Hwan Tae Lee, M.D., Jeong-Yeal Ahn, M.D., Ja Young Seo, M.D., Yiel-Hea Seo, M.D., Pil-Whan Park, M.D., Kyung-Hee Kim, M.D., Jaehyung You, Wookwan Park, Woo Jin Kim, M.D., Young Ho Yoon, M.D., Gwiyoung Oh, M.D., Daewon Kim, M.D.. Lab Med Online 2024;14:118-26. https://doi.org/10.47429/lmo.2024.14.2.118

ABO incompatibility is not considered a contraindication for hematopoietic stem cell transplantation (HSCT). We hypothesized that recipient-derived isoagglutinin (RDI) levels could play a critical role in clinical outcomes. In this study, we compared clinical outcomes such as survival, GVHD, infection, relapse, transfusion, and engraftment, among ABO-compatible patients (ABOc), ABO-incompatible patients (ABOi) with low RDI, and ABOi patients with high RDI. The ABOi with high RDI group was defined as recipients with more than 1:16 RDI levels. We analyzed 103 recipients (ABOc, 53; ABOi with low RDI, 36; ABOi with high RDI, 14). The ABOi with high RDI group showed a decreased 1-year survival and increased acute GVHD grade IV and RBC transfusion (p = 0.017, 0.027, and 0.032, respectively). The ABOi with high RDI group was an independent risk factor for increased death, RBC transfusion, and poor platelet (PLT) engraftment (odds ratio (OR) = 3.20, p = 0.01; OR = 8.28, p = 0.02; OR = 0.18, p = 0.03, respectively). The ABOi with high RDI group showed significantly delayed PLT engraftment. In conclusion, this is the first study underscoring high RDI levels as a marker predicting unfavorable outcomes in ABOi HSCT.

OBJECTIVE:Modular workcells could be a better solution than total laboratory automation (TLA) in hemostasis laboratories. Here, we evaluated the impact of implementing a modular workcell (HemoCell) with an intelligent data management facility (HemoHub).METHODS:We compared the turnaround times (TATs), numbers of rerun samples, and rerun times pre- and postimplementation of the HemoCell at Gil Medical Center. Prothrombin time (PT), activated partial thromboplastin time (aPTT), D-dimer, and fibrinogen were evaluated.RESULTS:The TAT standard deviations (SDs) and maximum TAT values decreased after HemoCell implementation, although the mean TATs for PT, aPTT, and D-dimer were increased. Numbers of rerun samples were increased (18.1/day vs 44.7/day). However, rerun times were reduced, and SDs were decreased during the post-HemoCell period compared with pre-HemoCell. Additionally, technologists needed smaller working space and less labor.CONCLUSION:The modular workcell could improve quality and efficiency by providing more consistent TATs and shorter rerun times in the hemostasis laboratory.

While numerous studies have evaluated humoral responses to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccines, data on the cellular responses to these vaccines remain sparse. We evaluated T cell responses to ChAdOx1-nCoV-19 and BNT162b2 vaccinations using an interferon gamma (IFN-γ) release assay (IGRA). ChAdOx1-nCoV-19- and BNT162b2-vaccinated participants initially showed stronger T cell responses than unvaccinated controls. The T cell response decreased over time and increased substantially after the administration of a BNT162b2 booster dose. Changes in the T cell response were less significant than those in the anti-receptor-binding domain IgG antibody titer. The study results can serve as baseline data for T cell responses after SARS-CoV-2 vaccination and suggest that the IGRA can be useful in monitoring immunogenicity.

Eukaryotic translation initiation factor 2α (eIF2α) is a key mediator of the endoplasmic reticulum (ER) stress–induced unfolded protein response (UPR). In mammals, eIF2α is phosphorylated by overnutrition-induced ER stress and is related to the development of obesity. Here, we studied the function of phosphorylated eIF2α (p-eIF2α) in AgRP neurons using a mouse model (AgRPeIF2αA/A) with an AgRP neuron–specific substitution from Ser 51 to Ala in eIF2α, which impairs eIF2α phosphorylation in AgRP neurons. These AgRPeIF2αA/A mice showed decreases in starvation-induced AgRP neuronal activity and food intake and also revealed an increased responsiveness to leptin. Intriguingly, impairment of eIF2α phosphorylation produced decreases in the starvation-induced expression of UPR and autophagy genes in AgRP neurons. Collectively, these findings suggest that eIF2α phosphorylation regulates AgRP neuronal activity by affecting intracellular responses such as the UPR and autophagy during starvation, thereby participating in the homeostatic control of whole-body energy metabolism.

Background: The distribution of bacteria isolated from bloodstream infections and cumulative antimicrobial susceptibility data are the basis for empirical decisions regarding antibiotics as an initial treatment. Therefore, it is important to consistently collect blood culture results of individual patients and analyze them correctly. Methods: The blood culture results of patients at Gachon University Gil Medical Center from 2016 to 2020 were analyzed retrospectively to determine the bacterial distributions and antibiotic resistance patterns. Duplicates were eliminated by including only the first isolate of each species per patient. Results: Escherichia coli (27.1%) was the most commonly isolated bacterium from blood cultures, followed by Klebsiella pneumoniae (10.1%) and Staphylococcus aureus (8.6%). The methicillin resistance rate of S. aureus was 49.2%, and the vancomycin resistance rate of Enterococcus faecium was 39.5%; with no significant changes over the study period. The cefotaxime, ciprofloxacin, and ertapenem resistance rates of E. coli were 35.0%, 46.8%, and 0.7%, respectively. Seventeen carbapenem-resistant E. coli strains were isolated, of which 11 produced carbapenemase. The cefotaxime, ciprofloxacin, and ertapenem resistance rates of K. pneumoniae were 29.5%, 31.7%, and 5.4%, respectively. Forty-eight carbapenem-resistant K. pneumoniae strains were isolated, of which 37 produced carbapenemase. The imipenem resistance rates of Acinetobacter baumannii and Pseudomonas aeruginosa were 72.3% and 23.4%, respectively. Conclusion: In the blood culture results from 2016 to 2020, the isolation frequency of E. coli, K. pneumoniae, and E. faecium showed an increasing trend, whereas that of S. aureus was stable. Over the 5 year study period, the ciprofloxacin resistance rate of E. coli and P. aeruginosa and ampicillin/sulbactam resistance rate of A. baumannii significantly increased.

Background: Elevated soluble suppression of tumorigenicity 2 (sST2) levels may predict mortality in heart failure (HF) patients. The AFIAS ST2 assay (AFIAS ST2, Boditech Med Inc., Chuncheon, Korea) and ichroma ST2 assay (ichroma ST2, Boditech Med Inc.) are newly developed point-of-care (POC) assays for measuring sST2 level. We evaluated the performance of these assays, in terms of cut-off validation and prognosis, and compared them with that of the Presage ST2 assay (Presage ST2, Critical Diagnostics, San Diego, CA, USA). Methods: We validated the US FDA-claimed sST2 clinical cut-off of 35 ng/mL using 420 serum samples (298 samples from the universal sample bank of the American Association for Clinical Chemistry and 122 samples from reference individuals from Konkuk University Medical Center). We compared AFIAS ST2 and ichroma ST2 with Presage ST2, using 206 samples from patients with HF. We assessed prognosis using the three assays in 252 samples from the Barcelona ambulatory HF cohort subsets. Results: The upper reference limits of AFIAS ST2 and ichroma ST2 were within the clinical cut-off of Presage ST2. The results of AFIAS ST2 and ichroma ST2 were highly correlated with those of Presage ST2 (r = 0.82 and 0.81, respectively). Based on this cut-off, all three assays predicted cardiovascular death. Conclusions: The new POC assays, AFIAS ST2 and ichroma ST2, would be useful in clinical practice for managing HF patients, with performances equivalent to that of Presage ST2.

BACKGROUND:Immunity against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) wanes over time after vaccination.METHODS:We compared SARS-CoV-2 antibody levels in serial samples from 350 vaccinated individuals at 3 time points (3 weeks after the first or second dose and before the third dose) with 4 assays: GenScript cPASS SARS-CoV-2 neutralization antibody detection kits (cPASS), Siemens SARS-CoV-2 IgG (sCOVG), Abbott SARS-CoV-2 IgG II Quant (CoV-2 IgG II), and an Immuno-On™ COVID-19 IgG test (Immuno-On IgG). Antibody levels by time, concordance between assays, and values from other tests corresponding to the percent inhibition results in cPASS were assessed.RESULTS:The median values at three time points were 49.31%, 90.87%, and 53.38% inhibition for cPASS, 5.39, 13.65, and 2.24 U/mL for sCOVG, 570.25, 1279.65, and 315.80 AU/mL for CoV-2 IgG II, and 223.22, 362.20, and 62.20 relative units (RU) for Immuno-On IgG. The concordance with cPASS at each time point ranged from 0.735 to 0.984, showing the highest concordance in the second sample and lowest concordance in the third in all comparative tests. The values corresponded to 30% inhibition, and the cutoffs of cPASS, were 2.02 U/mL, 258.6 AU/mL, and 74.2 RU for each test. Those for 50%, 70%, and 90% inhibition were 3.16, 5.66, and 8.26 U/mL for sCOVG, while they were 412.5, 596.9, and 1121.6 AU/mL for CoV-2 IgG II and 141.8, 248.92, and 327.14 RU for Immuno-On IgG.CONCLUSIONS:This study demonstrated the dynamic changes in antibody values at different time points using four test systems and is expected to provide useful baseline data for comparative studies and standardization efforts in the future.

Spexin (SPX) is a recently identified neuropeptide that is believed to play an important role in the regulation of energy homeostasis. Here, we describe a mediating function of SPX in hypothalamic leptin action. Intracerebroventricular (icv) SPX administration induced a decrease in food intake and body weight gain. SPX was found to be expressed in cells expressing leptin receptor ObRb in the mouse hypothalamus. In line with this finding, icv leptin injection increased SPX mRNA in the ObRb-positive cells of the hypothalamus, which was blocked by treatment with a STAT3 inhibitor. Leptin also increased STAT3 binding to the SPX promoter, as measured by chromatin immunoprecipitation assays. In vivo blockade of hypothalamic SPX biosynthesis with an antisense oligodeoxynucleotide (AS ODN) resulted in a diminished leptin effect on food intake and body weight. AS ODN reversed leptin's effect on the proopiomelanocortin (POMC) mRNA expression and, moreover, decreased leptin-induced STAT3 binding to the POMC promoter sequence. These results suggest that SPX is involved in leptin's action on POMC gene expression in the hypothalamus and impacts the anorexigenic effects of leptin.

Objectives We examined the frequencies and sizes of glycosylphosphatidylinositol (GPI)-deficient cells as per the International Clinical Cytometry Society/European Society for Clinical Cell Analysis (ICCS/ESCCA) consensus guidelines for the high-sensitivity detection of GPI-deficient cells. Methods In 2018, the ICCS/ESCCA guidelines for the high-sensitivity detection of GPI-deficient cells were published. We evaluated frequencies and sizes of GPI-deficient red blood cells (RBCs), neutrophils, and monocytes as determined using the ICCS/ESCCA guidelines and Clinical and Laboratory Standards Institute (CLSI) guidelines in patients with a hematologic malignancy, aplastic anemia, or cytopenia. Results A total of 106 (38.7%) patients exhibited GPI deficiency in at least one blood cell lineage. GPI-deficient cells of one or more lineages were found in 62.7% of patients with a hematologic malignancy, 51.1% of patients with aplastic anemia, and 23.4% of patients with cytopenia. GPI-deficient monocytes were most frequently detected in all three groups. By population size, GPI-deficient clones (>1%) in monocytes were mostly detected in patients with a hematologic malignancy or aplastic anemia. Rare cells with GPI deficiency (<0.1%) in monocytes were most common among patients with cytopenia. Conclusion High-sensitive flow cytometry analysis including monocytes may be necessary for patients with a hematologic disorder.

The clinical demand for molecular tests is increasing and thus, the need for quality control is also increasing.The Korean Association of External Quality Assessment Service has operated a test proficiency program for molecular hepatitis virus testing twice a year (molecular microbiology, hepatitis virus 1, 2).The proficiency test for hepatitis virus molecular testing includes hepatitis B virus (HBV; quantitative and qualitative), hepatitis C virus (HCV; quantitative and qualitative), and HBV DNA drug resistance mutation tests.The HBV (quantitative) and HCV (quantitative) programs had the greatest number of participants, with 117 and 71 institutions, respectively, participating in the second evaluation in 2021.This represents 5.4% and 9.2% increases, respectively, compared to the first evaluation in 2016.For the remaining tests, less than 20 institutions participated, which was not sufficient to confirm an increase or decrease in the number of participants.The market share was high for the two major quantitative test manufacturers, Roche (Switzerland) and Abbott (USA), and Roche was the dominant manufacturer of the qualitative test.However, laboratory-developed tests were the main tests used to detect drug resistance mutations.The coefficients of variation for the HBV (quantitative) test for low-and high-concentration samples were 20.5% and 29.6%, respectively, while those for the HCV (quantitative) test were 37.1% and 32.3%, respectively.The percentage of intended responses for the qualitative test ranged from 85.7%-100.0%through the whole period, and was 100.0% for 2020 and 2021.The percentage of intended responses for the HBV DNA drug resistance test for three agents, adefovir, entecavir, and lamivudine, ranged from 78.6%-100.0%,84.

BACKGROUND Obesity represents a growing global health threat, which generally portends increased morbidity and mortality in the context of traumatic injuries. We hypothesized that there may exist a protective effect related to increased weight and truncal girth provided for obese patients in penetrating torso injuries, although this may not exert a significant positive impact overall upon clinical outcomes. METHODS A comprehensive review of the literature was conducted across five databases up to March 2021 (Medline, Pubmed, Embase, Web of Science and the Cochrane library) to examine the effect of obesity on penetrating thoracoabdominal injuries. The primary outcome was to determine the rate of nonsignificant injury and injury patterns. Secondary outcomes examined were lengths of stay, complications, and mortality. Comparisons were drawn by meta-analysis. The study protocol was registered with PROSPERO under CRD42020216277. RESULTS There were 2,952 publications assessed with 12 meeting the inclusion criteria for review. Nine studies were included for quantitative analysis, including 5,013 patients sustaining penetrating thoracoabdominal injuries, of which 29.6% were obese. Obese patients that sustained stab injuries underwent more nontherapeutic operations. Obese patients that sustained gunshot injuries had longer intensive care and total hospital length of stay. Obese patients suffered more respiratory complications and were at an increased risk of death during their admission. CONCLUSION The “armor phenomenon” does not truly protect obese patients, a population that experiences increased morbidity and mortality following penetrating thoracoabdominal injuries. LEVEL OF EVIDENCE Systematic Review and Meta-Analysis; Level IV.

differentials, or the presence of immature blood cells are known to be associated with hematologic disorders and a wide variety of cardiovascular diseases and even metabolic disorders [1,2].For these matters, laboratories need automated hematology analyzers (HA) that can process a urry of data rapidly, let alone with high precision.Such HA should be able to identify abnormal results, report them, and induce manual con rmation if required.Since the 1970s, when 3-part WBC differentials were rst available, recent technological advancements have led HA increasingly complex, and various analyzers were developed using new methods such as uorescent dye or ow cytometry in conjunction with traditional methods like electrical impedance and laser light scattering [3,4].As a result, HA progressed to the stage where they are capable of counting 5-part WBC differential, identifying ab-

Preproenkephalin (PPE) is a precursor molecule for multiple endogenous opioid peptides Leu-enkephalin (ENK) and MetENK, which are involved in a wide variety of modulatory functions in the nervous system. Despite the functional importance of ENK in the brain, the effect of brain-derived factor(s) on PPE expression is unknown. We report the dual effect of neural epidermal growth factor (EGF)-likelike 2 (NELL2) on PPE gene expression. In cultured NIH3T3 cells, transfection of NELL2 expression vectors induced an inhibition of PPE transcription intracellularly, in parallel with downregulation of protein kinase C signaling pathways and extracellular signal-regulated kinase. Interestingly, these phenomena were reversed when synthetic NELL2 was administered extracellularly. The in vivo disruption of NELL2 synthesis resulted in an increase in PPE mRNA level in the rat brain, suggesting that the inhibitory action of intracellular NELL2 predominates the activation effect of extracellular NELL2 on PPE gene expression in the brain. Biochemical and molecular studies with mutant NELL2 structures further demonstrated the critical role of EGF-like repeat domains in NELL2 for regulation of PPE transcription. These are the first results to reveal the spatio-specific role of NELL2 in the homeostatic regulation of PPE gene expression.

Abstract Objectives Vision Pro (West Medica, Perchtoldsdorf, Austria) is a recently developed digital morphology analyzer. We evaluated the performance of Vision Pro on white blood cell (WBC) differentials. Methods In a total of 200 peripheral blood smear samples (100 normal and 100 abnormal samples), WBC preclassification and reclassification by Vision Pro were evaluated and compared with manual WBC count, according to the Clinical and Laboratory Standards Institute guidelines (H20-A2). Results The overall sensitivity was high for normal WBCs and nRBCs (80.1–98.0%). The overall specificity and overall efficiency were high for all cell classes (98.1–100.0% and 97.7–99.9%, respectively). The absolute values of mean differences between Vision Pro and manual count ranged from 0.01 to 1.31. In leukopenic samples, those values ranged from 0.09 to 2.01. For normal WBCs, Vision Pro preclassification and manual count showed moderate or high correlations (r=0.52–0.88) except for basophils (r=0.34); after reclassification, the correlation between Vision Pro and manual count was improved (r=0.36–0.90). Conclusions This is the first study that evaluated the performance of Vision Pro on WBC differentials. Vision Pro showed reliable analytical performance on WBC differentials with improvement after reclassification. Vision Pro could help improve laboratory workflow.

The hypothalamic neuroendocrine system is strongly implicated in body energy homeostasis. In particular, the degree of production and release of arginine vasopressin (AVP) in the hypothalamus is affected by plasma osmolality, and that hypothalamic AVP is responsible for thirst and osmolality-dependent water and metabolic balance. However, the osmolality-responsive intracellular mechanism within AVP cells that regulates AVP synthesis is not clearly understood. Here, we report a role for tonicity-responsive enhancer binding protein (TonEBP), a transcription factor sensitive to cellular tonicity, in regulating osmosensitive hypothalamic AVP gene transcription. Our immunohistochemical work shows that hypothalamic AVP cellular activity, as recognized by c-fos, was enhanced in parallel with an elevation in TonEBP expression within AVP cells following water deprivation. Interestingly, our in vitro investigations found a synchronized pattern of TonEBP and AVP gene expression in response to osmotic stress. Those results indicate a positive correlation between hypothalamic TonEBP and AVP production during dehydration. Promoter and chromatin immunoprecipitation assays confirmed that TonEBP can bind directly to conserved binding motifs in the 5'-flanking promoter regions of the AVP gene. Furthermore, dehydration- and TonEBP-mediated hypothalamic AVP gene activation was reduced in TonEBP haploinsufficiency mice, compared with wild TonEBP homozygote animals. Therefore, our result support the idea that TonEBP is directly necessary, at least in part, for the elevation of AVP transcription in dehydration conditions. Additionally, dehydration-induced reductions in body weight were rescued in TonEBP haploinsufficiency mice. Altogether, our results demonstrate an intracellular machinery within hypothalamic AVP cells that is responsible for dehydration-induced AVP synthesis.

Dear Editor,Chronic myeloid leukemia (CML), BCR-ABL1-positive, typically shows leukocytosis due to neutrophilia in various stages of maturation [1], and basophilia is one of the characteristic find...

Background:The Sysmex DI-60 system (Sysmex, Japan) is an automated cell image analyzer.This study aimed to evaluate the performance of the DI-60 system for the differential analysis of leukocytes.Methods: A total of 220 samples were analyzed in this study.The agreement between DI-60 pre-classification and manual verification by experts was determined.The correlation between the differential leukocyte counts obtained using the DI-60 system and those manually obtained in the peripheral blood smears were determined. Results:The pre-classification agreement of DI-60 was 91.0%.The correlation coefficients of normal five-part differentials were 0.9163 (segmented neutrophils), 0.9017 (lymphocytes), 0.8533 (monocytes), 0.8345 (eosinophils), and 0.3505 (basophils).The sensitivity, specificity, positive predictive value, negative predictive value, and the efficiency of counting the abnormal cells, including blasts, promyelocytes, myelocytes, metamyelocytes, lymphocyte variants, and erythroblasts, were determined.The efficiency of the DI-60 system in counting the blasts, promyelocytes, myelocytes, metamyelocytes, lymphocyte variants, and erythroblasts was 99.5%, 100.0%, 95.9%, 96.5%, 98.6%, 100.0%, and 95.9%, respectively.Conclusions: The pre-classification agreement of DI-60 was higher than that of previous studies.The correlation between the differential leukocyte counts obtained with the DI-60 system and those of manual counting was acceptable.The performance of DI-60 as a screening tool in clinical laboratories may be good; however, it is yet to replace manual slide review.

Background Although plasma free hemoglobin (fHb) test is important for assessing intravascular hemolysis, it is still dependent on the gold standard Harboe method using manual and labor-intensive spectrometric measurements at the wavelength of 380-415-450 nm. We established an automated fHb assay using a routine chemistry autoanalyzer that can be tuned to a wavelength of 380-416-450 nm. Methods The linearity, precision, accuracy, correlation, and sample carryover of fHb measurement using TBA200FRneo method and manual Harboe method were evaluated, respectively. fHb values measured by manual Harboe method were compared with those measured by our new automated TBA200FRneo method. Results fHb measurements were linear in the range of 0.05~38.75 mu mol/L by TBA200FRneo and 0.05~9.69 mu mol/L by manual Harboe method. Imprecision analysis (%CV) revealed 0.9~2.8% for TBA200FRneo method and 5.3~13.6% for the manual Harboe method. Comparison analysis showed 0.9986 of correlation coefficient (TBA200FRneo = 0.970 x Harboe + 0.12). In analytical accuracy analysis, the manual Harboe method revealed about 4 times higher average total error % (12.2%) than the TBA200FRneo automated method (2.8%). The sample carryover was -0.0016% in TBA200FRneo method and 0.0038% in Harboe method. Conclusions In the measurement of fHb, the automated TBA200FRneo method showed better performance than the conventional Harboe method. It is expected that the automated fHb assay using the routine chemistry analyzer can replace the gold standard Harboe method which is labor-intensive and need an independent spectrophotometry equipment.

Abstract Background A growing body of evidence shows that hypothalamic inflammation is an important factor in the initiation of obesity. In particular, reactive gliosis accompanied by inflammatory responses in the hypothalamus are pivotal cellular events that elicit metabolic abnormalities. In this study, we examined whether MyD88 signaling in hypothalamic astrocytes controls reactive gliosis and inflammatory responses, thereby contributing to the pathogenesis of obesity. Methods To analyze the role of astrocyte MyD88 in obesity pathogenesis, we used astrocyte-specific Myd88 knockout (KO) mice fed a high-fat diet (HFD) for 16 weeks or injected with saturated free fatty acids. Astrocyte-specific gene expression in the hypothalamus was determined using real-time PCR with mRNA purified by the Ribo-Tag system. Immunohistochemistry was used to detect the expression of glial fibrillary acidic protein, ionized calcium-binding adaptor molecule 1, phosphorylated signal transducer and activator of transcription 3, and α-melanocyte-stimulating hormone in the hypothalamus. Animals’ energy expenditure was measured using an indirect calorimetry system. Results The astrocyte-specific Myd88 KO mice displayed ameliorated hypothalamic reactive gliosis and inflammation induced by injections of saturated free fatty acids and a long-term HFD. Accordingly, the KO mice were resistant to long-term HFD-induced obesity and showed an improvement in HFD-induced leptin resistance. Conclusions These results suggest that MyD88 in hypothalamic astrocytes is a critical molecular unit for obesity pathogenesis that acts by mediating HFD signals for reactive gliosis and inflammation.