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Biomedical subjects

Carl-David Agardh

Publications and source records attributed to Carl-David Agardh.

At least 19 recordsLinked to original sources

Polymorphism in the MHC2TA gene is associated with features of the metabolic syndrome and cardiovascular mortality.

BACKGROUND: Recently, a -168A-->G polymorphism in the MHC class II transactivator gene (MHC2TA) was shown to be associated with increased susceptibility to myocardial infarction (MI). AIM: To confirm the association between the MHC2TA -168A-->G polymorphism and MI and to study its putative role for microalbuminuria, the metabolic syndrome (MetS) and cardiovascular mortality. MATERIALS AND METHODS: Using an allelic discrimination method we genotyped 11,064 individuals from three study populations: 1) 4,432 individuals from the Botnia type 2 diabetes (T2D) study, 2) 1,222 patients with MI and 2,345 control subjects participating in the Malmö Diet and Cancer study and comprising an MI case-control sample, and 3) 3,065 T2D patients from the Local Swedish Diabetes registry. RESULTS: No association between the -168A-->G polymorphism in MHC2TA and MI was observed. However, in the Botnia cohort the AG/GG genotypes were associated with cardiovascular mortality after MI (1.78 [1.09-2.92], p = 0.02). In addition, the AG/GG genotypes were more common in subjects with MetS (40.1% vs. 36.9%, p = 0.03) and in non-diabetic subjects with microalbuminuria (45.4% vs. 36.5%, p = 0.003) compared to control subjects. CONCLUSIONS: A polymorphism in MHC2TA was associated with cardiovascular mortality and predictors of cardiovascular mortality, microalbuminuria and MetS.

Adult↗

Modifying a standard method allows simultaneous extraction of RNA and protein, enabling detection of enzymes in the rat retina with low expressions and protein levels.

The aim of the study was to evaluate messenger RNA and protein expression in limited amounts of tissue with low protein content. The Chomczynski method was used for simultaneous extraction of RNA, and protein was modified in the protein isolation step. Template mass and cycling time for the complementary DNA synthesis step of real-time reverse transcription-polymerase chain reaction (RT-PCR) for analysis of catalase, copper/zinc superoxide dismutase, manganese superoxide dismutase, the catalytic subunit of glutamylcysteine ligase, glutathione peroxidase 1, and the endogenous control cyclophilin B (CypB) were optimized before PCR. Polymerase chain reaction accuracy and efficacy were demonstrated by calculating the regression (R2) values of the separate amplification curves. Appropriate antibodies, blocking buffers, and running conditions were established for Western blot, and protein detection and multiplex assays with CypB were performed for each target. During the extraction procedure, the protein phase was dissolved in a modified washing buffer containing 0.1% sodium dodecyl sulfate, followed by ultrafiltration. Enzyme expression on real-time RT-PCR was accomplished with high reliability and reproducibility (R2, 0.990-0.999), and all enzymes except for glutathione peroxidase 1 were detectable in individual retinas on Western blot. Western blot multiplexing with CypB was possible for all targets. In conclusion, connecting gene expression directly to protein levels in the individual rat retina was possible by simultaneous extraction of RNA and protein. Real-time RT-PCR and Western blot allowed accurate detection of retinal protein expressions and levels.

Animals↗

Increased serum levels of the specific advanced glycation end product methylglyoxal-derived hydroimidazolone are associated with retinopathy in patients with type 2 diabetes mellitus.

Advanced glycation end products (AGEs) are thought to play a major pathogenic role in diabetic retinopathy. The most important AGE is unknown, but as increased serum methylglyoxal-derived hydroimidazolone has been demonstrated in patients with type 2 diabetes mellitus, the aim of the present study was to elucidate possible associations between serum levels of hydroimidazolone and retinopathy in patients with type 2 diabetes mellitus. We recruited 227 patients with type 2 diabetes mellitus and retinopathy ranging from none to proliferative. Level of retinopathy was determined from 7 standard field stereo photographs per eye according to the Early Treatment Diabetic Retinopathy Study. The patients were 66 +/- 11 years old, with a known diabetes duration of 14 +/- 9 years. Serum levels of hydroimidazolone were determined with a competitive immunoassay. Serum levels of hydroimidazolone were increased in nonproliferative (median, 4.50 U/mL; interquartile range, 3.69-5.77 U/mL) and proliferative retinopathy (median, 4.88 U/mL; interquartile range, 3.70-6.52 U/mL) compared with patients without retinopathy (median, 4.02 U/mL; interquartile range, 3.47-4.88 U/mL) (P = .008 and .002, respectively). There was no association between hydroimidazolone and hemoglobin A1c (r = 0.04, P = .57). In addition, patients with proliferative retinopathy and a relatively short known duration of diabetes, that is, less than the median of 14 years, had increased serum levels of hydroimidazolone (median, 6.91 U/mL; interquartile range, 4.70-8.91 U/mL) compared with those with nonproliferative retinopathy (median, 4.34; interquartile range, 3.86-5.53U/mL, P = .015). Serum levels of hydroimidazolone are increased in type 2 diabetic patients with retinopathy. This association is independent of hitherto known associated factors, such as hemoglobin A1c.

Adult↗

Expression of antioxidant enzymes in rat retinal ischemia followed by reperfusion.

To evaluate the expression and protein levels of antioxidant enzymes in the rat retina exposed to oxidative stress induced by ischemia-reperfusion injury. Retinal ischemia was induced in female Wistar rats by ligation of the optic nerve and vessels behind the left eye bulb, and was followed by reperfusion for 0, 3, 6, or 24 hours. The right eye served as control. RNA and protein were extracted simultaneously from each retina. Expressions of the endogenous antioxidant enzymes glutathione peroxidase (GPx1), catalase (CAT), copper/zinc superoxide dismutase, manganese superoxide dismutase, and the catalytic subunit of glutamylcysteine ligase (GCLc) were analyzed with real-time reverse transcription polymerase chain reaction and related to the endogenous control cyclophilin B. Protein levels were measured with Western blot analysis. During the early phase (0 or 3 hours) of reperfusion, no changes were seen in enzyme expression. After 6 hours, GCLc expression increased by a factor of 1.14 (P = .034), followed by a decline of 0.80 after 24 hours (P = .00004), according to the comparative Ct method. After 24 hours of reperfusion, GPx1 expression increased by a factor of 1.14 (P = .028), and CAT had decreased by 0.82 (P = .022). Expressions of copper/zinc superoxide dismutase and manganese superoxide dismutase showed a tendency toward a decrease by factors of 0.86 (P = .055) and 0.88 (P = .053), respectively, after 24 hours. Protein levels did not differ for any of the antioxidants, regardless of reperfusion time. The slightly increased messenger RNA expression of GPx1 after 24 hours of reperfusion with a concomitant very modest decrease in CAT and GCLc expression and no change in protein levels indicate a very modest, if any, response to oxidative stress generated by ischemia followed by reperfusion in rat retina.

Animals↗

Colorectal cancer in middle-aged women in relation to hormonal status: a report from the Women's Health in the Lund Area (WHILA) study.

OBJECTIVE: To delineate a perceived association of estradiol versus estradiol plus norethisterone hormone therapy on the prevalence of colorectal cancer in postmenopausal women. METHODS: The Women's Health in the Lund Area (WHILA) project covers 10,766 women aged 50-60 years, living in the Lund area, Sweden. Out of this population, 6908 (64%) women completed questionnaires, underwent physical and laboratory assessments and had self-reported information regarding colorectal cancer. Four hundred and twenty-two (6%) were premenopausal (PM), 3600 (52%) were postmenopausal without hormone therapy (PM0), 2452 (36%) were postmenopausal with combined hormone therapy (PMT-HT) and 364 (5%) were postmenopausal with estrogen monotherapy (PMT-E). RESULTS: There were 21 cases of colorectal cancer (0.3%), one in the PM group, 16 in the PM0 group, two in the PMT-HT group and another two in the PMT-E group. Colorectal cancer prevalence was lower in the PMT-HT than in the PM0 group (odds ratio (OR) = 0.18, 95% confidence interval (CI) = 0.04-0.80). However, for the PMT-E group, the OR (95% CI) was 1.02 (0.86-1.20). There was a positive association between low physical activity (p = 0.04), low parity (p = 0.02) and risk of colorectal cancer. CONCLUSION: Combined hormone therapy seemed to be associated with a lower risk of colorectal cancer in postmenopausal women in contrast to estrogen monotherapy. Hence the progestin might have a protective role.

Adenocarcinoma↗

Obese women and the relation between cardiovascular risk profile and hormone therapy, glucose tolerance, and psychosocial conditions.

OBJECTIVE: To evaluate the relation between cardiovascular disease (CVD) risk factors and hormone therapy, serum hormone levels, glucose tolerance, and psychosocial and psychological conditions in subjectively healthy obese female subjects. RESEARCH DESIGN AND METHODS: The study included 606 women, aged 50-64 years, with BMI 30-40 kg/m(2) and no history of cardiovascular or other severe diseases. One group with a CVD risk profile (n = 473) (i.e., cholesterol >7.0 mmol/l, HDL cholesterol <1.2 mmol/l, triglycerides >2.0 mmol/l, systolic or diastolic blood pressure >140/90 mmHg, or waist-to-hip ratio >0.85) was compared with women without such risk (n = 133). Steroid hormones, leptin, insulin, and oral glucose tolerance tests (OGTTs) were analyzed. A subgroup of women with baseline impaired glucose tolerance (IGT) completed a 2.5-year follow-up OGTT. RESULTS: Fewer obese postmenopausal women with CVD risk had ever used hormone therapy (odds ratio 0.24 [95% CI 0.07-0.75]), after multivariate adjustments. Furthermore, women with CVD risk had a higher testosterone index (1.07 [1.01-1.13]) and more had insulin resistance (1.04 [1.00-1.08]) and IGT (2.92 [1.50-5.69]), while OGTT was similar at follow-up. No differences were observed regarding family history or lifestyle, except that fewer women with CVD risk consumed fruits, boiled vegetables, or whole-grain cereals. More women with CVD risk lived alone (3.26 [1.28-8.31]) and had more mental problems (1.16 [1.05-1.28]). CONCLUSIONS: Previously healthy obese women with a CVD risk profile seemed to have a high risk of diabetes, as well as psychosocial or psychological problems. Hormone therapy was associated with reduced CVD risk. Obesity's growing burden on society makes it more important to further target individuals that are at greatest risk of future health hazards.

Adult↗

Immunomodulation with human recombinant autoantigens.

The loss of beta cells in type 1 diabetes is the consequence of a T cell-dependent autoimmune attack. Autoantibodies against GAD65 (Mr 65.000 isoform of glutamic acid decarboxylase), IA-2 (insulinoma-associated protein IA-2) or insulin, alone or in combination, predict disease. Preclinical studies in spontaneously diabetic rodents suggest that immunomodulation with autoantigens might alter the course of autoimmune diabetes. Oral insulin reduces the development of diabetes in risk subjects with high insulin autoantibody levels. Giving alum-formulated GAD65 to patients classified with latent autoimmune diabetes of the adult (LADA) is safe and suggests possible immunomodulating effects of GAD65. Future immunomodulation trials might better ascertain subjects based on HLA genetic risk factors, the level of insulin that is still produced or by combining autoantigens with, for example, anti-CD3 antibodies, to induce antigen-specific tolerance and thereby a long-lasting protection for beta cells.

Animals↗

Albuminuria and hypertension are independently associated with circulating antipericyte autoantibodies in type 2 diabetic patients.

AIMS: To determine whether albuminuria, hypertension, or HbA 1c are independently associated with antipericyte autoantibodies (APAAs) in type 2 diabetes mellitus. METHODS: Two hundred ninety-nine subjects with different degrees of retinopathy according to the Early Treatment Diabetic Retinopathy Study Scale participated in this study. Albuminuria was defined as an albumin/creatinine ratio above the normal cutoff limit, that is, 2.0 g/mol for men and 2.8 g/mol for women. Hypertension was defined as a diastolic blood pressure more than 90 mm Hg, a systolic blood pressure more than 140 mm Hg, or pharmacological antihypertensive treatment. Serum APAAs were detected by immunofluorescence on tissue-cultured bovine retinal pericytes. Association analysis was performed using univariate and multivariate statistical tools. RESULTS: In type 2 diabetes, APAAs were independently associated with albuminuria (OR = 0.56; P < .04), hypertension (OR = 2.21; P < .01), as well as with proliferative retinopathy (OR = 0.39; P < .01). CONCLUSIONS: The increased prevalence of APAA in patients with hypertension may suggest that these antibodies are related to tissue damage and repair and that the decline in frequency with albuminuria may serve as a marker for more advanced angiopathy. Future longitudinal studies are needed to determine whether the frequency of APAA is associated with the progression of angiopathy, and to determine the biological activity and antigens recognized by the antibody.

Aged↗

Women living alone have an increased risk to develop diabetes, which is explained mainly by lifestyle factors.

OBJECTIVE: The purpose of this study was to assess the role of household conditions for the progression to diabetes in women with impaired glucose tolerance (IGT). RESEARCH DESIGN AND METHODS: A total of 461 women, aged 50-64 years, with IGT defined by an oral glucose tolerance test, had baseline advice on physical exercise, diet, smoking, and alcohol habits. Physical examination, blood tests, and questionnaires were completed at baseline and after 2.5 years. Household status was categorized into living alone or with a partner, other adults, or children. RESULTS: Women living alone had a 2.68-fold increased risk (95% CI 1.02-7.05) of developing diabetes after adjustments for biological risk factors. Further stepwise adjustments for education, occupation, subjective mental health, exercise, diet, and alcohol showed remaining significant odds ratios (ORs), decreasing from 3.26 (1.19-8.96) to 3.03 (1.02-8.99). However, when smoking status was added, the OR became nonsignificant, 2.07 (0.62-6.88). More women who lived alone smoked and did not reduce their daily cigarette consumption compared with women in other household conditions. At follow-up, women living alone had reduced their alcohol consumption and were more often abstainers and fewer had healthy dietary habits or had improved their diet. Physical exercise did not differ among the groups. Separate analyses of any other household status did not show any excess risk for development of diabetes. CONCLUSIONS: Women living alone had a higher risk to progress from IGT to diabetes, mostly explained by smoking, alcohol, and dietary habits. Household conditions should be accounted for when assessing future risk for diabetes.

Alcohol Drinking↗

Does hormone therapy increase allergic reactions and upper gastrointestinal problems? Results from a population-based study of Swedish woman. The women's health in the Lund area (WHILA) study.

OBJECTIVES: To delineate the use of various drugs particularly pertaining to allergy and upper gastrointestinal problems in relation to hormone status in middle aged women. METHODS: An analysis from a population-based study on women born between 1935 and 1945 and lived in the Lund area southern Sweden. Of 10,766 women, 6,917 provided complete data sets; in turn 5,673 were assessed for the use of medication in this study. Among the cohort, 9% of women were premenopausal (PM), 54% were postmenopausal without hormone replacement therapy (PM0) and 37% were current hormone replacement therapy users (PMT). RESULTS: There were 7 (1.3%) women in PM, 11 (0.4%) in PM0 and 21 (1.0%) in PMT group who used loratadine regularly. There was a significant difference between the PM and PM0 groups and also between the PM0 and PMT groups in the use of loratadine (P < 0.05 ). Among 21 loratadine users in PMT group 4 (19%) used transdermal patches and 17 (81%) used oral HRT. The result for omeprazole use was as follows: 4 (0.8%) of PM group, 39 (1.3%) of PM0 group and 42 (2.0%) of PMT group. The use of omeprazole was significantly higher in the PMT group than in the PM (P = 0.05 ) and PM0 group (P < 0.05 ). There was no relation between the use of omeprazole and smoking or alcohol consumption. CONCLUSIONS: Use of hormone replacement therapy seems to be related to a higher frequency of omeprazole and loratadine use, which implies that hormone replacement therapy, may be associated with more upper gastrointestinal symptoms as well as allergy.

Aged↗

Markers of inflammation in patients with coronary artery disease are also associated with glycosylated haemoglobin A1c within the normal range.

AIMS: Diabetes is a risk factor for atherosclerosis and low-degree inflammation may play a central role in both diseases. Glycosylated haemoglobin A(1c) (HbA(1c)) is an established measure of long-term glycaemic control but data on its correlation with markers of inflammation are limited, especially in patients with atherosclerotic manifestations. The aim of the present study was thus to investigate the associations between HbA(1c) and a panel of inflammation-sensitive parameters in patients with and without diabetes. METHODS AND RESULTS: This single centre cross-sectional study comprised 314 consecutive subjects who underwent coronary angioplasty for stable coronary artery disease. Sixty-six patients had diabetes mellitus. Haemoglobin A(1c) and markers of inflammation, i.e., plasma levels of CRP, fibrinogen, and albumin, erythrocyte sedimentation rate and white blood cell count were measured. All inflammation markers were altered in a more inflammatory direction in diabetic patients. Furthermore, when non-diabetic patients with HbA(1c) levels within the normal range were studied separately, all inflammation-sensitive parameters except albumin correlated significantly with HbA(1c). CONCLUSION: In subjects with known coronary atherosclerosis, low-degree inflammatory activity is not only increased in diabetic patients, but also increased with increasing HbA(1c) in non-diabetic individuals with HbA(1c) within the normal range, i.e., at a pre-diabetic level of glucose metabolism derangement.

Aged↗

A prospective study of different types of hormone replacement therapy use and the risk of subsequent breast cancer: the women's health in the Lund area (WHILA) study (Sweden).

OBJECTIVES: Reports suggest that combined estrogen plus progestin hormone replacement therapy (HRT) confers a higher breast cancer risk than estrogen alone. We aimed to establish whether breast cancer risk depends on the type of HRT formula. METHODS: The cohort consisted of 6586 women, aged 50-64 years, from the Lund area, Sweden, with no reported breast cancer upon inclusion. We obtained information such as HRT use through a questionnaire between December 1995 and February 2000. New breast cancers were identified through the South Swedish tumor registry. RESULTS: Between inclusion and December 2001, 101 women developed breast cancer. Only ever use of the continuous combined estrogen plus progestin (CCEP) formula differed between cases and controls (45.2% versus 23.5%; p = 0.000001). Compared with never users, exclusive CCEP users had the highest age-adjusted hazard ratio HR 3.3 (95% CI: 1.9-5.6; p < 0.001), followed by users of CCEP in addition to other HRT formulas HR 2.8 (95% CI: 1.4-5.5; p = 0.003). No significant increase was seen in women who exclusively used other HRT formulas. CONCLUSION: Women who used CCEP had over three times the risk of developing breast cancer compared with never users and twice the risk compared with users of other types of HRT.

Breast Neoplasms↗

Type 1 diabetes patients with severe non-proliferative retinopathy may benefit from panretinal photocoagulation.

PURPOSE: To examine whether panretinal photocoagulation for severe non-proliferative retinopathy in type 1 diabetes patients could halt the progression of retinopathy with subsequent vitreous haemorrhages and visual impairment. METHODS: During a 10-year follow-up study period of 344 type 1 diabetes patients, 81 subjects went through panretinal photocoagulation. Forty patients were treated for severe non-proliferative retinopathy (age at onset of diabetes 14 +/- 8 years, diabetes duration 18 + 10 years) and 41 for proliferative retinopathy (age at onset 15 +/- 10 years, diabetes duration 22 + 13 years). One randomly selected eye per patient forms the basis for the study. Metabolic control, systolic and diastolic blood pressure, serum creatinine and urinary albumin levels were measured and analysed yearly during the follow-up period. RESULTS: A total of 35% (14/40) of eyes treated for severe non-proliferative retinopathy developed neovascularizations during a mean time of 2.9 +/- 1.5 years. Vitreous haemorrhages were more frequent in eyes with proliferative retinopathy at treatment than in eyes with severe non-proliferative retinopathy (12/41 versus 2/40; p = 0.007). The number of vitrectomies due to vitreous haemorrhages in eyes treated for severe non-proliferative retinopathy tended to be lower (1/40 versus 6/41; p = 0.052). Before photocoagulation, visual acuity (VA) was similar in eyes with severe non-proliferative retinopathy and in those with proliferative retinopathy (1.0, 0.4-1.0 versus 1.0, 0.1-1.0; median and range). Visual impairment and blindness tended to develop more often in eyes treated for proliferative retinopathy compared to those treated for severe non-proliferative retinopathy (10/40 versus 4/40; p = 0.056). Eyes with neovascularizations at follow-up were more often visually impaired (VA < 0.5) than eyes without neovascularizations (15/55 versus 1/26; p = 0.016). CONCLUSION: In type 1 diabetes, panretinal photocoagulation may be beneficial even at the severe non-proliferative retinopathy stage in terms of preventing vitreous haemorrhage, subsequent vitrectomy and visual impairment.

Adolescent↗

Diabetes duration may modify the association between genetic variation in the glycoprotein Ia subunit of the platelet collagen receptor and risk of severe diabetic retinopathy: a working hypothesis.

Genetic factors appear to contribute to the severity and progression of diabetic retinopathy. We assessed the associations of the C807T and Glu505Lys variants of the glycoprotein Ia (alpha(2) integrin) subunit of the platelet/endothelial collagen receptor and risk of retinopathy in a population-based survey of 288 diabetic patients in one Swedish community. Neither variant was associated with retinopathy risk overall. However, the 807T variant was associated with increased risk of severe retinopathy, and the association was modified by diabetes duration. Among patients with diabetes of longer duration (>/=25 years), the 807T variant was strongly associated with risk of severe retinopathy (odds ratio 7.49, 95% confidence interval 1.75 to 32.1). There was no association between the 807T variant and risk of severe retinopathy among patients with diabetes duration <25 years. The Lys505 variant of glycoprotein Ia was associated with an odds ratio for severe retinopathy of 1.88 (95% confidence interval 0.83 to 4.24). Overall, there was a significant interaction between glycoprotein Ia genotype and duration of diabetes on the risk of retinopathy (P-value for interaction = 0.019). These results suggest the hypothesis that genetic variation of platelet glycoprotein Ia may play a particularly important role during the advanced stages of diabetic retinopathy.

Adult↗

A small number of older type 2 diabetic patients end up visually impaired despite regular photographic screening and laser treatment for diabetic retinopathy.

PURPOSE: The present study describes the prevalence of visual impairment and blindness in a geographically defined population 8 years after the introduction of a screening programme in 1987 for early detection of sight-threatening diabetic retinopathy. METHODS: Of 374 patients with diabetes, comprising 2.6% of the population in the study community, 72% were examined with fundus photography or biomicroscopy during 1994-95. These patients form the basis of this study. The screening programme was fulfilled by 93% of subjects, all of whom underwent ophthalmic examinations at least every other year. A total of 79 eyes in 52 patients received photocoagulation for macular oedema alone or in combination with severe non-proliferative or proliferative retinopathy. RESULTS: Eight years after the implementation of the programme, only three patients, all with type 2 diabetes (diabetes diagnosed at or after 30 years of age), had visual acuity < or = 0.1. The total number of eyes with visual acuity < or = 0.5 was higher in insulin-treated type 2 diabetic patients (n = 20) than in those on oral treatment (n = 5) or diet treatment only (n = 1) (p = 0.006 in both cases). The only independent risk factor for visual impairment in eyes with sight-threatening retinopathy was age. CONCLUSION: A small number of older type 2 diabetic patients end up with visual impairment due to unsuccessful photocoagulation of macular oedema.

Adolescent↗

The influence of hormonal status and features of the metabolic syndrome on bone density: a population-based study of Swedish women aged 50 to 59 years. The women's health in the Lund area study.

This study investigated whether there is an association between bone density and features of the metabolic syndrome in relation to hormonal status. All women aged 50 to 59 years living in a defined geographic area in Sweden were offered a health assessment program including blood glucose, lipid profile, blood pressure, and bone densitometry. Women were divided into 3 groups according to their hormonal status: premenopausal (PM), postmenopausal with hormone replacement therapy (PMT), and postmenopausal without hormone replacement therapy (PM0). Of the 6,886 women investigated, 7% were PM, 41% PMT, and 52% PM0. The overall prevalence of osteopenia and osteoporosis, according to the World Health Organization (WHO) definition, was 42.6% and 6.6%, respectively. T-score in the PM group was higher than in the PMT (P <.05) and PM0 groups (P <.001) and higher in the PMT group compared with the PM0 group (P <.001). Also, in the total cohort, the bone density was positively associated with body weight, body mass index (BMI), waist-to-hip ratio (WHR), systolic blood pressure (SBP), diastolic blood pressure (DBP), serum triglycerides, and blood glucose (P <.001 for all) and negatively associated with serum levels of cholesterol (P <.05) and high-density lipoprotein (HDL) (P <.001). This was most evident among the PMO women, suggesting that the influence of metabolic factors on bone density increases when the levels of hormones decrease. This indicates that hormone replacement therapy maintains treated women in a premenopausal status concerning the metabolic factors.

Absorptiometry, Photon↗

Application of quantitative competitive polymerase chain reaction for measurements of mRNA from antioxidative enzymes in the diabetic rat retina and kidney.

This present study applied quantitative competitive polymerase chain reaction (QC-PCR) in the analyses of mRNA expression of the endogenous antioxidative enzymes CuZn superoxide dismutase (SOD), MnSOD, catalase, and glutathione peroxidase in tissue samples from the retina and kidney cortex of diabetic rats. RNA was extracted from snap-frozen retinas and pieces of the kidney cortex of male Wistar rats with streptozotocin (STZ)-induced diabetes and control rats. The mRNA levels were analyzed using QC-PCR. The animals were kept in the laboratory for 1 and 6 months, respectively, and fed a normal or probucol- (1% wt/wt) enriched diet. By using QC-PCR, relative mRNA levels of all antioxidative enzymes could be estimated in the retina and kidney cortex. In the retina, the relative catalase mRNA concentration was about 1/10 that of the other enzymes. After 6 months of diabetes, there was a 100% increase of the catalase (median, 0.012 [range, 0.008 to 0.017] v 0.006 [0.005 to 0.010]; P =.011) and a 50% increase of the glutathione peroxidase mRNA levels (0.88 [0.44 to 1.12] v 0.52 [0.31 to 0.79]; P =.044). In the kidney cortex, the relative glutathione peroxidase mRNA level was 10 to 15 times higher, and catalase mRNA level about half of those of CuZnSOD and MnSOD. After 1 month of diabetes, there was an increase only of the glutathione peroxidase mRNA levels, by 170% (17.59 [6.19 to 29.49] v 6.96 [2.34 to 9.04]; P =.047). We conclude that quantification of mRNA can provide difficulties when the amount of sample RNA is limited and/or the gene expression is low. The present study shows QC-PCR to be useful as a tool for measuring expression of mRNA not only in the kidney cortex but also in small tissue samples like the retina. Our results indicate modestly increased mRNA expression of catalase and glutathione peroxidase in the retina and likewise modestly increased mRNA expression of glutathione peroxidase in the kidney cortex of rats with STZ-induced diabetes. Extended studies, also including enzyme activities, are needed before any effect of hyperglycemia on the overall enzyme activity can be established.

Animals↗