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Oleik Aside Bağlı Akut Akciğer Hasarı Modeli Oluşturulan Ratlarda Tocilizumab ve Deksametazon'un Karaciğer Üzerine Etkilerinin Histopatolojik Olarak İncelenmesi

Year 2022, Volume: 11 Issue: 2, 167 - 172, 14.07.2022
https://doi.org/10.53424/balikesirsbd.989313

Abstract

Amaç: Bu çalışmada, oleik asit ile akut akciğer hasarı modeli oluşturulan ratlarda Tocilizumab ve Deksametazon'un karaciğer üzerindeki etkilerinin incelenmesi hedeflenmiştir. Gereç ve Yöntem: Çalışma materyalini otuz altı erkek rat oluşturdu ve CN, OA, TCZ ve DEX olmak üzere altı gruba ayrıldı. Kontrol grubu dışındaki tüm gruplar intravenöz olarak OA verildi. OA enjeksiyonlarından sonra diğer bileşikler intravenöz olarak uygulandı (DEX 0.1 mg/kg ve DEX 10 mg/kg, TCZ 2 mg/kg ve TCZ 4 mg/kg). Bulgular: OA+DEX 10 grubu sıçanlarda hepatositlerde hidropik dejenerasyonun diğer gruplara göre arttığı belirlendi (p<0.05). OA+TCZ 2 ve 4 gruplarında hepatositlerde vakuol oluşumu kontrol grubuna benzer bulundu (p<0.05). Ayrıca OA+DEX (0,1 ve 10 mg/kg) gruplarında hepatositlerde nekroz ve sinüzoidlerde daralma daha şiddetli olduğu görüldü. Sonuç: Yüksek doz deksametazonun hepatotoksik etki gösterdiği belirlendi. COVID-19/ARDS hastalarının tedavisinde kullanılacak ilaçların hepatotoksik etkilerinin belirlenmesi için karaciğer fonksiyon testlerinin yapılması ve değerlendirilmesi gerektiği düşünülmektedir.

Supporting Institution

Kastamonu Üniversitesi Bilimsel Araştırma Projeleri Koordinasyon Birimi

Project Number

KÜ-BAP01/2019-12

References

  • Abdelhalim, M. A. K., & Jarrar, B. M. (2011). Gold nanoparticles induced cloudy swelling to hydropic degeneration, cytoplasmic hyaline vacuolation, polymorphism, binucleation, karyopyknosis, karyolysis, karyorrhexis and necrosis in the liver. Lipids in Health and Disease, 10(1), 1-6.
  • Akella, A., Sharma, P., Pandey, R., & Deshpande, S. B. (2014). Characterization of oleic acid-induced acute respiratory distress syndrome model in rat.
  • Beilman, G. (1995). Pathogenesis of oleic acid-induced lung injury in the rat: distribution of oleic acid during injury and early endothelial cell changes. Lipids, 30(9), 817-823.
  • Bleibel, W., Kim, S., D’Silva, K., & Lemmer, E. R. (2007). Drug-induced liver injury. Digestive diseases and sciences, 52(10), 2463-2471. Browning, J. D., & Horton, J. D. (2004). Molecular mediators of hepatic steatosis and liver injury. The Journal of clinical investigation, 114(2), 147-152.
  • Bulmuş, F. G., Gürsu, M. F., Muz, M. H., Yaman, İ., Bulmuş, Ö., & Sakin, F. (2013). Protective effects of alpha-lipoic acid on oleic acid-induced acute lung injury in rats. Balkan medical journal, 30(3), 309.
  • Canbay, A., Gieseler, R., Gores, G., & Gerken, G. (2005). The relationship between apoptosis and non-alcoholic fatty liver disease: an evolutionary cornerstone turned pathogenic. Zeitschrift für Gastroenterologie, 43(02), 211-217.
  • De Benedetti, F., Brunner, H. I., Ruperto, N., Kenwright, A., Wright, S., Calvo, I., Cuttica, R., Ravelli, A., Schneider, R., & Woo, P. (2012). Randomized trial of tocilizumab in systemic juvenile idiopathic arthritis. New England Journal of Medicine, 367(25), 2385-2395.
  • Drucker, C., Gewiese, J., Malchow, S., Scheller, J., & Rose-John, S. (2010). Impact of interleukin-6 classic-and trans-signaling on liver damage and regeneration. Journal of autoimmunity, 34(1), 29-37.
  • Ducheix, S., Montagner, A., Polizzi, A., Lasserre, F., Regnier, M., Marmugi, A., Benhamed, F., Bertrand-Michel, J., Mselli-Lakhal, L., & Loiseau, N. (2017). Dietary oleic acid regulates hepatic lipogenesis through a liver X receptor-dependent signaling. PLoS One, 12(7), e0181393.
  • Erol, N., Saglam, L., Saglam, Y. S., Erol, H. S., Altun, S., Aktas, M. S., & Halici, M. B. (2019). The protection potential of antioxidant vitamins against acute respiratory distress syndrome: a rat trial. Inflammation, 42(5), 1585-1594.
  • Herrero, R., Sánchez, G., Asensio, I., López, E., Ferruelo, A., Vaquero, J., Moreno, L., de Lorenzo, A., Bañares, R., & Lorente, J. A. (2020). Liver–lung interactions in acute respiratory distress syndrome. Intensive Care Medicine Experimental, 8(1), 1-13.
  • Kang, O., Kim, S., Seo, Y., Joung, D., Mun, S., Choi, J., Lee, Y., Kang, D., Lee, H., & Kwon, D. (2013). Curcumin decreases oleic acid-induced lipid accumulation via AMPK phosphorylation in hepatocarcinoma cells. Eur Rev Med Pharmacol Sci, 17(19), 2578-2586.
  • Koh, Y. (2014). Update in acute respiratory distress syndrome. Journal of intensive care, 2(1), 1-6.
  • Koksel, O., Kaplan, M. B., Ozdulger, A., Tamer, L., Degirmenci, U., Cinel, L., Bastürk, M., & Kanik, A. (2005). Oleic acid-induced lung injury in rats and effects of caffeic acid phenethyl ester. Experimental lung research, 31(5), 483-496.
  • Lee, J. Y., Moon, J. H., Park, J. S., Lee, B.-W., Kang, E. S., Ahn, C. W., Lee, H. C., & Cha, B. S. (2011). Dietary oleate has beneficial effects on every step of non-alcoholic Fatty liver disease progression in a methionine-and choline-deficient diet-fed animal model. Diabetes & metabolism journal, 35(5), 489.
  • Luo, P., Liu, Y., Qiu, L., Liu, X., Liu, D., & Li, J. (2020). Tocilizumab treatment in COVID‐19: a single center experience. Journal of medical virology, 92(7), 814-818.
  • Mahamid, M., Paz, K., Reuven, M., & Safadi, R. (2011). Hepatotoxicity due to tocilizumab and anakinra in rheumatoid arthritis: two case reports. International journal of general medicine, 4, 657.
  • Matute-Bello, G., Frevert, C. W., & Martin, T. R. (2008). Animal models of acute lung injury. American Journal of Physiology-Lung Cellular and Molecular Physiology, 295(3), L379-L399.
  • Mikiewicz, M., Otrocka-Domagała, I., Paździor-Czapula, K., & Rotkiewicz, T. (2017). Influence of long-term, high-dose dexamethasone administration on proliferation and apoptosis in porcine hepatocytes. Research in veterinary science, 112, 141-148.
  • Moore, B. B., & Hogaboam, C. M. (2008). Murine models of pulmonary fibrosis. American Journal of Physiology-Lung Cellular and Molecular Physiology, 294(2), L152-L160.
  • Muhović, D., Bojović, J., Bulatović, A., Vukčević, B., Ratković, M., Lazović, R., & Smolović, B. (2020). First case of drug‐induced liver injury associated with the use of tocilizumab in a patient with COVID‐19. Liver International, 40(8), 1901-1905.
  • Okun, J. G., Conway, S., Schmidt, K. V., Schumacher, J., Wang, X., de Guia, R., Zota, A., Klement, J., Seibert, O., & Peters, A. (2015). Molecular regulation of urea cycle function by the liver glucocorticoid receptor. Molecular metabolism, 4(10), 732-740.
  • Oldfield, V., Dhillon, S., & Plosker, G. L. (2009). Tocilizumab. Drugs, 69(5), 609-632.
  • Pascussi, J. M., Drocourt, L., Gerbal‐Chaloin, S., Fabre, J. M., Maurel, P., & Vilarem, M. J. (2001). Dual effect of dexamethasone on CYP3A4 gene expression in human hepatocytes: sequential role of glucocorticoid receptor and pregnane X receptor. European journal of biochemistry, 268(24), 6346-6358.
  • Patel, R., Patel, M., Tsai, R., Lin, V., Bookout, A. L., Zhang, Y., Magomedova, L., Li, T., Chan, J. F., & Budd, C. (2011). LXRβ is required for glucocorticoid-induced hyperglycemia and hepatosteatosis in mice. The Journal of clinical investigation, 121(1), 431-441.
  • Patterson, E., Yao, L., Ramic, N., Lewis, J., Cepinskas, G., McCaig, L., Veldhuizen, R., & Yamashita, C. (2013). Lung-derived mediators induce cytokine production in downstream organs via an NF-κB-dependent mechanism. Mediators of inflammation, 2013.
  • Roussel, D., Dumas, J.-F., Simard, G., Malthièry, Y., & Ritz, P. (2004). Kinetics and control of oxidative phosphorylation in rat liver mitochondria after dexamethasone treatment. Biochemical Journal, 382(2), 491-499.
  • Schuster, D. P. (1994). ARDS: clinical lessons from the oleic acid model of acute lung injury. American journal of respiratory and critical care medicine, 149(1), 245-260.
  • Schwartz, D. B., Bone, R. C., Balk, R. A., & Szidon, J. R. (1989). Hepatic dysfunction in the adult respiratory distress syndrome. Chest, 95(4), 871-875. Serviddio, G., Villani, R., Stallone, G., Scioscia, G., Foschino-Barbaro, M. P., & Lacedonia, D. (2020). <? covid19?> Tocilizumab and liver injury in patients with COVID-19. Therapeutic Advances in Gastroenterology, 13, 1756284820959183.
  • Slutsky, A. S., & Ranieri, V. M. (2013). Ventilator-induced lung injury. New England Journal of Medicine, 369(22), 2126-2136.
  • Sun, S., Liu, D., Zhang, H., Zhang, X., & Wan, B. (2019). Effect of different doses and time‑courses of corticosteroid treatment in patients with acute respiratory distress syndrome: A meta‑analysis. Experimental and therapeutic medicine, 18(6), 4637-4644.
  • Wan, E. S., Qiu, W., Baccarelli, A., Carey, V. J., Bacherman, H., Rennard, S. I., Agustí, A., Anderson, W. H., Lomas, D. A., & DeMeo, D. L. (2012). Systemic steroid exposure is associated with differential methylation in chronic obstructive pulmonary disease. American journal of respiratory and critical care medicine, 186(12), 1248-1255.
  • Wiener-Kronish, J., Albertine, K., & Matthay, M. (1991). Differential responses of the endothelial and epithelial barriers of the lung in sheep to Escherichia coli endotoxin. The Journal of clinical investigation, 88(3), 864-875.
  • Yang, P., Formanek, P., Scaglione, S., & Afshar, M. (2019). Risk factors and outcomes of acute respiratory distress syndrome in critically ill patients with cirrhosis. Hepatology Research, 49(3), 335-343.
  • Zhang, X., Morcos, P. N., Saito, T., & Terao, K. (2013). Clinical pharmacology of tocilizumab for the treatment of systemic juvenile idiopathic arthritis. Expert review of clinical pharmacology, 6(2), 123-137.
  • Zhang, X., Ouyang, J., & Thung, S. N. (2013). Histopathologic manifestations of drug-induced hepatotoxicity. Clinics in liver disease, 17(4), 547-564.
  • Zhang, X., & Peck, R. (2011). Clinical pharmacology of tocilizumab for the treatment of patients with rheumatoid arthritis. Expert review of clinical pharmacology, 4(5), 539-558.
  • Ziamajidi, N., Khaghani, S., Hassanzadeh, G., Vardasbi, S., Ahmadian, S., Nowrouzi, A., Ghaffari, S. M., & Abdirad, A. (2013). Amelioration by chicory seed extract of diabetes-and oleic acid-induced non-alcoholic fatty liver disease (NAFLD)/non-alcoholic steatohepatitis (NASH) via modulation of PPARα and SREBP-1. Food and Chemical Toxicology, 58, 198-209.

Histopathological Examination of the Effects of Tocilizumab and Dexamethasone on the Liver in Rats of Oleic Acid induced Acute Lung Injury

Year 2022, Volume: 11 Issue: 2, 167 - 172, 14.07.2022
https://doi.org/10.53424/balikesirsbd.989313

Abstract

Objective: This study, it was aimed to examine the effects of Tocilizumab and Dexamethasone on the liver in rats in which the acute lung injury model was created with Oleic acid. Materials and Methods: Thirty-six male rats were used as study material and were divided into six groups as Control, OA, TCZ and DEX. All groups except the control group received OA intravenously and after OA injections other compounds were administered intravenously (DEX 0.1 mg/kg and DEX 10 mg/kg, TCZ 2 mg/kg and TCZ 4 mg/kg). Results: It was determined that hydropic degeneration increased in hepatocytes in OA+ DEX 10 group rats compared to other groups (p<0.05). Vacuole formation in hepatocytes in OA+TCZ 2 and 4 groups was found to be similar to the control group (p<0.05). In addition, necrosis in hepatocytes and narrowing of sinusoids were detected more severely in OA+DEX (0.1 and 10 mg/kg) groups. Conclusion: As a result, it was determined that high dose Dexamethasone had hepatotoxic effect. It is thought that liver function tests should be performed and evaluated to determine the hepatotoxic effects of drugs to be used in the treatment of patients with COVID/19 ARDS.

Project Number

KÜ-BAP01/2019-12

References

  • Abdelhalim, M. A. K., & Jarrar, B. M. (2011). Gold nanoparticles induced cloudy swelling to hydropic degeneration, cytoplasmic hyaline vacuolation, polymorphism, binucleation, karyopyknosis, karyolysis, karyorrhexis and necrosis in the liver. Lipids in Health and Disease, 10(1), 1-6.
  • Akella, A., Sharma, P., Pandey, R., & Deshpande, S. B. (2014). Characterization of oleic acid-induced acute respiratory distress syndrome model in rat.
  • Beilman, G. (1995). Pathogenesis of oleic acid-induced lung injury in the rat: distribution of oleic acid during injury and early endothelial cell changes. Lipids, 30(9), 817-823.
  • Bleibel, W., Kim, S., D’Silva, K., & Lemmer, E. R. (2007). Drug-induced liver injury. Digestive diseases and sciences, 52(10), 2463-2471. Browning, J. D., & Horton, J. D. (2004). Molecular mediators of hepatic steatosis and liver injury. The Journal of clinical investigation, 114(2), 147-152.
  • Bulmuş, F. G., Gürsu, M. F., Muz, M. H., Yaman, İ., Bulmuş, Ö., & Sakin, F. (2013). Protective effects of alpha-lipoic acid on oleic acid-induced acute lung injury in rats. Balkan medical journal, 30(3), 309.
  • Canbay, A., Gieseler, R., Gores, G., & Gerken, G. (2005). The relationship between apoptosis and non-alcoholic fatty liver disease: an evolutionary cornerstone turned pathogenic. Zeitschrift für Gastroenterologie, 43(02), 211-217.
  • De Benedetti, F., Brunner, H. I., Ruperto, N., Kenwright, A., Wright, S., Calvo, I., Cuttica, R., Ravelli, A., Schneider, R., & Woo, P. (2012). Randomized trial of tocilizumab in systemic juvenile idiopathic arthritis. New England Journal of Medicine, 367(25), 2385-2395.
  • Drucker, C., Gewiese, J., Malchow, S., Scheller, J., & Rose-John, S. (2010). Impact of interleukin-6 classic-and trans-signaling on liver damage and regeneration. Journal of autoimmunity, 34(1), 29-37.
  • Ducheix, S., Montagner, A., Polizzi, A., Lasserre, F., Regnier, M., Marmugi, A., Benhamed, F., Bertrand-Michel, J., Mselli-Lakhal, L., & Loiseau, N. (2017). Dietary oleic acid regulates hepatic lipogenesis through a liver X receptor-dependent signaling. PLoS One, 12(7), e0181393.
  • Erol, N., Saglam, L., Saglam, Y. S., Erol, H. S., Altun, S., Aktas, M. S., & Halici, M. B. (2019). The protection potential of antioxidant vitamins against acute respiratory distress syndrome: a rat trial. Inflammation, 42(5), 1585-1594.
  • Herrero, R., Sánchez, G., Asensio, I., López, E., Ferruelo, A., Vaquero, J., Moreno, L., de Lorenzo, A., Bañares, R., & Lorente, J. A. (2020). Liver–lung interactions in acute respiratory distress syndrome. Intensive Care Medicine Experimental, 8(1), 1-13.
  • Kang, O., Kim, S., Seo, Y., Joung, D., Mun, S., Choi, J., Lee, Y., Kang, D., Lee, H., & Kwon, D. (2013). Curcumin decreases oleic acid-induced lipid accumulation via AMPK phosphorylation in hepatocarcinoma cells. Eur Rev Med Pharmacol Sci, 17(19), 2578-2586.
  • Koh, Y. (2014). Update in acute respiratory distress syndrome. Journal of intensive care, 2(1), 1-6.
  • Koksel, O., Kaplan, M. B., Ozdulger, A., Tamer, L., Degirmenci, U., Cinel, L., Bastürk, M., & Kanik, A. (2005). Oleic acid-induced lung injury in rats and effects of caffeic acid phenethyl ester. Experimental lung research, 31(5), 483-496.
  • Lee, J. Y., Moon, J. H., Park, J. S., Lee, B.-W., Kang, E. S., Ahn, C. W., Lee, H. C., & Cha, B. S. (2011). Dietary oleate has beneficial effects on every step of non-alcoholic Fatty liver disease progression in a methionine-and choline-deficient diet-fed animal model. Diabetes & metabolism journal, 35(5), 489.
  • Luo, P., Liu, Y., Qiu, L., Liu, X., Liu, D., & Li, J. (2020). Tocilizumab treatment in COVID‐19: a single center experience. Journal of medical virology, 92(7), 814-818.
  • Mahamid, M., Paz, K., Reuven, M., & Safadi, R. (2011). Hepatotoxicity due to tocilizumab and anakinra in rheumatoid arthritis: two case reports. International journal of general medicine, 4, 657.
  • Matute-Bello, G., Frevert, C. W., & Martin, T. R. (2008). Animal models of acute lung injury. American Journal of Physiology-Lung Cellular and Molecular Physiology, 295(3), L379-L399.
  • Mikiewicz, M., Otrocka-Domagała, I., Paździor-Czapula, K., & Rotkiewicz, T. (2017). Influence of long-term, high-dose dexamethasone administration on proliferation and apoptosis in porcine hepatocytes. Research in veterinary science, 112, 141-148.
  • Moore, B. B., & Hogaboam, C. M. (2008). Murine models of pulmonary fibrosis. American Journal of Physiology-Lung Cellular and Molecular Physiology, 294(2), L152-L160.
  • Muhović, D., Bojović, J., Bulatović, A., Vukčević, B., Ratković, M., Lazović, R., & Smolović, B. (2020). First case of drug‐induced liver injury associated with the use of tocilizumab in a patient with COVID‐19. Liver International, 40(8), 1901-1905.
  • Okun, J. G., Conway, S., Schmidt, K. V., Schumacher, J., Wang, X., de Guia, R., Zota, A., Klement, J., Seibert, O., & Peters, A. (2015). Molecular regulation of urea cycle function by the liver glucocorticoid receptor. Molecular metabolism, 4(10), 732-740.
  • Oldfield, V., Dhillon, S., & Plosker, G. L. (2009). Tocilizumab. Drugs, 69(5), 609-632.
  • Pascussi, J. M., Drocourt, L., Gerbal‐Chaloin, S., Fabre, J. M., Maurel, P., & Vilarem, M. J. (2001). Dual effect of dexamethasone on CYP3A4 gene expression in human hepatocytes: sequential role of glucocorticoid receptor and pregnane X receptor. European journal of biochemistry, 268(24), 6346-6358.
  • Patel, R., Patel, M., Tsai, R., Lin, V., Bookout, A. L., Zhang, Y., Magomedova, L., Li, T., Chan, J. F., & Budd, C. (2011). LXRβ is required for glucocorticoid-induced hyperglycemia and hepatosteatosis in mice. The Journal of clinical investigation, 121(1), 431-441.
  • Patterson, E., Yao, L., Ramic, N., Lewis, J., Cepinskas, G., McCaig, L., Veldhuizen, R., & Yamashita, C. (2013). Lung-derived mediators induce cytokine production in downstream organs via an NF-κB-dependent mechanism. Mediators of inflammation, 2013.
  • Roussel, D., Dumas, J.-F., Simard, G., Malthièry, Y., & Ritz, P. (2004). Kinetics and control of oxidative phosphorylation in rat liver mitochondria after dexamethasone treatment. Biochemical Journal, 382(2), 491-499.
  • Schuster, D. P. (1994). ARDS: clinical lessons from the oleic acid model of acute lung injury. American journal of respiratory and critical care medicine, 149(1), 245-260.
  • Schwartz, D. B., Bone, R. C., Balk, R. A., & Szidon, J. R. (1989). Hepatic dysfunction in the adult respiratory distress syndrome. Chest, 95(4), 871-875. Serviddio, G., Villani, R., Stallone, G., Scioscia, G., Foschino-Barbaro, M. P., & Lacedonia, D. (2020). <? covid19?> Tocilizumab and liver injury in patients with COVID-19. Therapeutic Advances in Gastroenterology, 13, 1756284820959183.
  • Slutsky, A. S., & Ranieri, V. M. (2013). Ventilator-induced lung injury. New England Journal of Medicine, 369(22), 2126-2136.
  • Sun, S., Liu, D., Zhang, H., Zhang, X., & Wan, B. (2019). Effect of different doses and time‑courses of corticosteroid treatment in patients with acute respiratory distress syndrome: A meta‑analysis. Experimental and therapeutic medicine, 18(6), 4637-4644.
  • Wan, E. S., Qiu, W., Baccarelli, A., Carey, V. J., Bacherman, H., Rennard, S. I., Agustí, A., Anderson, W. H., Lomas, D. A., & DeMeo, D. L. (2012). Systemic steroid exposure is associated with differential methylation in chronic obstructive pulmonary disease. American journal of respiratory and critical care medicine, 186(12), 1248-1255.
  • Wiener-Kronish, J., Albertine, K., & Matthay, M. (1991). Differential responses of the endothelial and epithelial barriers of the lung in sheep to Escherichia coli endotoxin. The Journal of clinical investigation, 88(3), 864-875.
  • Yang, P., Formanek, P., Scaglione, S., & Afshar, M. (2019). Risk factors and outcomes of acute respiratory distress syndrome in critically ill patients with cirrhosis. Hepatology Research, 49(3), 335-343.
  • Zhang, X., Morcos, P. N., Saito, T., & Terao, K. (2013). Clinical pharmacology of tocilizumab for the treatment of systemic juvenile idiopathic arthritis. Expert review of clinical pharmacology, 6(2), 123-137.
  • Zhang, X., Ouyang, J., & Thung, S. N. (2013). Histopathologic manifestations of drug-induced hepatotoxicity. Clinics in liver disease, 17(4), 547-564.
  • Zhang, X., & Peck, R. (2011). Clinical pharmacology of tocilizumab for the treatment of patients with rheumatoid arthritis. Expert review of clinical pharmacology, 4(5), 539-558.
  • Ziamajidi, N., Khaghani, S., Hassanzadeh, G., Vardasbi, S., Ahmadian, S., Nowrouzi, A., Ghaffari, S. M., & Abdirad, A. (2013). Amelioration by chicory seed extract of diabetes-and oleic acid-induced non-alcoholic fatty liver disease (NAFLD)/non-alcoholic steatohepatitis (NASH) via modulation of PPARα and SREBP-1. Food and Chemical Toxicology, 58, 198-209.
There are 38 citations in total.

Details

Primary Language English
Subjects Health Care Administration
Journal Section Articles
Authors

Funda Terzi 0000-0002-6184-5408

Hüseyin Serkan Erol 0000-0002-9121-536X

Project Number KÜ-BAP01/2019-12
Publication Date July 14, 2022
Submission Date August 31, 2021
Published in Issue Year 2022 Volume: 11 Issue: 2

Cite

APA Terzi, F., & Erol, H. S. (2022). Histopathological Examination of the Effects of Tocilizumab and Dexamethasone on the Liver in Rats of Oleic Acid induced Acute Lung Injury. Balıkesir Sağlık Bilimleri Dergisi, 11(2), 167-172. https://doi.org/10.53424/balikesirsbd.989313

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