Zdeněk Rušavý

Parameters of oxidative stress, DNA damage and DNA repair in type 1 and type 2 diabetes mellitus

Objectives: (i) to determine the extent of oxidative stress and DNA damage and repair using a panel of selected markers in patients with type 1 and type 2 diabetes mellitus (T1DM, T2DM), (ii) to find their possible relationships with diabetes compensation and duration, and finally (iii) to test for the effect of functional polymorphisms in the 8-oxoguanin DNA glycosylase (rs1052133), catalase (rs1001179) and superoxide dismutase (rs4880) genes on respective intermediate phenotypes. Methods: A total of 207 subjects (23 children and 44 adults with T1DM, 52 adult patients with T2DM and 88 healthy adult control subjects) were enrolled in the study. The following markers of redox state were determined in participants: erythrocyte superoxide dismutase (Ery-SOD), whole blood glutathione peroxidase (WB-GPx), erythrocyte glutathione (Ery-GSH), plasma total antioxidant capacity (P-tAOC) and plasma malondialdehyde (P-MDA). Furthermore, the extent of DNA damage and repair was ascertained using the following parameters: DNA single strand breaks (DNAssb), DNA repair capacity (DNArc) and DNA repair index (DNRI). Results: Comparison of T1DM vs. T2DM patients revealed significantly higher Ery-GSH content (P < 0.0001) and significantly lower Ery-SOD activity (P = 0.0006) and P-tAOC level (P < 0.0001) in T1DM subjects. T2DM diabetics exhibited a significant increase in DNAssb (P < 0.0001) and significant decrease in both DNArc (P < 0.0001) and DNRI (P < 0.0001) compared with T1DM patients. Patient’s age (irrespective of DM type) significantly correlated with DNAssb (r = 0.48, P < 0.0001), DNArc (r = −0.67, P < 0.0001) and DNRI (r = −0.7, P < 0.0001). Allele frequencies of all studied polymorphisms did not exhibit any significant association with the investigated parameters. Conclusion: We demonstrated significant age- and DM type-related changes of oxidative DNA modification and capacity for its repair in subjects with T1DM and T2DM.

Could we predict asymptomatic cardiovascular autonomic neuropathy in type 1 diabetic patients attending out-patients clinics?

ZusammenfassungHINTERGRUND UND ZIELE: Die kardiovaskuläre autonome Neuropathie (KAN) ist bei Patienten mit Typ 1 Diabetes mellitus (DM1) mit einer erhöhten Morbidität und Mortalität verbunden. Diese Komplikation kann über lange Zeit ohne Symptome einhergehen. Ziel dieser Studie war es die Prävalenz, den Schweregrad und mögliche Vorhersageparameter der asymptomatischen KAN zu erfassen. PATIENTEN UND METHODEN: 107 Patienten mit DM1 (52 Männer, 55 Frauen, Alter 39,8 ± 12,4 Jahre [18–72]; Dauer des DM1: 16,6 ± 9,5 Jahre [0,5–43], Alter bei Manifestation 23,5 ± 12,8 Jahre [1–54], BMI 25,1 ± 3,2 [1,9–33,91]) wurden untersucht. Das Vorliegen einer KAN wurde mit Hilfe von Standard-Reflex-Testen (Ewing Batterie) erhoben und die Patienten wurden entsprechend den Ergebnissen in drei Gruppen eingeteilt: Gruppe 0 ohne KAN, Gruppe I mit erstgradiger KAN und Gruppe 2 mit zweitgradiger KAN. Wir erhoben außerdem die mit KAN am häufigsten einhergehenden chronischen Komplikationen des DM1, Episoden schwerer Hypoglykämie, zeitliche Parameter des DM1 (Patientenalter, Dauer des DM1, Alter bei Manifestation) und außerdem DM1-spezifische Parameter wie glykosyliertes Hb, BMI, kardiovaskuläre Erkrankungen und Blutdruck. Die Vorhersagbarkeit einer KAN wurde entsprechend den erhobenen Korrelationen beurteilt. ERGEBNISSE: Nur 50 der 107 der Patienten (46%) zeigten keine KAN. Bei 38 Patienten (36%) fanden wir eine erstgradige , bei 19 (18%) eine zweitgradige KAN. KAN korrelierte mehr mit der Dauer des DM1 (p < 0,001), als mit dem Alter der Patienten (p < 0,05). Es bestand eine grenzwertig signifikante Korrelation (p = 0,053) mit dem glykosylierten Hb. Weiters haben wir eine signifikant positive Korrelation zwischen KAN und dem Auftreten von chronischen Komplikationen (periphere Neuropathie: p < 0,001), Retinopathie: p < 0,001, Nephropathie: erhöhtes Kreatinin: p < 0,03, Albuminurie: p < 0,01) gefunden. Obwohl der Blutdruck bei allen Patienten im Normalbreich (124,2/74,5 ± 11,5/7,8 mmHg) lag, wurde eine positive Korrelation mit der KAN (p < 0,05) bestätigt. Zu den akuten Komplikationen des DM1 bestand keine Korrelation. SCHLUSSFOLGERUNGEN: Unsere Studie zeigt, dass die asymptomatische KAN bei Patienten mit DM1 sehr häufig ist. Mit Hilfe von multifaktorieller logistischer Regression konnten wir zeigen, dass bei gleichzeitigem Vorliegen von Albuminurie, peripherer Neuopathie und erhöhtem systolischem Blutdruck eine hohe Wahrscheinlichkeit für das zusätzliche Vorhandensein einer KAN besteht.SummaryBACKGROUND AND AIMS: Diabetic cardiovascular autonomic neuropathy (CAN) is associated with increased morbidity and mortality. This complication may be asymptomatic for a long time. The aim of this study was to assess the prevalence, severity and predictors of asymptomatic CAN in patients with type 1 diabetes mellitus (DM1). PATIENTS AND METHODS: 107 patients with DM1 were enrolled: 52 men and 55 women aged 39.8 ± 12.4 years (18–72), duration of DM 16.6 ± 9.5 years (0.5–43), age at DM manifestation 23.5 ± 12.8 years (1–54) and BMI 25.1 ± 3.2 (18.9–33.91). CAN was assessed using standard cardiovascular reflex tests (Ewing battery) and the patients were divided into three groups according to the results: Group 0, without CAN; Group I, 1st degree CAN; Group II, 2nd degree CAN. We assessed the most frequent relationships between CAN and chronic complications, episodes of severe hypoglycemia, time-related parameters (age of patients, duration of diabetes, age at manifestation), glycosylated hemoglobin (HbA1c), BMI, cardiovascular diseases and blood pressure, and determined the predictability of CAN on the basis of these relationships. RESULTS: Only 50 of the 107 patients (46%) showed no CAN. We found 1st degree CAN in 38 patients (36%) and 2nd degree CAN in 19 (18%). CAN correlated more significantly with the duration of diabetes (p < 0.001) than with age (p < 0.05). The relationship between CAN and HbA1c was on the borderline of statistical significance (p = 0.053). We found a positive correlation between CAN and the presence of chronic complications [peripheral neuropathy (p < 0.001), retinopathy (p < 0.001), and some markers of nephropathy: creatinine (p < 0.03), albuminuria (p < 0.01)]. Although blood pressure was within the physiological range (124.2/74.5 ± 11.5/7.8 mmHg) in all patients, a positive correlation with CAN was confirmed (p < 0.05). No relationship with occurrence of severe hypoglycemia was found. CONCLUSIONS: According to our results, asymptomatic CAN is very frequent in patients with DM1. By using multifactorial logistic regression (step-wise) analysis we demonstrated that if albuminuria, peripheral neuropathy and elevated systolic BP are present simultaneously, there is a high probability that the patient also has CAN (84.9% of initial group correctly predicted, p < 0.001).

The insulin analog glargine during an unplanned pregnancy.

Sir, We have read with interest the papers by Leipold and coworkers on diabetic fetopathy and the editorial on ‘designer’ insulins [1–3]. The use of the insulin analog glargine during pregnancy is not recommended. What we do with this treatment during unplanned pregnancy? We present the cases of seven women with type 1 diabetes treated with the insulin analog glargine during pregnancy. The unplanned pregnancies were diagnosed at 5–10 weeks’ gestation. The patients were treated with long-acting glargine and multiple daily injections of shortacting analogs (lispro, aspart). Glycosylated hemoglobin levels before and during pregnancy are shown in Table 1. Patient E never become reconciled to her disease and did not cooperate, even during pregnancy. Slight progression of diabetic retinopathy occurred during pregnancy in patients B and F; both women had diabetes of 15–16 years duration with unsatisfactory glycemic control over a long period, and hyperglycemia went down too fast during pregnancy. Patient C (with treated autoimmune thyroiditis) was diagnosed with Addison’s disease at the end of pregnancy; severe hypoglycemias occurred repeatedly at very low insulin dose. The pregnancy of patient D was complicated by hypothyroidism at 14 weeks’ gestation. All the women delivered at 37–38 weeks: two vaginally and five by caesarean section. Seven children with mean weight 3370 g (2900–3700 g) were born without congenital malformations. The newborns of patients E, F and G had symptoms of considerable diabetic fetopathy, with dyspnea (two children, Apgar score 7-8-9) and hypoglycemia (one child). The other four newborns were without neonatal complications. Our patients were informed about the lack of experience in using glargine during pregnancy; they decided not to change the treatment and signed an informed consent. Rapid-acting analog insulins are more potent than regular insulin in reducing postprandial hyperglycemia in diabetic pregnant women [4]. The advantage of using the analog insulin glargine in pregnancy lies in reducing the number of nocturnal hypoglycemias, which may hinder the achievement of tight glycemic control [4]. Animal studies did not confirm a negative effect of insulin analogs (lispro, aspart and glargine) on embryo-fetal development [4, 5], and any evidence of side effects on the fetus (newborn) was not shown in studies on the use of lispro in more than 800 diabetic pregnant women [5]. The use of lispro during pregnancy has been approved. Results of a randomized trial with aspart (322 type 1 diabetic patients) were presented at the meeting of the American Diabetes Association in June 2006. Treatment with aspart was at least as safe as human insulin regarding perinatal outcome [6]. Only case reports on the use of glargine during pregnancy were found in the literature: a total of six diabetic type 1 patients were treated with glargine during the whole pregnancy [7, 8]; in five other patients it was used during [9] and in four patients after the first trimester [7, 10–11]. All the women delivered newborns without malformations. Two studies with glargine in type 1 diabetic women were presented at the ADA meeting: in a multi-center control trial with 47 patients, treatment with glargine during pregnancy appeared to be as safe and efficient as with Protaphan insulin [12]; in the other study of 115 women on glargine, no evidence of increased congenital malformations or adverse maternal or fetal outcomes was demonstrated [13]. In conclusion: the use of aspart during pregnancy has been approved in Europe from July 2006, but treatment with glargine during pregnancy is still not recommended. Further randomized trials are needed. Note: This work was supported by grant MSM 21620814 from the Ministry of Education to the Faculty of Medicine, Charles University, Prague.

Monogenic glucokinase diabetes and pregnancy: a case study

SummaryIt is estimated that up to 3 % of patients with gestational diabetes have glucokinase diabetes, termed also maturity-onset diabetes of the young type 2. The disorder has autosomal dominant inheritance. There is a 50 % risk of transmission of the gene to next generation. Two scenarios with different approach to the treatment may occur in pregnancy with glucokinase diabetes: either the fetus inherits the glucokinase mutation and the treatment of maternal hyperglycemia by insulin could increase the risk of fetal growth restriction, or the fetus is without glucokinase gene mutation and untreated hyperglycemia of the mother increases the risk of macrosomia and perinatal morbidity and insulin therapy is necessary. This article describes the outcome of two pregnancies in a patient with monogenic diabetes with glucokinase deficiency. A specific approach to the treatment is discussed.

Influence of physiological and supraphysiological hyperinsulinemia on skin microcirculation in healthy volunteers.

AIM To examine skin perfusion in dependency on insulinemia in healthy subjects. METHODS All volunteers were informed in detail about the procedures and signed informed consent. The protocol of this study was approved by the ethical committee. In our study, a two stage hyperinsulinemic euglycemic clamp was performed, with insulinemia 100 and 250 mIU/mL and glycemia 5.0 mmol/L (3% standard deviation). Before the clamp and in steady states, microcirculation was measured by laser Doppler flowmetry and transcutaneous oximetry and energy expenditure was measured by indirect calorimetry. Results (average and standard deviation) were evaluated with paired t-test. RESULTS Physiological (50 mIU/L) insulinemia led to higher perfusion in both tests; hyperemia after heating to 44%-1848% (984-2046) vs 1599% (801-1836), P < 0.05, half time of reaching peak perfusion after occlusion release 1.2 s (0.9-2.6) vs 4.9 s (1.8-11.4), P < 0.05. Supraphysiological (150 mIU/L) insulinemia led to even higher perfusion in both tests; hyperemia after heating to 44%-1937% (1177-2488) vs 1599% (801-1836), P < 0.005, half time to reach peak perfusion after occlusion release 1.0 s (0.7-1.1) vs 4.9 s (1.8-11.4), P < 0.005. A statistically significant increase occurred in tissue oxygenation in both insulinemia. The difference in perfusion and oxygenation between physiological and supraphysiological hyperinsulinemia was not statistically significant. CONCLUSION The post occlusive hyperemia test in accordance with heating test showed significantly increasing skin perfusion in the course of artificial hyperinsulinemia. This effect rises non-linearly with increasing insulinemia. Dependency on the dose was not statistically significant.