Assessment of subclinical left ventricular systolic dysfunction in patients with type 2 diabetes: Relationship with HbA1c and microvascular complications.

BACKGROUND: We aimed to examine the association between glycated hemoglobin (HbA1c), microvascular complications, and subclinical left ventricular (LV) systolic dysfunction, and to determine the strength of the correlation in asymptomatic patients with type 2 diabetes mellitus (T2DM). METHODS: Global longitudinal strain (GLS) was employed to assess the subclinical LV function of 152 enrolled T2DM patients with preserved LV ejection fraction, with the cutoff for subclinical LV systolic dysfunction predefined as GLS < 18%. RESULTS: According to univariate analysis, the reduced GLS exhibited association with the clinical features including HbA1c, triglyceride, systolic blood pressure, fasting glucose, heart rate, diabetic retinopathy, and urinary albumin creatinine ratio (UACR) (all p < .05). After the factors of gender, age, and related clinical covariables adjusted, multiple logistic regression analysis revealed the HbA1c (odds ratio [OR] 1.66; 95% confidence interval [CI] 1.30-2.13; p < .001), UACR (OR 2.48; 95% CI 1.12-5.47; p = .025) and triglyceride (OR 1.84; 95% CI 1.12-3.03; p = .017) as the independent risk factors for the reduced GLS. Receiver operating characteristic curve showed a predictive value of the HbA1c for the subclinical LV systolic dysfunction (area under curve: 0.74; p < .001). CONCLUSIONS: In asymptomatic T2DM patients, subclinical LV systolic dysfunction was associated with HbA1c, diabetic complications, and triglyceride. More prominently, HbA1c may exert a prognostic significance for the progression of myocardial damage.

Association of peripheral neuropathy with subclinical left ventricular dysfunction in patients with type 2 diabetes.

OBJECTIVES: The impacts of diabetic peripheral neuropathy (DPN) on clinical manifestations of left ventricular (LV) function in patients suffering from type 2 diabetes mellitus (T2DM) and the preserved LV ejection fraction (LVEF) lack a full evaluation. This study was carried out to investigate the correlation of peripheral neuropathy with subclinical LV systolic dysfunction, accompanied by the exploration of the relevant clinical features of peripheral neuropathy in these patients. METHODS: A retrospective analysis was conducted depending on the data of 101 consecutive inpatients with T2DM and preserved LVEF (all ≥ 50 %), without coronary artery disease and other histories of heart disease. All subjects received both a nerve conduction assessment and a speckle-tracking echocardiography examination. Global longitudinal strain (GLS) was conducted to assess the subclinical LV systolic function. RESULTS: Forty-six (46 %) patients were diagnosed as DPN according to electrophysiological examination and clinical assessment. A significant difference was revealed in GLS between patients with and without DPN (16.5 ± 2.8 vs. 19.3 ± 3.4, p < 0.001). Multiple logistic regression analysis indicated GLS as one of the independent determinative factors for DPN (odds ratio, 0.68; P < 0.001). In addition, motor-sensory nerve conduction exhibited a significant positive correlation with GLS, which may not be revealed between the types of peripheral nerve damage. CONCLUSIONS: Despite the preserved LVEF, the subclinical LV myocardial dysfunction may have occurred in T2DM patients with DPN. Peripheral nerve conduction was significantly correlated with GLS. An early assessment of nerve conduction may exert a dual warning significance for the progression of subclinical LV dysfunction in asymptomatic patients with T2DM.

SIRT1 rs10823108 and FOXO1 rs17446614 responsible for genetic susceptibility to diabetic nephropathy.

SIRT1 and FOXO1 play an important role in the pathogenesis of diabetic nephropathy (DN). However, the association between genetic polymorphisms and susceptibility to type 2 DN (T2DN) has not been explored. In this study, a total of 1066 patients with type 2 diabetes mellitus (T2DM) (413 without and 653 with DN) were enrolled. The genotypes of three htSNPs (rs3818292, rs4746720, rs10823108) within SIRT1 and two htSNPs (rs2721068, rs17446614) in FOXO1 were determined by PCR-RFLP. HbA1C, LDL, HDL, TC, and TG levels were also examined. SIRT1 rs10823108 AA genotype was significantly associated with a decreased risk of DN (OR = 0.60, 95%CI: 0.38-0.97), while GA genotype (OR = 1.77, 95%CI: 1.33-2.35) and AA genotype (OR = 2.32, 95%CI: 1.25-4.34) of FOXO1 rs17446614 was associated with an increased T2DN risk. The interactions among rs1744 6614, BMI and duration of diabetes (OR: 2.63, 95%CI: 1.23-4.31) were also observed. Subsequent haplotype analysis revealed that two haplotype defined by AC (OR: 1.50, 95%CI: 1.15-1.94) and AT (OR: 1.79, 95%CI: 1.06-2.80) within FOXO1 gene may increase the risk of T2DN. In conclusion, genetic variant rs10823108 in SIRT1 and variant rs17446614 in FoxO1 may contribute to the risk of DN in T2DM patients.

Lentiviral Vector-Mediated FoxO1 Overexpression Inhibits Extracellular Matrix Protein Secretion Under High Glucose Conditions in Mesangial Cells.

Diabetic nephropathy is characterized by inordinate secretion of extracellular matrix (ECM) proteins from mesangial cells (MCs), which is tightly associated with excessive activation of TGF-ß signaling. The forkhead transcription factor O1 (FoxO1) protects mesangial cells from hyperglycemia-induced oxidative stress, which may be involved in ameliorating the redundant secretion of ECM proteins under high glucose conditions. Here, we reported that high glucose elevated the level of p-Akt to attenuate endogenous FoxO1 bioactivities in MCs, accompanied with decreases in the mRNA expressions of catalase (CAT) and superoxide dismutase 2 (SOD2). Meanwhile, the expressions of major ECM proteins-FN and Col I-increased under high glucose conditions, in consistent with the activation of TGF-ß/Smad signaling. By contrast, overexpression of nucleus-localized FoxO1 (insensitive to Akt phosphorylation) directly up-regulated the expressions of anti-oxidative enzymes, accompanied with inactivation of TGF-ß/Smad3 pathway, as well as decreases of extracellular matrix proteins. Moreover, similar to those MCs overexpressed of nucleus-localized FoxO1 in high glucose conditions, MCs with down-regulation of FoxO1 by small interference-RNA under normal glucose conditions showed increased FN level and activated TGF-ß/Smad3 pathway. Our findings link the anti-oxidative activity of FoxO1 and the TGF-ß-induced secretion of ECM proteins, indicating the novel role of FoxO1 in protecting MCs under high glucose conditions.

Effects of FoxO1 on podocyte injury in diabetic rats.

OBJECTIVE: This study was designed to investigate the protective effect of forkhead transcription factor O1 (FoxO1) on podocyte injury in rats with diabetic nephropathy. METHODS: Streptozotocin-induced diabetic rats were served as DM group, while DM rats transfected with blank lentiviral vectors (LV-pSC-GFP) or lentiviral vectors carrying constitutively active FoxO1 (LV-CA-FoxO1) were served as LV-NC group or LV-CA group, respectively. The control group (NG) consisted of uninduced rats that received an injection of diluent buffer. At 2, 4, and 8 weeks after transfection, the levels of urine albumin, blood glucose, blood urea nitrogen, serum creatinine and urine podocalyxin were measured. Real-time PCR and western blotting were performed to measure mRNA and protein levels of FoxO1, podocalyxin, nephrin, and desmin in renal cortex. In addition, light and electron microscopy were used to detect structural changes in the glomerulus and podocytes. RESULTS: Compared with the rats in LV-NC and DM groups, LV-CA rats showed a significant increase in FoxO1 mRNA and protein levels and a distinct decrease in urine albumin, blood urea nitrogen, and serum creatinine (except at the two-week time point) levels (p < 0.05). Podocalyxin and nephrin mRNA and protein levels increased (p < 0.05), whereas desmin mRNA and protein levels decreased (p < 0.05). Pathological changes in glomerulus were also ameliorated in LV-CA group. CONCLUSIONS: Upregulating expression of FoxO1 by transduction with recombinant lentivirus ameliorates podocyte injury in diabetic rats.

Activation of FoxO1/ PGC-1α prevents mitochondrial dysfunction and ameliorates mesangial cell injury in diabetic rats.

The generation of hyperglycemia-induced mitochondrial reactive oxygen species (ROS) is a key event in diabetic nephropathy development. The forkhead-box class O1 (FoxO1) and peroxisome proliferator-activated receptor γ co-activator 1α (PGC-1α) proteins are implicated in oxidative stress. We investigated the in vivo association of FoxO1 and PGC-1α in renal cortices from streptozotocin-induced diabetic rats and in rat kidney mesangial cells (MCs) treated with high glucose, in vitro. High-glucose induced FoxO1 inhibition was associated with decreased PGC-1α expression in MCs. These changes were accompanied by mitochondrial dysfunction and increased ROS generation. However, constitutive FoxO1 activation increased PGC-1α expression and partially reversed these changes, which were significantly decreased by the treatment of PGC-1α-small interfering RNA. We identified PGC-1α as a direct FoxO1 transcriptional target by chromatin immunoprecipitation. In addition, lentiviral-mediated FoxO1 overexpression in diabetic-rat kidneys significantly increased PGC-1α, NRF-1, and Mfn2 expression, and decreased malondialdehyde production and proteinuria. These data suggest that FoxO1/PGC-1α activation protected rats against high-glucose-induced MC injury by attenuating mitochondrial dysfunction and cellular ROS production.

Overexpression of the FoxO1 Ameliorates Mesangial Cell Dysfunction in Male Diabetic Rats.

The dysfunction of mesangial cells (MCs) in high-glucose (HG) conditions plays pivotal role in inducing glomerular sclerosis by causing the imbalance between generation and degradation of extracellular matrix (ECM) proteins, which ultimately leads to diabetic nephropathy. This study was designed to determine the function of forkhead box protein O1 (FoxO1), an important transcription factors in regulating cell metabolism and oxidative stress, in MCs in HG conditions. Up-regulation of fibronectin, collagen type IV, and plasminogen activator inhibitor (PAI-1) was observed under HG conditions in vivo and in vitro, accompanied with elevation of protein kinase B (Akt) phosphorylation and reduction of FoxO1 bioactivity. After overexpression of constitutively active (CA) FoxO1 in vivo and in vitro by using lentivirus vector, in vivo and in vitro, FoxO1 expression and activity was increased, in accordance with up-regulation of antioxidative genes (catalase and superoxide dismutase, leading to alleviated oxidative stress as well as attenuated Akt activity, whereas overexpression of wild type-FoxO1 only expressed partial effect. Moreover, CA-FoxO1 decreased the expression of fibronectin, collagen type IV, and PAI-1, causing amelioration of renal pathological changes and decrease of ECM protein deposition in glomerulus. Overexpression of CA-FoxO1 in renal cortex also decreased activin type-I receptor-like kinase-5 levels and increased signaling mothers against decapentaplegic (Smad) 7 levels, and simultaneously inhibited Smad3 phosphorylation. Results from in vitro study indicated that increased combination of FoxO1 and Smad3 may interfere with the function of Smad3, including Smad3 phosphorylation and translocation, interaction with cAMP response element binding protein (CREB)-binding protein, and binding with PAI-1 promoter. Together, our findings shed light on the novel function of FoxO1 in inhibiting ECM deposition, which is beneficial to ameliorate MC dysfunction.

Nesfatin-1 correlates with hypertension in overweight or obese Han Chinese population.

We investigated blood nesfatin-1 levels in hypertension patients. We found that fasting plasma nesfatin-1 levels were significantly higher in hypertension patients than in control groups, especially in overweight/obese hypertension patients (4.5 ± 2.1 versus 3.3 ± 1.1 ng/ml, p < 0.01). Body mass index, systolic blood pressure and diastolic blood pressure were indeed positively correlated with fasting plasma nesfatin-1 levels (r = 0.234, p < 0.05; r = 0.304, p < 0.01; r = 0.251, p < 0.05; r = 0.461, p < 0.01; respectively). Logistic regression analysis revealed that the plasma level of nesfatin-1 could be independent of risk prediction over standard measures (OR = 1.547, 95% CI: 1.153-6.273, p = 0.026). Nesfatin-1 has the incremental contribution to hypertension risk prediction (IDI: 0.014, p = 0.018; NRI: 0.050, p = 0.043). The plasma nesfatin-1 level in hypertension patients with microalbuminuria are significantly higher than those without microalbuminuria patients (6.4 ± 2.1 ng/ml versus 3.9 ± 1.8 ng/ml, p < 0.01). Nesfatin-1 might play an important role in obesity hypertension, and its increase could be a risk factor for obesity-associated hypertension.

[Effects of over-expressed forkhead transcription factor O1 on high glucose induced extracellular matrix secretion in rat mesangial cells].

OBJECTIVE: To explore the role and molecular mechanism of forkhead transcription factor O1 (FoxO1) for extracellular matrix (ECM) protein accumulation in rat mesangial cells (MCs) cultured under high-glucose conditions. METHODS: The MCs were transfected with either lentiviral vectors containing the sequences of constitutively active FoxO1 (LV-CA-FoxO1) or empty vectors (LV-NC-GFP). The MCs were divided into 4 groups of normal glucose (5.6 mmol/L) (NG), high glucose (25 mmol/L) (HG), high glucose plus LV-CA-FoxO1 (LV-CA) and high-glucose plus empty lentiviral vectors (LV-NC). After 72 h treatment, the mRNA levels of MCs were measured through real-time polymerase chain reaction (PCR) in each group, including FoxO1, fibronectin (FN), collagen I(Col I), collagen IV (Col IV), transforming growth factor-ß1 (TGF-ß1), TGF-ß type Iand type II receptor (TGF-ßRI/II). The protein levels of FoxO1, phosphorylation FoxO1 (p-FoxO1), TGF-ß1 and TGF-ßRI/II were assessed by Western blot. And the distributions of TGF-ßRI in MCs were detected by immunofluorescence. RESULTS: Expression and bioactivity changes of FoxO1: Compared with NG group, there was no significant difference in either mRNA or protein levels of FoxO1 in HG group (P > 0.05) whereas the protein levels of p-FoxO1 markedly increased (P < 0.05). The levels of FoxO1 mRNA and protein increased in LV-CA group versus HG group (mRNA: 17.36 ± 1.13 vs 1.01 ± 0.12, protein: 4.38 ± 0.09 vs 0.93 ± 0.10, both P < 0.05). And there was a significant decrease of p-FoxO1/FoxO1 ratio (P < 0.05). Thus it suggested an enhancement in FoxO1 expression and transcriptional activity. Changes of ECM protein expression. The mRNA expression levels of FN, Col Iand Col IV in HG group were elevated compared with NG group (all P < 0.05) while the expressions of these indices in LV-CA group became attenuated compared with HG group (all P < 0.05). Changes of TGF-ß pathway activation: the expressions of TGF-ß1 and TGF-ßRI/IIboth increased in HG group (all P < 0.05), but decreased in LV-CA group (all P < 0.05). The results of immunofluorescence suggested that the staining of TGF-ßRIdecreased in MCs in LV-CA group versus HG group. All indices of LV-NC group had no statistical difference with those of HG group (all P > 0.05). CONCLUSION: FoxO1 plays a pivotal role in down-regulating the activation of TGF-ß pathway and thereby inhibiting the secretion of ECM.