Effect of quercetin on the IRE1-XBP1 apoptotic pathway in Cadmium-treated rat testes

Efecto de la quercetina en la vía apoptótica IRE1-XBP1 en testículos de rata tratados con Cadmio

Background: Cadmium (Cd) is a recognized toxic metal with serious reproductive toxicity in both animals and humans, while quercetin (Que) is considered the potent antioxidant among flavonoid compounds. However, little is known about the alleviating effect of Que on Cd-induced testicular cell apoptosis. This experiment aims to investigate the alleviating effect of Que on Cd-induced testicular cell apoptosis in rats. Methods: Twenty-four four-week-old male Sprague-Dawley rats were randomly divided into control group, Cd group, Cd + Que group and Que group. Following the assigned treatments, the testicular tissues of the rats were examined 28 days later. The levels of malondialdehyde (MDA) and reducing glutathione (GSH) in testicular tissue were measured via colorimetry, with HE (hematoxylin and eosin) and TUNEL (a TdT-UTP nick end labeling) staining employed to assess tissue damage and cell apoptosis. Total mRNA (messenger RNA) was extracted from testicular tissue using the Trizol method, and reverse transcription was followed by quantitative real-time PCR (Polymerase Chain Reaction) to measure the expression levels of relevant genes. The expression of related proteins was assessed using Western blot. Results: The data revealed that Cd exposure led to a decrease in body weight and a significant increase in MDA and GSH content in testicular tissue. In addition, histopathological examination of the tissue revealed extensive pathological changes, and TUNEL staining showed significant cell apoptosis in the tissue. Furthermore, Cd treatment promoted the expression of relevant genes, including IRE1α (inositol-requiring kinase 1), Caspase-12, XBP1 (X box-binding protein 1), GRP78 (glucose-regulated protein 78), Bax and Caspase-3, in the IRE1-XBP1 apoptosis pathway. Meanwhile, the anti-apoptotic gene Bcl-2 was significantly decreased. However, the application of Que significantly reduced these alterations and cellular damage induced by Cd. Conclusions: Our study suggests that Que can mitigate testicular tissue damage and cell apoptosis resulting from Cd exposure by suppressing oxidative stress and the IRE1-XBP1 pathway.

Resumen

Antecedentes: El cadmio (Cd) es un metal tóxico reconocido con grave toxicidad reproductiva tanto en animales como en humanos, mientras que la quercetina (Que) se considera el potente antioxidante entre los compuestos flavonoides. Sin embargo, poco se sabe sobre el efecto paliativo de Que en la apoptosis de células testiculares inducida por Cd. Este experimento tiene como objetivo investigar el efecto paliativo de Que en la apoptosis de células testiculares inducida por Cd en ratas. Métodos: Veinticuatro ratas macho Sprague-Dawley de cuatro semanas de edad fueron aclimatadas durante una semana y divididas aleatoriamente en grupo control, grupo Cd, grupo Cd + Que y grupo Que. Tras los tratamientos asignados, se examinaron los tejidos testiculares de las ratas 28 días después. Los niveles de malondialdehído (MDA) y glutatión reductor (GSH) en el tejido testicular se midieron mediante colorimetría, y se emplearon las tinciones HE (hematoxilina y eosina) y TUNEL (a TdT-UTP nick end labeling) para evaluar el daño tisular y la apoptosis celular. Se extrajo el ARNm total (ARN mensajero) del tejido testicular mediante el método Trizol y se realizó la transcripción inversa seguida de la PCR cuantitativa en tiempo real (reacción en cadena de la polimerasa) para medir los niveles de expresión de los genes relevantes. Resultados: Los datos revelaron que la exposición al Cd provocó una disminución del peso corporal y un aumento significativo del contenido de MDA y GSH en el tejido testicular. Además, el examen histopatológico del tejido reveló cambios patológicos extensos, y la tinción TUNEL mostró una apoptosis celular significativa en el tejido. Además, el tratamiento con Cd fomentó la expresión de genes relevantes, como IRE1α (inositol-requiring kinase 1), Caspasa-12, XBP1 (X box-binding protein 1), GRP78 (glucose-regulated protein 78), Bax y Caspasa-3, en la vía de apoptosis IRE1-XBP1. Mientras tanto, el gen anti-apoptótico Bcl-2 disminuyó significativamente. Sin embargo, la aplicación de Que redujo significativamente estas alteraciones y el daño celular inducido por el Cd. Conclusiones: Nuestro estudio sugiere que Que puede mitigar el daño testicular y la apoptosis celular resultantes de la exposición al Cd mediante la supresión del estrés oxidativo y de la vía IRE1-XBP1.

Cadmium; Testis; Toxicity; Apoptosis; Quercetin

Palabras Clave

Cadmio; Testículo; Toxicidad; Apoptosis; Quercetina

[1] Dharmadasa P, Kim N, Thunders M. Maternal cadmium exposure and impact on foetal gene expression through methylation changes. Food and Chemical Toxicology. 2017; 109: 714–720.

[2] Jigyasu AK, Siddiqui S, Jafri A, Arshad M, Lohani M, Khan IA. Biological synthesis of cdte quantum dots and their anti-proliferative assessment against prostate cancer cell line. Journal of Nanoscience and Nanotechnology. 2020; 20: 3398–3403.

[3] Liu Y, Yuan Y, Xiao Y, Li Y, Yu Y, Mo T, et al. Associations of plasma metal concentrations with the decline in kidney function: a longitudinal study of Chinese adults. Ecotoxicology and Environmental Safety. 2020; 189: 110006.

[4] Stayner L, Smith R, Thun M, Schnorr T, Lemen R. A quantitative assessment of lung cancer risk and occupational cadmium exposure. IARC Scientific Publications. 1992; 447–455.

[5] Zhu Q, Li X, Ge RS. Toxicological effects of cadmium on mammalian testis. Frontiers in Genetics. 2020; 11: 527.

[6] Wang M, Chen Z, Song W, Hong D, Huang L, Li Y. A review on cadmium exposure in the population and intervention strategies against cadmium toxicity. Bulletin of Environmental Contamination and Toxicology. 2021; 106: 65–74.

[7] Peng L, Huang YT, Zhang F, Chen JY, Huo X. Chronic cadmium exposure aggravates malignant phenotypes of nasopharyngeal carcinoma by activating the Wnt/β-catenin signaling pathway via hypermethylation of the casein kinase 1α promoter. Cancer Management and Research. 2019; 11: 81–93.

[8] Lan Y, Hu L, Feng X, Wang M, Yuan H, Xu H. Synergistic effect of PS-MPs and Cd on male reproductive toxicity: ferroptosis via Keap1-Nrf2 pathway. Journal of Hazardous Materials. 2024; 461: 132584.

[9] de Angelis C, Galdiero G, Menafra D, Garifalos F, Verde N, Piscopo M, et al. The environment and male reproductive system: the potential role and underlying mechanisms of cadmium in testis cancer. Critical Reviews in Toxicology. 2023; 53: 412–435.

[10] Zhang Z, Wang Q, Gao X, Tang X, Xu H, Wang W, et al. Reproductive toxicity of cadmium stress in male animals. Toxicology. 2024; 504: 153787.

[11] Gao X, Li G, Pan X, Xia J, Yan D, Xu Y, et al. Environmental and occupational exposure to cadmium associated with male reproductive health risk: a systematic review and meta-analysis based on epidemiological evidence. Environmental Geochemistry and Health. 2023; 45: 7491–7517.

[12] Pizent A, Tariba B, Živković T. Reproductive toxicity of metals in men. Archives of Industrial Hygiene and Toxicology. 2012; 63: 35–46.

[13] Machado-Neves M. Effect of heavy metals on epididymal morphology and function: an integrative review. Chemosphere. 2022; 291: 133020.

[14] Zhang Q, Yang Y, Liu J, Wu Y, Liu Y, Zhang J. Testicular dysfunction and “its recovery effect” after cadmium exposure. Food and Chemical Toxicology. 2024; 188: 114656.

[15] Ikokide EJ, Oyagbemi AA, Oyeyemi MO. Impacts of cadmium on male fertility: lessons learnt so far. Andrologia. 2022; 54: e14516.

[16] Fitzgerald R, Olsen A, Nguyen J, Wong W, El Muayed M, Edwards J. Pancreatic islets accumulate cadmium in a rodent model of cadmium-induced hyperglycemia. International Journal of Molecular Sciences. 2020; 22: 360.

[17] Templeton DM, Liu Y. Multiple roles of cadmium in cell death and survival. Chemico-Biological Interactions. 2010; 188: 267–275.

[18] Djulejic V, Petrovic B, Jevtic J, Vujacic M, Clarke BL, Cirovic A, et al. The role of cadmium in the pathogenesis of myeloid leukemia in individuals with anemia, deficiencies in vitamin D, zinc, and low calcium dietary intake. Journal of Trace Elements in Medicine and Biology. 2023; 79: 127263.

[19] Fujiwara Y, Lee JY, Banno H, Imai S, Tokumoto M, Hasegawa T, et al. Cadmium induces iron deficiency anemia through the suppression of iron transport in the duodenum. Toxicology Letters. 2020; 332: 130–139.

[20] Bi SS, Talukder M, Sun XT, Lv MW, Ge J, Zhang C, et al. Cerebellar injury induced by cadmium via disrupting the heat-shock response. Environmental Science and Pollution Research International. 2023; 30: 22550–22559.

[21] Winiarska-Mieczan A. Protective effect of tea against lead and cadmium-induced oxidative stress-a review. BioMetals. 2018; 31: 909–926.

[22] Zwolak I. The role of selenium in arsenic and cadmium toxicity: an updated review of scientific literature. Biological Trace Element Research. 2020; 193: 44–63.

[23] Bhardwaj JK, Panchal H, Saraf P. Cadmium as a testicular toxicant: a review. Journal of Applied Toxicology. 2021; 41: 105–117.

[24] Wang J, Ding L, Wang K, Huang R, Yu W, Yan B, et al. Role of endoplasmic reticulum stress in cadmium-induced hepatocyte apoptosis and the protective effect of quercetin. Ecotoxicology and Environmental Safety. 2022; 241: 113772.

[25] Liu J, Luo LF, Wang DL, Wang WX, Zhu JL, Li YC, et al. Cadmium induces ovarian granulosa cell damage by activating PERK-eIF2α-ATF4 through endoplasmic reticulum stress. Biology of Reproduction. 2019; 100: 292–299.

[26] Rafati Rahimzadeh M, Rafati Rahimzadeh M, Kazemi S, Moghadamnia AA. Cadmium toxicity and treatment: an update. Caspian Journal of Internal Medicine. 2017; 8: 135–145.

[27] Chen J, Pan T, Wan N, Sun Z, Zhang Z, Li S. Cadmium-induced endoplasmic reticulum stress in chicken neutrophils is alleviated by selenium. Journal of Inorganic Biochemistry. 2017; 170: 169–177.

[28] Li X, Ge M, Zhu W, Wang P, Wang J, Tai T, et al. Protective effects of astilbin against cadmium-induced apoptosis in chicken kidneys via endoplasmic reticulum stress signaling pathway. Biological Trace Element Research. 2022; 200: 4430–4443.

[29] Andres S, Pevny S, Ziegenhagen R, Bakhiya N, Schäfer B, Hirsch-Ernst KI, et al. Safety aspects of the use of quercetin as a dietary supplement. Molecular Nutrition & Food Research. 2018; 62: 1700447.

[30] Moskaug JØ, Carlsen H, Myhrstad M, Blomhoff R. Molecular imaging of the biological effects of quercetin and quercetin-rich foods. Mechanisms of Ageing and Development. 2004; 125: 315–324.

[31] D’Andrea G. Quercetin: a flavonol with multifaceted therapeutic applications? Fitoterapia. 2015; 106: 256–271.

[32] Badr GM, Elsawy H, Sedky A, Eid R, Ali A, Abdallah BM, et al. Protective effects of quercetin supplementation against short-term toxicity of cadmium-induced hematological impairment, hypothyroidism, and testicular disturbances in albino rats. Environmental Science and Pollution Research International. 2019; 26: 8202–8211.

[33] Ding L, Zhu H, Wang K, Huang R, Yu W, Yan B, et al. Quercetin alleviates cadmium-induced BRL-3A cell apoptosis by inhibiting oxidative stress and the PERK/IRE1α/ATF6 signaling pathway. Environmental Science and Pollution Research International. 2023; 30: 125790–125805.

[34] Wang J, Zhu H, Wang K, Yang Z, Liu Z. Protective effect of quercetin on rat testes against cadmium toxicity by alleviating oxidative stress and autophagy. Environmental Science and Pollution Research International. 2020; 27: 25278–25286.

[35] Huang R, Ding L, Ye Y, Wang K, Yu W, Yan B, et al. Protective effect of quercetin on cadmium-induced renal apoptosis through cyt-c/caspase-9/caspase-3 signaling pathway. Frontiers in Pharmacology. 2022; 13: 990993.

[36] Unsal V, Dalkıran T, Çiçek M, Kölükçü E. The role of natural antioxidants against reactive oxygen species produced by cadmium toxicity: a review. Advanced Pharmaceutical Bulletin. 2020; 10: 184–202.

[37] Yang W, Liu R, Sun Q, Huang X, Zhang J, Huang L, et al. Quercetin alleviates endoplasmic reticulum stress-induced apoptosis in buffalo ovarian granulosa cells. Animals. 2022; 12: 787.

[38] Iqbal T, Cao M, Zhao Z, Zhao Y, Chen L, Chen T, et al. Damage to the testicular structure of rats by acute oral exposure of cadmium. International Journal of Environmental Research and Public Health. 2021; 18: 6038.

[39] Wang YJ, Yan J, Yin F, Li L, Qin YG, Meng CY, et al. Role of autophagy in cadmium-induced testicular injury. Human & Experimental Toxicology. 2017; 36: 1039–1048.

[40] Shukla P, Sahu NK, Kumar R, Dhalla DK, Rakshit S, Bhadauria M, et al. Quercetin ameliorates acute acrylamide induced spleen injury. Biotechnic & Histochemistry. 2023; 98: 221–229.

[41] Oyovwi MO, Oghenetega OB, Victor E, Faith FY, Uchechukwu JG. Quercetin protects against levetiracetam induced gonadotoxicity in rats. Toxicology. 2023; 491: 153518.

[42] Burukoğlu D, Bayçu C. Protective effects of zinc on testes of cadmium-treated rats. Bulletin of Environmental Contamination and Toxicology. 2008; 81: 521–524.

[43] Beltrame FL, Cerri PS, Sasso-Cerri E. Cimetidine-induced Leydig cell apoptosis and reduced EG-VEGF (PK-1) immunoexpression in rats: evidence for the testicular vasculature atrophy. Reproductive Toxicology. 2015; 57: 50–58.

[44] Alizadeh B, Salehzadeh A, Ranji N, Arasteh A. Effects of N-Acetyl cysteine on genes expression of c-myc, and Ask-1, histopathological, oxidative stress, inflammation, and apoptosis in the liver of male rats exposed to cadmium. Biological Trace Element Research. 2022; 200: 661–668.

[45] Matović V, Buha A, Ðukić-Ćosić D, Bulat Z. Insight into the oxidative stress induced by lead and/or cadmium in blood, liver and kidneys. Food and Chemical Toxicology. 2015; 78: 130–140.

[46] Cirmi S, Maugeri A, Micali A, Marini HR, Puzzolo D, Santoro G, et al. Cadmium-induced kidney injury in mice is counteracted by a flavonoid-rich extract of bergamot juice, alone or in association with curcumin and resveratrol, via the enhancement of different defense mechanisms. Biomedicines. 2021; 9: 1797.

[47] Spiazzi CC, Manfredini V, Barcellos da Silva FE, Flores EM, Izaguirry AP, Vargas LM, et al. γ-Oryzanol protects against acute cadmium-induced oxidative damage in mice testes. Food and Chemical Toxicology. 2013; 55: 526–532.

[48] Francis Stuart SD, Villalobos AR. GSH and zinc supplementation attenuate cadmium-induced cellular stress and stimulation of choline uptake in cultured neonatal rat choroid plexus epithelia. International Journal of Molecular Sciences. 2021; 22: 8857.

[49] Capriglione F, Maiuolo J, Celano M, Damante G, Russo D, Bulotta S, et al. Quercetin protects human thyroid cells against cadmium toxicity. International Journal of Molecular Sciences. 2021; 22: 6849.

[50] Song Y, Zhang R, Wang H, Yan Y, Ming G. Protective effect of agaricus blazei polysaccharide against cadmium-induced damage on the testis of chicken. Biological Trace Element Research. 2018; 184: 491–500.

[51] Karri V, Schuhmacher M, Kumar V. Heavy metals (Pb, Cd, As and MeHg) as risk factors for cognitive dysfunction: A general review of metal mixture mechanism in brain. Environmental Toxicology and Pharmacology. 2016; 48: 203–213.

[52] Fayazipour D, Deckert J, Akbari G, Soltani E, Chmielowska-Bąk J. Mitochondria specific antioxidant, MitoTEMPO, modulates Cd Uptake and oxidative response of soybean seedlings. Antioxidants. 2022; 11: 2099.

[53] Fu X, Cui J, Meng X, Jiang P, Zheng Q, Zhao W, et al. Endoplasmic reticulum stress, cell death and tumor: association between endoplasmic reticulum stress and the apoptosis pathway in tumors (Review). Oncology Reports. 2021; 45: 801–808.

[54] Pincus D, Chevalier MW, Aragón T, van Anken E, Vidal SE, El-Samad H, et al. BiP binding to the ER-stress sensor Ire1 tunes the homeostatic behavior of the unfolded protein response. PLOS Biology. 2010; 8: e1000415.

[55] Bhardwaj M, Leli NM, Koumenis C, Amaravadi RK. Regulation of autophagy by canonical and non-canonical ER stress responses. Seminars in Cancer Biology. 2020; 66: 116–128.

[56] Urra H, Dufey E, Lisbona F, Rojas-Rivera D, Hetz C. When ER stress reaches a dead end. Biochimica et Biophysica Acta. 2013; 1833: 3507–3517.

[57] Li Z, Xu T, Peng L, Tang X, Chi Q, Li M, et al. Polystyrene nanoplastics aggravates lipopolysaccharide-induced apoptosis in mouse kidney cells by regulating IRE1/XBP1 endoplasmic reticulum stress pathway via oxidative stress. Journal of Cellular Physiology. 2023; 238: 151–164.

[58] Zhao L, Gu Q, Xiang L, Dong X, Li H, Ni J, et al. Curcumin inhibits apoptosis by modulating Bax/Bcl-2 expression and alleviates oxidative stress in testes of streptozotocin-induced diabetic rats. Therapeutics and Clinical Risk Management. 2017; 13: 1099–1105.