The impact of obesity and bariatric surgery on the immune microenvironment of the endometrium.

BACKGROUND: The incidence of endometrial cancer is rising in parallel with the obesity epidemic. Obesity increases endometrial cancer risk and weight loss is protective, but the underlying mechanisms are incompletely understood. We hypothesise that the immune microenvironment may influence susceptibility to malignant transformation in the endometrium. The aim of this study was to measure the impact of obesity and weight loss on the immunological landscape of the endometrium. METHODS: We conducted a prospective cohort study of women with class III obesity (body mass index, BMI ≥ 40 kg/m2) undergoing bariatric surgery or medically-supervised low-calorie diet. We collected blood and endometrial samples at baseline, and two and 12 months after weight loss intervention. Serum was analysed for inflammatory markers CRP, IL-6 and TNF-α. Multiplex immunofluorescence was used to simultaneously identify cells positive for immune markers CD68, CD56, CD3, CD8, FOXP3 and PD-1 in formalin-fixed paraffin-embedded endometrial tissue sections. Kruskal-Wallis tests were used to determine whether changes in inflammatory and immune biomarkers were associated with weight loss. RESULTS: Forty-three women with matched serum and tissue samples at all three time points were included in the analysis. Their median age and BMI were 44 years and 52 kg/m2, respectively. Weight loss at 12 months was greater in women who received bariatric surgery (n = 37, median 63.3 kg) than low-calorie diet (n = 6, median 12.8 kg). There were significant reductions in serum CRP (p = 3.62 × 10-6, r = 0.570) and IL-6 (p = 0.0003, r = 0.459), but not TNF-α levels, with weight loss. Tissue immune cell densities were unchanged except for CD8+ cells, which increased significantly with weight loss (p = 0.0097, r = -0.323). Tissue CD3+ cell density correlated negatively with systemic IL-6 levels (p = 0.0376; r = -0.318). CONCLUSION: Weight loss is associated with reduced systemic inflammation and a recruitment of protective immune cell types to the endometrium, supporting the concept that immune surveillance may play a role in endometrial cancer prevention.

PROgesterone Therapy for Endometrial Cancer Prevention in Obese Women (PROTEC) Trial: A Feasibility Study.

Obesity is the major etiologic driver for endometrial cancer. The levonorgestrel intrauterine system (LNG-IUS) reduces the risk of endometrial cancer and its precursor, atypical hyperplasia. We assessed feasibility and uptake of the LNG-IUS for primary prevention of endometrial cancer in high-risk women and its impact on endometrial tissue biomarkers. Women with class-III obesity [body mass index (BMI) > 40 kg/m2] and histologically normal endometrium were invited to participate in a clinical trial of the LNG-IUS for endometrial protection. Recruitment, successful LNG-IUS insertion, and adherence to trial procedures were recorded. We measured impact of the LNG-IUS on circulating biomarkers of endometrial cancer risk, endometrial proliferation (Ki-67, pAKT, PTEN), endometrial hormone receptor status [estrogen receptor and progesterone receptor (PR)], mental wellbeing, and menstrual function. At 6 months, women chose to keep their LNG-IUS or have it removed. In total, 103 women were approached, 54 were offered a participant information sheet, 35 agreed to participate, and 25 received a LNG-IUS. Their median age and BMI were 54 years [interquartile range (IQR) 52-57] and 47 kg/m2 (IQR 44-51), respectively. Three women (3/35, 9%) were ineligible due to atypical hyperplasia/endometrial cancer on their baseline biopsy. The LNG-IUS was well tolerated and had a positive overall effect on bleeding patterns and mental wellbeing. The LNG-IUS was associated with endometrial morphologic change, reduced Ki-67, and PR expression, but circulating biomarkers of endometrial cancer risk were unchanged. All but one woman (96%) kept her LNG-IUS. The LNG-IUS appears to be acceptable to some women with class-III obesity for primary prevention of endometrial cancer, which could provide a strategy for a prevention trial.Prevention Relevance: Novel strategies are urgently needed to prevent the rise in endometrial cancer diagnoses predicted by escalating obesity rates. Here, we show that women with class III obesity are willing to engage in risk reduction with a levonorgestrel intrauterine system, which could provide a strategy for an endometrial cancer prevention trial.

Baseline Serum HE4 But Not Tissue HE4 Expression Predicts Response to the Levonorgestrel-Releasing Intrauterine System in Atypical Hyperplasia and Early Stage Endometrial Cancer.

The levonorgestrel-releasing intrauterine system (LNG-IUS) is a conservative management option for atypical hyperplasia (AH) and low grade early stage endometrial cancer (EEC), but around 1 in 3 patients fail to respond to treatment. The aim of this study was to investigate if serum and/or tissue HE4 expression could predict response to LNG-IUS therapy. Patients with AH or presumed Stage I EEC had serum and endometrial samples taken at baseline and at 3-month intervals over 12 months post-insertion of LNG-IUS. 74 patients were recruited and baseline demographics recorded. Of 57 patients for whom response was histologically determinable, 39 (68%) were responders and 18 (32%) non-responders. Mean baseline serum HE4 was significantly lower in responders (62.1 ± 1.1 pM, 95% confidence interval (CI) 52.7-73.2), compared to non-responders (125.6 ± 1.3 pM, 95% CI 74.5-211.7, p = 0.014), including when considering age, BMI, menopausal status, smoking status, and histological grade as covariables (p = 0.005). Baseline tissue HE4 expression was not significantly different in responders compared to non-responders (p = 0.999). Responders showed a significant mean reduction (-9.8 ± 3.4%, 95% CI -16.7 to -2.8%, p = 0.008) in serum HE4 between baseline and 3 months (p = 0.008), whereas non-responders showed no significant change (p = 0.676). Neither responders nor non-responders showed a significant percentage change in serum HE4 from baseline beyond 3 months (p > 0.05). Change in serum HE4 between baseline and 3 and 6 months and tissue HE4 tissue expression between baseline and 3, 6, and 12 months was not significantly different in responders compared to non-responders (p > 0.05). This study suggests that baseline serum HE4, but not baseline tissue HE4 expression, is independently predictive of response to the LNG-IUS and could be used to guide management decisions.

The impact of obesity and bariatric surgery on circulating and tissue biomarkers of endometrial cancer risk.

Obesity is the strongest risk factor for endometrial cancer (EC). To inform targeted screening and prevention strategies, we assessed the impact of obesity and subsequent bariatric surgery-induced weight loss on endometrial morphology and molecular pathways implicated in endometrial carcinogenesis. Blood and endometrial tissue were obtained from women with class III-IV obesity (body mass index ≥40 and ≥50 kg/m2 , respectively) immediately prior to gastric bypass or sleeve gastrectomy, and at two and 12 months' follow up. The endometrium underwent pathological examination and immunohistochemistry was used to quantify proliferation (Ki-67), oncogenic signaling (PTEN, pAKT, pERK) and hormone receptor (ER, PR) expression status. Circulating biomarkers of insulin resistance, reproductive function and inflammation were also measured at each time point. Seventy-two women underwent bariatric surgery. At 12 months, the mean change in total and excess body weight was -32.7 and -62.8%, respectively. Baseline endometrial biopsies revealed neoplastic change in 10 women (14%): four had EC, six had atypical hyperplasia (AH). After bariatric surgery, most cases of AH resolved (5/6) without intervention (3/6) or with intrauterine progestin (2/6). Biomarkers of endometrial proliferation (Ki-67), oncogenic signaling (pAKT) and hormone receptor status (ER, PR) were significantly reduced, with restoration of glandular PTEN expression, at 2 and 12 months. There were reductions in circulating biomarkers of insulin resistance (HbA1c, HOMA-IR) and inflammation (hsCRP, IL-6), and increases in reproductive biomarkers (LH, FSH, SHBG). We found an unexpectedly high prevalence of occult neoplastic changes in the endometrium of women undergoing bariatric surgery. Their spontaneous reversal and accompanying down-regulation of PI3K-AKT-mTOR signaling with weight loss may have implications for screening, prevention and treatment of this disease.

PRE-surgical Metformin In Uterine Malignancy (PREMIUM): a Multi-Center, Randomized Double-Blind, Placebo-Controlled Phase III Trial.

PURPOSE: Endometrioid endometrial cancer is strongly associated with obesity and insulin resistance. Metformin, an insulin sensitizer, reduces endometrial tumor growth in vitro. Presurgical window studies allow rapid in vivo assessment of antitumor activity. Previous window studies found metformin reduced endometrial cancer proliferation but these lacked methodological rigor. PREMIUM measured the anti-proliferative effect of metformin in vivo using a robust window study design.Patients and Methods: A multicenter, double-blind, placebo-controlled trial randomized women with atypical hyperplasia or endometrioid endometrial cancer to receive metformin (850 mg daily for 3 days, and twice daily thereafter) or placebo for 1 to 5 weeks until surgery. The primary outcome was posttreatment IHC expression of Ki-67. Secondary outcomes investigated the effect of metformin on markers of the PI3K-Akt-mTOR and insulin signaling pathways and obesity. RESULTS: Eighty-eight women received metformin (n = 45) or placebo (n = 43) and completed treatment. There was no overall difference in posttreatment Ki-67 between the metformin and placebo arms, in an ANCOVA analysis adjusting for baseline Ki-67 expression (mean difference -0.57%; 95% CI, -7.57%-6.42%; P = 0.87). Metformin did not affect expression of markers of the PI3K-Akt-mTOR or insulin signaling pathways, and did not result in weight loss. CONCLUSIONS: Short-term treatment with standard diabetic doses of metformin does not reduce tumor proliferation in women with endometrioid endometrial cancer awaiting hysterectomy. This study does not support a biological effect of metformin in endometrial cancer and casts doubt on its potential application in the primary and adjuvant treatment settings.

Modification of the Beckman-Coulter second-generation enzyme-linked immunosorbent assay protocol improves the reliability of serum antimüllerian hormone measurement.

OBJECTIVE: To determine whether the modified Beckman-Coulter 2nd-generation (Gen II) antimüllerian hormone (AMH) assay (Gen IIm) provides more consistent results following storage at room temperature and on dilution than the original Gen II assay, to compare AMH results from the modified assay with those obtained from the original assay, and to assess the relationship between new AMH values and the antral follicle count (AFC). DESIGN: Cohort. SETTING: Hospital fertility clinic. PATIENT(S): A total of 678 consecutive women (21-46 years old) investigated for subfertility. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): AMH was measured by means of the Gen IIm assay protocol in women with known AFC. AMH values were obtained on a subset of serum samples by means of both original and modified assays. RESULT(S): Specimens analyzed by Gen IIm exhibited a proportional AMH response on dilution, and AMH values decreased by an average of 12.1% after 7 days at room temperature, in contrast to the steady increase seen with the use of the original Gen II assay. Gen IIm assay values were, on average, 51.4% higher than Gen II values. Population analysis suggested a conversion factor of 1.35 (95% CI 1.23-1.47) between the Gen IIm and historical data obtained for the Diagnostic Systems Laboratories AMH assay. The relationship between the Gen IIm AMH measurement and AFC was adequately represented by a linear function. CONCLUSION(S): The Gen IIm assay gave more reliable AMH results on sample dilution and storage than the original Gen II protocol. Findings obtained with the use of the original Gen II ELISA method should be treated with caution.

The measurement of anti-Müllerian hormone: a critical appraisal.

CONTEXT: Measurement of anti-Müllerian hormone (AMH) is perceived as reliable, but the literature reveals discrepancies in reported within-subject variability and between-method conversion factors. Recent studies suggest that AMH may be prone to preanalytical instability. We therefore examined the published evidence on the performance of current and historic AMH assays in terms of the assessment of sample stability, within-patient variability, and comparability of the assay methods. EVIDENCE ACQUISITION: We reviewed studies (manuscripts or abstracts) measuring AMH, published in peer-reviewed journals between January 1, 1990, and August 1, 2013, using appropriate PubMed/Medline searches. EVIDENCE SYNTHESIS: AMH levels in specimens left at room temperature for varying periods increased by 20% in one study and by almost 60% in another, depending on duration and the AMH assay used. Even at -20°C, increased AMH concentrations were observed. An increase over expected values of 20-30% or 57%, respectively, was observed after 2-fold dilution in two linearity-of-dilution studies, but not in others. Several studies investigating within-cycle variability of AMH reported conflicting results, although most studies suggest that variability of AMH within the menstrual cycle appears to be small. However, between-sample variability without regard to menstrual cycle as well as within-sample variation appears to be higher using the GenII AMH assay than with previous assays, a fact now conceded by the kit manufacturer. Studies comparing first-generation AMH assays with each other and with the GenII assay reported widely varying differences. CONCLUSIONS: AMH may exhibit assay-specific preanalytical instability. Robust protocols for the development and validation of commercial AMH assays are required.

Serum cystatin C is independently associated with renal impairment and high sensitivity C-reactive protein in systemic lupus erythematosus.

OBJECTIVES: In systemic lupus erythematosus (SLE) patients, glomerular filtration rate (GFR) is usually estimated using the modified Cockcroft-Gault (mCG) and Modification of Diet in Renal Disease (MDRD) equations. We aimed to study cystatin C (sCysC) in SLE to assess its agreement with standard renal indices and investigate factors affecting sCysC in SLE. METHODS: SLE patients (≥4 ACR criteria) and healthy women from Greater Manchester were recruited and clinical assessments were undertaken. SCysC was measured using R & D Systems' ELISA. Agreement between renal measures was assessed using Deming plots and factors associated with sCysC in SLE were examined by multiple linear regression analyses. RESULTS: 178 patients and 68 controls had median (IQR) ages of 53 (46-61) and 50 (39-60) years, respectively. In an age-adjusted analysis, SLE patients had higher sCysC (1.16 [0.98-1.36] vs. 0.950 [0.73-1.13] mg/l; p<0.0001) and within SLE those with a history of lupus nephritis had higher sCysC (1.31 [1.10-1.66] vs. 1.11 [0.95-1.29] mg/l; p<0.005). SCysC correlated positively with serum creatinine, and inversely to renal measures (r=-0.530; p<0.0001 [mCG], and r=-0.620; p<0.0001 [MDRD]). There was closer agreement between the two eGFR measures than between either eGFR measures and sCysC. In addition to age and serum creatinine, a multivariate analysis (ß, p) found that high-sensitivity C-reactive protein (hs-CRP) (0.03, 0.026) was also independently associated with sCysC in SLE. CONCLUSIONS: In SLE, sCysC may be influenced by low grade inflammation as well as by renal dysfunction. Therefore, SCysC should not supplant current assessment of renal dysfunction in SLE.

Anti-Mullerian hormone: poor assay reproducibility in a large cohort of subjects suggests sample instability.

STUDY QUESTION: What is the variability of anti-Müllerian hormone (AMH) concentration in repeat samples from the same individual when using the Gen II assay and how do values compare to Gen I [Diagnostic Systems Ltd (DSL)] assay results? SUMMARY ANSWER: The Gen II AMH assay displayed appreciable variability, which can be explained by sample instability. WHAT IS KNOWN ALREADY: AMH is the primary predictor of ovarian performance and is used to tailor gonadatrophin dosage in cycles of IVF/ICSI and in other routine clinical settings. Thus, a robust, reproducible and sensitive method for AMH analysis is of paramount importance. The Beckman Coulter Gen II ELISA for AMH was introduced to replace earlier DSL and Immunotech assays. The performance of the Gen II assay has not previously been studied in a clinical setting. STUDY DESIGN, SIZE AND DURATION: We studied an unselected group of 5007 women referred for fertility problems between 1 September 2008 and 25 October 2011; AMH was measured initially using the DSL AMH ELISA and subsequently using the Gen II assay. AMH values in the two assays were compared using a regression model in log(AMH) with a quadratic adjustment for age. Additionally, women (n = 330) in whom AMH had been determined in different samples using both the DSL and Gen II assays (paired samples) identified and the difference in AMH levels between the DSL and Gen II assays was estimated using the age-adjusted regression analysis. A subset of 313 women had repeated AMH determinations (n = 646 samples) using the DSL assay and 87 women had repeated AMH determinations using the Gen II assay (n = 177 samples) were identified. A mixed effects model in log(AMH) was utilized to estimate the sample-to-sample (within-subject) coefficients of variation of AMH, adjusting for age. Laboratory experiments including sample stability at room temperature, linearity of dilution and storage conditions used anonymized samples. MAIN RESULTS AND THE ROLE OF CHANCE: In clinical practice, Gen II AMH values were ∼20% lower than those generated using the DSL assay instead of the 40% increase predicted by the kit manufacturer. Both assays displayed high within-subject variability (Gen II assay CV = 59%, DSL assay CV = 32%). In the laboratory, AMH levels in serum from 48 subjects incubated at RT for up to 7 days increased progressively in the majority of samples (58% increase overall). Pre-dilution of serum prior to assay, gave AMH levels up to twice that found in the corresponding neat sample. Pre-mixing of serum with assay buffer prior to addition to the microtitre plate gave higher readings (72% overall) compared with sequential addition. Storage at -20°C for 5 days increased AMH levels by 23% compared with fresh samples. The statistical significance of results was assessed where appropriate. LIMITATIONS, REASONS FOR CAUTION: The analysis of AMH levels is a retrospective study and therefore we cannot entirely rule out the existence of differences in referral practices or changes in the two populations. WIDER IMPLICATIONS OF THE FINDINGS: Our data suggests that AMH may not be stable under some storage or assay conditions and this may be more pronounced with the Gen II assay. The published conversion factors between the Gen II and DSL assays appear to be inappropriate for routine clinical practice. Further studies are urgently required to confirm our observations and to determine the cause of the apparent instability. In the meantime, caution should be exercised in the interpretation of AMH levels in the clinical setting. CONFLICT OF INTEREST/STUDY FUNDING: S. Roberts is supported by the NIHR Manchester Biomedical Research Centre.

The reproducibility of serum anti-Müllerian hormone in subfertile women: within and between patient variability.

Serum anti-Müllerian hormone concentrations vary significantly over time and this should be taken into account when tailoring treatment protocols for patients undergoing controlled ovarian hyperstimulation (COH). Compared with FSH, serum anti-Müllerian hormone may have greater discriminatory power because of its modest intrapatient variation and the larger interpatient variation.

Explanations for the lower rates of diabetic neuropathy in Indian Asians versus Europeans.

OBJECTIVE: Risks of diabetes and cardiovascular disease are elevated worldwide in Indian Asians. However, risks of other diabetes-related complications, i.e., foot ulceration and amputation, also with a vascular basis, are substantially lower in Asians than in white Europeans in the U.K., possibly due to less neuropathy. We therefore compared signs, symptoms, and objective quantitative measures of diabetic neuropathy and their risk factors in Indian Asians and Europeans. RESEARCH DESIGN AND METHODS: This was a cross-sectional study of a population-based sample of age- and sex-matched adults with type 2 diabetes of European (95 male and 85 female) and Asian (96 male and 84 female) descent in the U.K. Patients were assessed for neuropathic symptoms, signs, nerve conduction, autonomic function, and quantitative sensory testing. Peripheral vascular function and other potential risk factors for neuropathy were measured. RESULTS Mean nerve conduction velocity Z scores were better in Asians (mean +/- SD 0.07 +/- 0.62) than in Europeans (-0.11 +/- 0.60; P = 0.007) and were explained by the shorter height, fewer pack-years smoked, and higher transcutaneous oxygen levels (TCpO(2)) in Indian Asians (P value for ethnic comparison attenuated to 0.2). Small fiber neuropathy was less prevalent in Indian Asians compared with Europeans (odds ratio 0.58 [95% CI 0.37-0.93]; P = 0.02) and was primarily accounted for by better TCpO(2) (0.70 [0.40-1.21]; P = 0.2). CONCLUSIONS: Asians with diabetes have substantially less large and small fiber neuropathy than Europeans, despite comparable traditional risk factors. Independent from smoking, the lower risk of neuropathy in Asians is due to better skin microvascularization and may help explain the substantially reduced Asian foot ulcer risk.

Biomarkers of oxidant stress, insulin sensitivity and endothelial activation in rheumatoid arthritis: a cross-sectional study of their association with accelerated atherosclerosis.

BACKGROUND: Women with rheumatoid arthritis (RA) have increased morbidity and mortality due to coronary heart disease. Chronic systemic inflammation is known to accelerate atherosclerosis and increase arterial stiffness in patients, but other mechanisms may also be involved. Biomarkers of oxidant stress, inflammation, insulinaemia and endothelial dysfunction were measured in blood and urine from 46 RA patients and 48 age-matched controls. Plaque formation and intima-medial thickness (IMT) were measured using B-mode carotid Doppler scan. FINDINGS: The prevalence of plaque was increased (p = 0.042) in RA patients between 50-59 years old compared to the same age group in controls. 8-isoprostane (p = 0.004), C-reactive protein (p < 0.001), interleukin-6 (p < 0.001), insulin (p = 0.035), adiponectin (p = 0.012), vascular cell adhesion molecule (VCAM) (p = 0.029) and E-selectin (p < 0.001) were all increased while selenium (p = 0.003) and LDL-cholesterol (p = 0.025) were both decreased in all RA patients. 8-isoprostane correlated with 10 year cardiac risk (r = 0.55, p < 0.001), VCAM with IMT (r = 0.37, p = 0.012) and E-selectin with rheumatoid factor titre (r = 0.43, p = 0.003) in RA patients. In the control group, age, carotid IMT, VCAM, systolic blood pressure and smoking status were all associated with plaque development whereas in RA patients only age was associated with plaque. CONCLUSION: The burden of atherosclerosis is particularly increased in middle-aged women with RA. Patients with RA have increased levels of oxidant stress, inflammation, insulin and soluble adhesion molecules. As the association between classical risk factors was much weaker in RA patients compared to controls, these additional factors may be more important in the accelerated development of atheroma in RA.

Local inflammation and hypoxia abolish the protective anticontractile properties of perivascular fat in obese patients.

BACKGROUND: Inflammation in adipose tissue has been implicated in vascular dysfunction, but the local mechanisms by which this occurs are unknown. METHODS AND RESULTS: Small arteries with and without perivascular adipose tissue were taken from subcutaneous gluteal fat biopsy samples and studied with wire myography and immunohistochemistry. We established that healthy adipose tissue around human small arteries secretes factors that influence vasodilation by increasing nitric oxide bioavailability. However, in perivascular fat from obese subjects with metabolic syndrome (waist circumference 111+/-2.8 versus 91.1+/-3.5 cm in control subjects, P<0.001; insulin sensitivity 41+/-5.9% versus 121+/-18.6% in control subjects, P<0.001), the loss of this dilator effect was accompanied by an increase in adipocyte area (1786+/-346 versus 673+/-60 mum(2), P<0.01) and immunohistochemical evidence of inflammation (tumor necrosis factor receptor 1 12.4+/-1.1% versus 6.7+/-1%, P<0.001). Application of the cytokines tumor necrosis factor receptor-alpha and interleukin-6 to perivascular fat around healthy blood vessels reduced dilator activity, resulting in the obese phenotype. These effects could be reversed with free radical scavengers or cytokine antagonists. Similarly, induction of hypoxia stimulated inflammation and resulted in loss of anticontractile capacity, which could be rescued by catalase and superoxide dismutase or cytokine antagonists. Incubation with a soluble fragment of adiponectin type 1 receptor or inhibition of nitric oxide synthase blocked the vasodilator effect of healthy perivascular adipose tissue. CONCLUSIONS: We conclude that adipocytes secrete adiponectin and provide the first functional evidence that it is a physiological modulator of local vascular tone by increasing nitric oxide bioavailability. This capacity is lost in obesity by the development of adipocyte hypertrophy, leading to hypoxia, inflammation, and oxidative stress.

Factors predicting anaphylaxis to peanuts and tree nuts in patients referred to a specialist center.

BACKGROUND: Although acute allergic reactions after ingestion of peanuts and tree nuts are common, fatalities are rare. Other than patients with coexisting asthma, it is currently not possible to predict which patients are most likely to develop severe reactions. OBJECTIVE: The aim of this study was to determine which clinical and laboratory parameters best predict the likelihood of severe allergic reactions. METHODS: From 1992 to 2004, we collected detailed information on the clinical severity and allergy test results of 1094 patients with peanut and tree nut allergy attending a regional allergy center. In a subgroup of 122 patients, sera were assayed for activity of enzymes involved in the catabolism of bradykinin. RESULTS: Severe pharyngeal edema was 3.8 (2.1-6.9) times more common in patients with severe rhinitis and 2.6 (1.8-3.7) more common after ingestion of tree nuts compared with peanuts. Patients with serum angiotensin-converting enzyme concentrations <37.0 mmol/L had a 9.6 (1.6-57)-fold risk of severe pharyngeal edema. Life-threatening bronchospasm was most likely in patients with severe asthma (relative risk, 6.8 [4.1-11.3]) and less so in patients with milder asthma (2.7 [1.7-4.0]). Altered levels of consciousness were more likely in patients with severe eczema (3.1 [1.1-8.4]). CONCLUSION: Severity of coexisting atopic diseases predicted which patients attending a tertiary referral clinic were most likely to develop life-threatening allergic reactions to peanuts and tree nuts. Patients with the lowest serum angiotensin-converting enzyme concentrations were more likely to develop life-threatening pharyngeal edema, suggesting that this complication may be partly mediated by bradykinin.

Ursodeoxycholic acid in primary biliary cirrhosis improves glutathione status but fails to reduce lipid peroxidation.

BACKGROUND: Ursodeoxycholic acid (UDCA) may slow progression in primary biliary cirrhosis (PBC), but its effect on survival is controversial. We have previously demonstrated that oxidant stress, with severely depressed plasma glutathione, is a feature of untreated PBC; this study examines the effect of UDCA on lipid peroxidation, antioxidant status and associated processes. PATIENTS AND METHODS: Markers of lipid peroxidation, antioxidant status, hepatic fibrogenesis, inflammation, cholestasis and synthetic function were measured at 0, 3, 6, 9 and 12 months in blood and urine from 35 PBC patients receiving UDCA. RESULTS: Plasma glutathione, reflecting intrahepatic levels, climbed steadily on UDCA; although still subnormal, the median value at 12 months was 2.4-fold higher than the untreated level. Liver enzyme markers and C-reactive protein also improved, whilst PIIINP improved steadily, but the change did not attain statistical significance. Serum bilirubin remained unchanged and total antioxidant capacity, albumin and vitamin E decreased after 12 months' UDCA treatment. 8-Isoprostane increased and malondialdehyde was unchanged. CONCLUSIONS: UDCA treatment partially corrected plasma glutathione status and some other biomarkers greatly improved, but lipid peroxidation was not reduced. UDCA may, therefore, require supplementation with glutathione precursors and/or antioxidant cocktails to reduce oxidant stress and thus delay disease progression to cirrhosis.

Non-invasive monitoring of oxidant stress in alcoholic liver disease.

OBJECTIVE: In alcoholic liver disease (ALD), progression from initial steatosis, through hepatitis to cirrhosis is well described, resulting in 20,000 deaths in the UK annually. However, pathological mechanisms are not well understood and drug trials have led to conflicting results. It has been established that alcohol consumption increases hepatic free radical production and oxidant stress has been implicated in the disease process. MATERIAL AND METHODS: Markers of lipid peroxidation, antioxidant status, hepatic fibrogenesis, inflammation and liver function were measured in blood and urine from 24 patients with established alcoholic cirrhosis and in 49 age- and sex-matched controls. RESULTS: In the ALD group, lipid peroxidation markers 8-isoprostane and malondialdehyde were significantly increased (p<0.001), as was the ratio of oxidized to reduced glutathione (p=0.027). The antioxidants selenium, glutathione (whole blood and plasma) and vitamins A, C and E were all significantly decreased (p<0.001); median plasma glutathione levels were only 19% of control levels. Type III procollagen peptide (PIIINP), a serum marker of hepatic fibrogenesis, and C-reactive protein (CRP) were both increased (p<0.001). Urinary 8-isoprostane correlated positively with PIIINP, CRP and markers of cholestasis (alkaline phosphatase and bilirubin) and negatively with glutathione (whole blood), vitamins A and E and albumin. CONCLUSIONS: Oxidant stress, as reflected in blood and urine by a wide range of pro- and antioxidant markers, is a significant feature of alcoholic cirrhosis, providing a mechanism by which alcohol intake may be linked to hepatic inflammation and fibrosis. Non-invasive markers could prove valuable in monitoring response to treatment during clinical trials.

Oxidant stress in type I autoimmune hepatitis: the link between necroinflammation and fibrogenesis?

Autoimmune hepatitis (AIH) is a chronic liver disease of unknown aetiology characterized by circulating autoantibodies, hyperglobulinaemia and interface hepatitis. The mechanisms of progression from initial autoimmune attack to fibrosis and cirrhosis are unclear but oxidant stress may be involved. Markers of lipid peroxidation, antioxidant status, hepatic fibrogenesis and liver function were measured in blood and urine in 35 controls and in 33 patients with type-1 AIH; histology was assessed in 18 patients. In AIH, markers of lipid peroxidation were significantly elevated (8-isoprostane in both plasma and urine P < 0.001; plasma malondialdehyde P = 0.017). Total antioxidant capacity in protein-free serum and total glutathione in both whole blood and plasma were significantly reduced (P = 0.007, P = 0.037, P < 0.001, respectively). The antioxidants selenium, vitamin A and vitamin E were significantly decreased (P = 0.007, P < 0.001, P = 0.025, respectively); vitamin C was unchanged. Urinary 8-isoprostane correlated positively with interface hepatitis and necroinflammatory score and with hepatic fibrogenesis (type III procollagen peptide). Interface hepatitis correlated negatively with vitamin A and whole blood total glutathione. Oxidant stress, as reflected in blood and urine by a wide range of pro- and antioxidant markers, is a significant feature of AIH and provides a probable mechanism linking hepatic necroinflammation to fibrogenesis and disease progression.

Oxidant stress is a significant feature of primary biliary cirrhosis.

Primary biliary cirrhosis (PBC) is a chronic cholestatic disorder characterised by an immunological, and often granulomatous, attack on bile ducts leading to fibrosis, cirrhosis, liver failure and death. Animal and human studies suggest that oxidant stress plays a key role in progression of other liver diseases, but no comprehensive investigation has been performed previously in PBC. A wide range of lipid peroxidation and antioxidant markers were measured in the blood and urine of 41 patients with histologically confirmed PBC. Lipid peroxidation markers were significantly elevated [plasma and urinary 8-isoprostane, P<0.001; plasma malondialdehyde (MDA), P=0.007] compared to age- and sex-matched controls. The most striking antioxidant depletion occurred with plasma total glutathione where levels were significantly reduced (30% of controls). Total serum antioxidant levels were decreased (P=0.013) and serum selenium and vitamin A were also lower (both P<0.001); vitamins C and E were normal. Most patients had early disease biochemically and were Child-Pugh grade A. Urinary 8-isoprostane correlated positively with Ludwig stage and markers of hepatic injury and cholestasis. This study clearly demonstrates that oxidant stress, as reflected in a comprehensive spectrum of lipid peroxidation and antioxidant markers, is a significant feature of early-stage PBC.

Oxidative stress in chronic hepatitis C: not just a feature of late stage disease.

BACKGROUND/AIMS: Chronic hepatitis C infection is a major world-wide problem, frequently progressing to cirrhosis, liver failure or hepatoma. The pathological mechanisms of disease progression are unclear but oxidant stress may play a role. METHODS: Markers of lipid peroxidation, antioxidant status, hepatic fibrogenesis and liver function were measured in blood or urine from 42 chronic hepatitis C patients. Fibrosis was graded histologically in a subgroup of 33 patients. RESULTS: The lipid peroxidation marker 8-isoprostane and the ratio of oxidized to reduced glutathione were significantly elevated (P<0.001, P=0.006). The antioxidants glutathione, selenium and vitamins A, C and E were significantly decreased (all P<0.001) compared to age and sex matched controls. Abnormal values were more marked in cirrhotics, but significant changes were also observed in the non-cirrhotic group. The fibrosis score correlated positively with urinary 8-isoprostane and type III procollagen peptide and negatively with vitamin A. CONCLUSIONS: Oxidant stress, as reflected in blood and urine by a wide range of pro- and antioxidant markers, is a significant feature of hepatitis C infection. Although more severe in the cirrhotic group, there was clear evidence of oxidant stress in non-cirrhotic patients. Antioxidant therapy may therefore have a role in slowing disease progression to cirrhosis.