THE EXPRESSION AND SIGNIFICANCE OF SERUM TRAb, TSAb, AND TSBAb IN PATIENTS WITH HASHIMOTO’S THYROIDITIS WITH HYPOTHYROIDISM

Objective To investigate the changes in serum thyrotropin receptor antibody (TRAb), thyroid stimulating antibody (TSAb), and thyroid-stimulating blocking antibody (TSBAb) and their significance in patients with Hashimoto’s thyroiditis with hypothyroidism (HT hypothyroidism). Methods A total of 133 patients with HT hypothyroidism (HT hypothyroidism group) and 125 healthy subjects (control group) were enrolled and their clinical data were collected. The serum levels of free triiodothyronine (FT3), free thyroxine (FT4), thyrotropin (TSH), and TRAb were measured and compared between the two groups. TSAb and TSBAb were also compared across the patients who were positive for TRAb (group B), who were negative for TRAb (group C), and the healthy subjects who were negative for TRAb (group A). A Pearson correlation analysis was used to examine the relationships between the serum levels of TSAb and TSBAb and the serum levels of FT3, FT4, TSH, and TRAb in patients with HT hypothyroidism. Results The serum TRAb level in the HT hypothyroidism group was significantly higher than that in the control group (Z=-5.434,P<0.05). Compared with group A, group B showed significantly higher serum levels of TRAb and TSBAb and positive rate of serum TSBAb (Z=6.228,F=17.985,t=7.767,χ2=15.273,P<0.05); compared with group C, group B had significantly higher serum levels of TRAb and TSBAb and positive rates of serum TSAb and TSBAb (Z=5.732,F=17.985,t=7.056,χ2=62.115,52.000,P<0.05). The Pearson correlation analysis showed that the serum TSBAb level in patients with HT hypothyroidism was positively correlated with their serum TRAb level (r=0.756,P<0.05). Conclusion The increased expression of TSBAb in serum may be an important factor in the pathogenesis of HT hypothyroidism. For HT hypothyroidism patients with positive serum TRAb, attention should be paid to the measurement of their serum TSBAb expression

Reverse-D-4F Increases the Number of Endothelial Progenitor Cells and Improves Endothelial Progenitor Cell Dysfunctions in High Fat Diet Mice

<div><p>Although high density lipoprotein (HDL) improves the functions of endothelial progenitor cells (EPCs), the effect of HDL ApoAI mimetic peptide reverse-D-4F (Rev-D4F) on EPC mobilization and repair of EPC dysfunctions remains to be studied. In this study, we investigated the effects of Rev-D4F on peripheral blood cell subpopulations in C57 mice treated with a high fat diet and the mechanism of Rev-D4F in improving the function of EPCs impaired by tumor necrosis factor-α (TNF-α). The high fat diet significantly decreased the number of EPCs, EPC migratory functions, and the percentage of lymphocytes in the white blood cells. However, it significantly increased the number of white blood cells, the percentage of monocytes in the white blood cells, and the level of vascular endothelial growth factor (VEGF) and TNF-α in the plasma. Rev-D4F clearly inhibited the effect of the high fat diet on the quantification of peripheral blood cell subpopulations and cytokine levels, and increased stromal cell derived factor 1α (SDF-1α) in the plasma. We provided in vitro evidence that TNF-α impaired EPC proliferation, migration, and tube formation through inactive AKT and eNOS, which was restored by Rev-D4F treatment. In contrast, both the PI3-kinase (PI3K) inhibitor (LY294002) and AKT inhibitor (perifosine) obviously inhibited the restoration of Rev-4F on EPCs impaired by TNF-α. Our results suggested that Rev-D4F increases the quantity of endothelial progenitor cells through increasing the SDF-1α levels and decreasing the TNF-α level of peripheral blood in high fat diet-induced C57BL/6J mice, and restores TNF-α induced dysfunctions of EPCs partly through stimulating the PI3K/AKT signal pathway.</p></div

EPCs number and migratory functions in different groups of mice.

<p>A, EPCs numbers in different groups of mice. Numbers per high-power field in percentage of control. B, Different groups of mice mononuclear cells were isolated and cultured for 10 days to detect DiI-acLDL/lectin double-positive cells. Merge: green lectin, red Dil-acLDL. C, Mononuclear cells were isolated and cultured for 10 days to detect the migratory functions of EPCs using the transwell method. Numbers per high-power field in percentage of control. Scale bar represented 20μm. Data are presented as mean±SD (n = 6). <i>**P</i> <0.01.</p

Correlations between SDF-1α and TNF-α with EPCs subsets in different groups of mice, and Rev-D4F inhibited TNF-α-induced impairment of EPCs functions.

<p>A, The increased level of SDF-1α significantly correlated with the increased number of the CD34/FLK-1 subset. B, TNF-α levels inversely correlated with CD34/FLK-1 subset numbers. Samples were pretreated with a PI3K inhibitor LY294002 (30μM) or AKT inhibitor perifosine (5μM) for 2 h and incubated with Rev-D4F (50μg/ml) for 6h, EPCs were treated with TNF-α for 24h to detect the functions of viability (C and D), migration (E and F) and tube formation (G and H). EPCs tube formation ability was calculated by the average of complete tube lengths. Data are means ± SD from at least three independent experiments. <i>**P</i> <0.01 versus control, ^<i>##</i><i>P</i> <0.01 versus TNF-α, ^ΔΔ<i>P</i> <0.01 versus LY294002.</p

The effect of Rev-D4F on plasma lipid level, NO, and cytokines (VEGF, TNF-α and SDF-1α) in different groups of mice.

<p>A, The level of total cholesterol was significantly increased in the high fat diet group of mice. B, C, and D show different groups of mice blood cytokine (SDF-1α, TNF-α and VEGF) levels. E, The blood NO level in different groups of mice was measured by an NO assay kit at 550nm. Data are presented as mean±SD (n = 6). <i>**P</i> <0.01.</p

The proportion of different leukocyte subsets in mice blood.

<p>A, Representative of the number of white blood cells per liter. B, C, and D Monocytes, neutrophils, and lymphocytes in percentage of white blood cells, respectively. Data are presented as mean±SD (n = 6). <i>**P</i> <0.01.</p

Rev-D4F decreased the expression of ICAM-1 and VCAM-1 in the arterial walls of C57BL/6J mice fed a high fat diet.

<p>Fluorescence intensity was calculated for the expression of ICAM-1 (A) and VCAM-1 (B). C, Representative of immunostained aortic sections with ICAM-1 and VCAM-1 antibodies. Relative fluorescence intensity in percentage of control. Scale bar represented 20μm. Data are presented as mean±SD (n = 6). <i>**P</i> <0.01.</p