Open Veterinary Journal, (2026), Vol. 16(4): 2232-2243

Research Article

10.5455/OVJ.2026.v16.i4.24

Reactive oxygen species and intrinsic apoptotic markers in thyroid dysfunction: Insights from experimental animal models

Abdelmoneim Abdelhai Mekki¹, Abdelkarim Abobakr Abdrabo¹ and Amar Mohamed Ismail^2,3*

¹Department of Clinical Chemistry, Faculty of Medical Laboratory Sciences, Al-Neelain University, Khartoum, Sudan

²Department of Biochemistry and Molecular Biology, Faculty of Science and Technology, Al-Neelain University, Khartoum, Sudan

³Department of Biomedical Science, Faculty of Pharmacy, Omar Al-Mukhtar University, Al Bayda, Libya

*Corresponding Author: Amar Mohamed Ismail. Department of Biomedical Science, Faculty of Pharmacy, Omar Al-Mukhtar University, Al Bayda, Libya. Email: amar.ismail [at] omu.edu.ly; amarqqqu [at] yahoo.com

Submitted: 18/11/2025 Revised: 01/03/2026 Accepted: 15/03/2026 Published: 30/04/2026

---

ABSTRACT

Background: Thyroid disorders are associated with elevated reactive oxygen species (ROS) levels that trigger apoptosis. Nevertheless, the precise connection between ROS levels and apoptotic markers in thyroid dysfunction remains unclear.

Aim: To explore the relationship between ROS levels and intrinsic apoptotic (IA) markers in thyroid homogenates derived from hypothyroidism and hyperthyroidism mouse models.

Methods: Eighteen male Wistar rats, each weighing 240 ± 10 g, were allocated to three groups of six rats. Hypothyroidism and hyperthyroidism were induced over 8 weeks using 0.05% Propylthiouracil (PTU) and 0.0012% Levothyroxine (L-Thy), respectively. T3, T4, and thyroid-stimulating hormone levels were measured, and thyroid size and body weights were recorded. The levels of ROS markers [MDA, glutathione (GSH), SOD-1, CAT, and GPX) and IA markers (Bax, Bcl-2, and caspase-3) were assessed in tissue homogenates.

Results: A gradual weight loss was observed in the hyperthyroidism group compared with the control group. The hypothyroid model showed elevated MDA levels and cleaved caspase-3, as well as a higher Bax/Bcl-2 ratio, whereas GSH, SOD-1, CAT, GPX, and Bcl-2 levels were lower than those in the control group (p < 0.05). In contrast, no changes were observed in the hyperthyroid models. Thyroid hormone levels are inversely correlated with ROS and positively correlated with antioxidant levels.

Conclusion: Hypothyroidism models exhibited increased oxidative stress and pro-apoptotic markers, suggesting the initiation of apoptosis and cellular damage. Conversely, the hyperthyroid models showed no such changes.

Keywords: Hypothyroidism, Hyperthyroidism, Intrinsic apoptotic markers, Rat models, ROS.

---

Introduction

Thyroid disorders, particularly among women, are a common global health issue. Hypothyroidism and hyperthyroidism are significant problems and are among the most frequent causes of death worldwide (Journy et al., 2017). The thyroid gland is an oxidative organ in which reactive oxygen species (ROS) are essential for thyroid hormone synthesis (Macvanin et al., 2023). The mechanism is governed by thyroid-stimulating hormone (TSH), iron-containing enzymes, thyroid peroxidase (TPO), iodine, iron, copper, and low levels of ROS (Kochman et al., 2021). Hyperthyroidism and hypothyroidism significantly dysregulate cell redox potential and ROS levels (Huang et al., 2019; Wróblewski et al., 2023). Studies have indicated that both conditions increase ROS production in various tissues through inflammation (Lasa and Contreras-Jurado, 2022). Hyperthyroidism induces ROS production in tissues, whereas hypothyroidism-related tissue damage reduces endogenous antioxidant levels (Mancini et al., 2016). In the thyroid, hydrogen peroxide, the principal ROS, serves as an electron acceptor for thyroid hormone synthesis and is produced by thyrocytes via dual oxidases in the NADPH oxidase family (Szanto et al., 2019; Faria and Fortunato, 2020; Kochman et al., 2021). Other ROS species reported during thyroid hormone synthesis include nitric oxide, tyrosine free radicals, diiodotyrosyl radical intermediates in thyroglobulin, iodine radicals, iodonium ions, and hypoiodous acid intermediates (Karbownik-Lewińska et al., 2022; Samardzic et al., 2023). Conversely, thyrocytes are regulated by high levels of enzymatic and non-enzymatic antioxidants, such as thioredoxin, glutathione (GSH), glutathione peroxidase (GSH-Px), superoxide dismutase (SOD), and catalase (El Hassani et al., 2019; Kochman et al., 2021), along with natural antioxidants such as selenium, α- and γ-tocopherols, ascorbic acid, and coenzyme Q (Paunkov et al., 2019; Sabatino, 2023). Elevated ROS production in the thyroid can cause oxidative damage to thyrocytes, potentially leading to disorders such as atherosclerosis and cancer (El Hassani et al., 2019; Kochman et al., 2021). Iodine, potassium bromate, and nitrobenzene treatments increase ROS levels and reduce thyroid size (Karbownik-Lewińska and Kokoszko-Bilska, 2012). ROS are essential for initiating cell signalling (Sinenko et al., 2021; Iqbal et al., 2024), but they can also damage organs by triggering inflammation and apoptosis through various mechanisms (García-Sánchez, et al., 2020; Vona et al., 2021; Chaudhary et al., 2023). ROS activate intrinsic apoptosis by triggering caspase activation. The balance between Bax and Bcl-2 is crucial for the mitochondrial apoptotic pathway, where an increase or decrease in Bcl-2 leads to either apoptosis or survival. This process is linked to several pathological conditions, including Alzheimer’s disease and autoimmune disorders (Bilous et al., 2020).

Despite numerous studies investigating the links between ROS and free radicals and thyroid disorders, the associations between these factors and intrinsic apoptotic markers in the thyroid gland remain unclear. In this study, we examined the relationships among ROS levels, lipid peroxides, antioxidants, and mitochondria-mediated apoptosis markers (Bax, Bcl-2, and caspase-3) in hypothyroid and hyperthyroid rat models.

---

Materials and Methods

Experimental animals

All experiments were conducted at the Faculty of Pharmacy, King Saud University, with animal welfare prioritized. Eighteen male Wistar rats (240 ± 10 g, 9 weeks old) from the KSU College of Pharmacy Animal House were used. Rats were selected based on age, weight, fitness, and health. They were housed under standard conditions with a 12-hour light/dark cycle at 22°C–26°C and fed a nutrient-rich diet, in accordance with the KSU Animal Ethics and Animal Research: Reporting of In vivo Experiments guidelines.

Induction of hyperthyroidism and hypothyroidism models

After a 7-day adaptation period, the rats were divided into three groups of six rats each. Group 1 (Control Group): Received daily normal saline. Group 2 (hypothyroidism): treated with 0.05% (w/v) Propylthiouracil (PTU) (Cat. No. P7629 (98%) (Sigma-Aldrich, Cambridge, UK) in normal saline (Xu et al., 2017, Celep et al., 2023). Group 3 (Hyperthyroidism): received L-Thy (Cat. T2376 (97%) (Sigma-Aldrich, Cambridge, UK). For 8 weeks, L-Thy was dissolved in drinking water at a concentration of 0.0012% (w/v) (Messarah et al., 2011; Nambiar et al., 2014). Activity responses to handling were recorded daily. Rat body weights were measured at baseline and at weeks 4, 6, and 8, and thyroid gland weights were recorded.

Rat euthanasia and sample collection

Before euthanasia, the rats were evaluated for activity, weight loss, tremors, ataxia, and skin lesions. Following an 18-hour fast, rats were anesthetized by intraperitoneal administration of pentobarbital sodium at a dosage of 50 mg/kg, in accordance with the American Veterinary Medical Association Guidelines for the Euthanasia of Animals: 2020 Edition. Blood samples were obtained through cardiac puncture, and the thyroid glands were collected, centrifuged, and stored at −20°C. Thyroid glands were isolated, weighed, and cryopreserved at −80°C for analysis.

Perpetration of the total number of cell homogenates

Total cell homogenates were prepared by homogenizing 50 mg of frozen thyroid tissue in 0.5 ml ice-cold phosphate-buffered saline (pH 7.4). Following centrifugation at 11,000 × g, the supernatant was collected. The tissues were then homogenized in 0.5 ml of RIPA buffer (Cat. No. 156034, Abcam, Cambridge, UK) with protease inhibitors for WB. The supernatants were stored at −80°C.

Estimation of serum thyroid hormone levels

Serum levels of TSH, T3, and T4 were measured using specific enzyme-linked immunosorbent assay kits. Cat. RTC007R (Biovender, CA, USA), No. MBS161170 and Cat. MBS261867 (MyBioSource, San Diego, CA, USA). Samples were analyzed in duplicate according to the manufacturer’s instructions. An enzyme-linked immunosorbent assay (ELISA) on the Dynex Best 2,000 analyzer was used to generate a standard curve by plotting absorbance at 450 nm against the standard concentration.

Measurement of reactive oxygen species markers

ELISA kits were used to assess thyroid homogenate levels. The GPX1 levels were determined using Cat. No. SEA295Ra (Houston, TX, USA). CAT, GSH, SOD-1, and MDA levels were measured using (Cat. No. MBS458146), (No. MBS1600441), (Cat. No. MBS036924), and (Cat. No. MBS9712310) was obtained from MyBioSource (San Diego, USA), following the methods previously published by D’Amico et al. (2024). Measurements were duplicated according to the manufacturer’s instructions. The color intensity was measured at 450 nm, and the ROS concentrations were determined using a Dynex Best 2,000 analyzer with a standard curve.

Western blotting

Bcl2, Bax, and cleaved caspase-3 levels in thyroid homogenates were quantified using a BCA protein determination kit (Cat. No. 23225, Thermo Fisher, USA) with minor modifications (Alegria-Schaffer et al., 2009) Figure S2. Proteins were diluted to 2 µg/µl in loading dye and heated at 100°C for 5 minutes. Equal protein concentrations (50 µg/well) from thyroid gland homogenates were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis at 100 V for 2 hours. The gels were transferred to nitrocellulose membranes and blocked with skim milk for 15 minutes at 15 mV. After washing with 1% TBST, the membranes were incubated for 2 hours with primary antibodies against β-actin, Cleaved Caspase-3 (Asp175), Bcl2, and Bax. Membranes were then washed and incubated with horseradish peroxidase-conjugated secondary antibodies. Three distinct high-resolution bands from each group were scanned using enhanced chemiluminescence and a C-DiGit Blot Scanner (LI-COR, NE, USA). Protein levels were quantified using β-actin as the reference.

Statistical analysis

Statistical analyses were performed using IBM SPSS Statistics for Windows, version 21 (SPSS, Armonk, NY, USA) and GraphPad Prism Version 8. The Shapiro-Wilk test was used to assess normality. Data are presented as mean ± standard deviation (M ± SD) and correlation coefficients (r²). Statistical significance for multiple comparisons was assessed using one-way analysis of variance, followed by post hoc Tukey’s HSD and Dunnett’s tests. Pearson’s correlation coefficient was used to evaluate the correlations between thyroid hormones and ROS parameters. Statistical significance was set at p ≤ 0.05.

Ethical approval

This study was approved by the Ethics Committee of Al-Neelain University’s Faculty of Medical Laboratory Science. (Ref. NU1309) 2019.

---

Results

Hypothyroid and hyperthyroid models induction

Rats treated with L-Thy exhibited reduced TSH levels (0.286 ± 0.079 ng/ml) and increased T3 (2,495 ± 268 pg/ml) and T4 (8.60 ± 1.32 ng/ml) compared to controls with TSH (0.895 ± 0.15 ng/ml), T3 (1,098 ± 251 pg/ml), and T4 (4.618 ± 0.65 ng/ml) levels (p < 0.05). PTU-treated rats exhibited decreased T3 (416 ± 130 pg/ml) and T4 (1.90 ± 0.24 ng/ml) levels and increased TSH (2.10 ± 0.34 ng/ml), indicating the successful induction of hypothyroid and hyperthyroid models (Fig. S1. A–C).

Body weight and thyroid gland size in the study groups

During the first 5 weeks of PTU treatment, the patient’s body weight remained unchanged. However, a significant increase was observed in weeks 6–8 compared with the control group. In contrast, rats treated with L-Thy exhibited weight loss from weeks 2 to 8 compared with controls (p < 0.05). PTU treatment reduced the size of the thyroid gland, whereas L-Thy treatment showed no change after 8 weeks (p <0.05) (Fig. 1A and B).

Fig. 1. A: Body weight and B: Thyroid weight comparisons among the hyperthyroidism, hypothyroidism, and control groups. a: Difference compared with the control group. b: Difference compared with the hyperthyroid group. Data are presented as mean ± SD, p ≤ 0.05.

Oxidative scavenging and oxidative stress markers

Fig. 2A–E shows that the hypothyroid homogenates had elevated MDA levels (2.00 ± 0.20 nmol/mg) and reduced GSH (0.20 ± 0.04 ng/mg), SOD (3.26 ± 0.59 U/mg), CAT (0.158 ± 0.04 ng/mg), and GPX (6.55 ± 1.06 ng/mg) compared with controls (p < 0.05). Levels of MDA (0.65 ± 0.12 nmol/mg), GSH (0.50 ± 0.06 ng/mg), SOD-1 (8.03 ± 2.38 U/mg), CAT (0.40 ± 0.06 ng/mg), and GPX (14.2 ± 1.90 ng/mg) in hyperthyroid homogenates did not differ from controls, which had MDA (0.56 ± 0.07 nmol/mg), GSH (0.60 ± 0.04 ng/mg), SOD-1 (10.6 ± 1.25 U/mg), CAT (0.40 ± 0.07 ng/mg), and GPX (17.2 ± 1.79 ng/mg) levels (Fig. 2. A–E).

Fig. 2. (A, B, C, D, and E). Comparison of oxidative stress/antioxidant markers in the hyperthyroidism, hypothyroidism, and control groups. a: Difference versus control. b: Difference versus the hyperthyroidism group. Data are presented as mean ± SD, p ≤ 0.05.

Expression of pro- and anti-apoptotic markers

Figure 3A–D demonstrate that hypothyroid models showed increased levels of Bax (0.35 ± 0.04), cleaved caspase-3 (0.37 ± 0.07), and Bax/Bcl-2 ratios (1.5 ± 0.5) relative to β-actin, along with decreased Bcl-2 levels (0.22 ± 0.03) relative to β-actin, compared to the control group, which exhibited values of (0.12 ± 0.03), (0.11 ± 0.04), (0.11 ± 0.01), and (0.58 ± 0.02) relative to β-actin, respectively (p < 0.05). In contrast, no significant changes were observed in the hyperthyroid models compared with the control group.

Fig. 3. (A, B, C, D) Comparison of pro-apoptotic and anti-apoptotic markers in the hyperthyroidism, hypothyroidism, and the control group. a: Difference compared with the control. b: Difference compared with the hyperthyroidism group. Data are presented as mean ± SD, p ≤ 0.05.

Relationships among antioxidants, oxidative stress, and thyroid hormone levels

The scatter plots revealed positive correlations between GSH, T3, and T4 levels and between SOD-1 and T3 and T4 levels, with r² values of 0.283, 0.140, 0.409, and 0.206, respectively. Inverse correlations were observed between MDA and thyroid hormone levels, with r² values of 0.460 and 0.526, respectively. The findings indicate that thyroid hormones are involved in the regulation and production of antioxidants (Figs. 4 and 5, panels A–D).

Fig. 4. (A, B, C, and D). The association of T3 with oxidative stress/antioxidant markers was assessed using Pearson’s correlation coefficient, where (r²) is the coefficient of determination and (y) is the correlation coefficient (−) negative and (+) positive correlations. Data are presented as r², p ≤ 0.05.

Fig. 5. (A, B, C, and D). The association between T4 and oxidative stress/antioxidant markers was assessed using Pearson’s correlation coefficient, where (r²) is the coefficient of determination, (y) is the correlation coefficient (−) negative, and (+) is positive. Data are presented as r², p ≤ 0.05.

---

Discussion

The relationship between ROS generation, intrinsic apoptosis, and cell damage induced by thyroid hormones remains unclear. This study examined ROS, lipid peroxides, antioxidants, and mitochondrial-mediated apoptosis markers (Bax, Bcl-2, and caspase-3) in hypothyroid and hyperthyroid animal models. L-Thy and PTU induce hyperthyroidism and hypothyroidism, respectively (Kandir and Keskin, 2016). The successful induction of these models aligns with the findings that chronic L-Thy administration induces hyperthyroidism, whereas PTU treatment causes hypothyroidism in laboratory animals. Studies have documented the effects of L-Thy on the thyroid gland (Ferreira et al., 2007; Jin and Sugitani, 2021), with evidence supporting the hypothyroid effect of PTU (Pantos et al., 2003; Pan et al., 2013).

Examination of body weight and thyroid gland size in hypothyroid and hyperthyroid models has substantial clinical significance. During weeks 6–8, hypothyroid rats gained weight, whereas hyperthyroid rats lost weight compared with controls. These findings are consistent with those of earlier studies (Shahid, et al., 2018), suggesting that weight fluctuations indicate thyroid dysfunction. In hypothyroidism, reduced catabolic rates cause water retention and fat accumulation, whereas elevated metabolic rates lead to weight loss in hyperthyroidism (Ye et al., 2017; Alidrisi et al., 2021). Meanwhile, the glands were smaller in hypothyroid models, whereas their size remained unchanged in hyperthyroid rats compared to the controls. These findings are similar to those of previous studies that showed that thyroid atrophy in primary idiopathic hypothyroidism results from elevated TSH levels, which are linked to simple goiter. This contrasts with the low TSH levels in hyperthyroidism, where TSH acts as a goitrogen (Žarković, 2012; Li et al., 2023).

Our study on thyroid disorders and ROS levels in patients with hypothyroidism and hyperthyroidism has important implications. Our findings showed no differences in MDA, GSH, SOD, CAT, or GPX levels between the hyperthyroid models and the control group, contradicting previous findings suggesting increased oxidative stress in thyrotoxicosis (Hsieh et al., 2023). This contradiction may be due to differences in dosage and treatment duration. The hypothyroid group had higher MDA levels and lower GSH, SOD-1, CAT, and GPX levels than the control group. These findings are consistent with previous studies linking hypothyroidism to reduced antioxidant production and increased ROS levels (Petrulea, et al., 2009; Kochman et al., 2021). Decreased activity in hypothyroidism impairs radical scavenging, leading to cellular damage via lipid peroxidation (Kochman et al., 2021).

Correlation analyses showed positive correlations between GSH, SOD-1, and thyroid hormone levels, whereas MDA levels showed an inverse correlation. These findings demonstrate the interactions among oxidative stress, antioxidant defenses, and thyroid hormone levels. Low and high thyroid hormone levels are associated with oxidative stress in hypothyroidism and hyperthyroidism, respectively (Venditti et al., 1997; Kochman et al., 2021). A recent study demonstrated that thyroid hormones facilitate liver bilirubin conjugation therapy, resulting in increased oxidative stress and enhanced antioxidant levels in hypothyroidism and hyperthyroidism, respectively (Shin et al., 2025).

This study examined apoptotic and anti-apoptotic markers in hyperthyroidism and hypothyroidism rat models. Hypothyroid models showed increased Bax and cleaved caspase-3, a higher Bax/Bcl-2 ratio, and decreased Bcl-2, indicating increased pro-apoptotic and decreased anti-apoptotic expression. Thyroid hormones inhibit apoptosis and promote cell proliferation (Oliva et al., 2013; Aldubayan, 2019). Increased oxidative stress triggers mitochondrial apoptosis in hypothyroidism, which may lead to greater thyroid cell damage (Villalpando-Rodriguez and Gibson, 2021). These findings suggest potential therapeutic targets for hypothyroidism. Hyperthyroid models showed no differences in pro-apoptotic and anti-apoptotic markers compared with controls, indicating a normal Bax/Bcl-2 ratio. To the best of our knowledge, this is the first study analyzing thyroid homogenates. Hyperthyroidism causes DNA damage in cardiac tissues and increases caspase levels in hepatic tissues, initiating apoptosis (Guler et al., 2022).

Our study elucidated the link between oxidative stress-induced apoptosis and cellular damage in hypothyroid models, whereas hyperthyroid models showed no changes. However, this study had some limitations, including the absence of clinical data from animal studies, the small sample size, and the lack of immunohistochemical analysis to confirm the Western blot results for cleaved caspase-3, which showed a single band in the frozen rat sections. Despite these limitations, our findings are significant and underscore the need for further studies.

---

Conclusion

However, the exact relationship between ROS levels and intrinsic mitochondrial apoptotic markers in thyroid disorders remains unclear. Our study offers experimental evidence that the hypothyroid model shows increased oxidative stress and pro-apoptotic levels, along with decreased antioxidant and anti-apoptotic levels. No significant changes were observed in the hyperthyroid mouse model. Thyroid hormone levels are inversely related to ROS levels and positively correlated with antioxidant levels. Increased pro-apoptotic markers and reduced antioxidant production in hypothyroidism cause thyroid cell damage, highlighting the need for effective management and therapeutic targets in future studies. Further prospective studies are needed to elucidate the relationship between ROS and apoptotic biomarkers in patients with hypothyroidism.

---

Acknowledgments

None.

Conflict of interest

None.

Funding

The authors have received no funding for this study.

Authors’ contributions

Conception and design: A.A.M., A.M.I.; supervision: A.M.I., A.A.A.; conducting experiments: A.A.M., A.A.A., A.M.I.; writing original research: A.A.M.; writing review and editing: A.A.M., A.A.A., A.M.I.; All authors have read and approved the manuscript.

Data availability

The datasets collected and analyzed in this study are available from the corresponding author upon reasonable request.

---

References

Aldubayan, M.A. 2019. Grape Seed Proanthocyanidin Alleviates Oxidative Stress and Apoptosis Involved in Liver of Hyperthyroid Mice. Annu. Res. Rev. Biol. 33(6), 1–11; doi: 10.9734/arrb/2019/v33i630138

Alegria-Schaffer, A., Lodge, A. and Vattem, K. 2009. Performing and optimizing Western blots with an emphasis on chemiluminescent detection. Methods Enzymology 463, 573–599.

Alidrisi, H.A., Odhaib, S.A., Altemimi, M.T. and Mansour, A.A. 2021. Patterns of bodyweight changes in patients with hypothyroidism, a retrospective study from Basrah, Southern Iraq. Cureus 13(6), 2–11; doi:10.7759/cureus.15408

Bilous, I., Pavlovych, L., Krynytska, I., Marushchak, M. and Kamyshnyi, A. 2020. Apoptosis and cell cycle pathway-focused genes expression analysis in patients with different forms of thyroid pathology. Open Access Macedonian J. Med. Sci. 8, 784–792; doi:10.3889/oamjms.2020.4760

Celep, N.A., Gedikli, S., Parlak, S.N. and Aliyev, E. 2023. ‘Experimental hypothyroidism increases oxidative stress and apoptosis in ovary of rats’. J. Clin. Nutr. 2(2), 21–29.

Chaudhary, P., Janmeda, P., Docea, A.O., Yeskaliyeva, B., Abdull Razis, A.F., Modu, B., Calina, D. and Sharifi-Rad, J. 2023. Oxidative stress, free radicals and antioxidants: potential crosstalk in the pathophysiology of human diseases. Front. Chem. 11, 1–24; doi:10.3389/fchem.2023.1158198

D'Amico, R., Cordaro, M., Fusco, R., Peritore, A.F., Genovese, T., Gugliandolo, E., Crupi, R., Mandalari, G., Caccamo, D., Cuzzocrea, S. and Di Paola, R. 2024. Correction to: consumption of cashew (Anacardium occidentale L.) Nuts Counteracts Oxidative Stress and Tissue Inflammation in Mild Hyperhomocysteinemia in Rats (Nutrients, Nutrients, 2022). Nutrients 16(1), 10; doi:10.3390/nu16010133

El Hassani, R.A., Buffet, C., Leboulleux, S. and Dupuy, C. 2019. Oxidativte stress in thyroid carcinomas: biological and clinical significance. Endocr. Relat. Cancer. 26(3), R131–R143; doi: 10.1530/ERC-18-0476

Faria, C.C.D. and Fortunato, R.S. 2020. The role of dual oxidases in physiology and cancer. Genet. Mol. Biol. 43(1), 1–9; doi:10.1590/1678-4685/GMB-2019-0096

Ferreira, E., Silva, A.E., Serakides, R., Gomes, A.E.S. and Cassali, G.D. 2007. Model of induction of thyroid dysfunctions in adult female mice. Arquivo Brasileiro De Medicina Veterinaria. E Zootecnia 59(5), 1245–1249; doi:10.1590/S0102-09352007000500022

García-Sánchez, A., Miranda-Díaz, A.G. and Cardona-Muñoz, E.G. 2020. The role of oxidative stress in physiopathology and pharmacological treatment with pro- and antioxidant properties in chronic diseases. Oxidative Med. Cellular Longevity 2020 (1), 2082145; doi: 10.1155/2020/2082145

Guler, G., Dasdelen, D., Baltaci, S.B., Sivrikaya, A., Baltaci, A.K. and Mogulkoc, R. 2022. The effects of thyroid dysfunction on DNA damage and apoptosis in liver and heart tissues of rats. Hormone. Mol. Biol. Clin. Invest. 43(1), 47–53.

Hsieh, C.T., Yen, T.L., Chen, Y.H., Jan, J.S., Teng, R.D., Yang, C.H. and Sun, J.M. 2023. Aging-associated thyroid dysfunction contributes to oxidative stress and worsened functional outcomes following traumatic brain injury. Antioxidants 12(2), 1–26; doi:10.3390/antiox12020217

Huang, Y., Cai, L., Zheng, Y., Pan, J., Li, L., Zong, L., Lin, W., Liang, J., Huang, H., Wen, J. and Chen, G. 2019. Association between lifestyle and thyroid dysfunction: a cross-sectional epidemiologic study in the She ethnic minority group of Fujian Province in China. BMC Endocrine Disord. 19(1), 1–9; doi:10.1186/s12902-019-0414-z

Iqbal, M.J., Kabeer, A., Abbas, Z., Siddiqui, H.A., Calina, D., Sharifi-Rad, J. and Cho, W.C. 2024. Interplay of oxidative stress, cellular communication and signaling pathways in cancer. Cell Communication Signaling 22(1), 1–16; doi:10.1186/s12964-023-01398-5

Jin, S. and Sugitani, I. 2021. Development of a rat model for evaluating thyroid-stimulating hormone suppression after total thyroidectomy. J. Nippon Med. School 88(4), 311–318.

Journy, N.M.Y., Bernier, M.O., Doody, M.M., Alexander, B.H., Linet, M.S. and Kitahara, C.M. 2017. Hyperthyroidism, hypothyroidism, and cause-specific mortality in a large cohort of women. Thyroid 27(8), 1001–1010; doi:10.1089/thy.2017.0063

Kandir, S. and Keskin, E. 2016. Hyperthyroidism on hematological parameters in rats: effects of hypothyroidism and. Ankara Universitesi. Veteriner. Fakultesi. Dergisi. 63(4), 371–376; doi:10.1501/vetfak_0000002755

Karbownik-Lewińska, M. and Kokoszko-Bilska, A. 2012. Oxidative damage to macromolecules in the thyroid—experimental evidence. Thyroid. Res. 5(1), 1–6; doi:10.1186/1756-6614-5-25

Karbownik-Lewińska, M., Stępniak, J., Iwan, P. and Lewiński, A. 2022. Iodine as a potential endocrine disruptor—a role of oxidative stress. Endocrine 78(2), 219–240; doi:10.1007/s12020-022-03107-7

Kochman, J., Jakubczyk, K., Bargiel, P. and Janda-Milczarek, K. 2021. The influence of oxidative stress on thyroid diseases. Antioxidants 10(9), 1442; doi:10.3390/antiox10091442

Lasa, M. and Contreras-Jurado, C. 2022. Thyroid hormones act as modulators of inflammation through their nuclear receptors. Front. Endocrinol. 13, 1–11; doi:10.3389/fendo.2022.937099

Li, S., Guo, W., Meng, Q., Zhu, M., Wei, H., Ji, F., Tan, L. and Zhang, W. 2023. The association between thyroid-stimulating hormone and thyroid nodules, goiter and thyroid antibody positivity. Front. Endocrinol. 14, 1–9; doi:10.3389/fendo.2023.1204552

Macvanin, M.T., Gluvic, Z., Zafirovic, S., Gao, X., Essack, M. and Isenovic, E.R. 2023. The protective role of nutritional antioxidants against oxidative stress in thyroid disorders. Front. Endocrinol. 13, 1–19; doi:10.3389/fendo.2022.1092837

Mancini, A., Di Segni, C., Raimondo, S., Olivieri, G., Silvestrini, A., Meucci, E. and Currò, D. 2016. Thyroid hormones, oxidative stress, and inflammation. Mediators Inflammation 2016(1), 6757154;, doi:10.1155/2016/6757154

Messarah, M., Saoudi, M., Boumendjel, A., Boulakoud, M.S. and Feki, A.E. 2011. Oxidative stress induced by thyroid dysfunction in rat erythrocytes and heart. Environ. Toxicol. Pharmacol. 31, 33–41; doi:10.1016/j.etap.2010.09.003

Nambiar, P.R., Palanisamy, G.S., Okerberg, C., Wolford, A., Walters, K., Buckbinder, L. and Reagan, W.J. 2014. Toxicities associated with 1-month treatment with propylthiouracil (PTU) and methimazole (MMI) in male rats. Toxicol. Pathol. 42(6), 970–983.

Oliva, F., Berardi, A.C., Misiti, S., Verga Falzacappa, C., Iacone, A. and Maffulli, N. 2013. Correspondence : thyroid hormones enhance growth and counteract apoptosis in human tenocytes isolated from rotator cuff tendons. Cell Death Dis. 4(7), 3–4; doi:10.1038/cddis.2013.229

Pan, T., Zhong, M., Zhong, X., Zhang, Y. and Zhu, D. 2013. Levothyroxine replacement therapy with vitamin E supplementation prevents oxidative stress and cognitive deficit in experimental hypothyroidism. Endocrine 43(2), pp.434–439.

Pantos, C., Malliopoulou, V., Mourouzis, I., Sfakianoudis, K., Tzeis, S., Doumba, P., Xinaris, C., Cokkinos, A.D., Carageorgiou, H., Varonos, D.D. and Cokkinos, D.V. 2003. Propylthiouracil-induce hypothyroidism is associated with increased tolerance of the isolated rat heart to ischemia-reperfusion. J. Endocrinol. 178(3), 427–435; doi:10.1677/joe.0.1780427

Paunkov, A., Chartoumpekis, D.V., Ziros, P.G., Chondrogianni, N., Kensler, T.W. and Sykiotis, G.P. 2019. Impact of antioxidant natural compounds on the thyroid gland and implication of the Keap1/Nrf2 signaling pathway. Curr. Pharm. Des. 25(16), 1828–1846; doi:10.2174/1381612825666190701165821

Petrulea, M.S., Duncea, I. and Muresan, A. 2009. Thyroid hormones in excess induce oxidative stress in rats. Acta. Endocrinologica. 5(2), 155–164; doi:10.4183/aeb.2009.155

Sabatino, L. 2023. Nrf2-Mediated Antioxidant Defense and Thyroid Hormone Signaling: a Focus on Cardioprotective Effects. Antioxidants 12(6), 12; doi:10.3390/antiox12061177

Samardzic, V.S., Macvanin, M.T., Zafirovic, S.S., Obradovic, M.M., Gluvic, Z.M., Grubin, J., Gao, X., Essack, M. and Isenovic, E.R. 2023. Nitric oxide, thyroglobulin, and calcitonin: unraveling the nature of thyroid nodules. Front. Endocrinol. 14, 1–11; doi:10.3389/fendo.2023.1241223

Shahid, M.A., Ashraf, M.A. and Sharma, S. 2018. Physiology, thyroid hormone. Treasure Island, FL: StatPearls Publishing. Available via http://europepmc.org/abstract/MED/29763182.

Shin, J.W., Sull, J.W., Minh, N.T. and Jee, S.H. 2025. Bilirubin Metabolism and Thyroid Cancer: insights from ALBI and PALBI Indices. Biomolecules 18(15), 1042.

Sinenko, S.A., Starkova, T.Y., Kuzmin, A.A. and Tomilin, A.N. 2021. Physiological Signaling Functions of Reactive Oxygen Species in Stem Cells: from Flies to Man. Front. Cell. Develop. Biol. 9, 1–21; doi:10.3389/fcell.2021.714370

Szanto, I., Pusztaszeri, M. and Mavromati, M. 2019. H2O2 metabolism in normal thyroid cells and in thyroid tumorigenesis: focus on NADPH oxidases. Antioxidants 8(5), 1–21; doi:10.3390/antiox8050126

Venditti, P., Balestrieri, M., Di Meo, S. and De Leo, T. 1997. Effect of thyroid state on lipid peroxidation, antioxidant defenses, and susceptibility to oxidative stress in rat tissues. J. Endocrinol. 155(1), 151–157; doi:10.1677/joe.0.1550151

Villalpando-Rodriguez, G.E. and Gibson, S.B. 2021. Reactive Oxygen Species (ROS) regulates different types of cell death by acting as a Rheostat. Oxidative. Med. Cellular. Longevity. 2021 (1), 9912436;, doi:10.1155/2021/9912436

Vona, R., Pallotta, L., Cappelletti, M., Severi, C. and Matarrese, P. 2021. The impact of oxidative stress in human pathology: focus on gastrointestinal disorders. Antioxidants 10(2), 1–26; doi:10.3390/antiox10020201

Wróblewski, M., Wróblewska, J., Nuszkiewicz, J., Pawłowska, M., Wesołowski, R. and Woźniak, A. 2023. The Role of Selected Trace Elements in Oxidoreductive Homeostasis in Patients with Thyroid Diseases. Int. J. Mol. Sci. 24(5), doi:10.3390/ijms24054840

Xu, Q.Y., Wang, X.L. and Peng, Y.F. 2017. Hypothyroidism induced by propylthiouracil decrease sirtuin1 content in rat heart. J. Lab. Precis. Med. 2, 67; doi:10.21037/jlpm.2017.08.03

Ye, J., Zhong, X., Du, Y., Cai, C. and Pan, T. 2017. Role of levothyroxine and vitamin E supplementation in the treatment of oxidative stress-induced injury and apoptosis of myocardial cells in hypothyroid rats. J. Endocrinol. Invest. 40(7), 713–719; doi:10.1007/s40618-017-0624-z

Žarković, M. 2012. The role of oxidative stress on the pathogenesis of Graves’ disease. J. Thyroid. Res. 2012(1), 302537; doi:10.1155/2012/302537/

---

Supplementary Materials

Fig. S1. A, B, and C. Comparison of thyroid hormone levels in the hyperthyroidism, hypothyroidism, and control groups after 8 weeks of L-Thy and PTU administration. Differences between groups were assessed using ANOVA and Tukey's HSD post hoc test. a: Difference compared to control. b: Difference compared to the hyperthyroidism group. Statistical significance was set at a p value of ≤ 0.05.

Fig. S2. Presents uncropped images of Bax, cleaved caspase-3, and Bcl2 arranged top to bottom. Control hyperthyroidism and hypothyroidism are displayed left to right using Western blot analysis.