Prevalence of Liver Steatosis in Tuberous Sclerosis Complex Patients: A Retrospective Cross-Sectional Study.

Introduction: Tuberous sclerosis complex (TSC) is a genetic disease caused by pathogenetic variants in either the TSC1 or TSC2 genes. Consequently, the mechanistic target of the rapamycin complex 1 (mTORC1) pathway, a regulator of cell growth, metabolism, and survival, becomes inappropriately activated, leading to the development of benign tumors in multiple organs. The role of mTORC1 in lipid metabolism and liver steatosis in TSC patients has not been well-studied, and clinical data on liver involvement in this population are scarce. Methods: We conducted a retrospective, cross-sectional study to compare liver steatosis in TSC patients with age-, sex-, BMI-, and diabetes status-matched controls. Participants with a definite diagnosis of TSC were recruited from the TSC clinic at UZ Brussel. Liver steatosis was quantified using the fat signal fraction from in-phase and out-of-phase MRI, with a threshold of ≥5% defining the presence of steatosis. We also evaluated the prevalence of liver angiomyolipomata in the TSC group and analyzed risk factors for both liver steatosis and angiomyolipomata. Results: The study included 59 TSC patients and 59 matched controls. The mean fat signal fraction was 4.0% in the TSC group and 3.9% in the controls, showing no significant difference (two-tailed Wilcoxon signed ranks test, p = 0.950). Liver steatosis was observed in 15.3% of TSC patients compared to 23.7% of the controls, which was not statistically significant (two-tailed McNemar test, p = 0.267). Liver angiomyolipomata were identified in 13.6% of the TSC cohort. Conclusions: Our study, describing in detail the liver phenotype of TSC patients, did not reveal a significant difference in the prevalence of MRI-assessed liver steatosis in a large cohort of TSC patients compared to a closely matched control group.

Clinical practice recommendations for kidney involvement in tuberous sclerosis complex: a consensus statement by the ERKNet Working Group for Autosomal Dominant Structural Kidney Disorders and the ERA Genes & Kidney Working Group.

Tuberous sclerosis complex (TSC) is an autosomal dominant disorder characterized by the presence of proliferative lesions throughout the body. Management of TSC is challenging because patients have a multifaceted systemic illness with prominent neurological and developmental impact as well as potentially severe kidney, heart and lung phenotypes; however, every organ system can be involved. Adequate care for patients with TSC requires a coordinated effort involving a multidisciplinary team of clinicians and support staff. This clinical practice recommendation was developed by nephrologists, urologists, paediatric radiologists, interventional radiologists, geneticists, pathologists, and patient and family group representatives, with a focus on TSC-associated kidney manifestations. Careful monitoring of kidney function and assessment of kidney structural lesions by imaging enable early interventions that can preserve kidney function through targeted approaches. Here, we summarize the current evidence and present recommendations for the multidisciplinary management of kidney involvement in TSC.

Understanding the impact of tuberous sclerosis complex: development and validation of the TSC-PROM.

BACKGROUND: Tuberous sclerosis complex (TSC) is a rare and complex genetic disorder, associated with tumor growth in various organ systems, epilepsy, and a range of neuropsychiatric manifestations including intellectual disability. With improving patient-centered care and targeted therapies, patient-reported outcome measures (PROMs) are needed to measure the impact of TSC manifestations on daily functioning. The aim of this study was to develop a TSC-specific PROM for adults that captures the impact of TSC on physical functions, mental functions, activity and participation, and the social support individuals with TSC receive, called the TSC-PROM. METHODS: COSMIN methodology was used to develop a self-reported and proxy-reported version. Development and validation consisted of the following studies: PROM development, content validity, structural validity, internal consistency, and construct validity. The International Classification of Functioning and Disability was used as a framework. Content validity was examined by a multidisciplinary expert group and cognitive interview study. Structural and construct validity, and internal consistency were examined in a large cohort, using confirmatory factor analysis, hypotheses testing, and Cronbach's alpha. RESULTS: The study resulted in an 82-item self version and 75-item proxy version of the TSC-PROM with four subscales (physical functions 18 and 19 items, mental functions 37 and 28 items, activities and participation 13 and 14 items, social support 13 items, for self version and proxy version respectively). Sufficient results were found for structural validity with sufficient unidimensionality for each subscale. With regard to construct validity, 82% of the hypotheses were met for the self version and 59% for the proxy version. The PROM showed good internal consistency (Cronbach's alpha 0.78-0.97). CONCLUSIONS: We developed a PROM for adults with TSC, named TSC-PROM, showing sufficient evidence for reliability and validity that can be used in clinical and research settings to systematically gain insight into their experiences. It is the first PROM in TSC that addresses the impact of specific TSC manifestations on functioning, providing a valuable, patient-centered addition to the current clinical outcomes.

Low agreement between various eGFR formulae in pediatric and young adult ADPKD patients.

BACKGROUND: Young autosomal dominant polycystic kidney disease (ADPKD) patients are becoming the new target population for the development of new treatment options. Determination of a reliable equation for estimated glomerular filtration rate (eGFR) from early stages is needed with the promising potential interventional therapies. METHODS: Prospective and longitudinal study on a cohort of 68 genotyped ADPKD patients (age range 0-23 years) with long-term follow-up. Commonly used equations for eGFR were compared for their relative performance. RESULTS: The revised Schwartz formula (CKiD) showed a highly significant decline in eGFR with aging (- 3.31 mL/min/1.73 m2/year, P < 0.0001). The recently updated equation by the Schwartz group (CKiDU25) showed a smaller (- 0.90 mL/min/1.73 m2/year) but significant (P = 0.001) decline in eGFR with aging and also showed a significant sex difference (P < 0.0001), not observed by the other equations. In contrast, the full age spectrum (FAS) equations (FAS-SCr, FAS-CysC, and the combined) showed no age and sex dependency. The prevalence of hyperfiltration is highly dependent on the formula used, and the highest prevalence was observed with the CKiD Equation (35%). CONCLUSIONS: The most widely used methods to calculate eGFR in ADPKD children (CKiD and CKiDU25 equations) were associated with unexpected age or sex differences. The FAS equations were age- and sex-independent in our cohort. Hence, the switch from the CKiD to CKD-EPI equation at the transition from pediatric to adult care causes implausible jumps in eGFR, which could be misinterpreted. Having reliable methods to calculate eGFR is indispensable for clinical follow-up and clinical trials. A higher resolution version of the Graphical abstract is available as Supplementary information.

On Methods for the Measurement of the Apelin Receptor Ligand Apelin.

Apelin exists in many isoforms, both in the circulation and in specific tissues. Apelin peptides have a short half-life but preservation before measurement is scarcely studied. Reproducible mass spectrometry methods to specifically measure a broad range of apelinergic peptide isoforms are currently lacking. A sample protocol to conserve apelinergic peptides in the preanalytical phase and a high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) method to measure apelinergic isoforms was developed. Apelin was measured in plasma. For validation, human embryonic kidney (HEK) cells transfected with cDNA for preproapelin were used. Results were compared with a validated radioimmunoassay (RIA) method. Acidifying plasma to pH 2.5 improves post-sampling stability of apelin. HPLC-MS/MS was unable to detect apelin isoforms in plasma of healthy volunteers (n = 16) and chronic kidney disease patients (n = 4). RIA could detect apelin in concentrations between 71 and 263 fmol/l in 10 healthy volunteers. An optimized preanalytical protocol was developed. A sensitive and specific HPLC-MS/MS method failed to detect apelin in human plasma. Apelin-36 was detected in HEK cells transfected with cDNA for preproapelin. Currently, RIA with relatively selective antibodies is the best alternative for the measurement of apelin but novel sensitive and specific methods are needed.

Interdependent Regulation of Polycystin Expression Influences Starvation-Induced Autophagy and Cell Death.

Autosomal dominant polycystic kidney disease (ADPKD) is mainly caused by deficiency of polycystin-1 (PC1) or polycystin-2 (PC2). Altered autophagy has recently been implicated in ADPKD progression, but its exact regulation by PC1 and PC2 remains unclear. We therefore investigated cell death and survival during nutritional stress in mouse inner medullary collecting duct cells (mIMCDs), either wild-type (WT) or lacking PC1 (PC1KO) or PC2 (PC2KO), and human urine-derived proximal tubular epithelial cells (PTEC) from early-stage ADPKD patients with PC1 mutations versus healthy individuals. Basal autophagy was enhanced in PC1-deficient cells. Similarly, following starvation, autophagy was enhanced and cell death reduced when PC1 was reduced. Autophagy inhibition reduced cell death resistance in PC1KO mIMCDs to the WT level, implying that PC1 promotes autophagic cell survival. Although PC2 expression was increased in PC1KO mIMCDs, PC2 knockdown did not result in reduced autophagy. PC2KO mIMCDs displayed lower basal autophagy, but more autophagy and less cell death following chronic starvation. This could be reversed by overexpression of PC1 in PC2KO. Together, these findings indicate that PC1 levels are partially coupled to PC2 expression, and determine the transition from renal cell survival to death, leading to enhanced survival of ADPKD cells during nutritional stress.

Simultaneous determination of allantoin and adenosine in human urine using liquid chromatography - UV detection.

We report a HPLC-UV method for the quantitative determination of allantoin and adenosine in human urine, validated according to the acceptance criteria of both the USA Food and Drug Administration (FDA) guideline for bioanalytical method validation and the European Medicines Agency (EMA) validation guidelines. Both allantoins and adenosine are compounds of the purine catabolic pathway. Adenosine is situated at the top as a uric acid (UA) precursor, while allantoin is the best-known degradation product of UA. These two compounds are endogenously present in human urine. Chromatographic separation was achieved with a gradient elution at 0.6 mL/min using a Zorbax SB-Aq column coupled to a Zorbax SB-Aq guard column. Three different mobile phases were used: mobile phase A consisted of 10 mM KH2PO4 (pH 4.7) in milli-Q water, mobile phase B was 12.5 mM KH2PO4 (pH 4.7) - ACN (80:20) and mobile phase C consisted of ACN - H2O (50:50). The linear response range in human urine was 14-800 µM for allantoin and 1.25-50 µM for adenosine. The recoveries of allantoin, adenosine and the internal standard were greater than 93.8%. The intra-day accuracy ranged between 99.5 and 104.9% for allantoin and between 96.6 and 107.3% for adenosine, while the inter-day accuracy ranged respectively from 91.2 to 103.0% and from 94.5 to 107.8%. The intra-day precision range was from 0.8 to 6.2% RSD for allantoin and from 0.6 to 15.0% for adenosine. The inter-day precision ranged from 2.1-17.5% for allantoin and from 2.9-17.9% for adenosine. This method was successfully applied to analyze both compounds in urine samples of healthy volunteers. In conclusion, an accurate, precise and stable HPLC-UV method was developed and validated to quantify the endogenously present compounds allantoin and adenosine in human urine samples.

Renal progression factors in young patients with tuberous sclerosis complex: a retrospective cohort study.

BACKGROUND: Renal pathology in tuberous sclerosis complex (TSC) is characterized by the growth of angiomyolipoma and renal cysts, and in rare cases renal cell carcinoma. Other consequences of renal involvement in TSC, including hypertension, proteinuria, and hyperfiltration, are not well studied. We aimed to analyze the early manifestations of the renal TSC phenotype in a young TSC cohort and to explore common, modifiable risk factors. METHODS: In this retrospective cohort study, TSC patients attending the TSC clinics of two tertiary hospitals were included. Data on demographics, history, genotype, kidney function, hematuria, proteinuria, blood pressure, and renal imaging were collected. RESULTS: Eighty patients were included, with a median age of 0.8 years (0.0-63.0) at first presentation, and a median follow-up time of 10.2 (0.4-41.0) years. Mutation analysis was available in 64 patients (80%). Renal lesions (cysts or angiomyolipoma) were observed in 55/73 (75%). Thirty-two percent (19/60) were hypertensive, 8/51 (16%) had proteinuria, and 18/71 (25%) had hyperfiltration (median eGFR 154 ml/min/m2). Six (7.5%) patients had developed end stage renal disease at the last follow-up. No association was found between hyperfiltration, hypertension, or proteinuria and CKD ≥ 3. Cox regression showed a significant positive association between the presence of a renal intervention and CKD ≥ 3 (Hazard-Ratio 3.91, P < 0.05). CONCLUSIONS: Besides renal cysts and angiomyolipoma, the modifiable progression factors hypertension, proteinuria, and hyperfiltration occur frequently and early in TSC patients. This represents a preventive treatment target.

Central nervous system gadolinium accumulation in patients undergoing periodical contrast MRI screening for hereditary tumor syndromes.

BACKGROUND: Patients with hereditary tumor syndromes undergo periodical magnetic resonance imaging (MRI) screening with Gadolinium contrast. Gadolinium accumulation has recently been described in the central nervous system after repeated administrations. The prevalence and rate of accumulation in different subgroups of patients are unknown. Neither are the mechanism nor clinical impact. This may cause uncertainty about the screening. To explore the prevalence and rate of Gadolinium accumulation in different subgroups, we retrospectively analyzed MRIs of patients with von Hippel-Lindau disease (VHL) and Tuberous Sclerosis Complex (TSC). METHODS: We determined the prevalence and rate of accumulation in the dentate nucleus and globus pallidus on unenhanced T1-weighted MRI from VHL and TSC patients. We compared the signal intensities of these regions to the signal intensity of the pons. We evaluated the impact of number of MRIs, kidney function and liver function on Gadolinium accumulation. RESULTS: Twenty eight VHL patients and 24 TSC patients were included. The prevalence of accumulation in the dentate nucleus and globus pallidus increased linearly according to number of Gadolinium enhanced MRIs and was higher in the VHL group (100%). A significant linear correlation between number of MRIs and increased signal intensity was observed in the VHL group. CONCLUSIONS: Gadolinium accumulation occurs in almost all patients undergoing contrast MRI screening after >5 MRIs. We advocate a screening protocol for patients with hereditary tumor syndromes that minimizes the Gadolinium dose. This can be accomplished by using a single administration to simultaneously screen for brain, spine and/or abdominal lesions, using an MRI protocol focused on either VHL- or TSC-specific lesions. Higher prevalence and rate of accumulation in VHL patients may be explained by the typical vascular leakage accompanying central nervous system hemangioblastomas.

Expanding the role of vasopressin antagonism in polycystic kidney diseases: From adults to children?

Polycystic kidney disease (PKD) encompasses a group of genetic disorders that are common causes of renal failure. The two classic forms of PKD are autosomal recessive polycystic kidney disease (ARPKD) and autosomal dominant polycystic kidney disease (ADPKD). Despite their clinical differences, ARPKD and ADPKD share many similarities. Altered intracellular Ca2+ and increased cyclic adenosine monophosphate (cAMP) concentrations have repetitively been described as central anomalies that may alter signaling pathways leading to cyst formation. The vasopressin V2 receptor (V2R) antagonist tolvaptan lowers cAMP in cystic tissues and slows renal cystic progression and kidney function decline when given over 3 years in adult ADPKD patients. Tolvaptan is currently approved for the treatment of rapidly progressive disease in adult ADPKD patients. On the occasion of the recent initiation of a clinical trial with tolvaptan in pediatric ADPKD patients, we aim to describe the most important aspects in the literature regarding the AVP-cAMP axis and the clinical use of tolvaptan in PKD.

An unexpected complication of chronic myelomonocytic leukemia: severe renal failure due to malignant tubulo-interstitial cell infiltration.

Acute renal failure may complicate the course of a hematologic malignancy but is a highly unusual finding in patients with chronic myelomonocytic leukemia. Kidney biopsy is rarely performed in this setting, and the pathologic substrate underlying kidney injury is not well identified. We present a case of a biopsy-proven acute tubulo-interstitial nephritis due to massive infiltration of neoplastic myelomonocytic cells. Since the leukemic process involving the kidney may respond favorably to treatment, a renal biopsy should be considered in any patient presenting with unexplained severe or evolving kidney disease.

Granulomatous nephritis and dermatitis in a patient with BRAF V600E mutant metastatic melanoma treated with dabrafenib and trametinib.

A 61-year-old man was diagnosed with stage IIIB BRAF V600E mutant melanoma in October 2012. He was treated with a combination therapy of dabrafenib and trametinib. He remained in complete remission for 18 months and the treatment was well tolerated after dose reduction because of pyrexia. In March 2013, he developed bilateral pitting edema of the legs with an erythematous, slightly infiltrated rash on his back and upper arms. His face was edematous, with a heliotrope rash-like aspect. Eye examination showed bilateral blepharitis. Additional blood test showed inflammation and acute kidney injury Rifle category failure. A skin and kidney biopsy indicated a granulomatous inflammation. A complete workup for other causes of granulomatous inflammation was negative. Treatment with dabrafenib and trametinib was stopped and corticosteroids were initiated, with a rapid beneficial effect on both the kidney function and skin rash. When corticosteroids were halted after 1 month, a rapid decline in the kidney function was observed. After reintroduction of corticosteroids, kidney function normalized and steroids could be tapered gradually over 6 months. To our knowledge, interstitial nephritis has not been described in patients on BRAF-targeted nor MEK-targeted therapy for melanoma, although it has been described in a melanoma patient treated with the immune checkpoint inhibitor, ipilimumab. Currently, the patient has no sign of local or distal recurrence of melanoma, notwithstanding that treatment with dabrafenib and trametinib has been stopped for 10 months and no other antimelanoma therapy was initiated.

Immune response in rabbit ovaries following infection of a recombinant myxoma virus expressing rabbit zona pellucida protein B.

In this study, we investigated the autoimmune response in rabbit ovaries following infection with a recombinant myxoma virus expressing rabbit zona pellucida protein B (MV-ZPB). A specific IgG antibody response to ZPB was elicited in the serum of infected rabbits and the antibody strongly bound to the zona pellucida of oocytes in secondary and tertiary follicles. T cell infiltration in the ovary was detected in a small proportion of the infected rabbits. In spite of this, the mean number of preovulatory and tertiary follicles in the ovary was significantly reduced at 30 days postinfection compared with that of the infected and uninfected controls. Histological analysis revealed that the cortex and medulla of these ovaries had accumulated a large number of probably luteinized cells and there were no follicles in these areas, indicating the ovaries were in a severe pathological condition. The data suggest that the delivery of ZP antigens using a recombinant myxoma virus is a prospective way to develop immunocontraceptive vaccines for rabbit population control, but that more understanding of the kinetics of the autoimmune response induced by viral delivery is needed.

Antibody response in the female rabbit reproductive tract to influenza haemagglutinin encoded by a recombinant myxoma virus.

The antibody response in serum and the reproductive tract of female rabbits to a model antigen, influenza virus haemagglutinin (HA), encoded by a recombinant myxoma virus was investigated. Strong and lasting IgG antibody responses to HA were induced in serum following intradermal, intranasal, and intravaginal immunisations. HA IgG was also detected in reproductive tract fluids but was only about 1% the titer of that in serum. HA IgA was not detected in serum of any infected groups and was occasionally detected in reproductive tract fluids at a low titer only after infections through mucosal sites. HA IgM was also detected only in some of the reproductive tract fluids at very low levels. Induction of ovulation did not change these patterns and B cell homing to the reproductive tract was not profound. In contrast, HA IgG and IgM titers in ovarian follicular fluids were comparable to that in serum. These data suggest that if this virus is used to deliver an immunocontraceptive vaccine that requires a high-level antibody response, the target antigen needs to be accessible to serum antibody or in the ovary.