Characterization of a Biochemical Mouse Model of Primary Aldosteronism for Thermal Therapies.

Introduction: Aldosterone-producing adenoma (APA) is the most common cause of endocrine-related hypertension but surgery is not always feasible. Current medical interventions are associated with significant side effects and poor patient compliance. New APA animal models that replicate basic characteristics of APA and give physical and biochemical feedback are needed to test new non-surgical treatment methods, such as image-guided thermal ablation. Methods: A model of APA was developed in nude mice using HAC15 cells, a human adrenal carcinoma cell line. Tumor growth, aldosterone production, and sensitivity to angiotensin II were characterized in the model. The utility of the model was validated via treatment with microwave ablation and characterization of the resulting physical and biochemical changes in the tumor. Results: The APA model showed rapid and relatively homogeneous growth. The tumors produced aldosterone and steroid precursors in response to angiotensin II challenge, and plasma aldosterone levels were significantly higher in tumor bearing mice two hours after challenge verses non-tumor bearing mice. The model was useful for testing microwave ablation therapy, reducing aldosterone production by 80% in treated mice. Conclusion: The HAC15 model is a useful tumor model to study and develop localized treatment methods for APA.

A cost-analysis of managing secondary and apparent treatment-resistant hypertension in a specialist multidisciplinary hypertension clinic.

OBJECTIVES: A knowledge gap exists around the costs and budget impact of specialist hypertension clinics. This study reports on the cost of providing care in a multidisciplinary hypertension clinic staffed by nephrologist, endocrinologist and cardiologist, which manages patients with suspected secondary hypertension and/or apparent treatment-resistant hypertension. The aim of this study is to provide the evidence required to inform policy and planning care pathways for this patient group. METHODS: A cost analysis from a healthcare provider perspective using micro-costing techniques was conducted to estimate the direct implementation costs of existing standard practice for the care pathway of patients attending the multidisciplinary hypertension clinic. Sixty-five patients originally recruited for a study of medication adherence in hypertension were included in the sample. RESULTS: The total care-pathway cost per patient, taking into account clinic visits, clinical reviews, investigations and MDT discussion, was estimated to be €3277, on average. For the patient subgroups, the average cost was €5644 for patients diagnosed with primary aldosteronism and €1446 for patients diagnosed with essential hypertension. CONCLUSION: There is significant cost associated with providing specialized hypertension care for patients with apparent treatment-resistant hypertension. Given the high rates of nonadherence in this population, it is likely that some of this cost could be avoided with better detection and management of medication adherence in this challenging population. Future studies should consider the cost-effectiveness of this or similar models of care by exploring the benefit to patients and the wider healthcare context of providing care of this type.

A Case of Aplasia Cutis Congenita in the Setting of Maternal Carbimazole Use in the First Trimester.

Aplasia cutis congenita (ACC) is one of several congenital malformations associated with antithyroid/thiourylene drug use in pregnancy. While uncommon among the general population (1-3/100 000 cases), the risk among those on thiourylenes is between 1.6% and 3%. The scalp is the most common site for this congenital anomaly. We present the case of a male infant with multifocal ACC of the scalp discovered at birth and born to a mother with Graves disease that was controlled during pregnancy using carbimazole. Thyroid function tests were normal throughout the pregnancy. There was no involvement of underlying subcutaneous tissue or structures. At age 18 months, the single largest lesion remained with only partial coverage. Prospective management involved periodic surveillance with planned 2-stage repair. This case reinforces the association between the antithyroid drugs carbimazole (CMZ) and methimazole (MMI) and supports the proposition of an MMI/CMZ embryopathy. It adds to a literature of case reports in which malformations arise in offspring of such mothers whose thyrotoxicosis is controlled antenatally, thereby challenging the suggestion that ACC is attributable to poorly controlled disease rather than thiourylenes. As yet the underlying mechanism is not understood, nor is it known why MMI and CMZ may cause potentially significant embryopathy while congenital defects attributable to the structurally similar propylthiouracil are typically less severe.

Major disparities in patient-reported adherence compared to objective assessment of adherence using mass spectrometry: A prospective study in a tertiary-referral hypertension clinic.

AIM: Many challenges exist in determining true rates of adherence to antihypertensive medications among individuals in a clinic setting. For the first time, we aimed to compare patient-reported antihypertensive adherence with objective evidence using mass spectrometry spot urinalysis in a tertiary referral clinic setting. METHODS: A prospective observational single-centre cohort study was performed in a tertiary referral hypertension clinic, encompassing antihypertensive initiation and persistence. Patients were referred with apparent treatment-resistant hypertension or for suspected secondary causes. Participants completed a self-reported assessment of antihypertensive adherence and provided a spot urine sample. The presence of antihypertensive medications and/or their respective metabolites was evaluated using high-performance liquid chromatography tandem mass spectrometry. Patients were determined to be adherent if they demonstrated both self-reported adherence and objective mass spectrometry evidence. RESULTS: Of all 105 eligible participants initially recruited, 73 (69.5%) met the eligibility criteria. Only 27.4% (95% confidence interval 0.2-0.4) of participants demonstrated true adherence to their self-reported antihypertensives, despite 75.3% (0.6-0.8) reporting adherence. Greatest medication adherence was achieved with angiotensin II receptor blockers (61%), with calcium-channel blockers and mineralocorticoid antagonists demonstrating least adherence (38%). CONCLUSION: In patients attending a tertiary hypertension clinic, the combined use of spot urine mass spectrometry and self-reporting identifies higher rates of nonadherence when compared to either modality alone. Both techniques should be combined for more accurate detection of medication adherence.

Mitotane Targets Lipid Droplets to Induce Lipolysis in Adrenocortical Carcinoma.

INTRODUCTION: Adrenocortical carcinoma (ACC) is a rare aggressive cancer with low overall survival. Adjuvant mitotane improves survival but is limited by poor response rates and resistance. Mitotane's efficacy is attributed to the accumulation of toxic free cholesterol, predominantly through cholesterol storage inhibition. However, targeting this pathway has proven unsuccessful. We hypothesize that mitotane-induced free-cholesterol accumulation is also mediated through enhanced breakdown of lipid droplets. METHODOLOGY: ATCC-H295R (mitotane-sensitive) and MUC-1 (mitotane-resistant) ACC cells were evaluated for lipid content using specific BODIPY dyes. Protein expression was evaluated by immunoblotting and flow cytometry. Cell viability was measured by quantifying propidium iodide-positive cells following mitotane treatment and pharmacological inhibitors of lipolysis. RESULTS: H295R and MUC-1 cells demonstrated similar neutral lipid droplet numbers at baseline. However, evaluation of lipid machinery demonstrated distinct profiles in each model. Analysis of intracellular lipid droplet content showed H295R cells preferentially store cholesteryl esters, whereas MUC-1 cells store triacylglycerol. Decreased lipid droplets were associated with increased lipolysis in H295R and in MUC-1 at toxic mitotane concentrations. Pharmacological inhibition of lipolysis attenuated mitotane-induced toxicity in both models. CONCLUSION: We highlight that lipid droplet breakdown and activation of lipolysis represent a putative additional mechanism for mitotane-induced cytotoxicity in ACC. Further understanding of cholesterol and lipids in ACC offers potential novel therapeutic exploitation, especially in mitotane-resistant disease.

Thermal ablation in adrenal disorders: a discussion of the technology, the clinical evidence and the future.

PURPOSE OF REVIEW: To summarise the emerging role of thermal ablation as a therapeutic modality in the management of functioning adrenal tumours and metastases to the adrenal gland. RECENT FINDINGS: Observational evidence has demonstrated the benefit of thermal ablation in (i) resolving adrenal endocrinopathy arising from benign adenomas, (ii) treating solitary metastases to the adrenal and (iii) controlling metastatic adrenocortical carcinoma and phaeochromocytoma/paraganglioma. SUMMARY: Microwave thermal ablation offers a promising, minimally invasive therapeutic modality for the management of functioning adrenocortical adenomas and adrenal metastases. Appropriate technological design, treatment planning and choice of imaging modality are necessary to overcome technical challenges associated with this emerging therapeutic approach.

ENSAT registry-based randomized clinical trials for adrenocortical carcinoma.

Adrenocortical carcinoma (ACC) is an orphan disease lacking effective systemic treatment options. The low incidence of the disease and high cost of clinical trials are major obstacles in the search for improved treatment strategies. As a novel approach, registry-based clinical trials have been introduced in clinical research, so allowing for significant cost reduction, but without compromising scientific benefit. Herein, we describe how the European Network for the Study of Adrenal Tumours (ENSAT) could transform its current registry into one fit for a clinical trial infrastructure. The rationale to perform randomized registry-based trials in ACC is outlined including an analysis of relevant limitations and challenges. We summarize a survey on this concept among ENSAT members who expressed a strong interest in the concept and rated its scientific potential as high. Legal aspects, including ethical approval of registry-based randomization were identified as potential obstacles. Finally, we describe three potential randomized registry-based clinical trials in an adjuvant setting and for advanced disease with a high potential to be executed within the framework of an advanced ENSAT registry. Thus we, therefore, provide the basis for future registry-based trials for ACC patients. This could ultimately provide proof-of-principle of how to perform more effective randomized trials for an orphan disease.

Liver X receptor inhibition potentiates mitotane-induced adrenotoxicity in ACC.

Adrenocortical carcinoma (ACC) is a rare aggressive malignancy with a poor outcome largely due to limited treatment options. Here, we propose a novel therapeutic approach through modulating intracellular free cholesterol via the liver X receptor alpha (LXRα) in combination with current first-line pharmacotherapy, mitotane. H295R and MUC-1 ACC cell lines were pretreated with LXRα inhibitors in combination with mitotane. In H295R, mitotane (20, 40 and 50 µM) induced dose-dependent cell death; however, in MUC-1, this only occurred at a supratherapeutic concentration (200 µM). LXRα inhibition potentiated mitotane-induced cytotoxicity in both cell lines. This was confirmed through use of the CompuSyn model which showed moderate pharmacological synergism and was indicative of apoptotic cell death via an increase in annexinV and cleaved-caspase 3 expression. Inhibition of LXRα was confirmed through downregulation of cholesterol efflux pumps ABCA1 and ABCG1; however, combination treatment with mitotane attenuated this effect. Intracellular free-cholesterol levels were associated with increased cytotoxicity in H295R (r2 = 0.5210) and MUC-1 (r2 = 0.9299) cells. While both cell lines exhibited similar levels of free cholesterol at baseline, H295R were cholesterol ester rich, whereas MUC-1 were cholesterol ester poor. We highlight the importance of LXRα mediated cholesterol metabolism in the management of ACC, drawing attention to its role in the therapeutics of mitotane sensitive tumours. We also demonstrate significant differences in cholesterol storage between mitotane sensitive and resistant disease.

Using microwave thermal ablation to develop a subtotal, cortical-sparing approach to the management of primary aldosteronism.

Objective: To investigate the feasibility and efficacy of localized, subtotal, cortical-sparing microwave thermal ablation (MTA) as a potential curative management for primary aldosteronism. The study investigated with equal importance the selected ablation of small volumes of adrenal cortex while sparing adjacent cortex. Method: An in-vivo study was carried out in swine (n = 8) where MTA was applied under direct visualization, to the adrenal glands at 45 W or 70 W for 60 s, using a lateral, side-firing probe and a non-penetrative approach. Animals were survived for 48 h post-procedurally. Animals were investigated for markers of histological, immunohistochemical and biochemical evidence of adrenal function and adrenal damage by assessing samples drawn intra-operatively and at the time of euthanasia. Results: Selected MTA (70 W for 60 s) successfully ablated small adrenocortical volumes (∼0.8 cm3) characterized by coagulative necrosis and abnormal expression of functional markers (CYP11B1 and CYP17). Non-ablated, adjacent cortex was not affected and preserved normal expression of functional markers, without increased expression of markers of heat damage (HSP-70 and HMGB-1). Limited adrenal medullary damage was demonstrated histologically, clinically and biochemically. Conclusion: MTA offers potential as an efficient methodology for delivering targeted subtotal cortical-sparing adrenal ablation. Image-guided targeted MTA may also represent a safe future modality for curative management of PA, in the setting of both unilateral and bilateral disease.

Infliximab Selectively Modulates the Circulating Blood Monocyte Repertoire in Crohn's Disease.

BACKGROUND: Infliximab (IFX), an anti-tumour necrosis factor alpha (TNFα) monoclonal antibody, provides clinical benefits in treating Crohn's disease (CD) but its mechanisms of action are not fully elucidated. This study investigated blood monocyte repertoires and the acute effects of IFX infusion on monocyte subset phenotype and function in IFX-treated patients with CD. METHODS: Monocytes and monocyte subsets were enumerated and phenotypically characterized by multicolor flow cytometry in freshly isolated blood from healthy controls (n = 21) and patients with CD treated with (IFX, n = 24) and without (non-IFX, n = 20) IFX. For the IFX-CD group, blood was sampled immediately before (tough-IFX) and after (peak-IFX) infusion. Monocyte responses to lipopolysaccharide were analyzed by whole-blood intracellular cytokine staining. RESULTS: Non-IFX and IFX-CD patients had increased numbers of intermediate (CD14CD16) monocytes compared with healthy controls, whereas classical (CD14CD16) and nonclassical (CD14CD16) monocytes were numerically reduced in the IFX-CD group alone. In all groups, monocyte subsets expressed high surface levels of transmembrane (tm)TNFα. After IFX infusion, a significant reduction in monocyte numbers occurred. Post-IFX monocytopenia was proportionately greatest for classical and intermediate subsets, correlated with postinfusion IFX levels and was not associated with monocyte apoptosis. In contrast, lipopolysaccharide-induced production of TNFα and IL-12 by monocytes was significantly reduced in peak-IFX compared with trough-IFX blood samples. CONCLUSIONS: Actively managed CD is associated with monocyte repertoire skewing suggestive of chronic inflammatory stimulation. Infused IFX acutely targets monocytes, likely by binding to tmTNFα, resulting in a non-apoptosis-related decline in circulating monocyte numbers and blunting of the inflammatory response of monocytes remaining in the blood.

ß-Blocker withdrawal is preferable for accurate interpretation of the aldosterone-renin ratio in chronically treated hypertension.

OBJECTIVES: To evaluate the effects of ß-adrenoreceptor antagonists (ß-blockers) on the aldosterone-renin ratio (ARR) in the context of antihypertensive polypharmacy in chronic hypertension. To determine the optimal duration of ß-blocker withdrawal required to normalize the ARR. DESIGN: A prospective, longitudinal study design was employed investigating two groups whom either remained on or withdrew from ß-blocker therapy. METHODS: Hypertensive individuals taking ß-blockers and a combination of thiazide diuretics, α1-blockers, calcium channel antagonists and angiotensin-converting enzyme inhibitor/angiotensin receptor blocker were recruited and followed over 10-12 weeks. ß-Blockers were withdrawn at the first visit. Blood pressure (BP) was measured at each visit and blood drawn serially for measurement of plasma renin activity (PRA), direct renin concentration (DRC) and aldosterone. BP was optimized by maximizing non-renin-suppressing antihypertensives. Main outcomes were ARR, DRC, PRA and aldosterone. Plasma renin activity was calculated from angiotensin I measured using radioimmunoassay (RIA), DRC was measured using chemiluminescent immunoassay assay, and aldosterone was measured using both RIA and Chemilluminescence Assay (CIL). RESULTS: False-positive ARR for primary aldosteronism (PA) occurred in 31% of patients taking ß-blockers. ARR returned to normal following ß-blocker withdrawal resulting from an increase in the DRC and PRA without affecting aldosterone. The optimum time for ß-blocker withdrawal was 2 weeks when using DRC and 3 weeks for PRA. ß-Blocker withdrawal did not adversely affect blood pressure. CONCLUSION: Raised ARR consequent to ß-blocker therapy causes false-positive screening for PA. Where ß-blockers can be safely withdrawn, this effect is reversed within 2-3 weeks depending on whether DRC or PRA is used to calculate ARR.

ATLANTIC-DIP: raised maternal body mass index (BMI) adversely affects maternal and fetal outcomes in glucose-tolerant women according to International Association of Diabetes and Pregnancy Study Groups (IADPSG) criteria.

CONTEXT: Raised maternal body mass index (BMI) in association with hyperglycemia is associated with adverse pregnancy outcome. The contribution of raised BMI as an independent risk factor for adverse pregnancy outcome is of growing concern and increasing prevalence. OBJECTIVE: The aim of this study was to investigate the effects of raised maternal BMI on pregnancy outcome in glucose-tolerant women using the International Association of Diabetes and Pregnancy Study Groups criteria. PARTICIPANTS AND SETTING: We studied a cohort of glucose-tolerant, pregnant women (n = 3656) who were attending antenatal obstetric clinics and were recruited to a universal screening program for gestational diabetes under the ATLANTIC-DIP partnership. DESIGN: We conducted a prospective observational study of pregnancy outcome. Maternal outcomes include glucose, delivery mode, pregnancy-induced hypertension, preeclampsia, antepartum hemorrhage, and postpartum hemorrhage. Fetal outcomes included birthweight, congenital malformation, fetal death, neonatal jaundice, hypoglycemia, and respiratory distress. RESULTS: Increasing maternal BMI was associated with adverse pregnancy outcomes: higher cesarean section rates, preeclampsia, pregnancy-induced hypertension, increased birth weight, and congenital malformation. The association of glucose with adverse pregnancy outcome was weak and did not interact with raised BMI. A BMI threshold of 28 kg/m(2) was associated with a significant rise in adverse pregnancy outcome. CONCLUSIONS: Raised maternal BMI, within the overweight range, is associated with adverse pregnancy outcomes. These adverse effects of BMI occur independently of maternal glucose. It is apparent that pregnancy unmasks an underlying unhealthy metabolic milieu in obese and overweight women.

The maternal and fetal impacts of obesity and gestational diabetes on pregnancy outcome.

Obesity has reached pandemic proportions and is of growing concern worldwide. Adverse health outcomes associated with a raised body mass index present the greatest challenge currently facing clinicians across all disciplines. Obesity is a chronic illness which is associated with metabolic disease, nutritional deficiency, musculoskeletal complications and cancer. These obesity-related health issues extend to pregnancy where they are responsible for producing a variety of medical and obstetric complications resulting in an increased incidence of maternal and fetal adverse outcomes. Management of diet, gestational diabetes and gestational and inter-gestational weight may improve outcomes in women who are obese during pregnancy. Specific recommendations for the management of obesity in pregnancy have recently been published.

Investigation of patients with atypical or severe hyperandrogenaemia including androgen-secreting ovarian teratoma.

Approximately 7% of women of reproductive age manifest polycystic ovary syndrome (PCOS) and <0.5% have other causes of hyperandrogenism including congenital adrenal hyperplasia (CAH), androgen-secreting tumour of an ovary or an adrenal gland, Cushing's syndrome or hyperthecosis. The presence of features atypical of PCOS should prompt more extensive evaluation than that usually undertaken. Features atypical of PCOS include the onset of symptoms outside the decade of 15-25 years, rapid progression of symptoms, the development of virilization and a serum testosterone concentration in excess of twice the upper limit of the reference range. Ethnic background, family history and specific clinical findings, e.g. Cushingoid appearance, may inform a focused investigation. Otherwise, patients should have measurement of 17-hydroxyprogesterone (17-OHP) under basal conditions ideally in the early morning, and if abnormal, they should have measurement of 17-OHP one hour after the administration of synthetic ACTH, 250 microg i.v., to screen for CAH, which is present in approximately 2% of hyperandrogenic patients. The overnight cortisol suppression test employing 1 mg dexamethasone at midnight is a sensitive test for Cushing's syndrome. Coronal tomographic (CT) scanning of the adrenals and transvaginal ultrasonography of the ovaries are the investigations of choice when screening for tumours in these organs. Less frequently required is catheterization and sampling from both adrenal and ovarian veins, which is a technically demanding procedure with potential complications which may provide definitive diagnostic information not available from other investigations. Illustrative case reports highlight some complexities in the investigation of hyperandrogenic patients presenting with features atypical of PCOS and include only the ninth case report of an androgen-secreting ovarian teratoma.