The Relation between Thyroid Dysfunction and Metabolic Dysfunction-Associated Fatty Liver Disease in Egyptian Patients | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| Afro-Egyptian Journal of Infectious and Endemic Diseases | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| Article 5, Volume 13, Issue 3, September 2023, Pages 174-180 PDF (285.29 K) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| Document Type: Original Article | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| DOI: 10.21608/aeji.2023.223013.1302 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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| Shimaa M Mansour* ; Fatma A Elgebaly; Eslam A Habba | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| Department of Tropical Medicine &Infectious diseases , Faculty of Medicine ,Tanta University, Egypt. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| Abstract | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| Background and study aim: Metabolic-dysfunction Associated Fatty Liver Disease ( MAFLD) caused by multiple risk factors like obesity, hepatitis C virus infection, and diabetes mellitus . One of the most important risk factors which may be implicated in MAFLD is thyroid dysfunction due to its important effects on hepatic fatty acid and synthesis of cholesterol. So, in this study we aimed to assess the association between thyroid dysfunction and MAFLD in Egyptian patients. Patients and Methods: This study was carried out on 100 patients who attended to Tanta tropical medicine outpatient clinic. They were older than 18 years fulfilling the criteria of MAFLD and thyroid functions were done for them . Results: There were 67% cases euthyroidism, 29% cases subclinical hypothyroidism(SCH) and 4% cases overt hypothyroidism. The number of females with subclinical hypothyroidism were 22(40%) but males were 7 (15.6%). Overt hypothyroidism was present in 4(7.3%) females ,but in males was 0. Conclusion: One of the most common endocrine abnormalities which may be present in MAFLD patients is thyroid dysfunction particularly subclinical hypothyroidism especially in females . | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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| MAFLD; subclinical hypothyroidism; overt hypothyroidism; euthyroidism; body mass index | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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INTRODUCTION One of the most common causes of chronic liver diseases worldwide is Metabolic dysfunction-associated fatty liver disease (MAFLD), it affects about 30% of the adults [1,2]. MAFLD means fatty infiltration of the liver (FLD) associated with metabolic dysfunctions. Some studies reported that the complications of MAFLD made it one of the most common causes of liver transplantation [3]. MAFLD has complex risk factors such as obesity, hypertension, atherogenic dyslipidaemia, Prothro-mbotic disorders, and Pro-inflammatory conditions and diabetes [4]. Dysfunctions of thyroid gland have been implicated as one of the most important risk factors of MAFLD due to its important role on hepatic synthesis of fatty acid and cholesterol [5, 6]. Maintaining liver metabolism needs thyroid function to be normal ; while thyroid disorders may lead to liver disease progression [7]. All thyroid hormones give an image for the decrease of thyroid function, while high TSH level more than the normal range usually means hypoactivity of the thyroid gland , as in subclinical hypothyroidism (SCH) [8]. SCH means elevated plasma level of TSH while FT4 level is normal, with a prevalence about 4–20% [9]. A previous study reported that the patients suffering from SCH with fatty liver diseases (FLD) had metabolic abnormalities more than those with SCH and a normal liver [10]. The studies that investigate the prevalence of thyroid dysfunction among the patients with MAFLD are not sufficient [9, 11, 12] . So In this study, we tried to study the thyroid function in MAFLD patients and to assess the prevalence of thyroid dysfunction in patients with MAFLD in Egyptian patients. PATIENTS AND METHODS This cross-sectional study carried out on 100 patients who attended to Tanta tropical medicine outpatient clinic, faculty of medicine. From October 2021 until October 2022. This study included male or female patients more than 18 years old with MAFLD which is diagnosed when hepatic steatosis was present in addition to the presence of three or more of the following risk determinants: 1) increased waist circumference (>102 cm [>40 in] for men, >88 cm [>35 in] for women); 2) elevated triglycerides (≥150 mg/dl); 3) low HDL cholesterol ( [8]. While we excluded patients who aged < 18 years , taking medication that may affect thyroid functions or lipid levels , had alcoholic liver disease taking more than 40g of alcohol (or four units) per day , had viral hepatitis ,Pregnant women and who unwilling to participate in our study. All the patients were subjected to; Personal history (name, age, sex, occupation, residence and marital state),Anthropometric measures: weight, length, BMI, waist circumference and complaint. In addition to FreeT3, Free T4, TSH. ,Lipid profile (triglycerides, HDL, LDH, cholesterol level) , Fasting Blood glucose level and or HBA1C, PCR for HCV for exclusion of viral hepatitis , liver enzymes , ultrasound on abdomen and pelvis for evaluation of liver condition and Fibroscan were done for all patients. Fibroscan was used to assess the stages of fibrosis and steatosis using Dimensional ultrasound TE (transient elastography). Dimensional ultrasound TE (transient elastography) was used for staging liver fibrosis and steatosis by measuring the velocity of a low-frequency (50 Hz) elastic shear wave propagating through the liver. This velocity has direct relation to the stiffness of the tissue, called the elastic modules (expressed as E=3qv2, where v is the shear velocity and q is the density of tissue, assumed to be constant). When the tissue is stiffer, the shear wave is propagating faster. The results are expressed in Kilopascals (KPa) and range from 1.5 to 75 KPa with 5 KPa as normal value higher in men and in patients with low or high body mass index (BMI) (U-shaped distribution). The CAP score (Controlled attenuation parameter) is measured in decibels per meter (dB/m) and ranges from 100-400 as following , S0 :292.3 . The liver stiffness is measured by the fibrosis score which is an indication of scarring. A fibrosis score F0 to F1 ( 2 to 7 kPa) means there is little or no scarring on the liver, while F2 (7.5 to 10 kPa) means moderate scarring that has spread outside the liver, while F3 (10 to 14 kPa) indicates severe scarring which has spread and disrupts normal blood flow, finally F4 ( 14 kPa or higher) means late-stage scarring or cirrhosis, where the scarring is permanent and the damage is irreversible. Statistical analysis: The organization, tabulation, presentation, and analysis of data were performed by using SPSS IBM Chicago, version 23. Qualitative data was divided into categories and presented as frequency number and percentage, with the chi square test used to determine the relationship between groups. Quantitative data was presented as mean ± SD and the relationship between groups was done by using independent student t test. The level of significance adopted was p < 0.05. RESULTS Study population consisted of 100 subjects including 45 men and 55 women. There were 50 patients with diabetes mellitus and 64 patients were hypertension, as demonstrated in table (1). There were 67% cases Euthyroidism , 29% cases Subclinical hypothyroidism and 4% cases Overt hypothyroidism. as demonstrated in table (2). Subclinical hypothyroidism, defined as elevated TSH with free T4 concentrations at the lower end of the euthyroid range. In overt primary hypothyroidism, TSH levels are high and T4 and T3 levels are low. The number of females with Subclinical hypothyroidism are 22(40%) but the number of males with subclinical hypothyroidism was 7 (15.6%). Overt hypothyroidism was present in 4(7.3%) female cases , with female predominance ( p= 0.002), as demonstrated in table (3). Regarding steatosis grades ,there were 62 patients grade 1 , 29 patients grade 2 and 9 patients grade 3 . while regarding fibrosis stages ,there were 5 patients stage 0 , 67 patients stage 1 , 26 patients stage 3 and 2 patients stage 4 . Mean and standard deviation of THS among studied group (4.4±1.9 uIU/mL) with normal reference range (0.3-6 uIU/mL), Mean and standard deviation of T3 among studied group (5.1±1.3 pg/ml) with normal reference range (3-7.8 pg/ml) Mean and standard deviation of T4 among studied group (16.5±3.2 ng/dl) with normal reference range (12-22 ng/dl), as demonstrated in table (4). There was a significant difference in waist circumference, HDL and triglyceride among different thyroid dysfunction groups. There was a difference in LDL and FBS, but not statistically significant ,as demonstrated in table (5) . There was a significant positive correlation between TSH (waist circumference and TG), but there was a significant negative correlation between TSH and HDL. There was a significant negative correlation between T3 (waist circumference, TG and HDL), There was a significant negative correlation between T4 (waist circumference, TG and HDL) and regarding steatosis grade there was a significant positive correlation between the grade of steatosis and the level of TSH , as demonstrated in table (6). But there was no significant difference between the grade of steatosis and T3, T4 levels. Table (1): General characteristics of the studied patients.
DM: diabetes mellitus, SD: standard deviation Table (2): Thyroid Status of the study Population.
Table (3): Relation between gender and thyroid functions groups.
p: p value for comparing between the two studied groups *: Statistically significant at p ≤ 0.05 , chi-square test Table (4): Clinical data among the studied group.
ALT :alanine aminotransferase AST :aspartate transaminase ,HDL : high-density lipoproteins ,LDL: low-density Lipoproteins, TG: triglyceride, FBS: fasting blood sugar , HbA1C: hemoglobin A1C, S: steatosis, TSH: thyroid stimulating hormone , Free T3 :Free triiodothyronine , Free T4 : Free thyroxine SD: standard deviation Table (5): Components of metabolic syndrome among different thyroid dysfunction groups.
DM : diabetes mellitus * Statistically significant at p ≤ 0.05 , chi- square test ,F test Table (6): Correlation between metabolic data and thyroid function tests.
TSH: thyroid stimulating hormone , Free T3 :Free triiodothyronine , Free T4 : Free thyroxine * Statistically significant at p ≤ 0.05, R test DISCUSSION In 2020 the MAFLD is prepared to replace NAFLD definition . So, many studies had been made to find the associations between thyroid dysfunction and NAFLD, reported that NAFLD patients are always characterized by low T4, FT4 levels and high TSH levels [13]. The prevalence of thyroid dysfunction and hypothyroidism in metabolic syndrome patients is higher than the prevalence in the normal population, which is 5.9% for thyroid dysfunction and 4.6% for hypothyroidism (0.3% overt and 4.3% subclinical hypothyroidism) [14,15] . low-normal thyroid function was much more prevalent than SCH (21.70% vs. 4.16% in the general population and 21.97% vs. 4.22% in patients with MAFLD) [16]. There are several possible mechanisms which can explain the relationship between hypothyroidism and MAFLD such as insulin resistance , dyslipidemia and obesity and they have important roles in the development of MAFLD, both insulin resistance and obesity are vital factors in the development of MAFLD, which are also common in hypothyroidism patients comparing with general population, in our study thyroid dysfunction prevalence was 33% among MAFLD patients [17]. Metabolism has a sex dimorphism, which results in different disease risks for men and women, there are several studies including the general population reported that females had thyroid dysfunction more than males [18] this is in agreement with our result as the percentage of females with subclinical hypothyroidism were (40%) but males with subclinical hypothyroidism were (15.6%). Thyroid hormones are recognized as catabolic hormones and they regulate various processes of metabolism, including the synthesis, mobilization, and breakdown of lipids. Hypothyroidism had been reported to be associated with an increased risk for dyslipidemia and atherosclerotic cardiovascular disease, in our study there was a significant negative correlation between T4 and TG and this was in agreement with Jang J, et al. that found a negative association between triglycerides and FT4 [19] . waist circumference was significantly different between patients with and without thyroid dysfunction this was in agreement with Khatiwada S, et al[20]. The positive association of TSH with TC and LDL-c may be due to autoimmune activation involving lipoprotein(a). In our study HDL cholesterol had a significant negative association with TSH level and this was reported also by Khatiwada S, et aland Rajendra Kc ,et al [20,21]. The study done in India founded that subclinical hypothyroidism had a significant association between the levels of TSH and cholesterol level ,triglycerides, LDL, and HDL across the metabolic syndrome group [22]. In our study there was a significant positive correlation between TSH and waist circumference, a significant positive correlation between TSH and TG, and significant negative correlation between TSH and HDL. However, a study done in Turkey showed that TSH was not correlated with any metabolic syndrome parameters [23]. But the study done by Fan et al , reported that patients suffered from thyroid hypofunction were more likely to be old age , females , and total cholesterol, , low-density lipoprotein (LDL), triglycerides were high in them [16]. Higher TSH levels can induce steatosis via TSH receptor (TSHR) signaling. TSH not only contributes to the negative feedback regulation of T4 secretion via TSHR in the thyroid gland but may also increase hepatic gluconeogenesis , decrease hepatic bile acid synthesis, and cause hypercholesterolemia by decreasing 3-hydroxy-3-methylglutaryl-CoAreductase phosphorylation, In this study we reported that the grades of steatosis in MAFLD patients were positively correlated with the level of TSH and this was the same results found by Chung et al and Xu et al,who reported thatgrades of fatty liver increased more in patients who had high TSH and this relationship was statistically significant (p = 0.001) ,this was reported by other studies that identified a positive association between NAFLD and TSH [24,25]. Thestudies done by Carulli et al. and Pagadala et al, suggested that the serum TSH concentration is correlated with the severity of the hepatic steatosis [26,27] . On the other hand Ittermann et al, detected no consistent association between serum TSH concentrations and hepatic steatosis[28]. While other study showed highly significant relationship between serum TSH levels and increasing the grades of fatty liver [29]. This study had some limitations: firstly, fatty liver disease was diagnosed by ultrasound and fibroscan, secondly, the sample size in our study is relatively small and finally, the study population was from one hospital and cannot represent the general population. Therefore, findings need to be validated using more sophisticated techniques as liver biopsies and a more representative population. CONCLUSION In conclusion, keeping in mind that thyroid dysfunction particularly subclinical hypothyroidism is one of the most common endocrine abnormalities that may occur in MAFLD especially in females .hence, thyroid function tests may be included as a routine investigation in all MAFLD patients especially females. Abbreviation MAFLD: Metabolic associated fatty liver diseases. SCH: Subclinical hypothyroidism . NAFLD: non alcoholic fatty liver disease . DM : diabetes mellitus ALT :alanine aminotransferase AST :aspartate transaminase HDL : high-density lipoproteins LDL: low-density Lipoproteins TG: triglyceride FBS: fasting blood sugar HbA1C: hemoglobin A1C, S: steatosis CAP: Controlled attenuation parameter TSH: thyroid stimulating hormone Free T3 :Free triiodothyronine Free T4 : Free thyroxine SD: standard deviation Funding: None. Conflict of interest: None. Ethical consideration: A written informed consent was signed by all patients and a code number for each patient was used, The study was approved by the Ethical Committee of Faculty of Medicine Tanta University (approval code 34913/9/21)
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| References | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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1.Le MH, Yeo YH, Li X, Li J, Zou B, Wu Y, et al 2019 global NAFLD prevalence: a systematic review and metaanalysis. Clin Gastroenterol Hepatol. 2021; 20(12):2809–17.
2.Estes C, Anstee QM, Arias-Loste MT, Bantel H, Bellentani S, Caballeria J, et al. Modeling NAFLD disease burden in China, France, Germany, Italy, Japan, Spain, United Kingdom, and United States for the period 2016–2030. J Hepatol. 2018;69(4):896–904.
3.Pais R, Barritt ASt, Calmus Y, Scatton O, Runge T, Lebray P, et al. NAFLD and liver transplantation: current burden and expected challenges. J Hepatol. 2016;65(6):1245–57.
4.Younossi Z, Anstee QM, Marietti M, Hardy T, Henry L, Eslam M,et al. Global burden of NAFLD and NASH: trends, predictions, risk factors and prevention. Nat Rev Gastroenterol Hepatol. 2018;15(1):11–20.
5.Sinha RA, Singh BK and Yen PM. Direct effects of thyroid hormones on hepatic lipid metabolism. Nat Rev Endocrinol. 2018; 14(5):259–69.
6.Ritter MJ, Amano I and Hollenberg AN. Thyroid hormone signalling and the liver. Hepatology. 2020; 72(2):742–52.
7.Jones DD, May KE, Geraci SA. Subclinical thyroid disease. Am J Med.2010;123(6):502–4.
8.Zhou W, Brumpton B, Kabil O, Gudmundsson J, Thorleifsson G, Weinstock J, et al. GWAS of thyroid stimulating hormone highlights pleiotropic effects and inverse association with thyroid cancer. Nat Commun. 2020;11(1):3981.
9.Cooper DS and Biondi B. Subclinical thyroid disease. Lancet. 2012;379(9821):1142–54.
10.Posadas-Romero C, Jorge-Galarza E, Posadas-Sánchez R, Acuña-Valerio J, Juárez-Rojas JG, Kimura-Hayama E, et al . Fatty liver largely explains associations of subclinical hypothyroidism with insulin resistance, metabolic syndrome, and subclinical coronary atherosclerosis. Eur J Endocrinol. 2014; 171(3):319–25.
11.van Tienhoven-Wind LJ and Dullaart RP. Low-normal thyroid function and the pathogenesis of common cardio-metabolic disorders. Eur J Clin Invest. 2015; 45(5):494–503.
12.Mitchell F.Thyroid function: low T4 levels a risk factor for fatty liver? Nat Rev Endocrinol. 2012;8(8):447.
13.van den Berg EH, van Tienhoven-Wind LJ, Amini M, Schreuder TC, Faber KN, Blokzijl H and Dullaart RP. Higher free triiodothyronine is associated with non-alcoholic fatty liver disease in euthyroid subjects: the lifelines cohort study. Metabolism. 2017;67:62–71.
14.Gyawali P, Takanche JS, Shrestha RK, Bhattarai P, Khanal K, Risal P, et al. Pattern of thyroid dysfunction in patients with metabolic syndrome and its relationship with components of metabolic syndrome. Diabetes Metab J. 2015; 39(1):66–73.
15.Hak AE, Pols HA, Visser TJ, Drexhage HA, Hofman A and Witteman JC. Subclinical hypothyroidism is an independent risk factor for atherosclerosis and myocardial infarction; The Rotterdam study. Ann Intern Med 2000; 132:270-278.
16.Fan H, Li L, Liu Z , Zhang P , Wu S, HanX, et al. Low thyroid function is associated with an increased risk of advanced fibrosis in patients with metabolic dysfunction-associated fatty liver disease. BMC Gastroenterology 2023; 23:3
17.Gluvic Z, Zaric B, Resanovic I, Obradovic M, Mitrovic A, Radak D, et al. Link between metabolic syndrome and insulin resistance. Curr Vasc Pharmacol 2017; 15(1):30–9.
18.Shantha GP, Kumar AA, Jeyachandran V, Rajamanickam D, Rajkumar K, Salim S, et al. Association between primary hypothyroidism and metabolic syndrome and the role of C reactive protein: a cross-sectional study from South India. Thyroid Res. 2009; 2(1):2. doi:10.1186/1756-6614-2-2.
19.Jang J, Kim Y, Shin J, Lee S, Choi Y and Park E: Association between thyroid hormones and the components of metabolic syndrome, BMC Endocrine Disorders 2018. 21; 18(1):29.
20.Khatiwada S, Sah S, Rajendra K, Baral N and Lamsal M: Thyroid dysfunction in metabolic syndrome patients and its relationship with components of metabolic syndrome, Clinical Diabetes and Endocrinology. 2016 1;2:3.
21.Rajendra Kc , Khatiwada S, Mehta K, Pandey P, Lamsal M and Majhi S. Cardiovascular Risk Factors in Subclinical Hypothyroidism: A Case Control Study in Nepalese Population. J Thyroid Res. 2015; 2015:305241
22.Meher L, Raveendranathan S, Kota S, Sarangi J and Jali S. Prevalence of hypothyroidism in patients with metabolic syndrome. Thyroid Res Pract. 2013;10:60–4.
23.Tarcin O, Abanonu GB, Yazici D and Tarcin O. Association of metabolic syndrome parameters with TT3 and FT3/FT4 ratio in obese Turkish population. Metab Syndr Relat Disord. 2012;10(2):137–42
24.Chung GE, Kim D, Kim W, Yim JY, Park MJ, Kim YJ, et al. Non-alcoholic fatty liver disease across the spectrum of hypothyroidism. J Hepatol 2012;57(1):150-156.
25.Xu C, Xu L, Yu C, Miao M and Li Y Association between thyroid function and nonalcoholic fatty liver disease in euthyroid elderly Chinese. Clin Endocrinol 2011; 75(2):240-246.
26.Carulli L, Ballestri S, Lonardo A, Lamiet F, Violi E , Losi L, et al. Is nonalcoholic steatohepatitis associated with a high-though-normal thyroid stimulating hormone level and lower cholesterol levels? Int Emerg Med 2013; 8(4):297-305.
27.Pagadala MR, Zein CO, Dasarathy S, Yerian L, Lopez R and McCullough A Prevalence of hypothyroidism in nonalcoholic fatty liver disease. Digest Dis Sci 2012;57(2):528-34
28.Ittermann T, Haring R, Wallaschofski H, Baumeister S , Nauck M , Dörr M ,et al. Inverse association between serum free thyroxine levels and hepatic steatosis: results from the Study of Health in Pomerania. Thyroid 2012;22(6):568-574.
29.Bano A, Chaker L, Plompen EPC, Hofman A, Dehghan A . Franco O, et al. Thyroid function and the risk of nonalcoholic fatty liver disease: the Rotterdam study. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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