Associations between Serum Kallistatin Levels and Markers of Glucose Homeostasis, Inflammation, and Lipoprotein Metabolism in Patients with Type 2 Diabetes and Nondiabetic Obesity.

Kallistatin is an endogenous serine proteinase inhibitor with various functions, including antioxidative, anti-inflammatory, and anti-atherosclerotic properties. To date, associations between kallistatin and lipoprotein subfractions are poorly investigated. In this study, we enrolled 62 obese patients with type 2 diabetes (T2D), 106 nondiabetic obese (NDO) subjects matched in gender, age, and body mass index, as well as 49 gender- and age-matched healthy, normal-weight controls. Serum kallistatin levels were measured with ELISA, and lipoprotein subfractions were analyzed using Lipoprint® (Quantimetrix Corp., Redondo Beach, CA, USA) gel electrophoresis. Kallistatin concentrations were significantly higher in T2D patients compared to NDO and control groups. We found significant positive correlations between very-low-density lipoprotein (VLDL), small high-density lipoprotein (HDL) subfractions, glucose, hemoglobin A1c (HbA1c), betatrophin, and kallistatin, while negative correlations were detected between mean low-density lipoprotein (LDL) size, large and intermediate HDL subfractions, and kallistatin in the whole study population. The best predictor of kallistatin was HbA1c in T2D patients, high-sensitivity C-reactive protein (hsCRP) and betatrophin in NDO patients, and hsCRP in controls. Our results indicate that kallistatin expression might be induced by persistent hyperglycemia in T2D, while in nondiabetic subjects, its production might be associated with systemic inflammation. The correlation of kallistatin with lipid subfractions may suggest its putative role in atherogenesis.

Gender-Dependent Associations between Serum Betatrophin Levels and Lipoprotein Subfractions in Diabetic and Nondiabetic Obese Patients.

Betatrophin, also known as angiopoietin-like protein 8 (ANGPTL8), mainly plays a role in lipid metabolism. To date, associations between betatrophin and lipoprotein subfractions are poorly investigated. For this study, 50 obese patients with type 2 diabetes (T2D) and 70 nondiabetic obese (NDO) subjects matched in gender, age, and body mass index (BMI) as well as 49 gender- and age-matched healthy, normal-weight controls were enrolled. Serum betatrophin levels were measured with ELISA, and lipoprotein subfractions were analyzed using Lipoprint gel electrophoresis. Betatrophin concentrations were found to be significantly higher in the T2D and NDO groups compared to the controls in all subjects and in females, but not in males. We found significant positive correlations between triglyceride, very low density lipoprotein (VLDL), large LDL (low density lipoprotein), small LDL, high density lipoprotein (HDL) -6-10 subfractions, and betatrophin, while negative correlations were detected between betatrophin and IDL, mean LDL size, and HDL-1-5. Proportion of small HDL was the best predictor of betatrophin in all subjects. Small LDL and large HDL subfractions were found to be the best predictors in females, while in males, VLDL was found to be the best predictor of betatrophin. Our results underline the significance of serum betatrophin measurement in the cardiovascular risk assessment of obese patients with and without T2D, but gender differences might be taken into consideration.

Impaired Organokine Regulation in Non-Diabetic Obese Subjects: Halfway to the Cardiometabolic Danger Zone.

Altered organokine expression contributes to increased cardiometabolic risk in obesity. Our aim was to evaluate the associations of serum afamin with glucose homeostasis, atherogenic dyslipidemia, and other adipokines in severe obesity to clarify the early metabolic alterations. 106 non-diabetic obese (NDO) subjects and 62 obese patients with type 2 diabetes matched for age, gender, and body mass index (BMI) were enrolled in this study. We compared their data with 49 healthy, lean controls. Serum afamin and retinol-binding protein 4 (RBP4), as well as plasma plasminogen activator inhibitor-1 (PAI-1), were measured with ELISA, and lipoprotein subfractions were analyzed using Lipoprint gel electrophoresis. Afamin and PAI-1 found to be significantly higher in the NDO and T2M group (p < 0.001 and p < 0.001, respectively) than in the controls. In contrast, RBP4 was unexpectedly lower in the NDO and T2DM group compared to controls (p < 0.001). Afamin showed negative correlations with mean LDL size and RBP4, but positive correlations with anthropometric, glucose/lipid parameters, and PAI-1 in both the overall patients and the in NDO + T2DM groups. BMI, glucose, intermediate HDL, and small HDL were predictors of afamin. Afamin may serve as a biomarker for the severity of cardiometabolic disturbances in obesity. The complexity of organokine patterns in NDO subjects draws attention to the diverse spectrum of obesity-related comorbidities.

Glucocorticoids Directly Affect Hyaluronan Production of Orbital Fibroblasts; A Potential Pleiotropic Effect in Graves' Orbitopathy.

Orbital connective tissue expansion is a hallmark of Graves' orbitopathy (GO). In moderate-to-severe active GO, glucocorticoids (GC) are the first line of treatment. Here we show that hydrocortisone (HC), prednisolone (P), methylprednisolone (MP), and dexamethasone (DEX) inhibit the hyaluronan (HA) production of orbital (OF) and dermal (DF) fibroblasts. HA production of GO OFs (n = 4), NON-GO OFs (n = 4) and DFs (n = 4) was measured by ELISA. mRNA expression of enzymes of HA metabolism and fibroblast proliferation was examined by RT-PCR and BrdU incorporation, respectively. After 24 h of GC treatment (1µM) HA production decreased by an average of 67.9 ± 3.11% (p < 0.0001) in all cell cultures. HAS2, HAS3 and HYAL1 expression in OFs also decreased (p = 0.009, p = 0.0005 and p = 0.015, respectively). Ten ng/mL PDGF-BB increased HA production and fibroblast proliferation in all cell lines (p < 0.0001); GC treatment remained effective and reduced HA production under PDGF-BB-stimulated conditions (p < 0.0001). MP and DEX reduced (p < 0.001, p = 0.002, respectively) PDGF-BB-induced HAS2 expression in OFs. MP and DEX treatment decreased PDGF-BB stimulated HAS3 expression (p = 0.035 and p = 0.029, respectively). None of the GCs tested reduced the PDGF-BB stimulated proliferation rate. Our results confirm that GCs directly reduce the HA production of OFs, which may contribute to the beneficial effect of GCs in GO.

Serum afamin and its implications in adult growth hormone deficiency: a prospective GH-withdrawal study.

Introduction: Adult growth hormone deficiency (AGHD) is associated with a high prevalence of metabolic syndrome (MS), which contributes to the unfavorable cardiovascular risk profile in these patients. Insulin like growth factor-1 (IGF-1) is a widely used biomarker, however it does not always reflect the cardiometabolic risk and has a poor relationship with clinical efficacy endpoints. Consequently, there is an unmet need for biomarkers to monitor responses to GH-replacement. Afamin is a hormone-like glycoprotein, expressed in the liver. Higher afamin levels are strongly associated with MS and insulin resistance (IR). Although both MS and IR are very common in AGHD, afamin has not been investigated in these patients. Purpose: To investigate afamin as a potential biomarker in patients with AGHD. Materials and methods: Participants included 20 AGHD patients (11 GH-substituted and 9 GH-unsubstituted) and 37 healthy controls. Subjects underwent routine laboratory examinations, anthropometric measurements, body composition analysis using multi-frequency bioelectrical impedance analysis (InBody720) and measurement of serum afamin concentrations. In GH-substituted subjects, GH-substitution was withdrawn for 2 months. Measurements were carried out right before GH-withdrawal, at the end of the 2-month withdrawal period, and 1 month after reinstituting GH-replacement therapy (GHRT). Results: GH-unsubstituted patients demonstrated higher afamin levels compared to controls (p=0.03). Afamin positively correlated with skeletal muscle mass, bone mineral content, total body water, extracellular- and intracellular water content, insulin (all, p<0.01), HOMA-IR (p=0.01) and C-peptide (p=0.03) levels in AGHD but not in healthy controls. In GH-substituted patients 2-month of GH-withdrawal caused significant changes in body composition, including decreased fat-free mass, skeletal muscle mass, total body water, and intracellular water content (all, p<0.01); but these changes almost fully recovered 1 month after reinstituting GHRT. Unexpectedly, afamin levels decreased after GH-withdrawal (p=0.03) and increased with reinstitution (p<0.01). Changes of afamin levels during GH-withdrawal positively correlated with changes of HOMA-IR (r=0.80; p<0.01) and changes of insulin (r=0.71; p=0.02). Conclusion: Higher afamin levels in unsubstituted AGHD patients might indicate severe metabolic dysregulation. Significant changes accompanying GH-withdrawal and reinstitution, along with strong correlations with measures of IR, suggest that afamin could be a promising biomarker to monitor GHRT-associated changes of insulin sensitivity.

Low Serum Fibroblast Growth Factor 21 Level and Its Altered Regulation by Thyroid Hormones in Patients with Hashimoto's Thyroiditis on Levothyroxine Substitution.

BACKGROUND/OBJECTIVES: Fibroblast growth factor 21 (FGF21) is a hormonal regulator of lipid and glucose metabolism exerting protection against atherosclerosis by multiple actions on the blood vessels, liver, and adipose tissues. We aimed to investigate serum FGF21 level and its relation to thyroid hormones and metabolic parameters among patients with Hashimoto's thyroiditis (HT). METHODS: Eighty patients with HT on levothyroxine treatment and eighty-two age- and BMI-matched adults without thyroid disease serving as controls were enrolled. Serum FGF21 concentrations were determined with an enzyme-linked immunosorbent assay. RESULTS: Median serum FGF21 level was significantly lower in HT patients compared with controls (74.2 (33.4-148.3) pg/mL vs. 131.9 (44.8-236.3) pg/mL; p = 0.03). We found a positive correlation between FGF21 and age, triglyceride, total cholesterol, and low-density lipoprotein cholesterol in both groups, while thyroid stimulating hormone and C-reactive protein showed a positive correlation, and thyroxine had an inverse correlation with FGF21 only in control subjects. According to multiple regression analyses, thyroid status is the main predictor of FGF21 in healthy controls, while it is not a significant predictor of FGF21 among HT patients on levothyroxine supplementation therapy. CONCLUSIONS: Our results indicate that the physiological role of thyroid function in the regulation of FGF21 synthesis is impaired in HT patients, which may contribute to the metabolic alterations characteristic of HT patients.

Advanced glycation end products and their soluble receptor (sRAGE) in patients with Hashimoto's thyroiditis on levothyroxine substitution.

Background: Advanced glycation end products (AGEs) are heterogenous group of irreversible chemical moieties originated from non-enzymatic glycation and oxidation of proteins, nucleic acids, and lipids. The engagement of AGEs with their chief cellular receptor (RAGE) activates a myriad of signaling pathways contributing to the progression of chronic diseases like autoimmune thyroiditis, type 2 diabetes mellitus and its complications. Soluble RAGE (sRAGE) prevents AGE-RAGE interaction in a competitive manner. Objective: We investigated the association between serum AGE, sRAGE and thyroid function in 73 Hashimoto thyroiditis patients (HT) on levothyroxine substitution, and in 83 age, BMI and gender-matched healthy controls. Methods: The serum AGEs levels were determined by autofluorescence on a multi-mode microplate reader, and the serum sRAGE levels by ELISA method. Results: Mean AGE level was lower (10.71 vs 11.45 AU/µg protein; p=0.046), while mean sRAGE level was higher (923 vs 755 pg/mL; p<0.0005) in the serum of HT patients than the controls. AGE correlated with age, while sRAGE correlated negatively with BMI in both groups. We found negative correlation between AGE and fT3 levels (r=-0.32; p=0.006) and sRAGE and TSH levels (r=-0.27; p=0.022) in HT patients, while we failed to find association between AGE, sRAGE and parameters of thyroid function in the control group. Median AGE/sRAGE ratio was lower in HT patients than in controls (2.4, IQR 1.9 - 3.1 vs 3.3, IQR 2.3 - 4.1 AU/pg; p < 0.001). In HT patients, the AGE/sRAGE ratio correlated positively with BMI and correlated negatively with fT3. Conclusion: According to our results in HT patients lower TSH and higher fT3 levels within the reference range is accompanied by a favorable AGE/RAGE balance. Further investigations are needed to confirm these results.