Decreased Expression of Heat Shock Protein 72 In Skeletal Muscle of Patients With Type 2 Diabetes Correlates With Insulin Resistance
Article Figures & Tables
Figures
- FIG. 1.
Quantitation of Hsp-72 mRNA in human muscle biopsies and heat-shocked NIH-3T3 cells, using competitive RT-PCR. A: Equal amounts of total RNA isolated from human muscles was added to titrated quantities (as indicated) of sRNA (competitor). RT-PCR of this sRNA yielded a product that was 80 nucleotides (nt) shorter (210 nt) than the amplification output generated from the native human Hsp-72 mRNA (290 nt, target), and that could therefore be distinguishable on the gel. B: The ratios of the amplification products from the competitor and target (C/T) were plotted against the number of copies of competitor added, and the plot was used to determine the equivalence point (indicated by arrows) between sRNA and target RNA. C: Kinetics of Hsp-72 expression after heat treatment of mammalian cells. NIH-3T3 cells were heat-treated at 43°C for 30 min and then incubated at 37°C. Total RNA was isolated at the indicated time points after heat treatment, and the amount of Hsp-72-specific message was determined with the competitive quantitative method. Results from two independent experiments (open or filled circles) are shown. Arrows indicate the beginning and the end of heat treatment. Basal Hsp-72 levels are shown by the two triangles. *106 copy/μg total RNA.
- FIG. 2.
Hsp-72 mRNA concentration in muscle tissues of control and type 2 diabetic patients. Amount of Hsp-72 messenger RNA was determined in muscle biopsies of six nondiabetic (control) and six type 2 diabetic patients (T2DM) using the competitive RT-PCR as described in research design and methods. Individual data are expressed, using log10 scale, as copies of Hsp-72 mRNA in 1 μg total RNA. The difference between the two groups is statistically significant (Mann-Whitney U test, P = 0.039). *106 copy/ μg total RNA.
- FIG. 3.
Hsp-72 mRNA levels in muscle biopsies of control subjects and identical monozygotic twins discordant for type 2 diabetes. The amount of Hsp-72 messenger RNA was determined using the competitive RT-PCR as described in research design and methods. A: Individual data are shown, using log10 scale, as copies of Hsp-72 mRNA in 1 μg total RNA. Statistical differences between the different groups were calculated with the nonparametric Mann-Whitney U test. B: Comparison of the expression of Hsp-72 mRNA between nondiabetic and diabetic individuals in twin-pairs. ○, twins with type 2 diabetes; ▴, twins with IGT; ▵, twins with NGT. Statistical differences (Wilcoxon-paired test) between nondiabetic and diabetic twins: P = 0.0431 for type 2 diabetes vs. NGT; P = 0.1755 (NS) for type 2 diabetes vs. IGT; P = 0.028for type 2 diabetes vs. NGT+IGT. *106 copy/μg total RNA. Tw, twin; T2DM, type 2 diabetes.
- FIG. 4.
Correlation between muscle Hsp-72 mRNA expression and rates of glucose and lipid turnover, as determined in protocol 2 (n = 36). All of the values are expressed as milligrams per kilogram FFM per minute. ○, control subjects; ▵, nondiabetic twins; ⧫, twins with type 2 diabetes. A: Insulin-stimulated total glucose uptake (RdI): control subjects: R = 0.531, P = 0.078 NS; nondiabetic subjects: R = 0.581, P = 0.0053; twins: R = 0.519, P = 0.0128; twins with type 2 diabetes: R = 0.441, NS. B: Insulin-stimulated glucose oxidation (GoxI): controls: R = 0.364, NS; nondiabetic subjects: R = 0.253, NS; twins: R = 0.564, P = 0.0068; twins with type 2 diabetes: R = 0.594, P = 0.0487. C: Net glycogen synthesis during insulin infusion (GsI): controls: R = 0.322, NS; nondiabetic subjects: R = 0.322, NS; twins: R = 0.514, P = 0.0137; twins with type 2 diabetes: R = 0.487, NS. D: lipid oxidation during insulin infusion (LipoxI): control subjects: R = −0.634, P = 0.0355; nondiabetic subjects: R = −0.425, P = 0.0416; twins: R = −0.389, P = 0.0623 (NS); twins with type 2 diabetes: R = 0.643, P = 0.0329 for. *106 copy/μg total RNA.
Tables
- TABLE 1
Clinical characteristics of study subjects
Parameters Protocol 1 Protocol 2 Type 2 diabetic subjects Control subjects Type 2 diabetic co-twins Nondiabetic co-twins Control subjects n (F/M) 6 (1/5) 6 (0/6) 12 (5/7) 12 (5/7) 12 (6/6) Age (years) 51 ± 4 48 ± 9 64 ± 3 64 ± 3 61 ± 2 BMI (kg/m2) 25.1 ± 2.7 27.1 ± 4.2 30.1 ± 1.3 27.5 ± 1.3 26.0 ± 1.0 Fasting plasma glucose (mmol/l) 8.9 ± 0.8* 5.7 ± 0.2 11.4 ± 1.1* 6.0 ± 0.2‡ 5.4 ± 0.2 Fasting plasma insulin (μU/ml) ND ND 12.7 ± 1.6* 7.3 ± 2.2 6.9 ± 0.9 2-h glucose (mmol/l) ND ND 19.5 ± 1.7* 8.1 ± 1.6‡ 5.7 ± 0.3 Glucose uptake (mg · kg FFM−1 · min−1) 2.8 ± 1.2† 6.8 ± 2.2 5.2 ± 0.7* 8.5 ± 0.8‡ 11.3 ± 0.9 Glucose storage (mg · kg FFM−1 · min−1) ND ND 2.8 ± 0.6* 4.8 ± 0.6‡ 7.6 ± 0.9 Glucose oxidation (mg · kg FFM−1 · min−1) ND ND 2.4 ± 0.2* 3.7 ± 0.2 3.8 ± 0.2 Lipid oxidation (mg · kg FFM−1 · min−1) ND ND 1.23 ± 0.13* 0.76 ± 0.18 0.65 ± 0.07 Data are means ± SD. ND, not determined in the study.
- *
* P < 0.01 compared with control subjects
- †
† P < 0.02 compared with control subjects;
- ‡
‡ P < 0.05 compared with control subjects
- TABLE 2
Correlation between skeletal muscle Hsp-72 mRNA and plasma fasting glucose or insulin levels, and 2-h glucose levels during oral glucose tolerance testing in different subgroups of patients from protocol 2
Parameters Control subjects without family background of diabetes Nondiabetic subjects Twins (all) Diabetic subjects n 12 24 24 12 Fasting glucose −0.106 (NS) −0.397 (NS) −0.391 (NS) −0.455 (NS) Fasting insulin −0.091 (NS) −0.197 (NS) −0.129 (NS) −0.210 (NS) 2-h glucose −0.744 (0.0236)* −0.716 (0.0006)* −0.327 (NS) −0.263 (NS) Data are R (P).
- *
* Statistically significant correlations.
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