Abnormal Glucose Metabolism and Insulin Resistance Are Induced via the IRE1α/XBP-1 Pathway in Subclinical Hypothyroidism

Abstract

Subclinical hypothyroidism (SCH) and diabetes mellitus are closely related and often occur together in individuals. However, the underlying mechanism of this association is still uncertain. In this study we re-analyzed the data of a mature database (NHANES, 1999 ~ 2002) and found that both fasting plasma glucose levels and the proportion of hyperglycemic subjects among SCH patients were higher than that found in euthyroid controls. SCH was also associated with a 2.29-fold increased risk for diabetes. Subsequently, we established an SCH mouse model and subjected it to an oral glucose tolerance test (OGTT) and an insulin tolerance test (ITT). SCH mice exhibited impaired glucose and insulin tolerance. Increased HOMA-IR and decreased ISI indexes, indicating insulin resistance (IR), were also observed in the SCH state. Hepatic ERp29 and Bip, as well as IRE1α and XBP-1s, were induced significantly in SCH mice, suggesting the induction of endoplasmic reticulum (ER) stress, particularly involving the IRE1α/XBP-1s pathway. Interestingly, when we relieved ER stress using 4-phenyl butyric acid, abnormal glucose metabolism, and IR status in SCH mice were improved. Our findings suggest that ER stress, predominantly involving the IRE1α/XBP-1s pathway, may play a pivotal role in abnormal glucose metabolism and IR in SCH that may help develop potential strategies for the prevention and treatment of diabetes.

Keywords: ER stress; IRE1α/XBP-1; Subclinical hypothyroidism; glucose impairment; insulin resistance.

Figure 1

Abnormal Glucose metabolism in SCH patients. (A) Plasma glucose distributions in different group (euthyroid controls and SCH patients), data were expressed as: median (P25, P75). (B) The proportion of the glucose categories in different group. (C) The prevalence of diabetes in different group. The error bars represent the 95% CI of the ratio. (D) The odds ratio of diabetes in different group.

Figure 2

SCH mice exhibited abnormal glucose metabolism and IR. Male C57/BL6 mice were given methimazole (MMI, 0.08 mg/kg·BW·d, SCH group) or a corresponding volume of vehicle (control group). (A) Oral glucose tolerance test (OGTT) was implemented to detect glucose tolerance at the 13th week of administering MMI to mice in the drinking water. (B) Insulin tolerance test (ITT) was implemented to assess insulin-stimulated glucose disposal at the 14th week of administering MMI to mice in the drinking water. (C) The fasting plasma glucose level was assayed at the 14th week (n = 4–6). (D) The fasting plasma insulin level was assayed at the 14th week (n = 4–6). (E) Homeostasis Model Assessment-Insulin Resistance (HOMA-IR) index was calculated. (F) Insulin sensitivity index (ISI) was calculated. The data are presented as the mean ± SD. ^*p < 0.05 compared with control.

Figure 3

Hepatic ER stress was induced in SCH mice. Male C57/BL6 mice were given methimazole (MMI, 0.08 mg/kg·BW·d, SCH group) or a corresponding volume of vehicle (control group) for 14 weeks, the expression of proteins in ERS pathway were detected by western blot (n = 4–6). (A) The protein expression of ERq29 was detected by western blot. (B) The mRNA expression of ERq29 was detected by Real-time PCR. (C) The expression of proteins in the Bip/IRE1α/XBP-1 pathway were detected by western blot. The data are presented as the mean ± SD. ^*p < 0.05 compared with control.

Figure 4

4-PBA alleviated hepatic ERS in SCH mice. We established SCH mouse model using MMI. When MMI was applied for 14 weeks, 4-PBA was given at a dose of 100 mg/kg·BW·d for 4 weeks, and mice were divided into four subgroups (as shown in Supplementary Figure 1): vehicle treated control mice group (con + veh group), 4-PBA treated control mice group (con + 4-PBA group), vehicle treated SCH mice group (SCH + vehicle group), and 4-PBA treated SCH mice group (SCH + 4-PBA group). (A) The protein expression of ERq29 was detected by western blot (n = 4–6). (B) The mRNA expression of ERq29 was detected by Real-time PCR. (C) The expression of proteins in the Bip/IRE1α/XBP-1 pathway were detected by western blot (n = 4–6). The data are presented as the mean ± SD. ^*p < 0.05 compared with control. ^#p < 0.05 compared with SCH.

Figure 5

4-PBA alleviated abnormal glucose metabolism and IR in SCH mice. (A) Oral glucose tolerance test (OGTT) was implemented to detect glucose tolerance at the 17th week of administering MMI to mice in the drinking water (n = 4–6). (B) Insulin tolerance test (ITT) was implemented to assess insulin-stimulated glucose disposal at the 18th week of administering MMI to mice in the drinking water (n = 4–6). (C) The fasting plasma glucose level was assayed at the 18th week (n = 3–4). (D) The fasting plasma insulin level was assayed at the 18th week (n = 3–4). (E) Homeostasis Model Assessment-Insulin Resistance (HOMA-IR) index was calculated. (F) Insulin sensitivity index (ISI) was calculated. The data are presented as the mean ± SD. ^*p < 0.05 compared with vehicle treated mice group. #p < 0.05 compared with 4-PBA treated SCH mice group.