Identifying patient-related predictors of permanent growth hormone deficiency.

Objective: Isolated childhood growth hormone deficiency (GHD) can persist into adulthood, and re-testing at the transition period is needed to determine whether continued growth hormone therapy is indicated. Here, our objective was to identify predictors of permanent GHD. Design: Retrospective single-centre study of patients with childhood-onset GHD who were re-tested after adult height attainment. Methods: Auxological, clinical, laboratory, and MRI data throughout follow-up were collected. Results: We included 101 patients. At GH treatment initiation, age was 8.1 ± 0.4 years, height -2.25 ± 0.8, and BMI -0.27 ± 0.1 SDS. The 29 (28.7%) patients with persistent GHD had lower height SDS (-2.57 ± 0.1 vs. -2.11 ± 0.1, p<0.001) and mean GH peaks (8.4 ± 1.0 vs.13.2 ± 0.5 mIU/L, p<0.001) at GHD diagnosis; at adult height, they had lower IGF1 (232 ± 19.9 vs. 331 ± 9.1 ng/mL, p<0.001) and higher BMI SDS (-0.15 ± 0.27 vs. -0.73 ± 0.13, p<0.005). By multivariate analysis, the best predictive model included height and BMI SDS, both GH peaks, and MRI findings at diagnosis. Patients with height at diagnosis <-3 SDS had a 7.7 (95% IC 1.4-43.1, p=0.02) fold higher risk of persistent GHD after adjustment on BMI SDS. An abnormal pituitary region by MRI was the strongest single predictor (7.2 times, 95% CI 2.7-19.8) and after multivariate analysis adjustment for GH peaks and height SDS at diagnosis, the risk increased to 10.6 (1.8 - 61.3) times. Conclusions: Height <-3 SDS at GHD diagnosis and pituitary MRI abnormalities should lead to a high index of suspicion for persistent GHD.

Hormone Immunoassay Interference: A 2021 Update.

Immunoassays are powerful qualitative and quantitative analytical techniques. Since the first description of an immunoassay method in 1959, advances have been made in assay designs and analytical characteristics, opening the door for their widespread implementation in clinical laboratories. Clinical endocrinology is closely linked to laboratory medicine because hormone quantification is important for the diagnosis, treatment, and prognosis of endocrine disorders. Several interferences in immunoassays have been identified through the years; although some are no longer encountered in daily practice, cross-reaction, heterophile antibodies, biotin, and anti-analyte antibodies still cause problems. Newer interferences are also emerging with the development of new therapies. The interfering substance may be exogenous (e.g., a drug or substance absorbed by the patient) or endogenous (e.g., antibodies produced by the patient), and the bias caused by interference can be positive or negative. The consequences of interference can be deleterious when clinicians consider erroneous results to establish a diagnosis, leading to unnecessary explorations or inappropriate treatments. Clinical laboratories and manufacturers continue to investigate methods for the detection, elimination, and prevention of interferences. However, no system is completely devoid of such incidents. In this review, we focus on the analytical interferences encountered in daily practice and possible solutions for their detection or elimination.

Influence of everolimus-based treatment on circulating regulatory T cells after liver transplantation: Comparative study with tacrolimus-based therapy.

Liver transplantation remains the only treatment for terminal liver diseases. However, immunosuppressive drugs required for allograft acceptance are toxic and may be responsible for severe side effects. Modulating the immune system to induce tolerance is a promising approach to reduce immunosuppressive regimen. More particularly, promoting natural CD4+ CD25+ FoxP3+ Tregs could be crucial in achieving tolerance. Contrary to calcineurin inhibitors, reports indicate that mTOR inhibitors may have a positive impact on Tregs. Here we present the first randomized prospective clinical study where Tregs levels from liver transplanted patients receiving either tacrolimus or everolimus were monitored for 6 months, starting from the day of transplantation. A total of 30 patients from four centers were monitored. Blood samples were obtained at day 0, day 14, one month, three months and six months post-transplantation. Flow-cytometry immunophenotyping of Tregs (CD4+ CD25+ CD127- FoxP3+) and functional assays with Tregs were performed to assess their immunosuppressive capacity. Levels of Tregs were significantly reduced after one month of standard tacrolimus-based immunosuppressive regimen (p<0.05). Four months after conversion, levels of Tregs from patients treated with everolimus was significantly higher than patients under tacrolimus (p<0.02). Functional assays demonstrated that Tregs conserved their capacity to suppress the proliferation of activated PBMC.

Exome sequencing for diagnosis of congenital hemolytic anemia.

BACKGROUND: Congenital hemolytic anemia constitutes a heterogeneous group of rare genetic disorders of red blood cells. Diagnosis is based on clinical data, family history and phenotypic testing, genetic analyses being usually performed as a late step. In this study, we explored 40 patients with congenital hemolytic anemia by whole exome sequencing: 20 patients with hereditary spherocytosis and 20 patients with unexplained hemolysis. RESULTS: A probable genetic cause of disease was identified in 82.5% of the patients (33/40): 100% of those with suspected hereditary spherocytosis (20/20) and 65% of those with unexplained hemolysis (13/20). We found that several patients carried genetic variations in more than one gene (3/20 in the hereditary spherocytosis group, 6/13 fully elucidated patients in the unexplained hemolysis group), giving a more accurate picture of the genetic complexity of congenital hemolytic anemia. In addition, whole exome sequencing allowed us to identify genetic variants in non-congenital hemolytic anemia genes that explained part of the phenotype in 3 patients. CONCLUSION: The rapid development of next generation sequencing has rendered the genetic study of these diseases much easier and cheaper. Whole exome sequencing in congenital hemolytic anemia could provide a more precise and quicker diagnosis, improve patients' healthcare and probably has to be democratized notably for complex cases.

Treatment with mTOR inhibitors after liver transplantation enables a sustained increase in regulatory T-cells while preserving their suppressive capacity.

BACKGROUND: The mammalian targets of rapamycin (mTOR) inhibitors (sirolimus [SRL] and everolimus [EVR]) are used after transplantation for their immunosuppressive activity. Regulatory T-cells (Tregs) play a crucial role in immune tolerance. mTOR inhibitors appear to preserve Tregs, unlike Tacrolimus (Tac). AIM: The aim of this study was to evaluate the number and function of Tregs in liver transplant recipients before and after conversion from Tac to mTOR inhibitors. METHODS: Fifteen patients with stable graft function where converted to SRL (n=5) or EVR (n=10). Tregs (CD4+ CD25+ FoxP3+ CD127low) number and activity were analysed prospectively in blood cells using flow cytometry, and functional assay. RESULTS: Patients of both groups displayed a sustained rise in Treg levels after introduction of mTOR inhibitors (Treg levels at 3 months: 6.45±0.38% of CD4 T-cells, vs. baseline level of 3.61±0.37%, P<0.001; mean fold increase 2.04±0.73). In SRL group, 3-month Treg levels were 6.01±0.53 vs. 3.79±0.39; P=0.037, while in EVR group they were 6.63±0.67 vs. 3.54±0.51; P=0.001. By contrast, no statistical change was observed in an unconverted Tac control group. Tregs also preserved their functional ability to suppress activated T-cells. CONCLUSION: These results suggest that mTOR inhibitors induce a significant increase in Tregs while maintaining suppressive activity after LT.

Troponin elevation in other conditions than acute coronary syndromes.

Troponin is a specific cardiac infarction isoform (TnIc, TnTc) and its determination is used for the diagnosis of myocardial infarction even with normal Electrocardiography. The increase of cardiac troponins occurs in a variety of clinical situations without an acute coronary syndrome (ACS), cardiologists and emergency physicians are often confronted with positive troponins that are difficult to interpret. Few data exist about the occurrence, the clinical characteristics and the predictive value in case of absence of ACS. The objective of this study is to present the main extracardiac causes responsible of the increase of TnIc. We present some clinical cases that illustrate this diagnostic problem. A troponin elevation is observed in myopericarditis, renal failure, heart failure, pulmonary embolism, septic shock, rhabdomyolysis, stroke and others where there is a myocardial damage unrelated to coronary occlusion. Many cases of false positives, which raise the possibility of analytical interferences, must be identified.

Overexpression of Regulatory T Cells Type 1 (Tr1) Specific Markers in a Patient with HCV-Induced Hepatocellular Carcinoma.

Hepatitis C virus (HCV) is an important causative agent of liver disease, but factors that determine the resolution or progression of infection are poorly understood. In this study, we suggested that existence of immunosuppressive mechanisms, supported by regulatory T cells and especially the regulatory T cell 1 subset (Tr1), may explain the impaired immune response during infection and thus the fibrosis aggravation to hepatocellular carcinoma (HCC). Using quantitative real-time PCR, we investigated the intra-hepatic presence of Tr1 cells in biopsies from a genotype 1b infected patient followed for an 18-year period from cirrhosis to HCC. We described a significant increase of gene expression in particular for the cytokines IL-10, TGF-ß, and their receptors that were perfectly correlated with an increased expression of the Tr1 specific markers (combined expression of CD4, CD18, and CD49b). This was strongly marked since the patient evolved in the pathology and could explain the failure of the treatment. In conclusion, evidence of regulatory T cell installation in the liver of chronically infected patient with cirrhosis and HCC suggests for the first time a key role for these cells in the course of HCV infection.

In vitro effects of cyclosporine A and tacrolimus on regulatory T-cell proliferation and function.

BACKGROUND: Liver transplantation is the treatment of end-stage liver diseases, including hepatitis C. Immunosuppression prevents graft rejection but seems to accelerate the recurrence of hepatitis C. Regulatory T cells (Tregs) may be beneficial in tolerance but deleterious in recurrent hepatitis C. We evaluated the effects of cyclosporine or tacrolimus, the principal immunosuppressive drugs, on Treg proliferation and function. METHODS: Human Tregs were isolated from healthy donors and cultured with cyclosporine, tacrolimus, or NIM811, a cyclosporine analog devoid of calcineurin-inhibiting activity. Treg proliferation and suppressive activity were assessed. The phenotype, cytokine production, and phosphorylation profile of nuclear factor of activated T cell of Tregs were also analyzed. RESULTS: Cyclosporine and tacrolimus both decreased Treg proliferation, but only low doses of cyclosporine reduced Treg activity, by inducing the production of interleukin 2 proinflammatory cytokines in these cells. Moreover, NIM811 also inhibited Treg activity. The phosphorylation of nuclear factor of activated T cell in Tregs was not altered by cyclosporine, suggesting that the effects of this drug are independent of the calcineurin pathway. CONCLUSION: In summary, low doses of cyclosporine inhibit Treg activity, a finding that might explain the beneficial effect of this drug on hepatitis C recurrence. In contrast, by maintaining Treg activity, tacrolimus could be more helpful than cyclosporine in controlling rejection.

IgG from patients with pulmonary arterial hypertension and/or systemic sclerosis binds to vascular smooth muscle cells and induces cell contraction.

OBJECTIVES: Pulmonary arterial hypertension (PAH) is characterised by remodelling of pulmonary arteries with enhanced vascular smooth muscle cell (VSMC) contraction, migration and proliferation. The authors investigated the presence of antibodies to human VSMCs in the serum of patients with systemic sclerosis with or without PAH and idiopathic PAH (iPAH). METHODS AND RESULTS: Antibodies to VSMCs were detected by immunofluorescence in sera from healthy controls and patients with scleroderma without PAH, scleroderma-associated PAH and iPAH. Serum IgG from these patients induced contraction of VSMCs in a collagen matrix in contrast with IgG from healthy controls. Several protein spots of interest and target antigens were identified by two-dimensional immunoblotting and MS, including stress-induced phosphoprotein 1 and α-enolase. Finally, antibodies to stress-induced phosphoprotein 1 were detected by ELISA in sera from 84%, 76% and 24% of patients with scleroderma without PAH, scleroderma-associated PAH and iPAH, respectively, compared with only 3% of healthy controls. CONCLUSION: The authors have identified IgG that binds to VSMCs in the serum of patients with scleroderma and iPAH. These antibodies may be pathogenic by modulating vascular contraction. The target antigens of these antibodies are stress-induced phosphoprotein 1 and α-enolase.