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Anthracene-Induced Breast Cancer in Female Rats

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Anthracene-Induced Breast Cancer in Female Rats Veterinary World, EISSN: 2231-0916 RESEARCH ARTICLE Available at www.veterinaryworld.org/Vol.14/March-2021/3.pdf Open Access Histopathological evaluation of Senecio rhizomatus Rusby in 7,12-dimethylbenz(α) anthracene-induced breast cancer in female rats Jorge Luis Arroyo-Acevedo1 , Oscar Herrera-Calderon2 , Juan Pedro Rojas-Armas1 , Roberto Chávez-Asmat3 , James Calva4 and Tapan Behl5 1. Laboratory of Experimental Pharmacology, Faculty of Medicine, Universidad Nacional Mayor de San Marcos, Av. Miguel Grau 755, Cercado de Lima 15001, Peru; 2. Department of Pharmacology, Bromatology and Toxicology, Faculty of Pharmacy and Biochemistry, Universidad Nacional Mayor de San Marcos, Jr Puno 1002, Lima 15001, Peru; 3. Association for the Development of Student Research in Health Sciences, Faculty of Medicine. Universidad Nacional Mayor de San Marcos, Av. Miguel Grau 755, Cercado de Lima 15001, Peru; 4. Departamento de Química y Ciencias Exactas, Universidad Técnica Particular de Loja, San Cayetano s/n, 1101608 Loja, Ecuador; 5. Department of Pharmacology, Chitkara College of Pharmacy, Chitkara University, Punjab 140401, India. Corresponding author: Oscar Herrera-Calderon, e-mail: [email protected] Co-authors: JLA: [email protected], JPR: [email protected], RC: [email protected], JC: [email protected], TB: [email protected] Received: 08-10-2020, Accepted: 25-01-2021, Published online: 06-03-2021 doi: www.doi.org/10.14202/vetworld.2021.569-577 How to cite this article: Arroyo-Acevedo JL, Herrera-Calderon O, Rojas-Armas JP, Chávez-Asmat R, Calva J, Behl T (2021) Histopathological evaluation of Senecio rhizomatus Rusby in 7,12-dimethylbenz(α) anthracene-induced breast cancer in female rats, Veterinary World, 14(3): 569-577. Abstract Background and Aim: Senecio rhizomatus Rusby (SrR) is a medicinal plant of the Asteraceae family and traditionally consumed as infusion in the Andean region from Peru for inflammatory disorders. This study aimed to determine the histopathological changes afforded by SrR in 7, 12-dimethylbenz[α]anthracene (DMBA)-induced breast cancer (BC) in rats. Materials and Methods: An ethanolic extract of SrR aerial parts was prepared by maceration with 96% ethanol, and the chemical components were identified by gas chromatography coupled to mass spectrometry; the antioxidant activity was determined by 1,1-diphenyl-2-picril-hidrazil (DPPH) assay; and the acute toxicity was assessed according to the OCED 423 guidelines. In a pharmacological study, 30 female Holztman rats were distributed randomly into five groups, as follows. Group I: Negative control (physiological serum, 2 mL/kg); Group II. DMBA (80 mg/Kg body weight); and Groups III, IV, and V: DMBA + ethanol extract of SrR at doses of 10, 100, and 200 mg/kg, respectively. Results: The antioxidant activity of the SrR extract against DPPH was 92.50% at 200 µg/mL. The oral administration of SrR at doses of 50, 300, 2000, and 5000 mg/kg did not show any clinical evidence of toxicity or occurrence of death. The groups that received SrR presented a lower frequency of tumors and a cumulative tumor volume compared with the DMBA group (p<0.05); the DMBA group exhibited a higher incidence of necrosis and moderate mitosis, up to 66.67% and 100.00%, respectively. Finally, infiltrating carcinoma with extensive tumor necrosis was evidenced. Conclusion: In experimental conditions, the ethanolic extract of SrR had a protective effect in DMBA-induced BC in female rats. Furthermore, the antioxidant activity of its main phytochemicals could be responsible for the effect observed, and SrR seems to be a safe extract in the preclinical phase. Keywords: breast cancer, carcinogenic, experimental pharmacology, phytotherapy, Senecio rhizomatus. Introduction because of its late diagnosis; thus, the establishment Epidemiological studies of cancer revealed that of a comprehensive plan that implements strategies breast cancer (BC) has a high prevalence in women for early diagnosis and timely medical treatment of and, together with lung and colorectal cancer, contrib- positive cases is necessary to reduce mortality [2]. Alternative and Complementary Medicine utes to more than 50% of all cases of cancer. BC rep- (ACM) is part of the folk tradition in several coun- resents 30% of the rate of all new cancer diagnoses, tries and its effectiveness in medical practice is scien- thus being the type of cancer in women and being the tifically proven; thus, ACM is rising worldwide. The second most common main cause of death [1]. In Peru, World Health Organization recommends the incorpo- BC is the second most frequent neoplasm, which has ration of ACM into national health systems [3,4]. In a great economic impact, and has a poor survival rate Peru, a complementary medicine service was imple- Copyright: Arroyo-Acevedo, et al. Open Access. This article is mented in EsSalud (a social health insurance) in 1998 distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/ and offered a wide variety of alternative complemen- by/4.0/), which permits unrestricted use, distribution, and tary therapies, such as herbal therapy [5]. reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the In Northern Peru, 47 species of plants, mostly Creative Commons license, and indicate if changes were made. Asteraceae, followed by Gentianaceae [6,7], are The Creative Commons Public Domain Dedication waiver (http:// creativecommons.org/publicdomain/zero/1.0/) applies to the data traditionally used to manage cancerous conditions. made available in this article, unless otherwise stated. In the American continent, Asteraceae is the richest Veterinary World, EISSN: 2231-0916 569 Available at www.veterinaryworld.org/Vol.14/March-2021/3.pdf family in species in the Southern region (15%), North Nacional Mayor de San Marcos (UNMSM), Lima, America (14%), and Mexico (13%). Moreover, it is Peru. the second most diverse plant family in most of the Plant material Andean and Central American tropical countries [8]. The aerial parts of SrR were collected from After the publication of the catalog of flowering plants Tambo district, Huancayo (Junín- Peru) at the coor- and gymnosperms, 222 genera and 1432 Asteraceae dinates 12°03′01″S - 75°13′17″W at 3260 m.a.s.l. species were registered in Peru; subsequently, the in April 2018. A voucher specimen is kept in the registry was updated to 245 genera and 1530 species, Herbarium of the Universidad Nacional Mayor de San with 14 genera being endemic, that is, Ascidiogyne, Marcos, Peru (Id. 059-USM-2018). The plant was rec- Caxamarca, Ellenbergia, Holoschkuhria, Hughesia, ognized by Hamilton Beltran-Santiago, Biology pro- Nothobaccharis, Uleophytum, Syncretocarpus, fessor and expert in Botany. Bishopanthus, Chionopappus, Pseudonoseris, Chucoa, Schizotrichia, and Aynia [9]. In Lima, the Extraction procedure capital of Peru, about 385 species are known, 343 of The aerial parts of SrR were dried at room tem- which correspond to native and/or naturalized spe- perature (20°C) and then subjected to successive cies and 42 are exclusively from crops, with the pre- extractions using 96% ethanol, similar to a procedure dominant families being Senecioneae, Heliantheae, previously [15]. The extraction of SrR was completed Asteraceae, and Inuleae [10,11]. within 72 h and replicated three times. The extract The Asteraceae family comprises species obtained was concentrated using a rotary evaporator such as Senecio rhizomatus Rusby (SrR; Known as at 40°C and 80 rpm, and finally stored at 4°C in an Llankahuasi), Bidens pilosa L., and Chuquiraga spi- amber flask until further use. nosa Lessing. The previous studies have indicated GC-MS analysis of the volatile components of that the methanolic fraction of B. pilosa stopped the S. rhizomatus progression of 7, 12-dimethylbenz[α]anthracene The analysis of the extract was performed in an (DMBA)-induced BC in rats [12]; moreover, C. spi- Agilent Technologies Chromatograph 6890 N series nosa also exerted a protective effect against DMBA- coupled to a mass spectrometer-detector (Agilent induced BC in rats because of its anti-tumorigenic, series 5973) operated in the electronionization mode hypolipidemic, hypoglycemic, antioxidant, and antig- at 70 eV and fitted with a 5% diphenyl and 95% enotoxic properties [13]. dimethylpolysiloxane capillary column (DB-5 MS, BC is pathology with a high frequency in 30 m × 0.25 mm × 0.25 µm). Helium was used as the women; thus, it is necessary to continue seeking thera- carrier gas (1.00 mL/min in the constant flow mode). peutic alternatives, such as natural products. Thus, the The injection system was operated in split mode (40:1) availability of previous studies of plants in the same at 220°C. The GC oven temperature was 40°C, which family may lead to the exploration of their probable was kept for 3 min, then increased to 270°C with a effects as antitumor agents. Therefore, the aims of this gradient rate of 8°C/min. The ion source temperature study were to (1) determine the protective effect of the was 250°C. One microliter of a solution of the extract ethanolic extract of the aerial parts of SrR in DMBA- diluted in CH2Cl2 (1:100, v/v) was injected into the induced BC in rats; (2) identify its main chemical apparatus. The extract components were identified components using gas chromatography coupled to through a computer search using the Wiley Registry mass spectrometry GC-MS; (3) determine its in vitro of Mass Spectral Data (6th edition), and through com- antioxidant activity against the 1,1-diphenyl-2-pic- parison of the calculated linear retention indices with ril-hidrazil (DPPH) radical; (4) to assess its acute tox- data from the literature NIST. icity at a single dose according to the Organization Quantitative data were obtained from peak areas for Economic Cooperation and Development (OECD) using a flame ionization detector [16].
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