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Biological Activity of Echinops Spinosus on Inhibition of Paracetamol- Induced Renal Inflammation

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Biological Activity of Echinops Spinosus on Inhibition of Paracetamol- Induced Renal Inflammation Biochemistry and Cell Biology Biological Activity of Echinops spinosus on Inhibition of Paracetamol- Induced Renal Inflammation Journal: Biochemistry and Cell Biology Manuscript ID bcb-2018-0212.R2 Manuscript Type: Article Date Submitted by the 20-Sep-2018 Author: Complete List of Authors: Hegazy, Marwa; Ain Shams University Faculty of Science Emam, Manal; Ain Shams University Faculty of Science Khattab, Hemmat; Ain Shams University Faculty of Science Helal, Nesma;Draft Ain Shams University Faculty of Science Nephrotoxicity, Phytochemicals, oxidative stress, gas chromatography Keyword: mass spectrometry, acetaminophen Is the invited manuscript for consideration in a Special Not applicable (regular submission) Issue? : https://mc06.manuscriptcentral.com/bcb-pubs Page 1 of 47 Biochemistry and Cell Biology 1 Biological Activity of Echinops spinosus on Inhibition of Paracetamol- 2 Induced Renal Inflammation 3 Short title: Echinops spinosus attenuate nephrotoxicity 4 Marwa Hegazya*, Manal Emama, Hemmat Khattabb, Nesma Helalb 5 a Biochemistry Department, Faculty of Science, Ain Shams University, Abbassia, 11566, Cairo, 6 Egypt. 7 b Botany Department, Faculty of Science, Ain Shams University, Abbassia, 11566, Cairo, Egypt. 8 *Corresponding author: Draft 9 Marwa Hegazy 10 Email: [email protected], [email protected] 1 https://mc06.manuscriptcentral.com/bcb-pubs Biochemistry and Cell Biology Page 2 of 47 11 12 Abstract 13 This study was designed to evaluate the possible mechanisms through which Echinops Spinosus 14 (ES) extract demonstrates nephroprotective effect on paracetamol (APAP)- 15 induced nephrotoxicity in rat. Twenty-Four Swiss albino rats were divided into four groups (six 16 rats each). Placebo group was orally administered sterile saline; APAP group received APAP (200 17 mg/kg/day i.p) daily; ES group was given orally ES extract (250 mg/kg); (APAP+ES) group: received 18 APAP as for APAP group and administrated ES extract as for ES group. Pretreatment of methyl 19 alcohol extract of ES reduced the protein expression of inflammatory parameters including 20 cyclooxygenase-2 (COX-2) and nuclearDraft factor kappa B (NF-κB) in kidney. It also reduced the 21 mRNA gene expression of tumor necrosis factor-α (TNF-α) and Interleukin-1β (IL-1β). ES 22 extract compensated deficits in the total antioxidant activity, suppressed lipid peroxidation and 23 amended the APAP induced histopathological kidney alterations. Moreover, ES treatment 24 restored the elevated levels of urea nitrogen in blood and creatinine in serum by acetaminophen. 25 ES extract attenuated the acetaminophen-induced elevations in renal nitric oxide levels. We 26 clarified that ES extract has the potential to defend kidney from APAP-induced inflammation, 27 and protection mechanism might by through decreasing oxidative stress and regulating the 28 inflammatory signaling pathway through modulating key signaling inflammatory biomarkers. 29 Key Words: Nephrotoxicity; Phytochemicals; oxidative stress; gas chromatography mass 30 spectrometry; acetaminophen. 31 Introduction 2 https://mc06.manuscriptcentral.com/bcb-pubs Page 3 of 47 Biochemistry and Cell Biology 32 A wide range of medicinal plants have been used in different countries and cultures as a 33 prophylactic and curative agent for urolithiasis (Gülçin et al., 2006). One of the most cited 34 families are Asteraceae (Ahmed et al. 2016). Genus Echinops family Asteraceae (Compositae) 35 comprises about 120 species distributed through Mediterranean region to central Asia 36 and Tropical Africa (Kadereit and Jeffrey 2007). It is common throughout the Sahara including 37 Sinai and the Red Sea coast. In Egypt, Echinops spinosus L is among five other species 38 representing this genus (Boulos 2009). Echinops spinosus (ES) is a perennial herb growing 1 39 meter or more and locally named Tassekra (Khedher 2014). Stems, leaves and roots of ES are 40 used as diuretic (Boulos 1983). 41 As a medicinal plant, ES is also used as a curative plant, it was frequently employed in 42 traditional medicine as an abortifacient,Draft a diuretic, for blood circulation, diabetes, gastric pain, 43 indigestion and spasmolytic problems (Khedher 2014). ES displayed several therapeutic 44 properties like anti-oxidant, anti-inflammatory and anti-microbial activities (Mujawar et al. 2015; 45 Maurya et al. 2015; Bouattour et al. 2016). 46 Secondary metabolites produced by plants are a great source of drug due to their safety, easy 47 availability, valuable effects on human body and high medicinal values. These metabolites have 48 been well-studied to not only their antioxidant properties but also, they have been proven to have 49 nephroprotective effects (Katanić et al. 2017; Gülçin, 2012). The Echinops characteristic 50 medicative uses could be due to the incidence of phenols (Khedher et al. 2014), quinoline 51 alkaloids Chevrier et al. 1976), flavanoids and sesquiterpenes (Boumaraf et al. 2016), acetylated 52 terpenoids and sterols (Bouattour et al. 2016), moreover as fatty acids and alkanes (Chevrier et 53 al. 1975). 3 https://mc06.manuscriptcentral.com/bcb-pubs Biochemistry and Cell Biology Page 4 of 47 54 Acetaminophen (N-acetyl-p-aminophenol; APAP), which is known as paracetamol, is the 55 most commonly used analgesic and antipyretic medication. However, its overdose leads to 56 kidney and liver damage. numerous scientific reports have focused majorly on APAP 57 hepatotoxicity. Alternatively, not many works approach APAP nephrotoxicity focusing on both 58 its mechanisms of action and therapeutic exploration (Karthivashan et al. 2016). 59 Acetaminophen toxicity generates acute tubular necrosis; one of the main causes of acute 60 renal failure. This toxicity initially happens by APAP metabolism to N-acetyl-p-benzoquinone 61 by the microsomal P-450 enzyme system that depletes reduced glutathione (GSH) and forms 62 APAP-protein adducts. Later on, the reactive nitrogen species peroxynitrite is created from nitric 63 oxide (NO) and superoxide resulting in 3-nitrotyrosine (Banerjee et al. 2017). Free radicals are 64 produced by exposure to drug toxicity inDraft an organism, and oxidative damage plays a significant 65 role in acetaminophen-induced hepatorenal injuries (Kandemir et al. 2017). Therefore, Medicinal 66 plants and phytomedicine are the prime choice of research as they possess better activity than 67 synthetic drugs and lesser side effects (Parameshappa et al. 2012). 68 Till date, no study has been dedicated to assess the protective efficacy of E. spinosus against 69 nephrotoxicity induced by APAP in rats. Keeping this visible, this study aimed primarily to 70 assess the nephroprotective effects and modulatory mechanisms of E. spinosus upon kidney of 71 APAP treated rats. 72 Materials and methods 73 Chemicals 74 Chemicals were purchased of high analytical grade from Biodiagnostic Company for 75 diagnostic and research reagents (Dokki, Giza, Egypt). Acetaminophen (APAP) and all 76 phenolic and flavonoid reagents were purchased from Sigma Co. USA. 4 https://mc06.manuscriptcentral.com/bcb-pubs Page 5 of 47 Biochemistry and Cell Biology 77 Plant Materials 78 E. spinosus shrubs with matched size and age were collected from Wadi Hagul arid habitat 79 throughout the flowering spring season (April 2017). The plant was then identified by Professor 80 Dr. Hemmat Khattab and documented in Botany Department, Ain Shams University (Egypt). 81 Plant name has been checked with http://www.theplantlist.org. Echinops spinosus L. is 82 a synonym of Echinops spinosissimus Turra. A voucher specimen was submitted at the 83 herbarium of botany department - faculty of science - Ain shams university for future reference. 84 Preparation of plant extract 85 The aerial parts (leaves/stem), roots and flower heads were cleansed, dried at room 86 temperature within the shade then ground,Draft to a powder by mechanical mills. The dried powder 87 (100 g) was extracted with distilled water, ethyl alcohol, methyl alcohol, petroleum ether, ethyl 88 acetate, and hexane at 4º C. After 72 h the extracts were filtered and concentrated on rotary 89 evaporator under reduced pressure at 30 °C. Then, the crude concentrated extracts for every 90 solvent were completed with methanol to final volume and then subjected to phytochemical 91 analysis (Harborne 1998) after discharging their colors by using active charcoal. The 92 phytochemical analysis was carried out qualitatively to determine the suitable solvent for the 93 maximum quantitative estimation of nutraceuticals secondary metabolites. 94 Preliminary phytochemical study 95 Preliminary qualitative phytochemical screening will give idea about the chemical 96 constituents present in the extract and will help for further investigation. Phytochemical 97 screening was done as explained in literature (Ikhiri et al. 1992; Silva et al. 1993; Harborne 98 1998; Houghton and Raman 1998). 5 https://mc06.manuscriptcentral.com/bcb-pubs Biochemistry and Cell Biology Page 6 of 47 99 Phytochemical properties of the extracts were studied using the following reagents and 100 methods: alkaloids with 10% acetic acid and ammonium hydroxide reagents; flavanoids with the 101 use of aluminum chloride colorimetric method; The total phenolics and tannins were measured 102 using Folin-Ciocalteu method; saponins by using vanillin -HC1 reagent; triterpnes and total 103 proanthocyanidins by using vanillin reagent; carbohydrate with Molish’s and anthrone reagents; 104 glycosides with Baljet’s test. Total antioxidant capacity was assessed
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