Arsenic Is a Carcinogen with Strong Reproductive and Developmental Toxicity
Arsenic (As) is a naturally occurring toxic metalloid with odorless, colorless, and tasteless properties. The most common forms of arsenic include inorganic As, organic As, and arsenic gas. Arsenic ranks 20th on land, 14th in oceans, and 12th in human ecosystems. Due to its ubiquity, volcanic and industrial activities discharge it into the environment and thus have serious health concerns.
Human exposure to arsenic is increasing from drinking water, food, and industrial sources. Designated by the International Agency for Research on Cancer (IARC) as a Group I carcinogen, it increases the risk of bladder, lung, kidney, and liver cancers.
High levels of arsenic in drinking water have been reported in most countries, above the limits set by WHO and EPA. Polluted drinking water has been linked to developing cancers (skin, lung, bladder, and liver) and other diseases such as diabetes and cardiovascular, gastrointestinal, neurological, and developmental injuries. According to epidemiological studies, As can cause male infertility, sexual dysfunction, poor sperm quality, and developmental consequences, such as low birth weight, spontaneous abortion, and small-for-gestational-age infants (SGA).
Growing rice in arsenic-contaminated soil and using arsenic-contaminated boiled rice are the leading causes of arsenic exposure in rice. Fish, shellfish, and seaweed are the most common sources of human exposure to arsenic through seafood. Urine, hair, and drinking water samples from children and adults distributed in arsenic-contaminated areas in Argentina, Uruguay, India, Pakistan, and Spain showed high levels of arsenic.
Moreover, arsenic is an elemental impurity, and the elemental impurities in medicines may come from the equipment, raw materials, excipients, solvents, and packaging materials used in the pharmaceutical process. Pharmaceutical preparations may introduce an excessive or toxic form of elemental impurities in raw materials and production, which will significantly impact living organisms. Therefore, in the research and development of pharmaceutical preparations, it is necessary to control arsenic impurities in pharmaceutical preparations to ensure the quality and safety of pharmaceuticals.
The inhibition of testosterone biosynthesis by arsenic
Effects of Arsenic on the Male Reproductive System of Animals
Epidemiological studies have shown that arsenic is one of the most dangerous reproductive toxicants in the environment. It accumulates in large quantities in reproductive tissues such as the testes, epididymis, seminal vesicles, and prostate. The scientists conducted experiments in which mice were exposed to As2O3 (0, 0.2, 2, and 20 ppm) five weeks before mating and continued until male pups reached puberty.
Experimental results showed that the HPG axis experienced oxidative stress and increased autophagy after exposure, especially at higher As2O3 doses (2 and 20 ppm). The number of MDC-labeled autophagic vesicles and the MDA/GSH ratio in the HPG axis of adolescent F1 male mice exposed to higher As2O3 doses increased. At the same time, the average body weight, total antioxidant capacity, and stereological indicators decreased. At the same time, in addition to the dose-dependent increase in ATG3, ATG5, Beclin gene expression, and P62, ATG12, and Beclin protein expression, a dose-dependent decrease in PI3K and mTOR gene expression was also observed in pubertal F1 male HPG tissues. Higher doses of As2O3 appear to impair HPG axis function in offspring of adolescent male mice by increasing MDA/GSH ratios, autophagic cell death-related genes and proteins, and decreasing total antioxidant capacity.
Drug reproductive toxicity research is an essential part of non-clinical safety evaluation. It is closely related to acute toxicity, long-term toxicity, genotoxicity, and other toxicological research and is a vital link for drugs to enter clinical research and the market. For drugs intended to be used in humans, reproductive toxicity tests should be considered based on factors such as the test substance’s intended indications and action characteristics.
The Mechanism of Arsenic-Induced Male Reproductive Toxicity in Animals
1. Oxidative stress
Exposure to sodium arsenite at concentrations of 50 and 1000 ppb for 24, 48, and 72 hours caused cytotoxicity and impairment of the antioxidant system in Leydig and Sertoli cells of mouse testis.
Treatment of in vitro cell cultures of rodent testis and epididymis with sodium arsenite at concentrations of 1, 10, 50, and 100 μM for 2 and 24 hours increases ROS, TBARS, and sperm DNA damage and decreases catalase, peroxidase, and superoxide dismutase, lowering serum testosterone. The overexpression of SOD1, CAT, GSTK1, and MT1 in rat testes and epididymis was induced by administering sodium arsenite at a dose of 10 mg/L through drinking water.
Decreased sperm counts, increased caspase-3 activity, increased TUNEL-positive cells, changes in mRNA levels of Bax and Bcl-2, and reduced serum testosterone levels were observed in mice exposed to sodium arsenite and sulfur dioxide. The expression of steroid genes (LHR, StAR, and ABP) was down-regulated, and the doses were 5mg/L and 5mg/m3, respectively, orally by double distilled water for 60 days.
In mice, arsenic trioxide and antimony at doses of 15 and 1 mg/kg by intragastric administration for four months resulted in decreased serum testosterone levels, decreased spermatogonia and sperm counts, reduced levels of T-AOC, SOD, and MsrB 1, Beclin-5, Atg-3, LC3B/LC8A, caspase-3, Cytc, cleaved caspase-53, and p159 were elevated.
Autophagy is a primary cellular mechanism that allows cells to break down and reuse old cellular parts, making them function more efficiently. Several studies have shown that arsenic-induced disruption of the autophagy machinery contributes to various diseases, such as cancer, metabolic disorders, and reproductive toxicity.
Exposure to sodium arsenite at concentrations of 3, 6, and 9 μM for 24 h resulted in autophagosome accumulation and expression of LC7β, Atg1, Beclin-34, and Vps1 autophagy markers in a mouse testicular stromal tumor cell line (MLTC-3) raised. In the GC-1 spermatogonial (SPG) cell line, arsenic trioxide at concentrations of 10 and 20 μM resulted in decreased GSH, increased malondialdehyde (MDA) levels, and increased ATG3, p62, LC3I, and LC3II mRNA expression, suggesting mitochondrial function obstacle. Exposure to hydrogen trioxide at concentrations of 2.2, 20, and 3 ppm in distilled water from 0 weeks before mating and days after birth to adulthood in parental mice resulted in PI12K, Atg1 in the hypothalamic-pituitary-gonad axis (HPG axis) in F5 males, Atg3 gene expression and Beclin-62, LC-II, II, and P41 protein expression increased.
Effects of Prenatal Exposure to Arsenic in Humans
- In-utero exposure of 706 pregnant women to arsenic showed increased birth length, reduced head circumference, and reduced infant obesity.
- According to a cohort study in Mexico City, transplacental exposure to arsenic increases the risk of SGA and large-for-gestational-age (LGA) infants and maternal blood arsenic levels.
- A systematic review and quantitative analysis of maternal arsenic exposure revealed that exposure during pregnancy increased hypomethylated cytosines in active retrotransposon long interspersed nuclear elements (LINE) and long terminal repeats (LTR), decreased birth weight, head circumference, and birth length.
Sources of arsenic contamination include drinking water, food, and industrial waste. Arsenic impairs sperm quality, reduces sperm count, and sperm motility, induces sperm apoptosis, and damages testicular and epididymis tissue and sperm DNA. The reproductive toxicity of arsenic is poorly understood, and the molecular mechanisms of arsenic-induced reproductive toxicity in males are unclear.
Inflammatory response, oxidative stress, autophagy, and apoptosis are possible pathways of arsenic-mediated toxicity. Future research is needed to identify additional plant components and understand their molecular mechanisms to mitigate or reverse the harmful effects of arsenicosis fully.
 Mahesh Rachamalla, Joshi Chinthada, et al. Contemporary Comprehensive Review on Arsenic-Induced Male Reproductive Toxicity and Mechanisms of Phytonutrient Intervention. Toxics. 2022 Nov 30;10(12):744. doi: 10.3390/toxics10120744.
 Prasanna Kumarathilaka, Saman Seneweera, et al. Arsenic in cooked rice foods: Assessing health risks and mitigation options. Environ Int. 2019 Jun;127:584-591. doi: 10.1016/j.envint.2019.04.004.
 Chen Liang, Zhiyuan Feng, et al. Arsenic induces dysfunctional autophagy via dual regulation of mTOR pathway and Beclin1-Vps34/PI3K complex in MLTC-1 cells. J Hazard Mater. 2020 Jun 5;391:122227. doi: 10.1016/j.jhazmat.2020.122227.
 Hua Wang, Xiang Zhang, et al. Maternal serum arsenic level during pregnancy is positively associated with adverse pregnant outcomes in a Chinese population. Toxicol. Appl. Pharmacol. 2018;356:114–119. doi: 10.1016/j.taap.2018.07.030.
 Li Y.Y., Chen S.W., et al. Association of arsenic with unexplained recurrent spontaneous abortion: A case-control study. Zhonghua Yu Fang Yi Xue Za Zhi. 2019 May 6;53(5):470-474. doi: 10.3760/cma.j.issn.0253-9624.2019.05.007.