[Downloaded free from http://www.ijpvmjournal.net on Tuesday, June 25, 2019, IP: 94.199.136.196] Original Article Effect of Harmine on Nicotine‑Induced Kidney Dysfunction in Male Mice Abstract Mohammad Reza Background: The nicotine content of cigarettes plays a key role in the pathogenesis of kidney Salahshoor, disease. Harmine is a harmal‑derived alkaloid with antioxidant properties. This study was Shiva Roshankhah, designed to evaluate the effects of harmine against nicotine‑induced damage to the kidneys of mice. Methods: In this study, 64 male mice were randomly assigned to eight groups: saline and Vahid Motavalian, nicotine‑treated groups (2.5 mg/kg), harmine groups (5, 10, and 15 mg/kg), and nicotine (2.5 mg/ Cyrus Jalili kg) + harmine‑treated groups (5, 10, and 15 mg/kg). Treatments were administered intraperitoneally Department of Anatomical daily for 28 days. The weights of the mice and their kidneys, kidney index, glomeruli characteristics, Sciences, Medical School, thiobarbituric acid reactive species, antioxidant capacity, kidney function indicators, and serum Kermanshah University of nitrite oxide levels were investigated. Results: Nicotine administration significantly improved kidney Medical Sciences, Daneshgah Ave., Taghbostan, malondialdehyde (MDA) level, blood urea nitrogen (BUN), creatinine, and nitrite oxide levels and Kermanshah, Iran decreased glomeruli number and tissue ferric reducing/antioxidant power (FRAP) level compared to the saline group (P < 0.05). The harmine and harmine + nicotine treatments at all doses significantly reduced BUN, kidney MDA level, creatinine, glomerular diameter, and nitrite oxide levels and increased the glomeruli number and tissue FRAP level compared to the nicotine group (P < 0.05). Conclusions: It seems that harmine administration improved kidney injury induced by nicotine in mice. Keywords: Harmine, kidney, nicotine Introduction metabolizing organ that discharges toxins in the urine. The primary or secondary Consumption of nicotine in the form of discharge of some toxins and drugs may cigarettes or other nicotine‑based products, result in chronic kidney disorders.[7] such as the hookah or pipe, is prevalent, Smoking cigarettes has a negative effect and nicotine addiction is the main reason on the kidneys. Cigarettes can increase [1] for tobacco consumption. As a member of albumin discharge and proteinuria and Solanaceae family, nicotine is an alkaloid cause kidney malfunction.[8] Antioxidant formed by pyridine and pyrrolidine loops supplements and antioxidant‑rich foods [2] found in the tobacco plant. Oxidative stress can reduce free radicals in the human body and an increase in lipid oxidase production and decrease oxidative damage.[9] Peganum occur following nicotine injection and may harmala is a member of the Zygophyllaceae lead to irreversible damage of the cellular family. It grows in many countries in membrane.[3] Increased levels of oxidative North Africa and the Middle East. It stress result in an imbalance in free radical contains alkaloids which are generally generation and antioxidant defense. This found in its seeds and root.[10] Harmine and Address for correspondence: imbalance, in turn, results in the oxidation harmaline are the most important alkaloids Dr. Cyrus Jalili, of biomolecules and changes their structure Medical Biology Research available in P. harmala that have positive Center, Kermanshah University [4] [11] and function. Gopaul et al. showed effects. The essence of this plant can be of Medical Sciences, that the lipid oxidase level increases used to treat bradycardia, decrease blood Kermanshah, Iran. in cigarette smokers.[5] The increased pressure, control angiogenesis, and has E‑mail: [email protected] level of reactive oxygen species (ROS) antiallergic, antispasm, and antiadrenergic effects.[12] Traditional medicine has listed causes oxidative stress and induces DNA Access this article online breakage. In turn, as particular proteins its positive effects as being a soporific Website: become deactivated, biological membranes emmenagogue, appetite inducer, and www.ijpvmjournal.net/www.ijpm.ir [13] degenerate.[6] Kidney is the main antiparasitic. Harmine is an active DOI: component of P. harmala and is a herbal 10.4103/ijpvm.IJPVM_85_18 alkaloid of the beta‑carboline family. Quick Response Code: This is an open access journal, and articles are distributed under the terms of the Creative Commons Attribution‑NonCommercial‑ShareAlike 4.0 License, which allows others to remix, tweak, and build upon the work How to cite this article: Salahshoor MR, non‑commercially, as long as appropriate credit is given and Roshankhah S, Motavalian V, Jalili C. Effect of the new creations are licensed under the identical terms. harmine on nicotine‑induced kidney dysfunction in For reprints contact: [email protected] male mice. Int J Prev Med 2019;10:97. © 2019 International Journal of Preventive Medicine | Published by Wolters Kluwer - Medknow 1 [Downloaded free from http://www.ijpvmjournal.net on Tuesday, June 25, 2019, IP: 94.199.136.196] Salahshoor, et al.: Effect of harmine on nicotine‑induced kidney dysfunction It is extracted from P. harmala and is known to have ether for 24 h. They were anesthetized due to inhalation pharmacologic effects, especially as an antioxidant.[14] It is of ether fume. Venipuncture from the animals’ hearts (right a strong controller of tyrosine phosphorylation‑regulated ventricle) was done. The blood sample was incubated kinase and shows cytotoxic activity against tumor cells for 15 min at 37°C to clot. Then the clotted blood was in the human body.[15] Harmine can induce apoptosis centrifuged for 15 min at 3000 rpm until the serum was and regulate transcription factors and preinflammatory separated. The separated serum was stored at a temperature cytokines.[16] In addition, it can suppress tumor necrosis of −70°C for measurement of the nitrite oxide, creatinine, factor‑α activity as well as nitrite oxide production.[17] and blood urea nitrogen (BUN). The mice were then Nitrite oxide is a signaling molecule that plays a significant sacrificed. The kidneys were removed and weighed on role in biological systems. Nicotine absorption in the body a microbalance sensitive up to 0.001 mg (Precisa 125A; appears to increase the serum levels of nitrite oxide and Switzerland) and the average weights of the kidneys were oxidative stress.[18] Nicotine has a toxic effect and harmine calculated and recorded. The renal index was derived from has advantageous properties.[19] No study has evaluated the the division of the total weight of the two left and right antioxidant effect of harmine on kidney damage induced by kidneys into the total weight of the mice.[9] nicotine. The current study was designed to evaluate the Histological and morphometric analysis effect of harmine on kidney disorders and damage induced by nicotine in animal samples. The kidney was divided into two equal parts from the middle by a cross‑section after the samples were fixed Methods by 10% formalin solution and washed. The central part was immersed in 70% alcohol, and the process of tissue Chemicals preparation was based on the conventional histology In this experimental study, harmine (7‑methoxy‑1 method. Serial sections were prepared (5 µm thick) using methyl‑9h‑pyrido (C10H14N2)) powder was purchased a microtome (EC350‑2). Hematoxylin and eosin staining (from Sigma, USA). The powder was diluted with methods were used to stain the nucleus purple and the saline (0.9%) to obtain the desired doses. Nicotine (CAS cytoplasm pink. The diameter and number of glomeruli Name:3‑[(2S)‑1‑methyl‑2‑pyrrolidinyl]pyridine) were examined under the Olympus BX‑51T‑32E01 was purchased from Sigma (USA) and dissolved in microscope linked to a DP12 camera with 3.34‑million saline (0.9%) for administration.[1,14] pixel resolution and Olysia Bio software (Olympus Optical, [6] Experimental animals Japan). A total of 64 male BALB/c mice weighting 27–30 g were Biochemical marker assays purchased from Razi Institute (Iran). All the mice were The blood taken from the heart was incubated at 37°C for housed in plastic cages in a room at 23 ± 2°C, under 15 min and centrifuged at 3000 rpm for 15 min to acquire controlled environmental conditions, a 12/12 h light/dark the serum. The serum samples were kept in a ‑20°C cycle, with free access to water and food. All investigations freezer. Plasma samples were assayed for concentrations conformed to the ethical and humane principles of of creatinine and BUN using an autoanalyzer (RA 1000; research and were approved by the Ethics Committee Technicon Instruments, USA).[21] of Kermanshah University of Medical Sciences (Ethics Griess technique Certificate No. 1395.38).[6] Griess technique uses zinc sulfate powder to eliminate Protocol and treatments the serum protein of the samples. Accordingly, The animals were randomly separated into eight groups zinc sulfate powder (6 mg) was mixed with serum (n = 8). Group 1 was for saline (normal saline; 1 ml DW/daily); samples (400 µl), and vortexed for 1 min. After group 2 was for nicotine (2.5 mg/kg);[1] groups 3 to 5 were centrifugation of the samples (10 min at 12,000 rpm), the treated with harmine in dosages of 5, 10, and 15 mg/kg, supernatant was used to measure the nitrite oxide. Briefly, 50 respectively;[20] groups 6 to 8 were treated with harmine in µl of sample was added to 100 µl of Griess reagent (Sigma, dosages of 5, 10, and 15 mg/kg, and nicotine (2.5 mg/kg). USA) and the reaction mixture was incubated for about Nicotine was administered intraperitoneally (IP) daily for 30 min at room temperature. The sample optical density 4 weeks. Harmine and harmine + nicotine were administered was measured by enzyme‑linked immunosorbent assay in the same manner. reader (Hyperion, USA) at a wavelength of 450 nm according to the manufacturer protocol.[1] Weight of mice, kidney, kidney index, and collection of blood serum Ferric reducing/antioxidant power (FRAP) and malondialdehyde (MDA) assays The body weight was measured at the onset and end of the study.