotic proteins and increased Bcl-2 expression. Conclusions: p-Coumaric acid ameliorates ethanol-induced kidney injury by restoring antioxidant production and suppressing cellular apoptosis and inhibiting NF-κB expression. p-Coumaric acid should be further investigated as a promising candidate for ethanol-induced kidney toxicity.
    2019(6):227-231, DOI: 10.4103/2221-1691.260394
    Abstract:
    Objective: To explore the seroprevalence, spatial distribution and risk factors for Leishmania seropositivity in Jordan. Methods: Blood samples from 872 apparently healthy participants were randomly selected from 11 governorates in Jordan and tested for anti- Leishmania K39 IgG. Risk factors (animal ownership and agriculture practices) and demographic data were also collected using pre-tested and validated questionnaire. Results: Overall, 2.52% of participants were seropositive for Leishmania spp. Participants living in the Jordan Valley plateau had significantly greater odds (adjusted odds ratio = 3.70, 95% CI 1.37-9.93) of seropositivity than those living in the Highlands after adjustment for age. Conclusions: This study supports the intermittent reports of cutaneous leishmaniasis outbreaks in the Jordan Valley. Vector control measures in the Jordan Valley should be considered, including insecticide treated bed nets, sugar baits and using flowering plants to attract and trap Phlebotomus papatasi sand flies. Active surveillance in the Jordan Valley is also recommended in light of this and other reports.
    2019(6):257-262, DOI: 10.4103/2221-1691.260398
    Abstract:
    Objective: To isolate, identify, and evaluate a new angiotensin-converting enzyme inhibitor from Peperomia pellucida (L.) Kunth herbs. Methods: A dried sample of Peperomia pellucida herb was successively macerated with n-hexane and ethyl acetate. The ethyl acetate extract solution was evaporated to obtain the crude extract. Vacuum liquid column chromatography and thin layer chromatography were performed to obtain two pure compounds. Then, both compounds were elucidated and identified using the spectroscopic method. Angiotensin-converting enzyme inhibitory activity studies of both compounds were determined using angiotensin-converting enzyme kit WST-1 with spectrophotometer microplate reader 96-well at 450 nm wavelength. Results: Two bioactive compounds were successfully isolated from Peperomia pellucida herb, including a new compound of 2,3,5-trimethoxy-9-(12,14,15-trimethoxybenzyl)-1H-indene and pellucidin A. Both compounds demonstrated angiotensin-converting enzyme inhibitory activity, with IC50 values of 72 μM (27.95 μg/mL) and 11 μM (4.4 μg/mL), respectively. Conclusions: In the present study, two active angiotensin-converting enzyme inhibitors were successfully isolated and purified from Peperomia pellucida which is used as an antihypertensive in traditional medicine, and support its use as an angiotensin-converting enzyme-inhibiting drug.
    2019(5):196-203, DOI: doi: 10.4103/2221-1691.258999
    Abstract:
    Objective: To explore the protective effect of Polygonum minus ethanolic extract on cisplatininduced neurotoxicity. Methods: In vitro test, total phenolic content assay and DPPH assay were performed to determine the antioxidant activity of Polygonum minus. For in vivo test, 30 male SpragueDawley rats were randomly divided into 5 groups: the control group, cisplatin 10 mg/kg, Polygonum minus 100 mg/kg, Polygonum minus 200 mg/kg and Polygonum minus 400 mg/kg. The control group and the cisplatin group were given distilled water whereas Polygonum minus groups received the respective dose of Polygonum minus extract orally for 14 d. On day 15, a single intraperitoneal administration of normal saline was given to the control group; while 10 mg/kg of cisplatin was given to the cisplatin group and Polygonum minus groups. Body weight, signs of illness, daily activity and mortality were observed at least once daily throughout the experimental period. On day 18, the anterior part of the brain was collected and processed for histological and ultrastructural analyses (right hemisphere). The remaining part (left hemisphere) of the brain was assayed to determine malondialdehyde and catalase levels for oxidative stress analyses. Results: Polygonum minus ethanolic extract possessed high phenolic content (977.6 mg GAE/g) and 95.9% DPPH radical scavenging activities. No mortality was observed in all groups. Rats in the cisplatin group were weak and less active compared to Polygonum minus treated rats. In the cisplatin group, disorganised cellular layers of the cerebral cortex were observed whereas rats treated with low and mid doses of Polygonum minus extract had normal cerebral cortex as in the control group. Mild ultrastructural changes were observed in rats treated with low and mid doses of Polygonum minus extract. Meanwhile, low and mid doses of Polygonum minus extract significantly reduced malondialdehyde level whereas low and mid doses of Polygonum minus extracts groups significantly increased catalase activity compared to the cisplatin group. Conclusions: Polygonum minus ethanolic extract at 100 and 200 mg/kg attenuates cisplatininduced oxidative stress in the cerebral cortex via its antioxidant activity.
    2019(6):232-239, DOI: 10.4103/2221-1691.260395
    Abstract:
    Objective: To establish an early warning system for cutaneous leishmaniasis in Fars province, Iran in 2016. Methods: Time-series data were recorded from 29 201 cutaneous leishmaniasis cases in 25 cities of Fars province from 2010 to 2015 and were used to fit and predict the cases using time-series models. Different models were compared via Akaike information criterion/ Bayesian information criterion statistics, residual analysis, autocorrelation function, and partial autocorrelation function sample/model. To decide on an outbreak, four endemic scores were evaluated including mean, median, mean + 2 standard deviations, and median + interquartile range of the past five years. Patients whose symptoms of cutaneous leishmaniasis began from 1 January 2010 to 31 December 2015 were included, and there were no exclusion criteria. Results: Regarding four statistically significant endemic values, four different cutaneous leishmaniasis space-time outbreaks were detected in 2016. The accuracy of all four endemic values was statistically significant (P<0.05). Conclusions: This study presents a protocol to set early warning systems regarding time and space features of cutaneous leishmaniasis in four steps: (i) to define endemic values based on which we could verify if there is an outbreak, (ii) to set different time-series models to forecast cutaneous leishmaniasis in future, (iii) to compare the forecasts with endemic values and decide on space-time outbreaks, and (iv) to set an alarm to health managers.
    2019(6):249-256, DOI: 10.4103/2221-1691.260397
    Abstract:
    Objective: To determine the anti-proliferative activity of Abrus precatorius (A. precatorius) leaf extracts and their effect on cell death. Methods: A. precatorius leaves were extracted successively with hexane, ethyl acetate and methanol by Soxhlet extraction. Aqueous extract was prepared by decoction at 50 ℃. Extracts of A. precatorius leaves were used to treat selected cancer and normal cell lines for 72 h. Furthermore, 3-(4,5-dimethyl thiazol-2-yl) 2,5-diphenyl tetrazolium bromide assay was performed to determine cell viability. Analysis of cell cycle arrest, apoptosis assay and apoptosis protein expressions were determined by flow cytometry. Results: Methanolic extract of A. precatorius leaves showed the lowest IC50 on MDA-MB-231 cells at (26.40±5.40) μg/mL. Flow cytometry analysis revealed that cell arrest occurred at G0/ G1 phase and the apoptosis assay showed the occurrence of early apoptosis at 48 h in MDA-MB-231 cells treated with methanolic extract of A. precatorius leaves. Methanolic extract of A. precatorius leaves induced apoptosis by upregulation of Bax, p53 and caspase-3 and downregulation of Bcl-2. Conclusions: Methanolic extract of A. precatorius leaves promotes MDA-MB-231 cell death by inducing cell cycle arrest and apoptosis possibly via the mitochondrial-related pathway.
    2019(6):240-248, DOI: 10.4103/2221-1691.260396
    Abstract:
    Objective: To investigate anti-dyslipidemic effects of hydroalcoholic fenugreek seed extracts, diosgenin, and 4-OH-Ile on HepG2 cell line. Methods: HepG2 cells were treated with hydroalcoholic fenugreek seed extracts, diosgenin, 4-OH-Ile, and orlistat. IC20 was calculated using the MTT method. The cells were then pre-treated with IC20 concentrations for 24 and 48 h. Real time PCR was employed to measure expression of liver X receptor alpha (LXR α ), sterol regulatory element-binding protein-1C (SREBP-1C), acetyl-CoA carboxylase (ACC), fatty acid synthase (FAS), fibroblast growth factor 21 (FGF21), peroxisome proliferator-activated receptor gamma (PPAR γ ), and low-density lipoprotein receptor (LDLR). Results: The results showed that LXR α (P=0.003, P<0.001), SREBP-1C (P<0.001, P<0.001), ACC (P=0.002, P=0.006), and FAS (P<0.001, P<0.001) were downregulated significantly, while FGF21 (P<0.001, P<0.001), PPAR 7 (P=0.004, P<0.001), and LDLR (P<0.001, P<0.001) were upregulated significantly in HepG2 cells treated with the IC20 of hydroalcoholic fenugreek seed extracts, diosgenin, 4-OH-Ile, and orlistat in 24 and 48 h, respectively. Conclusions: Hydroalcoholic fenugreek seed extracts, diosgenin, and 4-OH-Ile significantly modulate the expression of some important lipid metabolism related genes, which is similar to orlistat. Trigonella foenum-graecum seed extract or its derivatives should be further investigated for their dyslipidemia effects and its complications.
    2019(5):217-221, DOI: 10.4103/2221-1691.259002
    Abstract:
    Objective: To evaluate the neuropharmacological properties of Costus speciosus (C. speciosus) rhizome using different experimental mouse models. Methods: The anxiolytic effect was investigated by hole-board test, elevated plus maze and light/dark test, while central nervous system (CNS) depressant effect was evaluated by thiopental sodium-induced sleep test. Finally, antidepressant effect was evaluated by forced swimming test and tail suspension test. Results: In both elevated plus maze and hole board test, 400 mg/kg C. speciosus showed more significant CNS depressant effect than 1 mg/kg diazepam. Both 200 mg/kg and 400 mg/kg C. speciosus extract produced a significant dose-dependent decrease in onset of sleep. In forced swimming test, C. speciosus rhizome showed a decrease in duration of immobility in a dose-dependent manner. Imipramine (10 mg/kg) and C. speciosus extract at 400 mg/kg dose exhibited a significant reduction in duration of immobility in tail suspension test which provided additional evidence of antidepressant effect of C. speciosus rhizome. Conclusions: Our study indicates that C. speciosus rhizome possesses CNS depressant, anxiolytic and antidepressant-like activities. Further studies are warranted determine the exact phytoconstituents and mechanism of action responsible for the neuropharmacological effect.
    2019(6):263-270, DOI: 10.4103/2221-1691.260399
    Abstract:
    Objective: To identify alpha-glucosidase inhibitors from Ficus benghalensis and analyze gene set enrichment of regulated protein molecules. Methods: The phytoconstituents of Ficus benghalensis were queried for inhibitors of alpha-glucosidase, also identified as aldose reductase inhibitors. Druglikeness score, absorption, distribution, metabolism, excretion and toxicity profile, biological spectrum, and gene expression were predicated for each compound. Docking study was performed to predict the binding affinity with alpha-glucosidase and aldose reductase and compared with clinically proven molecules. Kyoto Encyclopedia of Genes and Genomes pathway analysis was performed for the regulated genes to identify the modulated pathways. Results: Apigenin, 3,4’,5,7-tetrahydroxy-3’-methoxyflavone, and kaempferol were identified as inhibitors of alpha-glucosidase and aldose reductase. Kaempferol was predicted to possess the highest binding affinity with both targets. The p53 signaling pathway was predicted to modulate the majority of protein molecules in diabetes mellitus. All the alpha-glucosidase inhibitors were also predicted as membrane integrity agonist and anti-mutagenic compounds. Conclusions: The current study indicates alpha-glucosidase inhibitors from Ficus benghalensis can act as aldose reductase inhibitors after absorption from the intestinal tract. Furthermore, these phytoconstituents are involved in the regulation of numerous protein molecules and pathways. Hence, the anti-diabetic efficacies of these compounds are due to their action on multiple protein molecules and synergistic effects which should be confirmed by future investigations.
    2019(5):204-208, DOI: 10.4103/2221-1691.259000
    Abstract:
    Objective: To evaluate antibacterial activity and the bioactive compounds of 50% hydroethanolic extract of Alpinia zerumbet (A. zerumbet) rhizomes. Methods: Eight reference microbial strains including two Gram-positive bacteria [Staphylococcus aureus (ATCC 29213) and Enterococcus faecalis (ATCC 29212)] and six Gram-negative bacteria [Escherichia coli (ATCC 25922), Klebsiella pneumoniae (ATTC 700603), Proteus mirabilis (DMST 8212), Salmonella enterica subsp. enterica serovar Vellore. (ATCC 15611), Shigella flexneri (ATCC 12022) and Pseudomonas aeruginosa (ATCC 27853)], were used to test antimicrobial susceptibility by the broth microdilution method. Bioactive compounds were analyzed by using HPLC. Results: The minimum inhibitory concentration values of A. zerumbet extract were 8 mg/mL for Staphylococcus aureus, Escherichia coli and Shigella flexneri and 16 mg/mL for Enterococcus faecalis and the other four Gram-negative bacilli. HPLC chromatograms revealed that the A. zerumbet extract contained hydroxybenzoic acids, hydroxycinnamic acids and flavonoids. Conclusions: The constituents of A. zerumbet rhizomes could be a potential source of antibacterial compounds, warranting further study of A. zerumbet extract.
    2019(11):449-455, DOI: 10.4103/2221-1691.270977
    Abstract:
    Objective: To evaluate spasmolytic mechanisms of aqueous and methanolic extracts from Distemonanthus benthamianus trunk-bark. Methods: Spasmolytic activities of extracts were evaluated in vitro on spontaneous and potassium chloride-induced jejunum contractions, or against cholinergic [acetylcholine (0.3 μmol/L)] stimulations. High performance liquid chromatography analysis of both extracts was performed in reference to standard compounds. Results: Extracts developed concentration-dependent inhibitory activities. The methanolic extract, which revealed better activity, produced spasmolytic and myorelaxant effects at concentrations of 0.01-0.30 mg/mL with EC50 of 0.06 and 0.09 mg/mL (95% CI: 0.03-0.3 mg/mL), respectively. Its anticholinergic effect was obtained at the same concentrations with EC50 of 0.11 mg/mL (95% CI: 0.03-0.3 mg/mL). Chromatograms showed the presence of gallic acid in both extracts, rutin being only detected in the aqueous extract. Conclusions: Distemonanthus benthamianus extracts exhibit verapamil and atropine-like activities, thus highlighting calcium channels and muscarinic receptors blocking potentials, which may be conveyed by some phenolic compounds. These results confirm the antidiarrheal activity of Distemonanthus benthamianus extracts.
    2019(5):222-226, DOI: 10.4103/2221-1691.259003
    Abstract:
    Objective: To compare the in vitro antiparasitic activity of aqueous extracts from Ziziphus joazeiro leaves and stem bark against Trypanosoma cruzi, Leishmania braziliensis, and Leishmania infantum, as well as to evaluate its cytotoxicity in mammalian cells, in addition to identifying the chemical composition of the extracts. Methods: Ziziphus joazeiro leaf and stem bark aqueous extracts were prepared by cold extraction maceration and subjected to ultra-efficient liquid chromatography coupled to a quadrupole/time of flight system. The susceptibility assays used Trypanosoma cruzi CLB5 strains and promastigote forms of Leishmania braziliensis and Leishmania infantum for antiparasitic activity of the extracts. Moreover, mammalian fibroblasts NCTC clone 929 were used for cytotoxicity analysis. Results: Terpenoid compounds, flavonoids and phenolic acid were identified in extracts. The stem bark aqueous extracts presented more significant results in terms of antiparasitic activity compared with the leaf aqueous extracts, especially against Leishmania braziliensis and Leishmania infantum promastigote forms with an IC50 < 500 μg/mL. The cytotoxicity evaluation showed moderate toxicity of the stem bark aqueous extracts, which is relevant information for the rational use of this plant part since it is widely used by the population. Conclusions: These preliminary results may contribute to the formulation of new therapeutic agents against this group of neglected diseases, so further investigations are required to delineate
    2019(5):181-187, DOI: 10.4103/2221-1691.258997
    Abstract:
    Objective: To determine the spatial distribution and infection rate of sand flies as vectors of Leishmania parasite in Ardabil province, northwest of Iran. Methods: This was a descriptive cross-sectional study. The sand flies were collected from 30 areas in all 10 districts of Ardabil province during 2017. The specimens were caught using the sticky traps. The head and genitalia of sand flies were separated and mounted in Berlese solution for microscopic identification. The Geographical Information System ArcMap10.4.1 software was used to provide the spatial maps. Results: A total of 2 794 sand flies specimens were collected and 22 species of sand flies were identified from the two genera: Phlebotomus and Sergentomyia from Ardabil province. The highest frequency was found in Phlebotomus papatasi (23.7%) followed by Phlebotomus kandelakii (13.0%). The promastigote form of Leishmania infantum parasite has been reported from the three main vectors of visceral leishmaniasis (Phlebotomus kandelakii, Phlebotomus perfiliewi and Phlebotomus tobbi) from Ardabil province, where the spatial distribution map of these visceral leishmaniasis vectors was prepared. Some important species of sand flies such as Phlebotomus kandelakii, Phlebotomus perfiliewi and Phlebotomus tobbi were reported and identified as main and probable vectors of visceral leishmaniasis in Ardabil. Conclusions: According to the Geographic Information System based maps, the frequency of the sand flies as leishmaniasis vectors, the leishmania parasite infection rate and the prevalence of the disease in the central areas of Ardabil province are higher than in other areas in Ardabil province.

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