High Insulin Group Research Articles

Role of M2 Macrophage Exosomes in Osteogenic Differentiation of Mouse Bone Marrow Mesenchymal Stem Cells under High-Glucose and High-Insulin

To study the role of M2 macrophage-derived exosomes (M2-exo) in osteogenic differentiation and Hedgehog signaling pathway of mouse bone marrow mesenchymal stem cells (BMSCs) under in vitro high-glucose and high-insulin conditions. RAW 264.7 cells were induced toward M2 macrophage polarization and then M2-exo were extracted and identified. Immunofluorescence assay was performed to detect the internalization of M2-exo by BMSCs. BMSCs were divided into the normal control group (Control group), the high-glucose and high-insulin group (HGI group), and the HGI with M2-exo intervention group (HGI+M2e group). BMSCs in the Control group were cultured in osteogenic inductive medium with 5.5 mmol/L glucose, but no insulin or M2-exo. BMSCs in the HGI group were cultured in osteogenic inductive medium with 25 mmol/L glucose and 174 nmol/L insulin. BMSCs in the HGI+M2e group were cultured in the same medium as that of the HGI group, with the additional treatment of 6, 30, 60 μg/mL M2-exo, respectively. After osteogenic induction for 7 days and 14 days, alkaline phosphatase (ALP) staining and alizarin red staining were performed respectively to assess the osteogenic differentiation potential of BMSCs from different groups. In addition, BMSCs in the Control group, HGI group, and HGI+M2e group treated with 30 μg/mL M2-exo were examined with qPCR after osteogenic induction for 14 days and Western blot after osteogenic induction for 21 days to assess the osteogenesis and the expression of Hedgehog pathway-related genes and proteins. M2 macrophage polarization was induced successfully, with highly positive expression of CD206, the M2 polarization surface marker. The M2-exo had the typical structure of round or oval-shaped bilayered-membrane vesicles. The diameter distribution of M2-exo ranged from 50 to 125 nm (accounting for 99.14% of all M2-exo). M2-exo samples showed positive expression of exosomal markers CD9, CD63 and CD81 proteins. Immunofluorescence staining showed that M2-exo were taken up and internalized by BMSCs. After osteogenic induction for 7 days, the ALP activity of BMSCs in the HGI group was lower than that of the Control group. After interventions of 6 μg/mL, 30 μg/mL, and 60 μg/mL M2-exo, the ALP activity of the HGI+M2-exo group was significantly increased compared with that of the HGI group ( P<0.05). After osteogenic induction for 14 days, the number of mineralized nodules in the HGI group was lower than that in the Control group, and after intervention, only the HGI+M2e group treated with 30 μg/mL M2-exo showed higher level of mineralization than that in the HGI group ( P<0.05). qPCR analysis revealed that the expression levels of the osteogenesis-related genes, including Runx2, Alp and Ocn, and Hedgehog pathway-related genes, including Gli1, Smo and Ptch1, were downregulated in the HGI group, all being lower than those of the Control group to varying degrees, while 30 μg/mL M2-exo treatment could promote the up-regulation of these genes, showing significant difference in comparison with their expression levels in the HGI group ( P<0.05). In addition, Western blot analysis showed that the expression of the osteogenesis-related proteins, including RUNX2 and COL1A1, and GLI1, the Hedgehog signaling pathway protein, was down-regulated in the HGI group, while the expression of COL1A1 and GLI1 was up-regulated after 30 μg/mL M2-exo treatment, showing significant difference when compared with that of the HGI group ( P<0.05). High glucose and high insulin had inhibitory effect on the osteogenic differentiation potential of BMSCs. After intervention with M2-exo, the Hedgehog signaling pathway in BMSCs was activated and the osteogenic differentiation potential was enhanced, suggesting that M2-exo might have therapeutic potentials for the treatment of diabetic bone disease.

Impact of Hyperinsulinemia on Long-Term Clinical Outcomes of Percutaneous Coronary Intervention in Patients without Diabetes Who Have Acute Myocardial Syndrome.

Background and ObjectivesHyperinsulinemia plays a key role in the development of cardiovascular impairment in patients with metabolic syndrome. The aim of this study was to evaluate the influence of hyperinsulinemia on long-term clinical outcomes of percutaneous coronary intervention (PCI) in patients without diabetes mellitus who have acute myocardial syndrome (ACS).MethodsBetween March 2016 and January 2019, we enrolled 468 patients with ACS and without diabetes mellitus who received primary PCI. Patients were divided into a low-insulin group (n = 157), medium-insulin group (n = 154), and high-insulin group (n = 157) according to tertiles of fasting insulin level. The primary endpoint was major adverse cardiac events (MACE; all-cause death, non-fatal myocardial infarction, target vessel revascularization [TVR]) at 24 months. The secondary endpoint was angina hospitalization.ResultsPatients in the high-insulin group had an unfavorable prognosis, with a higher rate of MACE (34.39%) than the low-insulin group (22.29%) and medium-insulin group (23.37%) at 24 months (P < 0.05). This difference was mainly driven by the increase in TVR. The high-insulin group also had a higher rate of angina hospitalization than the low-insulin group. Cox proportional hazards regression showed that high-insulin level (hazard ratio [HR] 1.870, 95% confidence interval [CI] 1.202–2.909), small-vessel lesion (HR 1.713, 95% CI 1.111–2.642), bifurcation lesion (HR 3.394, 95% CI 2.033–5.067), SYNTAX score (HR 1.084, 95% CI 1.039–1.130), and stent length (HR 1.017, 95% CI 1.002–1.032) increased the incidence of MACE in patients with ACS and without diabetes who underwent PCI.ConclusionHyperinsulinemia might be a valid predictor of clinical outcomes in patients with ACS undergoing PCI.

Metabolomics analysis reveals altered metabolites in lean compared with obese adolescents and additional metabolic shifts associated with hyperinsulinaemia and insulin resistance in obese adolescents: a cross-sectional study

IntroductionHyperinsulinaemia and insulin resistance (IR) are strongly associated with obesity and are forerunners of type 2 diabetes. Little is known about metabolic alterations separately associated with obesity, hyperinsulinaemia/IR and impaired glucose tolerance (IGT) in adolescents.ObjectivesTo identify metabolic alterations associated with obesity, hyperinsulinaemia/IR and hyperinsulinaemia/IR combined with IGT in obese adolescents.Methods81 adolescents were stratified into four groups based on body mass index (lean vs. obese), insulin responses (normal insulin (NI) vs. high insulin (HI)) and glucose responses (normal glucose tolerance (NGT) vs. IGT) after an oral glucose tolerance test (OGTT). The groups comprised: (1) healthy lean with NI and NGT, (2) obese with NI and NGT, (3) obese with HI and NGT, and (4) obese with HI and IGT. Targeted nuclear magnetic resonance-based metabolomics analysis was performed on fasting and seven post-OGTT plasma samples, followed by univariate and multivariate statistical analyses.ResultsTwo groups of metabolites were identified: (1) Metabolites associated with insulin response level: adolescents with HI (groups 3–4) had higher concentrations of branched-chain amino acids and tyrosine, and lower concentrations of serine, glycine, myo-inositol and dimethylsulfone, than adolescents with NI (groups 1–2). (2) Metabolites associated with obesity status: obese adolescents (groups 2–4) had higher concentrations of acetylcarnitine, alanine, pyruvate and glutamate, and lower concentrations of acetate, than lean adolescents (group 1).ConclusionsObesity is associated with shifts in fat and energy metabolism. Hyperinsulinaemia/IR in obese adolescents is also associated with increased branched-chain and aromatic amino acids.

Echocardiographic parameters according to insulin dose in young patients affected by type 1 diabetes.

Insulin dose has been found to associate to several cardiometabolic risk factors in type 1 diabetes. Changes over time in body weight and composition may partly explain this association. However, no data are available on the relationship between insulin dose and echocardiographic parameters of both systolic and diastolic function in type 1 diabetes. Therefore, the aim of the present study was to examine systolic and diastolic echocardiographic parameters in relation to insulin dose in young patients with type 1 diabetes. The study was carried out on 93 consecutive outpatients with type 1 diabetes with a mean age of 32.8 ± 9.8 years. All patients were examined with a transthoracic echocardiography. Clinical and laboratory data were collected. The median value of daily insulin dose was used to categorized patients in two groups: high and low insulin dose group. Patients belonging to the high insulin dose group showed higher levels of cardiometabolic risk factors such as BMI, triglycerides and TG/HDL cholesterol ratio. Indexes of both systolic and diastolic function were similar in both groups except isovolumetric relaxation time (IVRT), that was significantly prolonged in patients of the high insulin group (94.4 ± 15.0 vs 86.7 ± 13.1 ms, p = 0.008). In the multivariate regression analysis, insulin dose was positively and significantly associated with IVRT. In this study we report an association between insulin dose and impaired active diastolic myocardial relaxation. Future studies are needed to further explore this observation.

Association of peripartum plasma insulin concentration with milk production, colostrum insulin levels, and plasma metabolites of Holstein cows

The main objective of this study was to assess associations between plasma insulin concentration around parturition and production in Holstein cows. Primiparous and multiparous cows (n = 267) were enrolled. Blood samples were collected within 12 h after parturition (d 0), and on d 3 and 10 after calving. In addition, blood samples were collected 7 d before (-7 d) the expected date of parturition and colostrum samples were collected within 8 h after parturition from a subset of cows to measure insulin concentration (n = 47). All samples were harvested from 0630 to 1100 h and were used to quantify insulin, nonesterified fatty acids (NEFA), and β-hydroxybutyrate. The plasma concentrations of insulin on d -7 and 0 were not correlated with insulin levels in colostrum. Cows were grouped according to plasma insulin concentration based on the median as low insulin (L-INS) or high insulin (H-INS) on d 0 (median = 0.35 ng/mL; range 0.2 to 1.2), 3 (median = 0.32 ng/mL; range 0.2 to 1.6), and 10 (median = 0.30 ng/mL; range 0.2 to 0.8). We detected that cows in the L-INS group on d 0 (L-INS = 0.57 ± 0.02; H-INS = 0.49 ± 0.02 mmol/L), d 3 (L-INS = 0.56 ± 0.02; H-INS = 0.49 ± 0.02 mmol/L), and d 10 (L-INS = 0.61 ± 0.03; H-INS = 0.55 ± 0.03 mmol/L) had higher NEFA concentrations compared with cows in the H-INS group. Compared with H-INS cows, milk yield was higher for cows classified as L-INS on d 0 (L-INS = 40.75 ± 0.69; H-INS = 38.41 ± 0.64 kg) and d 10 (L-INS = 40.95 ± 0.74; H-INS = 38.66 ± 0.64 kg). Moreover, fat-corrected milk was higher for cows classified as L-INS on d 0 (L-INS = 40.59 ± 2.36; H-INS = 37.73 ± 2.31 kg) and d 10 (L-INS = 41.00 ± 2.42; H-INS = 38.65 ± 2.28 kg) compared with H-INS cows, and energy-corrected milk was higher for L-INS cows compared with H-INS cows regardless of the day (d 0, L-INS = 44.50 ± 0.70 vs. H-INS = 41.67 ± 0.64 kg; d 3, L-INS = 43.65 ± 0.74 vs. H-INS = 40.88 ± 0.72 kg; d 10, L-INS = 44.09 ± 0.73 vs. H-INS = 40.55 ± 0.68 kg). We conclude that low plasma insulin concentration during early lactation is associated with higher milk yield in the long term.

Metformin attenuates triglyceride accumulation in HepG2 cells through decreasing stearyl-coenzyme A desaturase 1 expression

BackgroundThe prevalence of nonalcoholic fatty liver disease (NAFLD) has increased worldwide. Metformin decreases triglyceride (TG) accumulation in hepatocytes in vivo and in vitro. Stearyl-coenzyme A desaturase 1 (SCD1) knockout mice also show decreased liver TG accumulation; however, whether SCD1 plays a role in the effect of metformin on TG accumulation is unknown. Therefore, the aim of this study was to investigate whether SCD1 mediated the effect of metformin on TG accumulation.MethodsHepG2 and AML12 cells were exposed to high glucose and high insulin with or without metformin. An adenovirus was used for the SCD1 knockdown and overexpression. The triglyceride level in cells was detected. The expression of related genes was detected by Western blot and quantitative real-time PCR. A dual-luciferase reporter assay was used to determine the effect of metformin on the transcriptional activity of the SCD1 promoter.ResultsMetformin decreased TG accumulation to normal level in HepG2 cells exposed to high glucose and high insulin. The expression of SCD1 and fatty acid synthetase (FAS) was also decreased to normal level by metformin. Knockdown of SCD1 mimicked the effect of metformin on decreasing TG levels in AML12 cells, and the overexpression of SCD1 attenuated the effect of metformin on decreasing TG accumulation in HepG2 cells. The dual-luciferase reporter assay showed that the transcriptional activity of the SCD1 promoter (− 550/+ 199) after metformin treatment was 2-fold lower compared to control group in HepG2 cells. Additionally, the phosphorylation of AMPK after metformin treatment was 2-fold higher, and the expression of sterol regulatory element-binding protein-1c (SREBP-1c) after metformin treatment was about 2-fold lower compared to high glucose and high insulin group in HepG2 cells.ConclusionsTogether, these results reveal that metformin reduces TG accumulation in HepG2 cells via inhibiting the expression of SCD1.

Association between high-fasting insulin levels and metabolic syndrome in non-diabetic middle-aged and elderly populations: a community-based study in Taiwan

ObjectivesWe aimed to determine the association between fasting insulin (FI) levels and metabolic syndrome (MetS) in non-diabetic middle-aged and elderly adults in a community in Taiwan.DesignCross-sectional observational study.SettingCommunity-based investigation in...

Insulin promotes preantral follicle growth and antrum formation through temporal expression of genes regulating steroidogenesis and water transport in the cat.

The aims of the present study were to determine the effects of insulin, invitro, on: (1) the viability and growth of domestic cat ovarian follicles; (2) mRNA expression of genes regulating steroidogenesis (cytochrome P450 family 17 subfamily, A polypeptide 1 (Cyp17a1), cytochrome P450 family 19 subfamily, A polypeptide 1 (Cyp19a1) and steroidogenic acute regulatory protein (Star)) and water transport (aquaporins (AQPs) Aqp1, Aqp3, Aqp7, Aqp9); and (3) steroid production (17β-oestradiol (E2), progesterone (P4), androstenedione (A4)). Cat secondary follicles were isolated from ovarian cortices and cultured in 0 (Control), 1 or 10µgmL-1 insulin for 14 days (Day 0=culture onset). Follicle and oocyte viability (based on neutral red staining), diameter and antrum formation were assessed every 72h and at the end of incubation (Day 14). Expression of steroidogenic and water transport genes was evaluated on Days 0, 6 and 12, and E2, P4 and A4 concentrations in the culture medium were determined on Day 12. By Day 14, 1 and 10µgmL-1 insulin had significantly promoted (P<0.05) both antrum formation in a mean (±s.e.m.) 26.9±9.0% and 78.0±10.0% of follicles respectively, and follicle growth (diameter 151.4±4.5 and 169.9±10.5µm respectively) compared with Control (antrum formation in 3.3±3.3% of follicles and follicle diameter 129.1±6.6µm). High insulin (10µgmL-1) treatment increased follicle viability compared with Control (86.0±9.8% vs 38.1±10.9% respectively; P<0.05). However, insulin had no beneficial effect (P>0.05) on oocyte diameter. Cyp17a1 expression on Days 6 and 12 was higher (P<0.05) in follicles cultured in the low (1µgmL-1) compared with high (10µgmL-1) insulin treatment, with no significant difference between low or high insulin vs Control groups. Star expression was higher (P<0.01) in the low insulin compared with Control group on Day 6, but Star was undetectable in the high insulin group by Day 12. Compared with high insulin, low insulin increased (P<0.05) Aqp1 expression on Day 6, but there were no significant differences between these two groups on Day 12. In contrast, high insulin decreased (P<0.05) Aqp9 transcript levels compared with Control. Only P4 production was affected by insulin, with P4 concentrations in the medium being higher (P<0.05) in the low compared with high insulin and Control groups. In summary, the findings indicate that insulin promotes cat ovarian follicle growth and survival invitro, including enhanced antrum formation, with the likely mechanism involving temporal expression of Cyp17a1, Star and Aqp9 genes.

Acute interaction between hydrocortisone and insulin alters the plasma metabolome in humans

With the aim of identifying biomarkers of glucocorticoid action and their relationship with biomarkers of insulin action, metabolomic profiling was carried out in plasma samples from twenty healthy men who were administered either a low or medium dose insulin infusion (n = 10 each group). In addition, all subjects were given metyrapone (to inhibit adrenal cortisol secretion) + /− hydrocortisone (HC) in a randomised crossover design to produce low, medium and high glucocorticoid levels. The clearest effects of insulin were to reduce plasma levels of the branched chain amino acids (BCAs) leucine/isoleucine and their deaminated metabolites, and lowered free fatty acids and acylcarnitines. The highest dose of hydrocortisone increased plasma BCAs in both insulin groups but increased free fatty acids only in the high insulin group, however hydrocortisone did not affect the levels of acyl carnitines in either group. The clearest interaction between HC and insulin was that hydrocortisone produced an elevation in levels of BCAs and their metabolites which were lowered by insulin. The direct modulation of BCAs by glucocorticoids and insulin may provide the basis for improved in vivo monitoring of glucocorticoid and insulin action.

Acute physiological effects of glucocorticoids on fuel metabolism in humans are permissive but not direct

Background and aimsThe effects of glucocorticoids on fuel metabolism are complex. Acute glucocorticoid excess promotes lipolysis but chronic glucocorticoid excess causes visceral fat accumulation. We hypothesized that interactions between cortisol and insulin and adrenaline account for these conflicting results. We tested the effect of cortisol on lipolysis and glucose production with and without insulin and adrenaline in humans both in vivo and in vitro.Materials and methodsA total of 20 healthy men were randomized to low and high insulin groups (both n = 10). Subjects attended on 3 occasions and received low (c. 150 nM), medium (c. 400 nM) or high (c. 1400 nM) cortisol infusion in a randomized crossover design. Deuterated glucose and glycerol were infused intravenously along with a pancreatic clamp (somatostatin with replacement of glucagon, insulin and growth hormone) and adrenaline. Subcutaneous adipose tissue was obtained for analysis. In parallel, the effect of cortisol on lipolysis was tested in paired primary cultures of human subcutaneous and visceral adipocytes.Results In vivo, high cortisol increased lipolysis only in the presence of high insulin and/or adrenaline but did not alter glucose kinetics. High cortisol increased adipose mRNA levels of ATGL, HSL and CGI‐58 and suppressed G0S2. In vitro, high cortisol increased lipolysis in the presence of insulin in subcutaneous, but not visceral, adipocytes.ConclusionsThe acute lipolytic effects of cortisol require supraphysiological concentrations, are dependent on insulin and adrenaline and are observed only in subcutaneous adipose tissue. The resistance of visceral adipose tissue to cortisol's lipolytic effects may contribute to the central fat accumulation observed with chronic glucocorticoid excess.

Relationship between Insulin Levels and Nonpsychotic Dementia: A Systematic Review and Meta-Analysis

Objectives To explore the relationship between insulin levels and nonpsychotic dementia. Methods Six electronic databases (PubMed, Cochrane, SCI, CNKI, VIP, and Wanfang) were searched from January 1, 2007, to March 1, 2017. Experimental or observational studies that enrolled people with nonpsychotic dementia or abnormal insulin levels in which insulin levels or MMSE scores (events in nonpsychotic dementia) were the outcome measures. Random-effects models were chosen for this meta-analysis. Sample size, mean, s.d., and events were primarily used to generate effect sizes (with the PRIMA registration number CRD42017069860). Results 50 articles met the final inclusion criteria. Insulin levels in cerebrospinal fluid were lower (Hedges' g = 1.196, 95% CI = 0.238 to 2.514, and P = 0.014), while the levels in peripheral blood were higher in nonpsychotic dementia patients (Hedges' g = 0.853 and 95% CI = 0.579 to 1.127), and MMSE scores were significantly lower in the high insulin group than in the healthy control group (Hedges' g = 0.334, 95% CI = 0.249 to 0.419, and P = 0.000). Conclusions Our comprehensive results indicate that blood insulin levels may increase in patients with nonpsychotic dementia.

The Relation between Serum Uric Acid and HbA1c Is Dependent upon Hyperinsulinemia in Patients with Newly Diagnosed Type 2 Diabetes Mellitus.

Objective. The aim of our study was to explore the dependent condition of the relationship between uric acid and blood glucose in type 2 diabetes. Research Design and Methods. We measured the HbA1c, serum uric acid, creatinine, lipids profiles, and so forth of 605 newly diagnosed type 2 diabetes patients, and oral glucose tolerance tests (OGTTs) were performed on each patient. The population was divided into high and low insulin groups. Multiple linear regression analyses were conducted to assess the relationship between uric acid and HbA1c. Results. Serum uric acid and HbA1c levels were low in newly diagnosed type 2 diabetes patients. However, we found no significant relationship between uric acid and HbA1c by regression analysis after adjusting total insulin. The concentration of uric acid was inversely correlated with HbA1c in the high insulin group, regardless of patient sex. However, no associations were found in low insulin group. Conclusions. The negative correlation between uric acid and HbA1c is conditional in newly diagnosed type 2 diabetes patients and is related to hyperinsulinemia. Therefore, uric acid is likely only useful as a biomarker of blood glucose in patients exhibiting hyperinsulinemia.

Body Composition and Metabolic Risk Variables for Obesity in Korean Healthy Women

Objective: To investigate which indices to relate with metabolic risk among body composition. Design and methods: As a cross sectional study, a total of 47 healthy Korean women aged 40.36 years, BMI 25.59 kg/m2, were assessed their BMI, percent of body fat (BIA), fat mass by DXA, adipose tissue (CT), fasting insulin, free fatty acid, cholesterol, triglyceride, HDL cholesterol, fasting plasma glucose, and HOMA IR index. Results: Neither dyslipidemia nor hyperglycaemia has any relation with central fatness indicators & BMI. Fasting insulin is related with %BF* after correction for age. Dyslipidemia and hyperglycaemia tend to be greater in low insulin group. Among body composition, BMI, ** fat mass (DXA) **, %BF** except VAT, SAT, and V/S ratio of high insulin group are significantly higher than those of low insulin group. Although not significant, V/S ratio is likely to be higher in low insulin group. (*: p<0.05, **:p<0.01) Conclusions: V/S ratio maybe a determinant to metabolic risks variables in healthy Korean women. Controlled lager cohort study would be considered in the future, to prove the association between SAT and metabolic derangement. In conclusion, %BF through BIA is considered as the major determinant to fasting insulin in Korean women.

Assessment of metabolic status in young Japanese females using postprandial glucose and insulin levels.

Lifestyle-related diseases develop through the accumulation of undesirable lifestyle habits both prior to the onset of disease as well as during normal healthy life. Accordingly, early detection of, and intervention in, metabolic disorders is desirable, but is hampered by the lack of an established evaluation index for young individuals. The purpose of this study was to investigate the utility of a biomarker of health in young female subjects. The subjects were young healthy Japanese females in whom energy expenditure was measured for a period of 210 min after a test meal. In addition, Δplasma glucose and Δserum insulin were calculated from the fasting and 30 min values. ΔPlasma glucose and Δserum insulin levels varied widely compared to fasting levels. Both the area under the curve of carbohydrate oxidation rate and serum free fatty acid levels were higher in individuals in the high Δplasma glucose group. Moreover, Δplasma glucose was higher in individuals in the high Δserum insulin group than in the low Δserum insulin group. We conclude that nutritional balanced liquid loading test using Δplasma glucose and Δserum insulin as the evaluation index is useful for the detection of primary metabolic disorders in young females.

The manner of the inflammation-boosting effect caused by acute hyperglycemia secondary to overfeeding and the effects of insulin therapy in a rat model of sepsis

BackgroundThe aim of the study was to investigate both the inflammation-boosting effect and the metabolic stress induced by acute hyperglycemia secondary to overfeeding with excessive glucose infusion and the effects of insulin therapy on those events in a rat model of sepsis. Materials and methodsSprague–Dawley rats underwent cecal ligation and puncture (CLP) or sham operation. Preestablished continuous intravenous glucose infusion was initiated immediately after surgery. First, rats with CLP-inducing sepsis were divided into three groups on the basis of the target blood glucose (BG) levels: high glucose (HG) group (overfed, >300 mg/dL), moderate glucose group (moderate hyperglycemia, 200–300 mg/dL), and no glucose group (100–150 mg/dL). The sham group received the same glucose infusion as that of the HG group. BG and plasma interleukin (IL) 6 levels were monitored over time. All rats were sacrificed 9 h after surgery to evaluate lung histology and measure hepatic total glutathione and malondialdehyde contents. Based on the results, the high glucose and insulin (HI) group was added to septic groups as a model of insulin therapy, in which insulin with the same HG dose as that in the HG group was administered to maintain moderate hyperglycemia. ResultsBG level in all groups remained in the preestablished target range throughout the experiment. Plasma IL-6 level in all septic groups increased in a time-dependent manner, whereas that in the sham group with moderate hyperglycemia hardly increased. Nine hours after CLP, plasma IL-6 level in the HG group rose to 7407.5 ± 1987.3 pg/mL, which was three times higher than that in the other septic groups. There was no significant difference among moderate glucose, no glucose, and HI groups, in which BG level remained constant at <300 mg/dL. The HG group showed the worst consequences of lung injury and oxidative stress in the liver, which were completely stable in HI group. ConclusionsAcute severe hyperglycemia in critical illness might excessively boost the existing systemic inflammatory response in a threshold-based manner. Insulin therapy under overfeeding could strongly inhibit such a boosting effect and oxidative stress in the liver.

Inhibitory effects of high glucose/insulin environment on osteoclast formation and resorption in vitro

Patients with type 2 diabetes mellitus (T2DM) exhibit hyperglycemia and hyperinsulinemia and increased risk of fracture at early stage, but they were found to have normal or even enhanced bone mineral density (BMD). This study was aimed to examine the molecular mechanisms governing changes in bone structure and integrity under both hyperglycemic and hyperinsulinemic conditions. Monocytes were isolated from the bone marrow of the C57BL/6 mice, induced to differentiate into osteoclasts by receptor activator of nuclear factor kappa-B ligand (RANKL) and macrophage colony-stimulating factor (M-CSF) and exposed to high glucose (33.6 mmol/L), high insulin (1 μmol/L), or a combination of high glucose/high insulin (33.6 mmol/L glucose and 1 μmol/L insulin). Cells cultured in α-MEM alone served as control. After four days of incubation, the cells were harvested and stained for tartrate resistant acid phosphatase (TRAP). Osteoclast-related genes including RANK, cathepsin K and TRAP were determined by using real-time PCR. The resorptive activity of osteoclasts was measured by using a pit formation assay. Osteoclasts that were derived from monocytes were of multinucleated nature and positive for TRAP, a characteristic marker of osteoclasts. Cell counting showed that the number of osteoclasts was much less in high glucose and high glucose/high insulin groups than in normal glucose and high insulin groups. The expression levels of RANK and cathepsin K were significantly decreased in high glucose, high insulin and high glucose/high insulin groups as compared with normal glucose group, and the TRAP activity was substantially inhibited in high glucose environment. The pit formation assay revealed that the resorptive activity of osteoclasts was obviously decreased in high glucose group and high glucose/high insulin group as compared with normal group. It was concluded that osteoclastogenesis is suppressed under hyperglycemic and hyperinsulinemic conditions, suggesting a disruption of the bone metabolism in diabetic patients.

Insulin-Mediated Oxidative Stress and DNA Damage in LLC-PK1 Pig Kidney Cell Line, Female Rat Primary Kidney Cells, and Male ZDF Rat Kidneys In Vivo

Hyperinsulinemia, a condition with excessively high insulin blood levels, is related to an increased cancer incidence. Diabetes mellitus is the most common of several diseases accompanied by hyperinsulinemia. Because an elevated kidney cancer risk was reported for diabetic patients, we investigated the induction of genomic damage by insulin in LLC-PK1 pig kidney cells, rat primary kidney cells, and ZDF rat kidneys. Insulin at a concentration of 5nM caused a significant increase in DNA damage in vitro. This was associated with the formation of reactive oxygen species (ROS). In the presence of antioxidants, blockers of the insulin, and IGF-I receptors, and a phosphatidylinositol 3-kinase inhibitor, the insulin-mediated DNA damage was reduced. Phosphorylation of protein kinase B (PKB or AKT) was increased and p53 accumulated. Inhibition of the mitochondrial and nicotinamide adenine dinucleotide phosphatase oxidase-related ROS production reduced the insulin-mediated damage. In primary rat cells, insulin also induced genomic damage. In kidneys from healthy, lean ZDF rats, which were infused with insulin to yield normal or high blood insulin levels, while keeping blood glucose levels constant, the amounts of ROS and the tumor protein (p53) were elevated in the high-insulin group compared with the control level group. ROS and p53 were also elevated in diabetic obese ZDF rats. Overall, insulin-induced oxidative stress resulted in genomic damage. If the same mechanisms are active in patients, hyperinsulinemia might cause genomic damage through the induction of ROS contributing to the increased cancer risk, against which the use of antioxidants and/or ROS production inhibitors might exert protective effects.

High insulin concentrations promote the in vitro growth and viability of canine preantral follicles

To determine whether the effects of different concentrations of insulin on the development of canine preantral follicles in vitro were associated or not with FSH, secondary follicles were isolated and cultured. In Experiment 1, follicles were cultured in the following media: modified minimum essential medium (CtrlMEM) alone; CtrlMEM plus 5 ng mL⁻¹ insulin (Ins5ng); CtrlMEM plus 10 ng mL⁻¹ insulin (Ins10ng); and CtrlMEM plus 10 μg mL⁻¹ insulin. In Experiment 2, follicles were cultured in the same media but in the presence of sequential FSH (i.e. CtrlFSH, Ins5ngF, Ins10ngF and 10μgF, respectively). Increasing concentrations of FSH (100, 500 and 1000 ng mL⁻¹) were added sequentially to the culture medium on Days 0, 6 and 12 of culture. Viability were assessed at the end of culture and follicular diameter and the antrum formation rate at four time points (Days 0, 6, 12 and 18). In Experiment 1, the high insulin concentration significantly increased follicular viability (P<0.05). In contrast, in Experiment 2, viability was not affected by the inclusion of insulin. In addition, viability was significantly better in follicles cultured in CtrlFSH (P<0.05). The diameter of follicles in the high-insulin group in Experiment 1 and high-insulin plus FSH group in Experiment 2 was superior to other groups tested. In experiment 2, the Ins10μg and Ins10μgF groups exhibited significantly higher antrum formation rates than the other groups. In conclusion, in the absence of FSH, high concentrations of insulin have beneficial effects on follicular viability. However, to promote the growth of canine preantral follicles in vitro, it is recommended that a combination of insulin and FSH be added to the medium.

The level of fasting serum insulin, but not adiponectin, is associated with the prognosis of early stage hepatocellular carcinoma

Impaired glucose tolerance influences the prognosis of hepatocellular carcinoma (HCC), but this mechanism is still not fully understood. We investigated the impact of the fasting serum levels of insulin and adiponectin on the prognosis of HCC and its recurrence. One hundred and forty patients with newly diagnosed HCC were enrolled in the prognosis study. Their fasting serum levels of insulin and adiponectin were determined. Of 140 patients, 59 patients who underwent curative treatment were subjected to analysis of the recurrence-free survival. The 140 patients were divided into two groups by the 50th percentile value of insulin (7.73 microIU/ ml) or total adiponectin (6.95 microg/ml). Kaplan-Meier analysis indicated that high insulin group (>7.73 microIU/ml) exhibited a significantly poorer prognosis than low insulin group (<7.73 microIU/ml) in early stage HCC (P=0.018). In contrast, the level of total adiponectin had no impact on the prognosis of HCC. Multivariate analysis indicated that fasting hyper-insulinemia was an independent risk factor for a poorer prognosis in early stage HCC (P=0.044). Likewise, Kaplan-Meier analysis indicated that the recurrence-free survival of high insulin group was significantly lower than that of low insulin group (P=0.017). The level of total adiponectin had no impact on the recurrence-free survival of HCC. Multivariate analysis indicated that fasting hyperinsulinemia was an independent risk factor for the lower recurrence-free survival of HCC (P=0.049). In conclusion, our study suggests that the fasting insulin level affects the clinical course of early stage HCC.

Insulin-dependent actions of pioglitazone in newly diagnosed, drug naïve patients with type 2 diabetes

The aim of this study was to study the effects of pioglitazone on several diabetic parameters with subjects possessing distinct levels of insulin. Treatment naive patients with type 2 diabetes received 15-30 mg/day pioglitazone monotherapy. At 3 months, levels of insulin, C-peptide, HbA1c, HOMA-R, HOMA-B and BMI were compared with those at baseline between the low (below 5.9 microU/ml, n = 48), medium (11.9-6 microU/ml n = 39) and high (above 12 microU/ml, n = 33) insulin groups. At baseline, differences existed in the levels of HbA1c, insulin, C-peptide, HOMA-R, HOMA-B, and BMI between these groups. In the high-insulin group significant reductions of insulin/C-peptide levels were observed, while in the low-insulin group significant increases of insulin/C-peptide were observed. In the medium-insulin group, no significant changes were observed. In contrast, the HbA1c levels significantly and similarly decreased in all the groups. Significant correlations between the changes of insulin/C-peptide levels with pioglitazone and the baseline insulin/C-peptide levels were observed. HOMA-R showed greater reductions in the high-insulin group, while HOMA-B showed greater increases in the low-insulin group in comparison to other groups. Multiple regression analysis revealed that the baseline insulin level is the predominant determinant of the changes of insulin levels with pioglitazone. These results suggest that pioglitazone appears to have two effects: to reduce insulin resistance (and lower insulin) and to improve beta-cell function (and increase insulin). The predominance of these effects appears to be determined by the insulin levels. Based on these data, a novel physiological model showing that pioglitazone may shift the natural history of diabetes toward an earlier stage (rejuvenation of beta-cell function) will be presented.