Correction to: CO2 and O2 removal during continuous veno-venous hemofiltration: a pilot study.

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CO2 and O2 removal during continuous veno-venous hemofiltration: a pilot study.

BACKGROUND: Carbon dioxide (CO2) accumulation is a challenging issue in critically ill patients. CO2 can be eliminated by renal replacement therapy but studies are scarce and clinical relevance is unknown. We prospectively studied CO2 and O2 behavior at different sample points of continuous veno-venous hemofiltration (CVVH) and build a model to calculate CO2 removal bedside. METHODS: In 10 patients receiving standard CVVH under citrate anticoagulation, blood gas analysis was performed at different sample points within the CVVH circuit. Citrate was then replaced by NaCl 0.9% and sampling was repeated. Total CO2 (tCO2), CO2 flow (V̇CO2) and O2 flow (V̇O2) were compared between different sample points. The effect of citrate on transmembrane tCO2 was evaluated. Wilcoxon matched-pairs signed rank test was performed to evaluate significance of difference between 2 data sets. Friedman test was used when more data sets were compared. RESULTS: V̇CO2 in the effluent (26.0 ml/min) correlated significantly with transmembrane V̇CO2 (24.2 ml/min). This represents 14% of the average expired V̇CO2 in ventilated patients. Only 1.3 ml/min CO2 was removed in the de-aeration chamber, suggesting that CO2 was almost entirely cleared across the membrane filter. tCO2 values in effluent, before, and after the filter were not statistically different. Transmembrane tCO2 under citrate or NaCl 0.9% predilution also did not differ significantly. No changes in V̇O2 were observed throughout the CVVH circuit. Based on recorded data, formulas were constructed that allow bedside evaluation of CVVH-attributable CO2 removal. CONCLUSION: A relevant amount of CO2 is removed by CVVH and can be quantified by one simple blood gas analysis within the circuit. Future studies should assess the clinical impact of this observation. TRIAL REGISTRATION: The trial was registered at https://clinicaltrials.gov with trial registration number NCT03314363 on October 192,017.

Combined Analysis of GAD65, miR-375, and Unmethylated Insulin DNA Following Islet Transplantation in Patients With T1D.

Aim: Several biomarkers have been proposed to detect pancreatic ß cell destruction in vivo but so far have not been compared for sensitivity and significance. Methods: We used islet transplantation as a model to compare plasma concentrations of miR-375, 65-kDa subunit of glutamate decarboxylase (GAD65), and unmethylated insulin DNA, measured at subpicomolar sensitivity, and study their discharge kinetics, power for outcome prediction, and detection of graft loss during follow-up. Results: At 60 minutes after transplantation, GAD65 and miR-375 consistently showed near-equimolar and correlated increases proportional to the number of implanted ß cells. GAD65 and miR-375 showed comparable power to predict poor graft outcome at 2 months, with areas under the curve of 0.833 and 0.771, respectively (P = 0.53). Using receiver operating characteristic analysis, we defined likelihood ratios (LRs) for rationally selected result intervals. In GADA-negative recipients (n = 28), GAD65 <4.5 pmol/L (LR = 0.15) and >12.2 pmol/L (LR = ∞) predicted good and poor outcomes, respectively. miR-375 could be used in all recipients irrespective of GAD65 autoantibody status (n = 46), with levels <1.4 pmol/L (LR = 0.14) or >7.6 pmol/L (LR = 9.53) as dual thresholds. The posttransplant surge of unmethylated insulin DNA was inconsistent and unrelated to outcome. Combined measurement of these three biomarkers was also tested as liquid biopsy for ß cell death during 2-month follow-up; incidental surges of GAD65, miR-375, and (un)methylated insulin DNA, alone or combined, were confidently detected but could not be related to outcome. Conclusions: GAD65 and miR-375 performed equally well in quantifying early graft destruction and predicting graft outcome, outperforming unmethylated insulin DNA.

Cystic Lymphangioma: Are Triglycerides Always Measurable?

The presence of chylous fluid with high triglycerides levels on endoscopic ultrasound- (EUS-) guided fine needle aspiration (FNA) is very pathognomonic for the diagnosis of cystic lymphangiomas of the pancreas. In our case report the puncture of the pancreatic cyst showed a typical milky fluid though measurable triglyceride concentrations were absent in the laboratory. Two possible explanations were found. First of all grossly lipemic samples show a slower rate of color development than do clear serums which can produce a false negative result if the sample is insufficiently diluted. Secondly, high lipase levels can divide triglycerides in glycerol and fatty acids, making the concentration of triglycerides undetectable.

An analytical comparison of three immunoassay platforms for subpicomolar detection of protein biomarker GAD65.

A disproportional increase of circulating GAD65 within hours from an intraportal islet allotransplantation has been validated as biomarker of beta cell loss and poor functional outcome. More sensitive assays are, however, needed to allow detection of episodes of subtle beta cell loss during late-stage graft rejection or in the peri-onset period of type 1 diabetes. We applied the same sandwich monoclonal antibody couple reactive towards the C- and N-terminus of GAD65 on three advanced immunoassay platforms-the Cytometric Bead Array (CBA, Becton, Dickinson and Company), ElectroChemiLuminescence ImmunoAssay (ECLIA, Meso Scale Discovery) and digital ELISA technology (Single Molecule Array-SIMOA, Quanterix. We then compared analytical performance (linearity, imprecision, limit of detection and functional sensitivity), correlation of results, and practicality. All evaluated techniques showed linearity up to at least 500 ng/dL (76.9 pmol/L). SIMOA achieved the lowest imprecision. The 3 platforms correlate well with each other and could all detect subpicomolar concentrations of GAD65 in plasma, but only SIMOA and CBA could quantify down to that range. SIMOA can achieve the highest sample throughput. The three methods tested allow sensitive detection of GAD65, but SIMOA appears best suited for automated quantification of subpicomolar concentrations.

Accelerated Progression to Type 1 Diabetes in the Presence of HLA-A*24 and -B*18 Is Restricted to Multiple Islet Autoantibody-Positive Individuals With Distinct HLA-DQ and Autoantibody Risk Profiles.

OBJECTIVE: We investigated the effect of HLA class I risk alleles on disease progression in various phases of subclinical islet autoimmunity in first-degree relatives of patients with type 1 diabetes. RESEARCH DESIGN AND METHODS: A registry-based group of siblings/offspring (aged 0-39 years) was monitored from single- to multiple-autoantibody positivity (n = 267) and from multiple-autoantibody positivity to clinical onset (n = 252) according to HLA-DQ, -A*24, -B*18, and -B*39 status. Genetic markers were determined by PCR sequence-specific oligotyping. RESULTS: Unlike HLA-B*18 or -B*39, HLA-A*24 was associated with delayed progression from single- to multiple-autoantibody positivity (P = 0.009) but not to type 1 diabetes. This occurred independently from older age (P < 0.001) and absence of HLA-DQ2/DQ8 or -DQ8 (P < 0.001 and P = 0.003, respectively), and only in the presence of GAD autoantibodies. In contrast, HLA-A*24 was associated with accelerated progression from multiple-autoantibody positivity to clinical onset (P = 0.006), but its effects were restricted to HLA-DQ8+ relatives with IA-2 or zinc transporter 8 autoantibodies (P = 0.002). HLA-B*18, but not -B*39, was also associated with more rapid progression, but only in HLA-DQ2 carriers with double positivity for GAD and insulin autoantibodies (P = 0.004). CONCLUSIONS: HLA-A*24 predisposes to a delayed antigen spreading of humoral autoimmunity, whereas HLA-A*24 and -B*18 are associated with accelerated progression of advanced subclinical autoimmunity in distinct risk groups. The relation of these alleles to the underlying disease process requires further investigation. Their typing should be relevant for the preparation and interpretation of observational and interventional studies in asymptomatic type 1 diabetes.

Twenty-Year Progression Rate to Clinical Onset According to Autoantibody Profile, Age, and HLA-DQ Genotype in a Registry-Based Group of Children and Adults With a First-Degree Relative With Type 1 Diabetes.

OBJECTIVE: We investigated whether islet autoantibody profile, HLA-DQ genotype, and age influenced a 20-year progression to diabetes from first autoantibody positivity (autoAb+) in first-degree relatives of patients with type 1 diabetes. RESEARCH DESIGN AND METHODS: Persistently islet autoAb+ siblings and offspring (n = 462) under 40 years of age were followed by the Belgian Diabetes Registry. AutoAbs against insulin (IAA), GAD (GADA), IA-2 antigen (IA-2A), and zinc transporter 8 (ZnT8A) were determined by radiobinding assay. RESULTS: The 20-year progression rate of multiple-autoAb+ relatives (n = 194) was higher than that for single-autoAb+ participants (n = 268) (88% vs. 54%; P < 0.001). Relatives positive for IAA and GADA (n = 54) progressed more slowly than double-autoAb+ individuals carrying IA-2A and/or ZnT8A (n = 38; P = 0.001). In multiple-autoAb+ relatives, Cox regression analysis identified the presence of IA-2A or ZnT8A as the only independent predictors of more rapid progression to diabetes (P < 0.001); in single-autoAb+ relatives, it identified younger age (P < 0.001), HLA-DQ2/DQ8 genotype (P < 0.001), and IAA (P = 0.028) as independent predictors of seroconversion to multiple positivity for autoAbs. In time-dependent Cox regression, younger age (P = 0.042), HLA-DQ2/DQ8 genotype (P = 0.009), and the development of additional autoAbs (P = 0.012) were associated with more rapid progression to diabetes. CONCLUSIONS: In single-autoAb+ relatives, the time to multiple-autoAb positivity increases with age and the absence of IAA and HLA-DQ2/DQ8 genotype. The majority of multiple-autoAb+ individuals progress to diabetes within 20 years; this occurs more rapidly in the presence of IA-2A or ZnT8A, regardless of age, HLA-DQ genotype, and number of autoAbs. These data may help to refine the risk stratification of presymptomatic type 1 diabetes.

Development of an Enhanced Sensitivity Bead-Based Immunoassay for Real-Time In Vivo Detection of Pancreatic ß-Cell Death.

There is a clinical need for plasma tests to detect and quantify the in vivo destruction of pancreatic ß-cells in type 1 diabetes. We previously developed a time-resolved fluorescence immunoassay (TRFIA) to glutamate decarboxylase 65 kDa (GAD65) (GAD65-TRFIA) that was able to detect the synchronous necrotic destruction of transplanted ß-cells in the hours after their infusion in the liver. This GAD65-TRFIA, however, lacked sensitivity to detect continued ß-cell rejection beyond this acute phase. The aim of present study was to gain at least an order of magnitude in analytical sensitivity by switching to Becton Dickinson cytometric bead array (CBA) (GAD65-CBA) enhanced sensitivity format, using the same couple of monoclonal antibodies. We compared the performances of GAD65-CBA and GAD65-TRFIA using Clinical and Laboratory Standards Institute protocols for linearity, imprecision, specificity, limit of detection, and functional sensitivity. We conducted a method comparison and assessed the biologic potential on samples from human recipients of islet grafts. The GAD65-CBA showed acceptable linearity and imprecision. Switching from TRFIA to CBA lowered functional sensitivity by a factor 35 and lowered limit of detection by a factor 11 with minimal need for method optimization. The enhanced sensitivity greatly expands the application domain of our biomarker and allowed for the first time to detect ongoing ß-cell destruction up to at least 1 day after islet transplantation. We conclude that the GAD65-CBA is suitable for biological and clinical assessment of the real-time destruction of ß-cells in intraportal transplantation.