Journal of Pharmacology and Texicology 2 (6): 5S ISSN 1816-496% © 2007 Academie Journals 8, 2007 Effect of Sesbania grandiflora on Membrane-bound ATPases in Cigarette Smoke Exposed Rats T.Ramesh, R. Mahesh and V. Hazeenea Begut Department of Siddha Medicine, Faculty of Sciences, Tamil University, ‘Thanjavus-613 005, Tamil Nadu, India Abstract: ‘The aim of present study to assesse the harm effects of ehronie eigaretie smoking on embrane-bouned ATPases a the protective effet of . grandiflora in rat ang, live, kidney and heart. Adult male WKY rats were exposed to cigarette smoke fora period ‘of 80 days and consecutively treated with aqueous suspension of S. grandffora (1000 me. Ke bawiday, po) fora petiod of 3 weeks. The levels of lipid peroxides as marker for evaluating the extent of mernbrane damage, the activities of Na-K-ATPase, Ca*-ATPase and Ma*-ATPase and associated cations sodium (Na), potassium (K"), ealeum (Ca) and ‘magnesium (Mg*) were investigated in rat mg, liver, kidney and heart, Membrane damage was evident ftom the increased levels of lipid peroxides, dercased aetivides of membrane bound ATPases and alterations in the levels of inorganic cations were observed in cigarette smoke exposed rats, Administration of aqueous suspension of $. grandiflora (ASSG) inhibited the levels of Lipid peroxides, ameliorated the activities of membrane-bound ATPases and maintained the ionic equilibrium in rats exposed to cigarete smoke. The results of our study indicate that ASSG protets the membrane-bound ATPases fiom cigarette smoking induced membrane damage Key words: Cigarette stoking lipid peroxidation, membrane-bound ATPases, Sesbania grandiflor INTRODUCTION Cigarette smoking is associated with various pathological conitions including pulmonary, cardio and cerebrovascular diseases, cancers and several others (US Department, 2001; Centers for Disease, 2004), Cigarette smoke is a complex mixture of over 4000 identified constituents (Genbacey-Krtoica, 2005) tha include numerous reactive substances such as a Tange quantity of reactive aldehydes (Pak ea, 1998), ee radical species, such as oxygen fee radeals and nitrogen species (Pryor and Stone, 1993) and diverse motals such as eadmium (Cé") (WHO, 1992), Free ‘radicals and other reactive oxygen and nitrogen species (ROS and NOS, respectively) are capable of initiating or promoting oxidative damage (Cross fa, 1993; Panda ofa, 1999), The membrane-bound ‘ATPases such a3 Na’- K” ATPase, Ca ATPase and Mg” ATPase contibutes the maintenance of ‘vascular homeostasis via facilitating transport of sodium, potassium, calcium and magnesium ions across the cell membranes (Stekhoren and Renting, 1981), It is reported that elronic exposure 10 cigarette smoke inhibited the activities of all these ATPases, alter the vascular homeostasis and hence ‘contribute to dovelopment of vascular disease (Anbarasi eta, 2005), Increased oxidative stress ‘coupled to increased production of superoxide anions may play a eritial tle in these processes (Pryor etal, 1983; Church and Pryor, 1985), Superoxide anions ean affect the membrare lipids by abstracting hydrogen atoms to form lpi free radicals, which in tum reaets with an oxygen molecule ‘Corresponding Autor: T. Romesh Depatinent o Fannactog, Schoo! of Deis Kyi Hee Unive Sea Sen 1-71 Te TSF 002295759J. Pharmacol Toxicol, ? (6): 359-566, 2 to give a lipid peroxy radical (Halliwell and Gutteridge, 1984), Peroxidation of membrane lipids initiates the loss of membrane integrity and membrane-bound enzyme activites and hence leads to impairment in cellular homeostasis CToskulkar and Glinsukon, 1992). Indeed, treatment with antioxidant micronutrients and vitanins modulated cigarette smoke iniuced lipid peroxidation, suggesting a pathological role foe the fre radicals in smoking related impairment in the atiities of ‘membrane bound ATPases and in cellular homeostasis (Tiwari, 2004). ‘Sesbania grandiflora L pers Febaceae), commonly known as “seshani an Sagathi’ has been ‘used as an important dietary nutritive source in Southeast Asian countries (Fetantinas, 1990-1991). S. grandiflora eaves are richest source of amino acids, minerals and antioxidant vitamins (The wealth, ‘of india, 1972; Govindan and Stanmgasinearam, 1987), Various parts of this plant are used in Indian ‘waditiona medicine for the treatment of a broad spectrum of illness insluding leprosy, gout, sheumatism and liver disorders Joshi, 2000; Vijayakumar ofa, 1907; Pasi and Uma, 2003). It also ‘has anxiolytic and anticonvulsive (Kasture era, 2002), ant-nflammatory, analgesic and antipyretic activity (Tamboli, 1996, 2000). Besides, S. grandorais mentioned as a potent antidote for tobacco and smoking-related diseases (Murugesan, 1988), Recently our study reported that S, grandiflora has hhypolipidemie property on cigarette smoke exposed rats (Ramesh and Hazeena Begum, 2006). Horever, the mechanisms underlying its beneficial effects against smoking associated diseases ars to ‘be Milly lucidlted. Inthe present study, we investigated the effets of $ grandiflora on activities ‘of membrane-bound ATPases in cigarette smoke exposed ats, MATERIALS AND METHODS: Chemicals ‘Adznosine-Tri-Phosphate (ATP) was obtained from the Sigma chemicals company (MO, StLouis, USA). All olher chemicals and solvents uilized inthis sty were purehused from (Glaxo Laboratories (P) Ltd. (Mumbai, india) Pant Matertal Frosh Sesbansa granalora leaves ware collected tem local plantation (Poovathur, Thanjavur, India), ‘The leaves were washed for any contaminants, died thoroughly under shade and powdered Finely, The powdsted leaves of S. grandiflora were reconstituted in disilled water to form a suspension, The aqueous suspension of Sezbanta grandiflora (ASSG) leaves was prepared Geshly every day piot to administation, Experimental Animats Male Wistar-Kyoto (WKY) rats weighing 125-150 g were obtained from Venkateshwara Animal Breeding Centre, Bangalore, Ini, All nim experiments and maintenance were carried out according tothe ethical guidelines suggested by the Institutional Animal Ethics Comittee. Animals were housed ‘inpolypropylene eages with ler tops under conzolled conditions of a 12h Hight’ 12 dark eyele andl 272°C, All the rats received standard pallet diet neut rat feed, Pune, India) and water ad libitt, Experimental Protocol! ‘The animals were divided ino four groups of six animals each, Group I; Control. Group II: Rats administred with ASSG (1000 mg kg and bw/day, p.0) fora period of three wooks, Group Ilr Rats expesed fo cigar smoke, Group IV; Rls expose to cigacit smoke an eonsccutvely administered ‘with ASSG (1000 mg kg! and b widay, po) fora period of tee weeks. Group TIL and Group IV rats were exposed to cigarette smoke by modified method of Eua-Mi etal. (1998) as fllows. ‘The rats were placed in « polypropylene cage with a lid made of polythene paper. A lighted igurett was placed ina Mask connected 10 the cage and wt Was supplied into the Mask foe 10 min by 360J. Pharmacol Toxicol, ? (6): 359-566, 2 small sir pump. A length of 39 cm ofeach cigarette was allowed tobe burned by clamping the butt ‘when it was placed in a flask. Each rat was subjected to inhalation of cigarette smoke seven times a lay at regular intervals of 1 (fom 11 AM to 5 PM) fora pericel of 90 days, Control rats were tested as similar exposed to ir instead of smoke. At the end ofthe experimental perio, the animals were sacrificed by cervical decapitation. Blood samples collected in plain tubes were centrifuged at 3000 xg (4°C) for 10min fo tain serum, Lugs, live, kidney and heart were isolated, cleaned of adhering fat and conneetive tissues, Known weight of tissues were homogenized in 0.1 M tris- HCI bulfer (pH 7.4) containing 0.25 M suerose and used for the Biochemical estimation Concentration of Conjugated Diens (CD) and hycroperoxides were estimated as a measure of pid peroxidation by the method of Recknagel and Ghost (1966) and Mairand Hall (1977), The achvites of Nat = KC ATPase (Boning, 1970), Ca®* ATPase (Hjerten and Pan, 1983) and Mg “ATPase (Ohnishi er al, 1982) were measured by the methods given previously. The amount of inorganic phosphomis was determined by the method of Fiske and Subbarow (1925). Sodium (Na) and potassium (K*) levels were analyzed in serum by the method of Butterworth (1951) and Barry and Rowland (1953). Concentrations of ealcium (Ca) and magnesium (Mg) inthe serum as well asin the tissues after digestion with nitic acid and petehlonic acid were measured by using atomic sorption spectrophotomter according to the method of Ballentine and Burford (1957). ‘Statistical Analysis Resulsare expressed as meaniSD (n=6). The observed differences were analyze! for statistical Significance by One-way of the analysis of variance with Tukey's multiple comparison as posttest. RESULTS: A significant increas in the concentration of hydreperaxides and conjugated diens were observed. in cigarette smoke exposed rats (Group 3) as compared to eatrol (Group 1) rats (Table 1). These levated levels were significantly reversed to near oontzl in rats expesed to cigarette smoke and treated ‘with ASSG (Group 4) when compared with Group 3 rats. ASSG alone treated rats (Group 2) did not show any Significant changes in the convention of hydroperoxides and conjugal dens ws computed to control (Group 1) rls. A significant decrease was observed in cigarette smoke exposed rats (Group 3) as compared to ccontral (Group 1) rats (Table 2). Group 4 rats showed significantly increased activities of Na’= K° ‘ATPase, Ca ATPase and Mg” ATPase when compared with Group 3 rats. Significant changes were nt observed inthe activtis of Na’- K” ATPase, Ca” ATPase and Mg ATPase in Group 2 rts as ‘compared to Group I ral, “Table Bcf § grr on Conjugated anes and Hydro contra experimental roy of rt Parmer Ta Live Kidney Hest Conjugated dees row? ss1e:081 erage asoio.as sora Growl sigT0 80 Gauss 8710.28 70.022 ro 43sost 12% reaped 1 Dxni0 29 a.so.dore Show TV sung eas oan. Hyeroperestaes Growl 213028 Sonus Sra Hop 16 ‘409e0.08 row 2am ong 050 Seow ieee 26 Senin ase 2 Vales are earesed x mea5D n= 6), Units Contd Denes wl vdropoxies anol 10g ts, Stati Comrganso are made between Grup Iv Grp I and Geo ls Gop x Gro "9-009, *pe00) 361