Identification of Silent Myocardial Ischemia in Patients with Long-Term Type 1 and Type 2 Diabetes.

(1) Background: This study aimed to analyze epidemiological data to identify risk factors for silent myocardial ischemia in patients with long-term type 1 and type 2 diabetes. (2) Methods: An analysis was performed on 104 patients with long-term type 1 and type 2 diabetes who had not previously been diagnosed with cardiovascular disease. During hospitalization, patients were subjected to a standard ECG exercise test on a treadmill. If the test could not be performed or the result was uncertain, a pharmacological exercise test with dobutamine was performed. In the case of a positive exercise ECG test or a positive dobutamine test, the patient underwent coronary angiography. (3) Results: Atherosclerotic lesions were found in 24 patients. Patients with silent ischemia were significantly older and had a lower mean left ventricular ejection fraction and a higher incidence of carotid atherosclerosis. The presence of microvascular complications did not increase the risk of silent ischemia. (4) Conclusions: Silent heart ischemia is more common in type 2 than type 1 diabetes. Predisposing factors include older age, coexistence of carotid atherosclerosis, lower left ventricular ejection fraction, and smoking in patients with type 1 diabetes. Concomitant microvascular complications are not a risk factor.

Energy reserves, oxidative stress and development traits of Spodoptera exigua Hübner individuals from cadmium strain.

Cadmium as a common environmental stressor may exert highly toxic effects on herbivorous insects. The question was whether possible elevation of an oxidative stress and imbalance of energetic reserves in insects may depend on developmental stage, sex and insect population's multigenerational history of exposure to cadmium. So, the aim of this study was to compare of the development traits, total antioxidant capacity, lipid peroxidation, RSSR to RSH ratio and the concentration of carbohydrates, glycogen, lipids and proteins in whole individuals (larvae or pupae) of Spodoptera exigua originating from two strains: control and selected over 120 generations with sublethal metal concentration (44 Cd mg per dry weight of diet). Generally, the increase of the protein, carbohydrates, glycogen concentration and lipid peroxidation decrease with age of the larvae were found. Revealed cases of a higher mobilisation of carbohydrates and proteins, and changes in total antioxidant capacity or lipid peroxidation, in individuals being under metal exposure, occurred in strain-depended mode. Short-term Cd exposure effect was connected with possible higher engagement of proteins and glycogen in detoxification processes, but also higher concentration of lipid peroxidation. In turn, for long-term Cd exposure effect lower lipids concentration and higher thiols usage seemed to be more specific.

Protective role of zinc in Spodoptera exigua larvae under 135-generational cadmium exposure.

The aim of this study was to investigate whether zinc supplementation modulates cadmium toxicity in the beet armyworm Spodoptera exigua selected for 135 generations towards cadmium tolerance. To achieve this, larvae originating from three laboratory populations of S. exigua (control strain - C; cadmium-intoxicated for 135 generations strain - Cd, and control strain intoxicated with Cd for 1 generation - CCd) were additionally exposed to zinc in three concentrations (Zn1, 400 µg Zn·g-1 dry mass of food; Zn2; 200 µg Zn·g-1 dry mass of food; Zn3, 100 µg Zn·g-1 dry mass of food). As the markers of toxicity, a life history traits (the duration of L4 and L5 stages), cellular (DNA damage indices) and biochemical parameters (ADP/ATP ratio and ATP and HSP70 concentrations) were chosen. The duration of larval stages of Zn supplemented larvae was prolonged, while cellular and biochemical indicators, in general, appeared to be lower in comparison to the insects from respective reference groups in each laboratory populations. Moreover, the range of the differences depended on zinc concentration in food. We can suspect that zinc supplementation contributed to the protection of S. exigua individuals against negative effects of cadmium intoxication, probably at the cost of growth rate. Significant differences in the response pattern between insects from different laboratory populations indicate that the influence of additional stress factors is dependent on the overall condition of animals and their previous adaptation to other stressors.

Reproduction and development of Spodoptera exigua from cadmium and control strains under differentiated cadmium stress.

The growth and development of living organisms is programmed in genes, but exogenous factors (e.g. cadmium) may modulate endogenous information. Heavy metals may disturb physiological functions and accumulate in the tissues. The insects under prolonged heavy metal stress show some modifications in their metabolism management. The aim of this study was to compare the reproduction and development between individuals of S. exigua from the strain, exposed over 130 generations to sublethal concentration of cadmium (44 mg Cd/kg dry weight of larval diet), and the individuals from the control strain, both additionally exposed to different concentration of cadmium (22-704 mg Cd/kg dry weight of larval diet). The exposure to various cadmium concentrations in the diet revealed survival difference between the cadmium and the control animals at the larvae stage. The differences between adults were not evident. The telomere length (responsible for the duration of a lifespan) in the cadmium strain was shorter in the females than in the males and the individuals from the control strain. TERF1 gene expression (indirectly responsible for the telomere length) was higher in the individuals from the cadmium strain 24 hrs after eclosion. The significant reduction in the larvae body mass was observed in both strains, when the metal concentration was equal to or higher than 264 mg/kg dry weight of larval diet. The EC50 values (defined as of body mass loss), calculated 48 hours after cadmium exposure of individuals from control and cadmium strains, were respectively 632 and 725 mg Cd/kg dry weight of diet. However, some difference in reproduction (the total number of eggs laid and the oviposition time) between the strains appeared only in the groups fed on the uncontaminated diet. The control females laid almost two times more eggs than those from the cadmium strain, and the control ones had more than two times longer oviposition time than the females from the cadmium strain. The fluctuation was also noted in the size of eggs and the hatching success on the following days when both strains were compared, while the hatching success was higher for the insects from the cadmium strain. In conclusion, the insects from the cadmium strain are more resistant to cadmium contamination, as it is evidenced by the EC50 parameter. However, the females from the cadmium strain start laying eggs statistically later, have shorter telomeres and slightly reduced TERF1 gene expression, but hutching success in the strain is significantly higher when compared with the control individuals.

Molecular changes in vitellogenin gene of Spodoptera exigua after long-time exposure to cadmium - Toxic side effect or microevolution?

The reproduction of pest insects is a continuously ongoing issue, especially in the environmental pollution context. Natural or artificial stressing factors enforce a kind of trade-off, most often between growth/survival and reproduction, which improves fitness of the organism. Harmful factors, such as cadmium, can affect the vitellogenesis leading to reduction of yolk synthesis and egg production. The aim of this study was to assess whether 130-generational selection to cadmium in food might have induced change in vitellogenesis of Spodoptera exigua. We analyzed the level of Vg gene expression in S. exigua from the control and the cadmium strain at regular time intervals within 48h after eclosion. The full sequence of Vg gene was also compared between strains. The vitellogenin gene expression in both strains was time-dependent. This dependence was more visible in the control strain. In the cadmium strain the vitellogenin expression was significantly lower, comparing with the control strain in the first day after eclosion but increased significantly in the second day. The sequenced CDS (5286bp long) of the control and the cadmium strains were translated into protein sequences containing both 1761 aa. The protein sequences comparison revealed that there is one amino acid change at aa position 1282. Multiple alignments of six orthologous proteins from different species showed that amino acid change is located in the conserved position. Long-lasting exposure to cadmium resulted in permanent mutation in vitellogenin gene. We do not know yet if the mutation can improve fitness of the cadmium-selected insects. However, we can suppose that the mutation is neutral or even beneficial. The mutation and most probably additional effects of cadmium exposure have an influence on the vitellogenin expression. Some modification in the expression of the vitellogenin receptor are also likely to be important.

Cross tolerance in beet armyworm: long-term selection by cadmium broadens tolerance to other stressors.

Long lasting exposure of animals to stressing factor may lead to the selection of population able to cope with the stressor at lower cost than unexposed individuals. The aim of this study was to assess whether 130-generational selection of a beet armyworm to cadmium in food might have induced tolerance also to other stressors. The potential tolerance was assessed by means of unspecific stress markers: HSP70 concentration, DNA damage level, and energy budget indices in L5 larval instars of beet armyworm. The animals originated from Cd-exposed and control strains exposed additionally in a short-term experiment to high/low temperature or pesticide-spinosad. The application of the additional stressors caused, in general, an increase in the levels of studied parameters, in a strain-dependent manner. The most significant increase was found in HSP70 level in the individuals from the Cd-strain exposed to various spinosad concentration. Therefore, multigenerational contact with cadmium caused several changes that enable the insect to survive under a chronic stress, preparing the organism to the contact with an additional, new stressor. This relationship may be described as a sort of cross tolerance. This may, possibly, increase the probability of population survivorship and, at the same time, decrease the efficiency of pesticide-based plant protection efforts.

Evaluation of Candidate Reference Genes for Quantitative Gene Expression Analysis in Spodoptera exigu a after Long-time Exposure to Cadmium.

Studies on the transcriptional control of gene expression play an important role in many areas of biology. Reference genes, which are often referred to as housekeeping genes, such as GAPDH, G3PDH, EF2, RpL7A, RpL10, TUBα and Actin, have traditionally been assumed to be stably expressed in all conditions, and they are frequently used to normalize mRNA levels between different samples in qPCR analysis. However, it is known that the expression of these genes is influenced by numerous factors, such as experimental conditions. The difference in gene expression underlies a range of biological processes, including development, reproduction and behavior. The aim of this study was to show the problems associated with using reference genes in the qPCR technique, in a study on inbred strains of Spodoptera exigua selected toward cadmium resistance. We present and discuss our results and observations, and give some recommendations concerning the use and limitations of housekeeping genes as internal standards, especially in research on insects. Our results suggest that holometabolism and poikilothermia, as well as time since metamorphosis and the level of exposure to the selective factor (cadmium in this case), have a significant effect on the expression of reference genes.

Phenotypic Plasticity, Epigenetic or Genetic Modifications in Relation to the Duration of Cd-Exposure within a Microevolution Time Range in the Beet Armyworm.

In the case of the pests inhabiting metal polluted or fields where the use of pesticides is common, a natural selection of resistant individuals can occur. This may pose serious problems for humans, agriculture, as well as the economies of many countries. In this study, the hypothesis that multigenerational (120 generations) exposure to cadmium of a beet armyworm population could be a selecting factor toward a more efficient DNA protection was verified. The hemocytes of individuals from two culture strains (control and Cd-exposed) were treated with H2O2 (a DNA-damaging agent) or PBS (reference). The level of DNA damage was assessed using the Comet assay immediately and 5, 15 and 30 min. after the treatment. The immediate result of the contact with H2O2 was that the level of DNA damage in the hemocytes of the insects from both strains increased significantly. However, in the cells of the Cd-exposed individuals, the level of DNA damage decreased over time, while in the cells from the control insects it remained at the same level with no evidence of repair. These results suggest that efficient defense mechanisms may exist in the cells of insects that have prolonged contact with cadmium. Some evolutionary and trade-off aspects of the phenomenon are discussed. In a wider context, comparing the results obtained in the laboratory with field studies may be beneficial for understanding basic mechanisms of the resistance of an organism. To summarize, the high potential for the repair of DNA damage that was observed in the insects from the cadmium strain may confirm the hypothesis that multigenerational exposure to that metal may possibly contribute to the selection of insects that have a wider tolerance to oxidative stress. However, our investigations of polymorphism using AFLP did not reveal differences between the two main insect strains.

Effect of intravenous versus subcutaneous insulin delivery on the intensity of neuropathic pain in diabetic subjects.

INTRODUCTION: The effectiveness of treatment of painful diabetic polyneuropathy remains unsatisfactory. The aim of this study was to compare effects of intravenous vs. subcutaneous insulin delivery in patients with diabetic symmetric sensorimotor polyneuropathy on pain relief, the quality of life, sleep disturbance, and the nerve conduction. MATERIAL AND METHODS: Thirty-four patients with diabetic polyneuropathy (mean age 62 ± 10 years, duration 17 ± 10 years), who reached a pain score over 40 mm on the VAS scale, HbA(1c) 7.5-10%, were randomly assigned to continuous intravenous insulin infusion (examined group) and multiple injections (control subjects). Before and after five days of the insulin treatment the effects on pain relief (SFMPQ-VAS), the quality of life improvement (EuroQol EQ-5D), and sleep disturbances (AIS) were assessed. RESULTS: Both groups experienced significant pain reduction, improvement of the quality of life, and reduction of sleep disturbances, i.e. a VAS in the study group of 69 ± 14 mm before treatment vs. 40 ± 19 mm after treatment (p < 0.001), and in control subjects 66 ± 16 mm vs. 47 ± 17 mm (p < 0.001). No difference in level of pain intensity reduction between the groups studied was found. CONCLUSIONS: Intensification of insulin treatment applied for five days results in improvement of the physical condition of patients with painful diabetic polyneuropathy, through pain relief, and improvement of the quality of life and sleep quality. The efficacy of insulin intravenous infusion and multiple injections is comparable.