Protective effect of afamin protein against oxidative stress related injury in human ovarian granulosa cells.

BACKGROUND: Ovarian granulosa cells (GCs) play crucial roles in follicular growth and development. Their normal function is influenced by various factors, including oxidative stress, which is a significant factor. Afamin protein is a vitamin E-specific binding protein that acts as a vitamin E carrier in follicular fluid. Although the mechanism of the protective effect of afamin on human ovarian GCs is still unclear, there is evidence it has an antioxidant effect in neuronal cells. METHODS: In this study, we investigated the protective effects of afamin proteins on testosterone propionate (TP)-induced ovarian GCs using a human ovarian tumor granulosa cell line (KGN). RESULTS: The results showed that afamin reduced TP-induced oxidative stress in KGN cells by decreasing the levels of oxidative damage markers, enhancing the activity of antioxidant enzymes, and exerting a protective effect on GCs. Supplementation with afamin repaired mitochondrial dysfunction by improving mitochondrial membrane potential damage and ATP levels. It counteracted TP-induced apoptosis by decreasing the activity of Caspase-3 and upregulating the expression of the anti-apoptotic gene (BCL-2) while downregulating the expression of the pro-apoptotic gene BCL-2-associated X protein (BAX). In addition, afamin regulated the expression of genes related to ovarian steroid hormone synthesis, ameliorating the endocrine dysfunction observed in TP-induced KGN cells. CONCLUSION: Therefore, Afamin proteins may serve as important complementary factors that protect GCs from other types of damage, such as oxidative stress, and may help improve ovarian follicle quality and female reproductive function.

Circulating afamin positively correlated with the miR-122 expression and type 2 diabetes mellitus-related phenotype according to the duration of diabetes.

Background: Afamin is a hepatokine that involves in glucose and lipids metabolism. miR-122 is mainly expressed in liver and involves in lipid and carbohydrate metabolism. This study aimed at investigating the circulating afamin, its correlation with type 2 diabetes mellitus (T2DM) and miR-122 gene expression in T2DM patients and healthy control subjects according to the duration of diabetes. Methods: This case-control study included 220 participants, with 100 individuals serving as controls and 120 individuals diagnosed with type 2 diabetes mellitus (T2DM). The miR-122 gene expression was assessed using real-time PCR. The serum concentration of biochemical parameters such as glucose levels, lipid profile, and small-dense low-density lipoprotein (sdLDL) were measured using colorimetric kits. Circulating afamin and insulin levels were assayed using an ELISA kit. Glycated hemoglobin (HbA1c) was measured using capillary electrophoresis. Results: Circulating afamin level was significantly higher in T2DM patients compared to the control group, (73.8 ± 10.8 vs. 65.9 ± 8.7, respectively; P < 0.001). Similarly, miR122 expression was significantly increased in T2DM patients compared to healthy control subjects (4.24 ± 2.01 vs. 1.00 ± 0.85, respectively; P < 0.001). Among patients diagnosed with T2DM, those with longstanding diabetes (>5 years) exhibited significantly higher levels of circulating afamin and miR-122 expression compared to individuals with a shorter duration of diabetes (≤5 years) (P < 0.05). Circulating afamin levels were significantly correlated with waist circumference, small-dense low-density lipoprotein (sdLDL), fasting blood sugar (FBS), insulin, resistance to insulin, and miR-122 expression, depending on the duration of the disease (P < 0.05). Furthermore, the performance of afamin as a diagnostic marker for T2DM was confirmed through receiver operating characteristic (ROC) analysis, yielding an area under the curve (AUC) of 0.7 (P < 0.001). Conclusions: Circulating afamin involved in the T2DM-related complications and its concentration is positively correlated to the miR-122 expression, especially in patient with longstanding diabetes.

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.

Clinical Significance of Adropin and Afamin in Evaluating Renal Function and Cardiovascular Health in the Presence of CKD-MBD Biomarkers in Chronic Kidney Disease.

AIM: The study aims to test the hypothesis that concentrations of adropin and afamin differ between patients in various stages of chronic kidney disease when compared with healthy controls. The study also investigates the association of the biomarkers (adropin and afamin) with CKD-MBD and traditional cardiovascular risk parameters in CKD patients. METHODOLOGY: The cross-sectional study includes the subjects divided into four groups comprising the control group (healthy volunteers = 50), CKD stages 1-2 patients (n = 50), CKD stages 3-4 patients (n = 50), CKD stage 5 patients (n = 50). Serum concentrations of adropin and afamin were determined using ELISA. Clinical variables (renal, lipid, and CKD-MBD parameters) were correlated to adropin and afamin concentrations. RESULTS: Afamin concentration was found to be higher in group IV, followed by groups III and II when compared to the control group, i.e., (83.243 ± 1.46, 64.233 ± 0.99, and 28.948 ± 0.72 vs. 14.476 ± 0.5) mg/L (p < 0.001), and adropin concentration was found to be lower in group IV as compared to groups III, II, and I (200.342 ± 8.37 vs. 284.682 ± 9.89 vs. 413.208 ± 12.32 vs. 706.542 ± 11.32) pg/mL (p < 0.001), respectively. Pearson correlation analysis showed that afamin was positively correlated with traditional cardiovascular risk biomarkers, while adropin showed a negative correlation. CONCLUSIONS: Adropin and afamin may potentially serve as futuristic predictors for the deterioration of renal function and may be involved in the pathological mechanisms of CKD and its associated complications such as CKD-MBD and high lipid levels.

Early Pregnancy Markers in the Serum of Ewes Identified via Proteomic and Metabolomic Analyses.

The diagnosis of ewes' pregnancy status at an early stage is an efficient way to enhance the reproductive output of sheep and allow producers to optimize production and management. The techniques of proteomics and metabolomics have been widely used to detect regulatory factors in various physiological processes of animals. The aim of this study is to explore the differential metabolites and proteins in the serum of pregnant and non-pregnant ewes by proteomics and metabolomics. The serum of ewes at 21, 28 and 33 days after artificial insemination (AI) were collected. The pregnancy stratus of the ewes was finally determined through ultrasound examination and then the ewes were grouped as Pregnant (n = 21) or N on-pregnant (n = 9). First, the serum samples from pregnant or non-pregnant ewes at 21 days after AI were selected for metabolomic analysis. It was found that the level of nine metabolites were upregulated and 20 metabolites were downregulated in the pregnant animals (p < 0.05). None of these differential metabolomes are suitable as markers of pregnancy due to their small foldchange. Next, the proteomes of serum from pregnant or non-pregnant ewes were evaluated. At 21 days after AI, the presence of 321 proteins were detected, and we found that the level of three proteins were upregulated and 11 proteins were downregulated in the serum of pregnant ewes (p < 0.05). The levels of serum amyloid A (SAA), afamin (AFM), serpin family A member 6 (SERPINA6) and immunoglobulin-like domain-containing protein between pregnant and non-pregnant ewes at 21-, 28- and 33-days post-AI were also analyzed via enzyme-linked immunosorbent assay (ELISA). The levels of SAA and AFM were significantly higher in pregnant ewes than in non-pregnant ewes, and could be used as markers for early pregnancy detection. Overall, our results show that SAA and AFM are potential biomarkers to determine the early pregnancy status of ewes.

Investigation of serum afamin concentration in pregnant women diagnosed with late fetal growth restriction or small for gestational age fetus.

OBJECTIVE: To investigate the afamin concentration in the serum of pregnant women diagnosed with late fetal growth restriction (FGR) or small for gestational age (SGA) in the third trimester. METHODS: This prospective case-control study was conducted on 126 pregnant women, 42 of whom were diagnosed with late FGR in the third trimester, 43 were SGA, and 41 were healthy controls. The groups were compared in terms of maternal serum afamin concentrations. RESULTS: Three groups were similar in terms of demographic characteristics and gestational age at blood sampling for afamin (p < .05). The median afamin concentration was determined as 199 ng/mL in the late FGR group, 153 ng/mL in the SGA group, and 108 ng/mL in the control group (p = .000). In the post-hoc analysis, while maternal serum afamin concentrations were found to be significantly higher in the late FGR group and SGA group compared to the control group but, this significance could not be shown between the FGR group and the SGA group (p = .00001, p = .005, p = .137, respectively). In the ROC analysis, the optimal cutoff value of serum afamin concentration to predict late FGR was determined as 141 ng/mL, with a sensitivity of 66.6% and a specificity of 85.3%. CONCLUSIONS: The serum afamin concentration in the third trimester was found to be higher in pregnant women with late FGR compared to the SGA and control groups. Although afamin is seen as a promising molecule in the clinical prediction of late FGR, this needs to be supported by large series of studies.

Serum afamin levels in predicting gestational diabetes mellitus and preeclampsia: A systematic review and meta-analysis.

Objectives: The association between biomarkers and the risk of gestational diabetes mellitus (GDM) or preeclampsia (PE) has been extensively studied. However, there is still a lack of convenient, specific, and sensitive indicators for early identification of GMD and PE. Therefore, we conducted a meta-analysis of published articles to investigate the value of afamin circulating levels in the early diagnosis of GDM and PE. Methods: We searched the PubMed, Embase, Cochrane Library, and Web of Science databases for English studies published before November 16, 2022, that examined the association between afamin and GDM or PE. In addition, we searched Clinicaltrials.gov for the relevant completed and ongoing clinical trials. Pooled standard mean differences (SMDs) and weighted mean differences (WMDs) with 95% confidence intervals (CIs) were used to compare the levels of afamin in different groups. Results: Eleven studies were included in our analysis (N = 3047 participants: 1195 GDM, 1407 non-GDM, 195 PE, and 250 non-PE). Subgroup analysis based on different blood collection periods found that the plasma afamin levels in pregnant women with GDM in the first trimester were significantly higher than those in healthy pregnant women (SMD = 0.481, 95% CI: 0.280-0.682), but the analysis showed the opposite results in the second and late stages (SMD = 0.292, 95% CI: -0.092-0.676). The plasma afamin levels of pregnant women with PE in the first trimester (SMD = 0.808, 95% CI: 0.558-1.059) and second/third trimesters (SMD = 0.904, 95% CI: 0.570-1.239) were significantly higher than those in healthy pregnant women. Conclusion: The plasma afamin levels in pregnant women with GDM in the first trimester were significantly higher than those in healthy pregnant women, but the analysis showed the opposite results in the second and third trimesters. The plasma afamin levels in pregnant women with PE in the first, second, and third trimesters were significantly higher than those in healthy pregnant women. Additional large-scale prospective studies are desired to verify these findings, and it is recommended that afamin should be included as a routine diagnostic test for women with GDM and PE. Systematic review registration: https://www.crd.york.ac.uk/PROSPERO/display_record.php?RecordID=339171, identifier CRD42022339171.

Evaluation of Afamin Level after Bariatric Surgery in Patient with Obesity.

BACKGROUND: The aim of this study is to evaluate afamin levels after weight loss in obese patients who underwent laparoscopic sleeve gastrectomy (LSG) and to investigate the relationship between them. In addition, after bariatric surgery, thyroid stimulating hormone (TSH), thyroxine (T4), low-density lipoprotein (LDL), very low-density protein (VLDL), total cholesterol, triglyceride (TG), high-density lipoprotein (HDL), insulin, and hemoglobin A1c (HgbA1c) levels were evaluated. METHODS: Preoperative and postoperative 6th month venous blood samples were obtained from 43 patients included in this study. The preoperative and postoperative 6th month body mass index (BMI), TG, total cholesterol, VLDL, HDL, insulin, HgbA1c, TSH, T4, and afamin levels of the patients who underwent bariatric surgery with obesity were compared. RESULTS: Serum afamin levels of patients decreased at 6 months postoperatively; however, it was not statistically significant. We observed a statistically significant decrease in patients' BMI, HDL, VLDL, TG, total cholesterol, TSH, T4, HgbA1c, and insulin values (p < 0.05). There were significant increases in HDL and T4 values. The change in LDL value was statistically insignificant. CONCLUSIONS: Recent studies have shown that there may be a cause-effect relationship between afamin and obesity. In our study, we observed a decrease in serum afamin levels after weight loss following bariatric surgery. In addition, we think that afamin may be used as a potential marker of metabolic syndrome in the future and may lead to improvements in the medical treatment of obesity.

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.

Novel Biomolecules in the Pathogenesis of Gestational Diabetes Mellitus 2.0.

Gestational diabetes mellitus (GDM) has become a major public health problem and one of the most discussed issues in modern obstetrics. GDM is associated with serious adverse perinatal outcomes and long-term health consequences for both the mother and child. Currently, the importance and purposefulness of finding a biopredictor that will enable the identification of women with an increased risk of developing GDM as early as the beginning of pregnancy are highly emphasized. Both "older" molecules, such as adiponectin and leptin, and "newer" adipokines, including fatty acid-binding protein 4 (FABP4), have proven to be of pathophysiological importance in GDM. Therefore, in our previous review, we presented 13 novel biomolecules, i.e., galectins, growth differentiation factor-15, chemerin, omentin-1, osteocalcin, resistin, visfatin, vaspin, irisin, apelin, FABP4, fibroblast growth factor 21, and lipocalin-2. The purpose of this review is to present the potential and importance of another nine lesser known molecules in the pathogenesis of GDM, i.e., 3-carboxy-4-methyl-5-propyl-2-furanpropanoic acid (CMPF), angiopoietin-like protein-8 (ANGPTL-8), nesfatin-1, afamin, adropin, fetuin-A, zonulin, secreted frizzled-related proteins (SFRPs), and amylin. It seems that two of them, fetuin-A and zonulin in high serum levels, may be applied as biopredictors of GDM.

Afamin Levels and Their Correlation with Oxidative and Lipid Parameters in Non-diabetic, Obese Patients.

Background: Afamin is a liver-produced bioactive protein and features α- and γ-tocopherol binding sites. Afamin levels are elevated in metabolic syndrome and obesity and correlate well with components of metabolic syndrome. Afamin concentrations, correlations between afamin and vitamin E, afamin and lipoprotein subfractions in non-diabetic, obese patients have not been fully examined. Methods: Fifty non-diabetic, morbidly obese patients and thirty-two healthy, normal-weight individuals were involved in our study. The afamin concentrations were measured by ELISA. Lipoprotein subfractions were determined with gel electrophoresis. Gas chromatography-mass spectrometry was used to measure α- and γ tocopherol levels. Results: Afamin concentrations were significantly higher in the obese patients compared to the healthy control (70.4 ± 12.8 vs. 47.6 ± 8.5 µg/mL, p < 0.001). Positive correlations were found between afamin and fasting glucose, HbA1c, hsCRP, triglyceride, and oxidized LDL level, as well as the amount and ratio of small HDL subfractions. Negative correlations were observed between afamin and mean LDL size, as well as the amount and ratio of large HDL subfractions. After multiple regression analysis, HbA1c levels and small HDL turned out to be independent predictors of afamin. Conclusions: Afamin may be involved in the development of obesity-related oxidative stress via the development of insulin resistance and not by affecting α- and γ-tocopherol levels.

Afamin predicts the prevalence and incidence of nonalcoholic fatty liver disease.

OBJECTIVES: In the general population, increased afamin concentrations are associated with the prevalence and incidence of metabolic syndrome as well as type 2 diabetes. Although metabolic syndrome is commonly associated with nonalcoholic fatty liver disease (NAFLD), there exist no information on afamin and NAFLD. METHODS: Afamin concentrations were cross-sectionally measured in 146 Austrian patients with NAFLD, in 45 patients without NAFLD, and in 292 age- and sex-matched healthy controls. Furthermore, the feasibility of afamin to predict incident NAFLD was evaluated in 1,434 adult participants in the population-based Cardiovascular Risk in Young Finns Study during a 10-year follow-up. RESULTS: Median afamin concentrations were significantly higher in NAFLD patients (83.6 mg/L) than in patients without NAFLD (61.6 mg/L, p<0.0001) or in healthy controls (63.9 mg/L, p<0.0001). In age- and sex-adjusted logistic regression analyses a 10 mg/L increase of afamin was associated with a 1.5-fold increase of having NAFLD as compared with patients without NAFLD and the risk was even two-fold when compared with healthy controls. In the population-based cohort, afamin concentrations at baseline were significantly lower in participants without NAFLD (n=1,195) than in 239 participants who developed NAFLD (56.5 vs. 66.9 mg/L, p<0.0001) during the 10-year follow up, with highest afamin values observed in individuals developing severe forms of NAFLD. After adjustment for several potentially confounding parameters, afamin remained an independent predictor for the development of NAFLD (OR=1.37 [95% CI 1.23-1.54] per 10 mg/L increase, p<0.0001). CONCLUSIONS: Afamin concentrations are increased in patients with NAFLD and independently predict the development of NAFLD in a population-based cohort.

Relationship between Serum Kallistatin and Afamin and Anthropometric Factors Associated with Obesity and of Being Overweight in Patients after Myocardial Infarction and without Myocardial Infarction.

Extensive clinical and epidemiological evidence has linked obesity to a broad spectrum of cardiovascular disease (CVD), including coronary disease, heart failure, hypertension, cerebrovascular disease, atrial fibrillation, ventricular arrhythmias, and sudden death. In addition, increasing knowledge of regulatory peptides has allowed an assessment of their role in various non-communicable diseases, including CVD. The study assessed the concentration of kallistatin and afamin in the blood serum of patients after a myocardial infarction and without a cardiovascular event, and determined the relationship between the concentration of kallistatin and afamin and the anthropometric indicators of being overweight and of obesity in these groups. Serum kallistatin and afamin were quantified by ELISA tests in a cross-sectional study of 160 patients who were divided into two groups: study group (SG) (n = 80) and another with no cardiovascular event (CG) (n = 80). Serum kallistatin concentration was significantly higher in the SG (p < 0.001), while the level of afamin was significantly lower in this group (p < 0.001). In addition, a positive correlation was observed in the SG between the afamin concentration and the waist to hip ratio (WHR), lipid accumulation product (LAP) and the triglyceride glucose index (TyG index). In the CG, the concentration of kallistatin positively correlated with the LAP and TyG index, while the concentration of afamin positively correlated with all the examined parameters: body mass index (BMI), waist circumference (WC), hip circumference (HC), waist to hip ratio (WHtR), visceral adiposity index (VAI), LAP and TyG index. Serum kallistatin and afamin concentrations are associated with the anthropometric parameters related to being overweight and to obesity, especially to those describing the visceral distribution of adipose tissue and metabolic disorders related to excessive fatness.

The Clinical Values of Afamin, Triglyceride and PLR in Predicting Risk of Gestational Diabetes During Early Pregnancy.

Objective: To establish a model to predict gestational diabetes mellitus (GDM) based on the clinical characteristics, early pregnancy (10-12 weeks gestation) peripheral blood routine, and biochemical indicators, and to explore its predictive efficiencies. Methods: Data from 607 pregnant women with GDM were compared to the data from 833 pregnant women without GDM admitted to the Obstetrics Department of Fujian Maternity and Child Health Hospital (affiliated to Fujian Medical University) from May 2018 to December 2018 were retrospectively included. The ages of the pregnant women, paternal ages, number of pregnancies, number of deliveries, pre-pregnancy heights/weights, and the calculated body mass indexes (BMI) were recorded. In all participants, 10-12 weeks of pregnancy, afamin concentration, routine blood work, prenatal aneuploidy screening, and biochemical testing were performed. At weeks 24-28 of gestation, patients underwent oral glucose tolerance test (OGTT) for GDM screening. Results: Multivariate logistic regression analysis showed that maternal age, early pregnancy afamin level, triglycerides, and platelet/lymphocyte ratio (PLR) were independent risk factors for gestational diabetes. The formula for predicting GDM probability was as follows: P = 1/1 + exp( - 6.054 + 0.774 × triglycerides + 0.002 × afamin + 0.155 × age - 0.012 × PLR)]. From the established ROC curve, the area under the curve (AUC) was 0.748, indicating that the model has a good degree of discrimination. When the predictive probability cut-off value was set on 0.358, sensitivity, specificity, positive predictive value, and negative predictive value were 69.2%, 68.3%, 42.5%, and 86.2%, respectively, and the accuracy rate was 70.2%. The Hosmer-Lemeshow test results showed that the goodness of the model fit has a good calibration ability (χ2 = 12.269, df=8, P=0.140). Conclusions: Maternal age, early pregnancy afamin level, triglycerides, and PLR are independent risk factors for gestational diabetes. When combined, the above indicators are helpful for prediction, early diagnosis, and intervention of gestational diabetes.

Serum Afamin a Novel Marker of Increased Hepatic Lipid Content.

Aim: Afamin is a liver-produced glycoprotein, a potential early marker of metabolic syndrome. Here we investigated regulation of afamin in a course of the metabolic disease development and in response to 3-month exercise intervention. Methods: We measured whole-body insulin sensitivity (euglycemic hyperinsulinemic clamp), glucose tolerance, abdominal adiposity, hepatic lipid content (magnetic resonance imaging/spectroscopy), habitual physical activity (accelerometers) and serum afamin (enzyme-linked immunosorbent assay) in 71 middle-aged men with obesity, prediabetes and newly diagnosed type 2 diabetes. Effects of 3-month exercise were investigated in 22 overweight-to-obese middle-aged individuals (16M/6F). Results: Prediabetes and type 2 diabetes, but not obesity, were associated with increased serum afamin (p<0.001). Afamin correlated positively with hepatic lipids, fatty liver index and liver damage markers; with parameters of adiposity (waist circumference, %body fat, adipocyte diameter) and insulin resistance (fasting insulin, C-peptide, HOMA-IR; p<0.001 all). Moreover, afamin negatively correlated with whole-body insulin sensitivity (M-value/Insulin, p<0.001). Hepatic lipids and fasting insulinemia were the most important predictors of serum afamin, explaining >63% of its variability. Exercise-related changes in afamin were paralleled by reciprocal changes in insulinemia, insulin resistance and visceral adiposity. No significant change in hepatic lipid content was observed. Conclusions: Subjects with prediabetes and type 2 diabetes had the highest serum afamin levels. Afamin was more tightly related to hepatic lipid accumulation, liver damage and insulin resistance than to obesity.

Novel metabolic marker Afamin: A predictive factor for Large-for-Gestational-Age (LGA) fetus estimation in pregnancies with gestational diabetes mellitus?

OBJECTIVE: Gestational diabetes mellitus (GDM) affects both maternal and fetal/infant outcomes during and after pregnancy. The reason for the high incidence of large-for-gestational-age (LGA) infants in GDM patients despite close monitorization of glucose levels with early detection of the disease remains unclear to date. Our study aims to investigate the levels of the third-trimester novel marker afamin in GDM versus non-GDM pregnancies in terms of glycemic control status and their utility in the prediction of LGA fetuses. MATERIAL AND METHODS: This prospective case-control study analysis involved 49 pregnant women with GDM diagnosed using the 75-g oral glucose tolerance test (75-g OGTT) and 40 randomly selected women with a similar body mass index (BMI) and gestational age (GA). Blood samples were collected in the third trimester of pregnancy. The afamin level was determined using a human afamin ELISA kit according to the manufacturer's procedure. RESULTS: There was no significant difference found in BMI or GA of patients. Third-trimester afamin levels were 93.91 mg/L and 83.87 mg/L in the GDM and non-GDM groups, respectively (p=0.625). Afamin values of patients were not correlated with age, BMI, GA, HgA1c, 75-g OGTT fasting and 75-g OGTT 1-hour, or 75-g OGTT 2-hour values (p>0.05). GDM patients with LGA fetuses had significantly higher afamin values than patients with appropriate-for-gestational-age (AGA) fetuses (120.8 mg/L versus 91.26 mg/L, respectively). Between GDM patients with either LGA or AGA fetuses, there was no statistically significant difference found for age, BMI, GAs, insulin dose, 75-g OGTT results, or HgA1c values. CONCLUSION: Our findings conclude that novel marker afamin levels could predict the risk of LGA infants independently of glycemic control status and provide insight into the pathogenesis of LGA fetuses, thus helping to reduce the risk of associated complications.

Identification and Validation of Combination Plasma Biomarker of Afamin, Fibronectin and Sex Hormone-Binding Globulin to Predict Pre-eclampsia.

The purpose of the present study was to identify a plasma protein biomarker able to predict pre-eclampsia (PE). Comprehensive quantitative proteomics using mass spectrometry with sequential window acquisition of all theoretical fragment ion spectra (SWATH-MS) was applied to plasma samples of 7 PE and 14 healthy pregnant women (for PE subjects, plasma samples were taken before onset of PE), and 11 proteins were selected as candidates potentially able to differentiate the two groups. Plasmas collected at gestational weeks 14-24 from 36 PE and 120 healthy pregnant women (for PE subjects, plasma samples were taken before onset of PE) were used to conduct selected reaction monitoring quantification analysis, optimize protein combinations and conduct internal validation, which consisted of 30 iterations of 10-fold cross-validation using multivariate logistic regression and receiver operating characteristic (ROC) analysis. The combination of afamin, fibronectin, and sex-hormone-binding globulin was selected as the best candidate. The 3-protein combination predictive model (predictive equation and cut-off value) generated using the internal validation subjects was successfully validated in another group of validation subjects (36 PE and 54 healthy (for PE subjects, plasma samples were taken before onset of PE)) and showed good predictive performance, with the area under the curve (AUC) 0.835 and odds ratio 13.43. In conclusion, we newly identified a 3-protein combination biomarker and established a predictive equation and cut-off value that can predict the onset of PE based on analysis of plasma samples collected during gestational weeks 14-24.

The positive effect of short-term nano-curcumin therapy on insulin resistance and serum levels of afamin in patients with metabolic syndrome.

OBJECTIVE: Metabolic syndrome (MS) is a cluster of cardio-metabolic risk factors. MS is known as a highly prevalent disease worldwide. According to the existing evidence, consuming curcumin has positive effects on lipids profile, glucose, and body weight. This study aimed to evaluate the effects of nano-curcumin therapy on insulin resistance and serum level of afamin in patients with MS. MATERIALS AND METHODS: Thirty MS patients (15 males and 15 females) received 80 mg/daily nano-curcumin for two months. The samples of fasting blood were collected from the participants at the beginning and 60 days after initiation of the intervention to measure biomarkers. RESULTS: Comparing pre- and post-treatment with nano-curcumin values revealed a significant decrease in fasting plasma glucose (FPG) (p=0.017), insulin, homeostatic model assessment of insulin resistance (HOMA-IR) (p=0.006), and afamin (p=0.047). Moreover, there was a significantly negative relationship between afamin and high-density lipoprotein cholesterol (HDL-C) (p=0.044), as well as a significantly positive relationship between afamin and systolic (SBP) (p<0.001) and diastolic (DBP) (p<0.001) blood pressures. CONCLUSION: Results suggest that taking nano-curcumin for 60 days may have positive effects on afamin, FPG, insulin, and HOMA-IR in patients with MS, but would not significantly affect other metabolic profiles. More studies with larger sample sizes are required to confirm these findings.

Seminal Plasma and Serum Afamin Levels Are Associated with Infertility in Men with Oligoasthenoteratozoospermia.

The plasma glycoprotein afamin has been previously identified as an alternative carrier protein for vitamin E in extravascular fluids such as plasma and cerebrospinal, ovarian follicular, and seminal fluids. However, to date, no study has established a relationship between afamin levels and infertility in women or men. The purposes of our study were (i) to assess the level of afamin in serum and seminal fluids in infertile men compared to healthy controls and (ii) to study the association between polymorphisms in afamin genes and male infertility. This observational, prospective study evaluated the afamin levels in serum and seminal fluids from infertile men (n = 39) and compared them to those in healthy controls (n = 30). We studied the association between single-nucleotide polymorphisms (SNPs) in the 5`-untranslated region (5`-UTR) of the afamin gene and infertility and analyzed a total of 1000 base pairs from the untranslated region of the afamin gene. Subjects with low sperm motility and low sperm concentration had higher median seminal afamin (18.9 ± 2.9 ng/mg of proteins) and serum afamin concentrations (24.1 ± 4.0 ng/mg of proteins) than subjects with normal sperm parameters (10.6 ± 1.4 ng/mg of proteins) (p < 0.02) (15.6 ± 1.4 ng/mg of proteins) (p < 0.002). A total of five different polymorphisms were found, including one deletion and four single-nucleotide polymorphisms (SNPs). A new transversion (A/T) (position 4:73481093) was identified in an oligoasthenoteratozoospermic patient and was associated with high levels of afamin in plasma and seminal fluids. The prevalence of this variant in our study in the case homozygous for TT is 0.985 (98.5%), and in the case heterozygous for TA is 0.015 (1.5%). Our results suggest that genetic variations in afamin might be associated with male infertility. These findings could significantly enhance our understanding of the molecular genetic causes of infertility.

The Role of Afamin and Other Trace Elements in the Prediction of GDM: a Tertiary Center Experience.

The objective of this study was to evaluate the utility of first trimester maternal serum afamin levels together with vitamin E and various elements (zinc, copper, selenium, and magnesium) for the prediction of gestational diabetes mellitus (GDM). All pregnant women between 11th and 14th weeks of gestation admitted for combined test were asked to participate in the study. A total of 797 women gave permission to participate in the study between January and September 2019. Serum was obtained by centrifugation and samples were frozen and kept at - 80 °C. For final analysis, forty-three GDM patients and forty-four healthy controls were selected. Levels of afamin, vitamin E, zinc, copper, selenium, and magnesium were compared between groups. The mean levels of afamin were found to be higher in women with GDM without statistical significance (63.69 ± 82.33 vs 44.25 ± 32.25 mg/L, p = 0.149). Vitamin E levels were found to be higher in women with GDM compared to controls without any statistical significance (5.04 ± 5.33 vs 4.47 ± 3.83 µg/mL, p = 0.568). While first trimester copper concentrations were higher among diabetic women (187.26 ± 34.78 vs 175.17 ± 30.40 µg/L, p = 0.088), this was not statistically significant. The other element levels (zinc, selenium, and magnesium) were found to be similar between the two groups (p = 0.624, p = 0.088, p = 0.254, p = 0.872, respectively). The results of our study demonstrated that mean levels of afamin, vitamin E, and copper were higher in women with GDM compared to controls. Additionally, first trimester maternal zinc, selenium, and magnesium levels were similar between diabetic and healthy pregnant women. However, more studies are needed to clarify the relationship between blood trace concentrations and GDM.