Evaluation of Roche Hemolysis Index for Reporting of Plasma Hemoglobin, and Considerations Regarding Hemolysis Detection in Use of Ventricular-Assist Devices

Introduction/Objective Our cardiac intensive care unit (CICU) asked the laboratory to provide plasma hemoglobin testing to monitor effects of ventricular-assist devices (VAD). Roche hemolysis index (HI) has been reported to be suitable for this purpose. Use of HI as a reportable test, however, must be validated in-house as a laboratory- developed test (LDT), according to CLIA regulations. We describe results of our evaluation, and caveats for intended use. Methods/Case Report Concentrated hemolysate was produced by osmotic lysis of packed red blood cells in water. Hemolysis in plasma was by dilution of concentrated hemolysate. Verification of HI as a measure of hemoglobin (mg/dL) used the Sigma-Aldrich spectrophotometric hemoglobin assay as a gold standard. Linearity, linear range, sensitivity, were investigated by dilution measurements; interferences by admixture experiments; reproducibility (precision, intra- and inter-assay) in each case by replicate measurements (n = 20). Results (if a Case Study enter NA) HI was confirmed to correspond to hemoglobin in mg/dL. Linearity was between 2-1000 mg/dL (r2 >0.99). Intra-assay precisions were <=2.5% (hemoglobin = 74,148 mg/dL); inter-assay precisions were <=4.9% (hemoglobin = 69,139 mg/dL). HI variability was ±2 at very low values (0-10 mg/dL). There were no interferences observed among common therapeutic drugs. Hyperlipemia (evaluated up to lipemic index (LI) = 225) and hyperbilirubinemia (evaluated up to icteric index (II) = 7) demonstrated no significant interference. Conclusion Analytically, HI exhibited acceptable performance for reporting of plasma hemoglobin. Deployment would require establishment of quality control (QC) and proficiency testing procedures for HI. Clinically, a caveat for use is that HI cannot distinguish between hemolysis in circulation vs. that due to sample collection. For the CICU, HI >10 mg/dL is characteristic of more than 20% of samples drawn in general, irrespective of use of VAD. Temporal patterns of HI must therefore be carefully evaluated in parallel with haptoglobin measurements to assist in VAD management.

Carboxyhemoglobin levels in children during extracorporeal membrane oxygenation support: a retrospective study

Introduction: Hemolysis is a common complication of extracorporeal membrane oxygenation (ECMO). There are few data on whether carboxyhemoglobin (COHb), a potential marker of hemolysis, are elevated during ECMO support. Methods: We conducted a single-center, retrospective study comparing peak COHb levels of children pre-, during, and post-ECMO from January 2017 to August 2020. Results: There were 154 ECMO runs in 147 children (154 PICU admissions) included in the study. The median age was 3.5 (IQR 0.2, 39.2) months. Veno-arterial ECMO was the predominant mode: 146/154 (94.8%). Eighty-seven children (56.5%) underwent cardiac surgery. Peak COHb levels during ECMO were statistically significantly higher compared to pre ECMO (COHb 1.8% (IQR 1.4, 2.6) vs COHb 1.2% (IQR 0.7, 1.7), p < 0.001) and post ECMO (COHb 1.6% (IQR 1.3, 2.2), p = 0.009). Children with COHb ⩾2% were younger and had longer duration of ECMO support. Plasma hemoglobin weakly correlated with COHb level ( r = 0.14; p = 0.04). Conclusions: Carboxyhemoglobin levels increased during ECMO support compared to the pre and post ECMO period. Younger age and longer ECMO duration were associated with COHb levels ⩾2%. Plasma hemoglobin weakly correlated with COHb level.

Profile of hematological parameters in plasmodium falciparum malaria: A study from West Bengal

Objectives: Anemia is one of the main clinical presentations of severe malaria caused by P. falciparum and one of the major morbidities of malaria. This study was undertaken to explore the burden of anemia and hematological derangement in patients with P. falciparum malaria. Material and Methods: A cross sectional study was conducted on 186 patients of all age groups suffering from falciparum malaria. Complete blood count (CBC) with peripheral blood smear, reticulocyte count, liver function test (LFT) and plasma hemoglobin were done in all patients. Direct Coombs test (DCT) and urine for hemoglobin was estimated in 19 patients where hemolytic anemia was suspected. Results: Anemia was seen in majority (78.7%) of the patients; 82.7% of males, 70.6% of females and 87.2% of children had anemia. Thrombocytopenia was seen in 9.1% cases. Unconjugated hyperbilirubinemia was seen in 33.8% patients though plasma hemoglobin was raised in only 1.1% patients. DCT was positive in 3 patients (15.7%) and hemoglobinuria was seen in 2 patients (10.5%). Conclusion: The present study revealed that anemia is one of the common manifestations of falciparum malaria and requires special attention to reduce the burden of this morbidity.

MO1046DOPING POLYSULFONE DIALYSIS MEMBRANES WITH HUMAN NEUTROPHIL ELASTASE INHIBITORS - A PILOT STUDY

Background and Aims During hemodialysis (HD) therapy, the long-term intradialytic contact of blood with large surfaced artificial materials (HD membranes) leads to continuous neutrophil activation, with the release of neutrophil elastase (NE), among other products. NE appears to contribute to enhance inflammation favoring the development of atherosclerosis, which is the main cause of mortality and morbidity in End-Stage Renal Disease patients. The modification of polysulfone (PSF) HD membranes by incorporating selective human NE inhibitors (NEIs) might reduce the inflammatory response and prevent HD associated complications. Thus, the present study aimed to dope PSF membranes with NEIs and assess their bioactivity and biocompatibility. Method As NEIs, it was used the commercially available Sivelestat (SIV), from Abcam, and an in house synthetized 4-oxo-β-lactam (D4L-3) based compound, selected from our library, and prepared as described elsewhere [1]. The PSF membranes were prepared according to [2] and further doped with each NEI by adsorption. Three independent assays were performed (in triplicates) where PSF membrane circles (2 cm in diameter) were incubated with ultrapure water (blank), NEIs vehicle (2.5% DMSO) or with 10 - 2000 nM SIV or D4L-3. The bioactivity of these modified PSF membranes was evaluated by a human NE activity assay [1]. The same method was used to determine the IC50 of both NEIs in solution. Biocompatibility assays (n = 3) were carried-out using duplicates of modified PSF membranes circles with Ø 2 cm incubated with 1.0 mL of whole-blood. After incubation, platelet (PLT) poor and PLT rich plasma were used to assess the levels of plasma hemoglobin (Hb) and PLT activation, respectively [2]. Results The IC50 of SIV and D4L-3 in solution were 30.9 and 87.6 nM, respectively. For PSF membranes doped with NEIs, their bioactivity increased in a concentration-dependent manner, with the highest NE inhibition of 44 and 22 % at 2000 nM SIV and D4L-3, respectively. The blank membranes showed the highest hemolytic capacity, whereas SIV and D4L-3 PSF membranes presented lower plasma Hb levels when compared with the blank or the vehicle; on average, SIV-PSF membranes presented 31% less plasma Hb than vehicle, while in D4L-3-PSF this decrease was of 51%. Regarding the thrombotic potential of these biomaterials, blank membranes presented slightly increased PLT activation levels when compared to vehicle, and modified SIV-PSF membranes showed, on average, 58% more PLT activation than the vehicle, while D4L-3-PSF membranes displayed 25% less, on average. Conclusion The successful adsorption of NEIs into PSF membranes was achieved and the NE inhibition ability was directly dependent on the concentration of the inhibitor utilized. Moreover, the bioactivity of SIV and D4L-3 when immobilized into PSF membranes it appears to follow their inhibitory capacity occurring in solution, with SIV showing a greater affinity for NE. However, concerning biocompatibility, D4L-3 displayed a greater safety performance than SIV, especially regarding the potential for triggering platelet activation. To ensure the applicability of these modified PSF membranes as a medical device, extensive further studies have to be carried-out by tweaking the conditions of the immobilization process in order to obtain an optimal equilibrium between bioactivity and biocompatibility, as well test new NEIs. Acknowledgments This work was supported by UIDB/50006/2020 and UIDB/04378/2020 with funding from FCT/MCTES through national funds, and by the project Dial4Life co-financed by FCT/MCTES (PTDC/MEC-CAR/31322/2017) and FEDER/COMPETE 2020 (POCI-01-0145-FEDER-031322).

Association of Plasma Hemoglobin A1c with Improvement of Cognitive Functions by Probiotic Bifidobacterium breve Supplementation in Healthy Adults with Mild Cognitive Impairment

We demonstrated the benefit of the probiotic strain, Bifidobacterium breve MCC1274 (synonym B. breve A1), at improving cognition in our previous double-blind, placebo-controlled clinical study. Analysis of the association of blood parameters changes with the improvement of cognitive function revealed an inverse correlation of HbA1c with total RBANS score amelioration after the study only in the probiotic group (ρ= –0. 4218, p = 0.0067). A stratified analysis based on baseline HbA1c with a median value showed a more remarkable benefit by the probiotic supplementation in the higher median subgroup. These data support the mechanism of anti-inflammation in improving cognition by the probiotic strain.

Gemtuzumab-Ozogamicin-Related Impaired Hemoglobin-Haptoglobin Scavenging as On-Target/Off-Tumor Toxicity of Anti-CD33 AML Therapy: A Report of Two Cases

Gemtuzumab-ozogamicin (GO) is a humanized anti-CD33 antibody, which is conjugated to a cytotoxic calicheamicin. It is used to treat acute myeloid leukemia (AML) in combination with chemotherapy. We describe here two GO-treated acute myeloid leukemia (AML) cases: both patients suffered from a toxic syndrome, which manifested as impaired hemoglobin-haptoglobin scavenging and accumulation of hemolysis-related products. Our observations and earlier reports indicated that the reaction was caused by GO-targeted destruction of CD33 + CD163+ monocytes/macrophages, which are responsible for the clearance of hemoglobin-haptoglobin complexes. The rise of plasma lactate dehydrogenase was an early sign of the reaction, and both patients had high levels of free plasma hemoglobin, but plasma haptoglobin and bilirubin levels were paradoxically normal. Symptoms included septic fever and abnormalities in cardiac tests and in the case of the first patient, severe neurological symptoms which required intensive care unit admittance. Therapeutic plasma exchanges supported the patients until the recovery of normal hematopoiesis. The symptoms may be easily confounded with infectious complications-related organ damage. Regarding the increasing use of gemtuzumab-ozogamicin and other emerging CD33-targeted cell therapies, we want to highlight this mostly unknown and probably underdiagnosed toxicity.

Iron Overload in Functional Hyperandrogenism: In a Randomized Trial, Bloodletting Does Not Improve Metabolic Outcomes

Context Functional hyperandrogenism may be associated with a mild increase in body iron stores. Iron depletion exerts a beneficial effect on metabolic endpoints in other iron overload states. Objectives (i) To determine the effect of iron depletion on the insulin sensitivity and frequency of abnormal glucose tolerance in patients with functional hyperandrogenism submitted to standard therapy with combined oral contraceptives (COC). ii) To assess the overall safety of this intervention. Design Randomized, parallel, open-label, clinical trial. Setting Academic hospital. Patients Adult women with polycystic ovary syndrome or idiopathic hyperandrogenism. Intervention After a 3-month run-in period of treatment with 35 μg ethinylestradiol plus 2 mg cyproterone acetate, participants were randomized (1:1) to 3 scheduled bloodlettings or observation for another 9 months. Main outcome measures Changes in insulin sensitivity index and frequency of prediabetes/diabetes, and percentage of women in whom bloodletting resulted in plasma hemoglobin &lt;120 g/L and/or hematocrit &lt;0.36. Results From 2015 to 2019, 33 women were included by intention-to-treat. During the follow-up, insulin sensitivity did not change in the whole group of women or between study arms [mean of the differences (MD): 0.0 (95%CI: −1.6 to 1.6)]. Women in the experimental arm showed a similar odds of having prediabetes/diabetes than women submitted to observation [odds ratio: 0.981 (95%CI: 0.712 to 1.351)]. After bloodletting, 4 (21.1%) and 2 women (10.5%) in the experimental arm had hemoglobin (Hb) levels &lt;120 g/L and hematocrit (Hct) values &lt;0.36, respectively, but none showed Hb &lt;110 g/L or Hct &lt;0.34. Conclusions Scheduled bloodletting does not improve insulin sensitivity in women with functional hyperandrogenism on COC.