Persistence of Human Papillomavirus, Overexpression of p53, and Outcomes of Patients After Endoscopic Ablation of Barrett's Esophagus.

We investigated the role of high-risk human papillomavirus (hr-HPV) in patients with Barrett's dysplasia and adenocarcinoma (EAC). Clearance vs persistence of HPV (DNA, E6 or E7 mRNA, and p16INK4A protein) and overexpression or mutation of p53 were determined for 40 patients who underwent endotherapy for Barrett's dysplasia or EAC. After ablation, dysplasia or neoplasia was eradicated in 34 subjects (24 squamous, 10 intestinal metaplasia). Six patients had detectable lesions after treatment; 2 were positive for transcriptionally active hr-HPV, and 4 had overexpression of p53. Before endotherapy, 15 patients had biologically active hr-HPV, 13 cleared the infection with treatment, and dysplasia or EAC was eliminated from 12 patients. One patient who cleared HPV after ablation acquired a p53 mutation, and their cancer progressed. Of 13 patients with overexpression of p53 before treatment, 10 cleared the p53 abnormality after ablation with eradication of dysplasia or neoplasia, whereas 3 of 13 had persistent p53 mutation-associated dysplasia after endotherapy (P = .004). Immunohistochemical and sequence analyses of p53 produced concordant results for 36 of 40 samples (90%). Detection of dysplasia or neoplasia after treatment was associated with HPV persistence or continued p53 overexpression.

Effects of atorvastatin on oxidative stress in chronic kidney disease.

AIM: Statins have pleiotropic effects that include attenuation of oxidative stress that may be relevant for chronic kidney disease (CKD) patients. We investigated the effect of long-term atorvastatin therapy on oxidative stress biomarkers in CKD patients. METHODS: This was a pre-specified secondary analysis of data from a randomized, double-blind, placebo-controlled trial (Lipid lowering and Onset of Renal Disease, LORD) in CKD patients. Participants received 10 mg/day atorvastatin (n = 47) or placebo (n = 39) for 3 years. Plasma measures (total F2-isoprostanes, malondialdehyde. protein carbonyls, uric acid, glutathione peroxidase (GPx) activity and total antioxidant capacity (TAC) ) were performed at baseline and at 3 years. Age and sex matched participants (n = 34) with normal kidney function were controls. RESULTS: CKD patients had significantly (P < 0.05) increased F2-isoprostanes and uric acid and decreased GPx activity compared with controls. When comparing the treatment (atorvastatin (A) vs placebo (P) ) change from baseline to 3 years, there were no significant differences (P > 0.05) in the group difference of the change values: (mean (95% CI), F2-isoprostanes = 5.3 (-29.2 to 39.8) pg/mL, protein carbonyls = 0.03 (-0.13 to 0.19) nmol/mg, GPx activity = -0.10 (-4.73 to 4.52) (U/L), uric acid = 8.8 (-33.9 to 51.6) µmol/L or TAC = -0.03 (-0.10 to 0.04) mmol/L. A significant difference (P = 0.04) in the change in malondialdehyde between groups, 1.52(0.09 to 2.96) µmol/L, was due to a large decrease in the placebo group. CONCLUSION: CKD patients had elevated oxidative stress that was not attenuated by atorvastatin 10 mg/day for 3 years.

Effects of atorvastatin on biomarkers of inflammation in chronic kidney disease.

BACKGROUND: Chronic kidney disease (CKD) is associated with inflammation. The effects of atorvastatin on biomarkers of inflammation were assessed in CKD patients in the LORD trial. METHODS: 117 patients with serum creatinine >120 µmol/L were randomized to receive atorvastatin 10 mg/day (56) or placebo (61) and followed for a mean of 2.5 years. 33 individuals with normal kidney function were controls. Outcomes included comparison of changes in pentraxin-3 (PTX3), TNF-α, CRP, IL-6, IL-8, and IL-10 between atorvastatin and placebo-treated patients. RESULTS: At baseline, compared with controls, CKD patients had increased PTX3 (mean, 1.08 vs. 0.58 ng/mL; p < 0.001), CRP (4.9 vs. 1.5 mg/L; p < 0.001), IL-8 (6.00 vs. 4.58 pg/mL; p = 0.001), IL-10 (59.0 vs. 17.6 pg/mL; p = 0.007), and TNF-α (18.0 vs. 5.6 ng/mL; p < 0.001). In patients with raised baseline plasma IL-6/8/10 and/or PTX3 the eGFR decline during the trial was significantly less in those treated with atorvastatin compared to placebo (mean change, -3.36; vs. + 1.25 mL/min/1.73 m2/year; difference, 4.61 95% CI 0.98 - 8.25; p = 0.002), whilst those without raised inflammatory biomarkers showed no difference. Placebo treated patients with raised TNF-α levels had no eGFR decline (p > 0.90), whereas in atorvastatin-treated patients eGFR declined (p = 0.05). CONCLUSIONS: CKD patients with inflammation treated with atorvastatin had significantly less eGFR decline. Larger studies using statin therapy, specifically enrolling CKD patients with inflammation, may be worthwhile exploring.

Transcriptionally active human papillomavirus is strongly associated with Barrett's dysplasia and esophageal adenocarcinoma.

OBJECTIVES: The role of human papillomavirus (HPV) in Barrett's esophagus (BE) remains unclear. The few studies that have previously investigated HPV and esophageal adenocarcinoma (EAC) or BE have produced either negative data or positive results of doubtful clinical/etiological significance or have detected only low-risk HPV types. We therefore prospectively determined the prevalence of biologically active HPV in esophageal epithelium of patients representing the Barrett's metaplasia-dysplasia-adenocarcinoma sequence. METHODS: HPV DNA was estimated by nested PCR and viral transcriptional activity detected by E6/7 oncogene mRNA expression and p16INK4A immunohistochemistry in fresh frozen and paraffin-embedded esophageal biopsies of patients with BE, Barrett's dysplasia (BD), and EAC, as well as controls. Biopsies were obtained from the transformation zone (squamocolumnar junction (SCJ)) and the lesion, or corresponding site in controls, i.e., 2 cm above the gastroesophageal junction (GEJ). RESULTS: Of the 261 patients, 81 were positive for HPV DNA. In controls and BE, the virus was mostly detected at the transformation zone. Compared with controls (18.0%), HPV positivity was significantly more common in BD (68.6%, incidence rate ratio (IRR) 2.94, 95% confidence interval (CI) 1.78-4.85, P<0.001) and EAC (66.7%, IRR 2.87, 95% CI 1.69-4.86, P<0.001), but not in BE (22.1%, IRR 1.06, 95% CI 0.60-1.85, P=0.85). Of the patients, 92.6% were high-risk (HR) HPV, i.e., types 16 and 18. Again, p16INK4A positivity was greatest in BD and EAC and much less in BE patients (44.1%, IRR 17.0 (95% CI 4.86-59.6, P<0.001), 44.4%, 17.0 (95% CI 4.87-59.4, P<0.001), and 10.6%, 3.93 (95% CI 1.01-15.3, P=0.048) respectively). In 66 HPV DNA-positive patients tested for E6/E7 mRNA, none of the control (n=16) or BE (n=13) individuals were positive, whereas 9/22 BD and 9/15 EAC patients demonstrated oncogene expression (P<0.001). When HPV DNA, p16INK4A, and E6/E7 mRNA were all positive, there was a very strong association with disease severity (SCJ: odds ratio (OR) 104, 95% CI 20.3-529, P<0.001; lesion: OR 62.2, 95% CI 12.4-311, P<0.001) than when all were negative. CONCLUSIONS: Transcriptionally active HR-HPV was strongly associated with BD and EAC, but was largely biologically irrelevant in BE and controls, suggesting a potential role in esophageal carcinogenesis. These data provide robust justification for further detailed longitudinal, interventional, and molecular studies.

Effect of hereditary haemochromatosis genotypes and iron overload on other trace elements.

PURPOSE: Hereditary haemochromatosis is a common genetic disorder involving dysregulation of iron absorption. There is some evidence to suggest that abnormal iron absorption and metabolism may influence the status of other important trace elements. In this study, the effect of abnormal HFE genotypes and associated iron overload on the status of other trace elements was examined. METHODS: Dietary data and blood samples were collected from 199 subjects (mean age = 55.4 years; range = 21-81 years). Dietary intakes, serum selenium, copper and zinc concentrations and related antioxidant enzymes (glutathione peroxidase and superoxide dismutase) in subjects with normal HFE genotype (n = 118) were compared to those with abnormal HFE genotype, with both normal iron status (n = 42) and iron overload (n = 39). RESULTS: For most dietary and biochemical variables measured, there were no significant differences between study groups. Red cell GPx was significantly higher in male subjects with normal genotypes and normal iron status compared to those with abnormal genotypes and normal iron status (P = 0.03) or iron overload (P = 0.001). Red cell GPx was also highest in normal women and significantly lower in the abnormal genotype and normal iron group (P = 0.016), but not in the iron overload group (P = 0.078). CONCLUSION: Although it may not be possible to exclude a small effect between the genotype groups on RBC GPx, overall, haemochromatosis genotypes or iron overload did not appear to have a significant effect on selenium, copper or zinc status.

Serum [25(OH)D] status, ankle strength and activity show seasonal variation in older adults: relevance for winter falls in higher latitudes.

BACKGROUND: seasonal variation exists in serum [25(OH)D] and physical activity, especially at higher latitudes, and these factors impact lower limb strength. This study investigates seasonal variation in leg strength in a longitudinal repeated measures design concurrently with serum vitamin D and physical activity. METHODS: eighty-eight community-dwelling independently mobile older adults (69.2 ± 6.5 years) were evaluated five times over a year, at the end of five consecutive seasons at latitude 41.1°S, recruited in two cohorts. Leg strength, serum [25(OH)D] and physical activity levels were measured. Time spent outside was recorded. Monthly falls diaries recorded falls. Data were analysed to determine annual means and percentage changes. RESULTS: significant variation in [25(OH)D] (±15%), physical activity (±13%), ankle dorsiflexion strength (±8%) and hours spent outside (±20%) (all P < 0.001) was demonstrated over the year, with maximums in January and February (mid-summer). Low mean ankle strength was associated with increased incidence of falling (P = 0.047). Quadriceps strength did not change (±2%; P = 0.53). CONCLUSION: ankle dorsiflexor strength varied seasonally. Increased ankle strength in summer may be influenced by increased levels of outdoors activity over the summer months. Reduced winter-time dorsiflexor strength may predispose older people to increased risk of tripping-related falls, and warrants investigation in a multi-faceted falls prevention programme.

Development and experimental application of an HPLC procedure for the determination of capsaicin and dihydrocapsaicin in serum samples from human subjects.

Total capsaicins are extracted from 2 mL aliquots of serum or plasma using methyl-isobutyl ketone, evaporation of the extract to dryness and reconstitution with 200 µL of acetonitrile. The HPLC mobile phase is 40:60 water:acetonitrile. The absorbance of the eluent is monitored at 205 nm. Standardisation uses a known mixture of pure capsaicin and dihydrocapsaicin. Accuracies are 98.9 and 100.6 % for capsaicin and dihydrocapsaicin respectively. Inter batch reproducibility for both is 15 %. The limits of detection are 2.6 and 3.8 ng/mL for capsaicin and dihydrocapsaicin respectively. Analyses of sera obtained previously from human subjects who had eaten chilli containing meals showed that in those that absorbed capsaicins (N = 30) then the median, mean and SD of their serum capsaicin were: 13.4, 18.9 and 16.3 ng/mL. The corresponding data for those sera (N = 13) that had measurable levels of dihydrocapsaicin were: 6.9, 7.5 and 3.6 ng/mL. This procedure is suitable for use in prospective studies of the metabolism of orally ingested chilli.

Effects of atorvastatin on NGAL and cystatin C in chronic kidney disease: a post hoc analysis of the LORD trial.

BACKGROUND: Neutrophil gelatinase-associated lipocalin (NGAL) and cystatin C are biomarkers of kidney injury and function, respectively. This study assessed whether plasma NGAL and/or serum cystatin C predicted baseline estimated glomerular filtration rate (eGFR) and urinary protein excretion, rate of change of eGFR and urinary protein excretion and whether atorvastatin influenced changes in these biomarkers in patients with chronic kidney disease (CKD). METHODS: This is a post hoc analysis of the Lipid Lowering and Onset of Renal Disease trial, a randomized double-blind, placebo-controlled trial where 88 patients with Stages 2-4 CKD received atorvastatin 10 mg/day (48) or placebo (40). Stored blood samples were analysed for NGAL and cystatin C at baseline and a mean of 1.5 and 2.9 years later. Serum creatinine and Modification of Diet in Renal Disease (MDRD) eGFR were obtained three monthly. RESULTS: There were negative associations between NGAL and cystatin C and eGFR (P = 0.025 and P < 0.001, respectively) at all time points. There were no associations between baseline NGAL and cystatin C and rate of change of eGFR (P = 0.44 and P = 0.49, respectively). Baseline NGAL but not cystatin C (P = 0.043 and P = 0.35, respectively) predicted rate of change of urinary protein excretion. In atorvastatin-treated patients, NGAL decreased (mean, -7.4 ng/mL/year; SD 128.4), whereas it increased in the placebo group [mean, 4.6 ng/mL/year; SD 56.6), the difference being statistically significant (P = 0.049). CONCLUSIONS: NGAL is a biomarker of existing CKD but did not predict CKD progression. Atorvastatin reduced plasma NGAL but the significance and mechanisms require further investigation. Atorvastatin had no significant effect on cystatin C.

Relationship between antioxidant enzyme genotype and activity and kidney function: a case-control study.

BACKGROUND: Oxidative stress is associated with the progression of chronic kidney disease (CKD). Links between antioxidant enzyme SNPs such as superoxide dismutase (SOD) Ala16Val, glutathione peroxidase (GPx) Pro197Leu and catalase C- 262T and CKD have not been investigated. This study compared antioxidant genotypes and activities of CKD patients with population controls, and determined their relationship to kidney function. METHODS: CKD patients (n = 230) and controls (n = 224) were screened for the GPx, SOD and catalase SNPs. Plasma and red blood cell (RBC) GPx, RBC SOD and RBC catalase activities, and estimated glomerular filtration rate (eGFR) were measured. RESULTS: Significantly more CKD patients (n = 5) had the GPx Leu/Leu genotype compared to controls (n = 0), and had lower eGFR (p = 0.054). CKD patients had significantly lower plasma GPx and RBC catalase activities compared to controls, whereas RBC GPx and RBC SOD activities were significantly higher in CKD patients (p < 0.001). CONCLUSIONS: CKD is associated with reduced plasma GPx and catalase activities and enhanced RBC GPx and SOD activities. Although, genotype frequencies were similar for both groups, lower eGFR was associated with the GPx Leu/ Leu genotype.

Glutathione peroxidase, superoxide dismutase and catalase genotypes and activities and the progression of chronic kidney disease.

BACKGROUND: Oxidative stress has been linked to the progression of disease, including chronic kidney disease (CKD). The aim of the present study was to determine the association between single-nucleotide polymorphisms (SNPs) of the antioxidant enzymes, superoxide dismutase (SOD), glutathione peroxidase (GPx) and catalase and their activities and the progression of CKD. METHODS: This is a prospective cohort study of 185 CKD patients (Stages 2-4), followed for up to 12 months. All patients were genotyped for SNPs of SOD (SOD Ala16Val), GPx (GPx Pro197Leu) and catalase (C-262T). The rate of change over the study period of estimated glomerular filtration rate (eGFR), plasma and red blood cell (RBC) GPx, RBC SOD and RBC catalase activities were determined. RESULTS: CKD patients with the SOD Ala/Val and Val/Val genotypes had a significantly greater eGFR decline compared to those with the Ala/Ala genotype (Ala/Val compared with Ala/Ala odds ratio (OR) 0.35, 95% CI 0.19 to 0.64, P = 0.001; Val/Val compared with Ala/Ala OR 0.25, 95% CI 0.10 to 0.65, P = 0.005). The progression of CKD was not associated with SNPs of the GPx or catalase genes studied but there was a direct relationship between the rate of change of plasma GPx activity and the rate of change of eGFR over 12 months (P = 0.025). CONCLUSION: CKD patients with the SOD Ala/Val and Val/Val genotypes have a greater decline in kidney function than those with the Ala/Ala genotype.