Cholesterol lowering treatment restores blood global DNA methylation in chronic kidney disease (CKD) patients.

BACKGROUND AND AIMS: Chronic kidney disease (CKD) is characterized by increased oxidative stress (OS). In consideration of the well-known link between OS and DNA methylation we assessed DNA methylcytosine (mCyt) concentrations in CKD patients at baseline and during cholesterol lowering treatment. METHODS AND RESULTS: DNA methylation and OS indices (malonyldialdehyde, MDA; allantoin/uric acid ratio, All/UA) were measured in 30 CKD patients randomized to three cholesterol lowering regimens for 12 months (simvastatin 40 mg/day, ezetimibe/simvastatin 10/20 mg/day, or ezetimibe/simvastatin 10/40 mg/day) and 30 age- and sex-matched healthy controls. DNA methylation was significantly lower in CKD patients vs. controls (4.06 ± 0.20% vs. 4.27 ± 0.17% mCyt, p = 0.0001). Treatment significantly increased mCyt DNA concentrations in all patients (4.06 ± 0.04% at baseline; 4.12 ± 0.03% at 4 months; 4.17 ± 0.03% at 8 months; and 4.20 ± 0.02% at 12 months, p = 0.0001 for trend). A trend for a greater effect on DNA methylation was observed with combined treatment ezetimibe/simvastatin 10/40 mg/day (+5.2% after one year treatment). The treatment-associated mCyt increase was significantly correlated with the concomitant reduction in MDA concentrations and All/AU ratios. CONCLUSION: Our results demonstrate that CKD patients have a lower degree of DNA methylation and that cholesterol lowering treatment restores mCyt DNA concentrations to levels similar to healthy controls. The treatment-associated increase in DNA methylation is correlated with a concomitant reduction in OS markers. The study was registered at clinicaltrials.gov (NCT00861731).

Impact of cholesterol lowering treatment on plasma kynurenine and tryptophan concentrations in chronic kidney disease: relationship with oxidative stress improvement.

BACKGROUND AND AIM: Tryptophan (Trp) degradation via indoleamine (2,3)-dioxygenase (IDO), with consequent increased in kynurenine (Kyn) concentrations, has been proposed as marker of immune system activation. Oxidative stress (OS) might contribute to the pro-inflammatory state in chronic kidney disease (CKD) through the activation of NF-kB, with consequent activation and recruitment of immune cells. METHODS AND RESULTS: Serum concentrations of Trp and Kyn, oxidative stress indices malondialdehyde (MDA) and allantoin/uric acid (All/UA) ratio and anti-oxidant amino acid taurine were measured in 30 CKD patients randomized to 40 mg/day simvastatin (group 1), ezetimibe/simvastatin 10/20 mg/day (group 2) or ezetimibe/simvastatin 10/40 mg/day (group 3) and treated for 12 months. Baseline Kyn and Kyn/Trp ratio were higher in CKD patients vs. healthy controls (1.67 ± 0.62 µmol/L vs 1.25 ± 0.40 µmol/L, p < 0.01 and 0.036 ± 0.016 vs 0.023 ± 0.010, p < 0.001 respectively). Both Kyn and Kyn/Trp ratio significantly decreased after cholesterol lowering treatment, to values comparable with healthy controls after one year treatment (1.67 ± 0.62 µmol/L vs 1.31 ± 0.51 µmol/L, p < 0.0001 and 0.036 ± 0.016 vs 0.028 ± 0.012 p < 0.0001, respectively). This was paralleled by a significant decrease in MDA (218 ± 143 nmol/L vs 176 ± 123 nmol/L, p < 0.01) and All/UA ratio (1.47 ± 0.72 vs 1.19 ± 0.51, p < 0.01) in CKD patients. CONCLUSIONS: Amelioration of both oxidative and inflammation status after cholesterol lowering treatment in CKD might be mediated by restoration of antioxidant taurine concentrations during therapy (from 51.1 ± 13.3 µmol/L at baseline to 63.1 ± 16.4 µmol/L, p < 0.001 by ANOVA), suggesting that improvement of both oxidative and inflammation status in CKD patients could be explained, at least partly, by the cholesterol lowering effects.

High dietary taurine inhibits myocardial apoptosis during an atherogenic diet: association with increased myocardial HSP70 and HSF-1 but not caspase 3.

BACKGROUND AND AIM: Apoptosis is a major cause of myocyte death, and taurine is anti-apoptotic. Heat shock protein 70 (HSP70) (which is regulated by heat shock factor-HSF-1) is also anti-apoptotic, and caspase 3 stimulates the apoptotic pathway. This study investigated whether taurine affects atherogenic diet-induced myocardial apoptosis, and whether HSP70, HSF-1 and caspase 3 are involved. METHODS: New Zealand white rabbits were divided into 3 groups for 4 weeks according to their diet. Group 1 (control) was fed a normal rabbit diet; Group 2 (MC) received a normal rabbit diet with 1% methionine plus 0.5% cholesterol. Group 3 received MC diet + 2.5% taurine (MCT). RESULTS: The atherogenic diet did not affect myocardial HSP70 or HSF-1 protein, but increased myocardial apoptotic nuclei to 40% (p < 0.01) versus 7% in con and 12% in MCT (p < 0.01). However, in MCT, myocardial HSP70 expression increased by 42.7% versus con and MC (p = 0.016), HSF-1 by 12% versus con and MC (p < 0.05), and total nuclei count increased by 37% versus MC (p < 0.05). Caspase 3 subunits remained unchanged in all groups, and HSP70 was increased approximately twofold in endothelial layer of arterioles (p = 0.01). CONCLUSION: This study shows that taurine could reduce myocardial apoptotic nuclei and thus confer myocardial cytoprotection via stimulating myocardial HSP70 via HSF-1 and caspase 3-independent mechanisms.

A meta-regression study of the clinical significance of serum aminotransferases in COVID-19.

OBJECTIVE: The aim of the study was to appraise the capacity of serum aminotransferases to discriminate between hepatic and other extra-pulmonary COVID-19-related manifestations and, potentially, to serve as predictors of poor clinical outcomes. MATERIALS AND METHODS: Ninety-eight studies were identified (79% from China), including 43,554 patients (57% males), 9,983 (62% males) with poor outcomes and 33,571 (50% males) with favorable outcomes. After splitting studies depending on whether serum alanine aminotransferase (ALT) concentrations were statistically different between patients with poor vs. favorable outcomes, the 35 'hepatic involvement' articles (p<0.05) included 28,510 patients (51% males), 5,279 (66% males) and 23,231 subjects (48% males) with poor and favorable outcomes, respectively. The 63 'extra-hepatic involvement' studies (p>0.05) included 15,044 patients (54% males), 4,704 (60% males) with poor outcomes and 10,340 (51% males) with favorable outcomes. RESULTS: The meta-analysis shows that serum aspartate aminotransferase (AST) concentrations were significantly higher in patients with poor outcomes than those with favorable outcomes (WMD 12.5 UI/L, 95% CI 10.9 to 14.1 p<0.001). Similarly, AST concentrations were significantly higher in the 'hepatic involvement' studies (WMD 16.3 UI/L, 95% CI 13.4 to 19.2 p<0.001) and in the 'extra-hepatic involvement' studies (WMD 10.3 UI/L, 95% CI 8.6 to 12.0 p<0.001). CONCLUSIONS: The different association of serum AST concentrations with some clinical, demographic, and biochemical factors in the two clusters suggests that in COVID-19 patients, serum AST elevation is not necessarily linked to real liver damage.

Diagnostic Accuracy of Anticarbamylated Protein Antibodies in Established Rheumatoid Arthritis: A Monocentric Cross-Sectional Study.

OBJECTIVE: To evaluate the diagnostic accuracy of anticarbamylated protein antibodies (CarP), alone and in combination with traditional biomarkers (rheumatoid factor [RF] and anticitrullinated peptide antibodies [ACPA]), in established rheumatoid arthritis (RA). METHODS: A commercially available enzyme-linked immunosorbent assay (ELISA) kit was used to assess CarP concentrations in serum samples of 200 established RA and 206 controls (115 healthy donors and 55 patients with other rheumatic diseases). Main outcome measures were sensitivity, specificity, and area under the curve (AUC; 95% confidence interval [CI]). Difference in accuracy was evaluated by comparison of the respective AUCs. RESULTS: A serum CarP cut-off of 1.47 ng/ml or more differentiated patients with RA from controls with 30% sensitivity, 97.1% specificity, and good accuracy (AUC[95%CI] = 0.83[0.79-0.86], P < 0.0001). However, it showed moderate diagnostic accuracy in seronegative RA patients: sensitivity 17.9%, specificity 96.9%, and AUC (95% CI) = 0.69 (0.63-0.75). The diagnostic accuracy of CarP_ACPA and CarP_RF combinations was significantly superior to that of ACPA and RF alone (P < 0.0001 and P = 0.015, respectively), but not to that of ACPA_RF combination (P = 0.089) In addition, the CarP_ACPA_RF combination did not improve the diagnostic accuracy of the ACPA_RF combination (AUC mean difference [95% CI] = 0.006 [-0.001 to 0.015], P = 0.10). The number of positive autoantibodies (0, 1, 2, or 3) was not significantly associated with moderate-severe disease (Disease Activity Score-28 [DAS-28] > 3.2) in adjusted multiple regression analysis. CONCLUSION: CarP has good diagnostic accuracy in established RA but not in seronegative RA. The addition of CarP to ACPA and RF alone or in combination does not significantly enhance the diagnostic accuracy of ACPA_RF combination.

High-throughput capillary electrophoresis method for plasma cysteinylglycine measurement: evidences for a clinical application.

Increased levels in plasma homocysteine and cysteine, and more recently, decreased levels in cysteinylglycine have been indicated as a risk factor for vascular diseases. Most assays focused their attention only on homocysteine determination and when also other thiols were measured, analytical times drastically increased. By modifying our previous method for thiols detection, we set up a rapid capillary electrophoresis method for the selective quantification of plasma cysteinylglycine, cutting the analysis time of about 50%. Samples were treated with tri-n-butylphosphine as reducing agent, proteins were precipitated with trichloroacetic acid and released thiols were successively derivatized by the selective thiol laser-induced fluorescence-labeling agent 5-iodoacetamidofluorescein and separated by capillary electrophoresis. A baseline separation between peaks was obtained in about 2 min using 3 mmol/L sodium phosphate/2.5 mmol/L boric acid as electrolyte solution with 75 mmol/L N-methyl-D-glucamine at pH 11.25 in a 47 cm long capillary with a cartridge temperature of 45 degrees C. The method application was checked by measuring plasma Cys-Gly levels in a group of patients affected by retinal vein occlusion (RVO), an important cause of visual loss in the elderly. The low levels of Cys-Gly found in the RVO patients suggest that these small thiols may have importance in the disease development.

Sex Differences in the Associations between L-Arginine Pathway Metabolites, Skeletal Muscle Mass and Function, and their Responses to Resistance Exercise, in Old Age.

OBJECTIVES: The current study was designed to explore the associations between L-arginine metabolites and muscle mass and function in old age, which are largely unknown. DESIGN: The study used a randomised, double-blind, placebo-controlled design. SETTING: The study was carried out in a laboratory setting. PARTICIPANTS: 50 healthy older adults [median age 70 years (IQR 67-73); 27 males]. INTERVENTION: Participants undertook an 18-week resistance exercise program, and a nutritional intervention (fish oil vs. placebo). MEASUREMENTS: Serum homoarginine, ornithine, citrulline, asymmetric dimethylarginine (ADMA), NG-monomethyl-L-arginine (L-NMMA), and symmetric dimethylarginine (SDMA), maximal voluntary contraction (MVC) and isokinetic torque of the knee extensors at 30° s-1 (MIT), muscle cross sectional area (MCSA) and quality (MQ) were measured at baseline and after the intervention. RESULTS: No significant exercise-induced changes were observed in metabolite concentrations. There were significant sex differences in the associations between metabolites and muscle parameters. After adjusting for age, glomerular filtration rate and fish oil intervention, citrulline (P=0.002) and ornithine (P=0.022) were negatively associated with MCSA at baseline in males but not females. However, baseline citrulline was negatively correlated with exercise-induced changes in MVC (P=0.043) and MQ (P=0.026) amongst females. Furthermore, amongst males, baseline homoarginine was positively associated with exercise-induced changes in MVC (P=0.026), ADMA was negatively associated with changes in MIT (P=0.026), L-NMMA (p=0.048) and ornithine (P<0.001) were both positively associated with changes in MCSA, and ornithine was negatively associated with changes in MQ (P=0.039). CONCLUSION: Therefore, barring citrulline, there are significant sex differences in the associations between L-arginine metabolites and muscle mass and function in healthy older adults. These metabolites might enhance sarcopenia risk stratification, and the success of exercise programs, in old age.

Peripheral blood CD8+ T-cell profiles in patients with psoriatic arthritis: a cross-sectional case-control study.

OBJECTIVE: While CD4+ T-cells are traditionally regarded as the main pathogenic T-cell subpopulation in psoriatic arthritis (PsA), the role of circulating CD8+ T-cells remains poorly characterized. We evaluated the differential representation of CD8+ T-cell subpopulations in peripheral blood (PB) of PsA patients. PATIENTS AND METHODS: CD8+IL-17+, CD8+IFNγ+ and CD8+IL-17-IL-22+ T-cells were evaluated by flow-cytometry in 25 consecutive PsA patients, 7 rheumatoid arthritis (RA) patients, 16 patients with psoriasis, and 26 healthy controls (HC). RESULTS: We observed a significant expansion of circulating IFN-γ producing CD8+ T-cells in PsA when compared to psoriasis [21.2 (6.9-55.8)% vs. 3.8 (0.7-11.8)%, p < 0.0001] and HC samples [21.2 (6.9-55.8)% vs. 4.05 (0.44-19.8)%, p < 0.0001]. A frequency of circulating IFN-γ producing CD8+T-cells ≥ 9% distinguished PsA from psoriasis patients with a specificity of 84% and a sensitivity of 87.5% [AUC = 0.9 (0.80-0.99), p < 0.0001]. In addition, we found a significant expansion of circulating IL-17 producing CD8+ T cells in RA patients when compared to PsA, psoriasis and HC samples. By contrast, there were no significant between-group differences in the prevalence of circulating IL-22 producing CD8+ T-cells. In PsA patients there was a significant correlation between number of swollen joints and frequency of circulating IFN-γ producing CD8+ T-cells, and between extent and severity of psoriasis and frequency of circulating IL-17 producing CD8+ T-cells. CONCLUSIONS: Circulating IFNγ-producing CD8+ T-cells are raised in PsA when compared to psoriasis, suggesting a potential pathogenetic involvement of CD8+ T-cells and IFNγ production in chronic joint inflammation and damage. The significant enrichment of circulating IL-17 producing CD8+ T-cells in RA when compared to PsA warrants functional characterization and confirmation in larger studies. We found no significant enrichment of circulating IL-22 producing CD8+ T-cells in PsA, RA and psoriasis.

Metals in plasma of nonagenarians and centenarians living in a key area of longevity.

The concentration of calcium (Ca), cobalt (Co), copper (Cu), iron (Fe), magnesium (Mg), manganese (Mn) and selenium (Se) in plasma of 76 nonagenarians (mean age, 89.0±6.3 years), 64 centenarians (mean age, 101±1 years) and 24 middle-aged subjects as controls (mean age 61.2±1.1 years), was determined by sector field inductively coupled plasma mass spectrometry. All the subjects lived in Sardinia, an Italian island, that has the higher prevalence of centenarians than in other European countries. A comparison among the three classes of age showed a significant depletion of Ca, Co, Fe, Mn and Se (all p<0.001) in nonagenarians and centenarians with respect to controls. In particular, the geometric mean (GM) values of Ca, Co, Fe, Mn and Se were: 94.1 µg/ml, 0.46 ng/ml, 1314 ng/ml, 2.47 ng/ml and 111 ng/ml in controls; 87.6 µg/ml, 0.22 ng/ml, 815 ng/ml, 1.07 ng/ml and 88.9 ng/ml in nonagenarians; 87.0 µg/ml, 0.29 ng/ml, 713 ng/ml, 1.27 ng/ml and 81.9 ng/ml in centenarians. The highest inverse relationship with age was observed for Fe (p<0.001; ρ=-0.352) and Se (p<0.001; ρ=-0.417). This trend was also observed when data were sorted by gender. On the other hand, Cu and Mg levels in plasma remained substantially unchanged during aging. As regards Cu, it was significantly higher in females than in males in controls (GM, 1294 ng/ml vs. 1077 ng/ml; p=0.012), in nonagenarians (GM, 1216 ng/ml vs. 1081 ng/ml; p=0.011) as well as in centenarians (GM, 1226 ng/ml vs. 1152 ng/ml; p=0.045) and in hypertensive subjects with respect to healthy people (GM, 1215 ng/ml vs. 1129 ng/ml; p=0.021). These data can be used to enhance knowledge and support the research on: i) metals involved in aging in areas with high rates of human longevity; ii) variables (gender, lifestyle habits and health status) as critical determinants in aging; and iii) mineral intake and supplementation at older age affecting the healthy aging.

Differences in semen freezability and intracellular ATP content between the rooster (Gallus gallus domesticus) and the Barbary partridge (Alectoris barbara).

This study aimed to compare viability, ATP content, and DNA integrity of rooster (Gallus gallus domesticus) and Barbary partridge (Alectoris barbara) fresh and frozen spermatozoa in order to identify factors possibly related to differences in semen freezability. Ejaculates were obtained from March to May by the abdominal massage method from 3 adult roosters and 12 adult Barbary partridges. Semen was frozen with different cryoprotectants using Lake's diluents as a base medium: 1) glycerol 11%; 2) glycerol 11% and trehalose 70 mmol/L; 3) dimethylacetamide (DMA) 6%; 4) DMA 6% and trehalose 70 mmol/L. Both fresh and frozen semen showed a lower viability and higher intracellular ATP concentrations in the Barbary partridge compared with the rooster (P < 0.05). In the Barbary partridge, semen viability after thawing did not differ among the 4 media used, but glycerol showed positive effects in avoiding a significant loss of ATP after thawing, compared with DMA containing media (P < 0.05). On the other hand, in the rooster a higher viability was recorded when semen was frozen in glycerol containing media compared to DMA (P < 0.0001), while ATP values significantly decreased after thawing (P < 0.05) without showing any differences among the semen frozen in the 4 different media. DNA integrity, as evaluated by the comet assay, was assessed only in frozen semen. In the Barbary partridge, mean scored parameter did not differ significantly among semen frozen in the 4 different media. In the rooster DNA fragmentation was higher in DMA ctr medium compared with the other media and with values found in Barbary partridge semen frozen in the same medium (P < 0.001). In both species, the addition of trehalose did not show any positive effects on viability, ATP levels and DNA integrity after thawing. In conclusion, species-related differences in semen features exist between the rooster and the Barbary partridge and the wide variation observed in ATP levels may account for differences in semen freezability between the two species.

Effect of acute exercise on low molecular weight thiols in plasma.

An important defence against free radicals is represented by plasma low molecular weight (LMW) thiols that compose a dynamic system of reduced and oxidized forms able to act as a buffer redox system. This study examined the effect of an acute graded exercise bout on LMW thiols in 16 young subjects (six sedentaries and 10 athletes). Blood analysis was performed before and immediately after the exercise and total and reduced thiols were measured in order to evaluate the thiol redox status. Findings suggested that the exercise test proposed was not enough to imbalance the redox status of all LMW thiols. However, when the redox status was evaluated for each thiol, it was evident that homocysteine (Hcy) redox status was significantly different after physical activity. In particular, we found a lower level of reduced Hcy after the exercise test both in sedentaries and in athletes. We concluded that duration and intensity of the proposed exercise were not enough to promote a reactive oxygen species production able to imbalance the redox thiols status and that the lowering of the reduced Hcy form may be due to the effect produced during the effort on the synthesis and/or removal processes of Hcy.