Evaluation of the Accuracy of Cr and BUN Using the ABL90 FLEX PLUS Blood Gas Analyzer and the Equivalence of Candidate Specimens for Assessment of Renal Function.

BACKGROUND: The ABL90 FLEX PLUS (Radiometer) is a blood gas analyzer that also provides creatinine (Cr) and blood urea nitrogen (BUN) results. We assessed the accuracy of the ABL90 FLEX PLUS to measure Cr and BUN and find suitable candidate specimens against primary specimens (heparinized whole-blood (H-WB)). METHODS: Paired H-WB, serum, and sodium-citrated whole-blood (C-WB) samples (105) were collected. The Cr and BUN levels in the H-WB using the ABL90 FLEX PLUS were compared with those of the serum using four automated chemistry analyzers. The suitability of the candidate specimens was assessed at each medical decision level according to the CLSI guideline EP35-ED1. RESULTS: The respective mean differences of the ABL90 FLEX PLUS for the Cr and BUN were below -0.10 and -3.51 mg/dL compared to the other analyzers. The systematic differences between the serum and the H-WB at the low, medium, and high medical decision levels were all 0% for Cr, but those of the C-WB were -12.96%, -11.81%, and -11.30%, respectively. Regarding imprecision, the SDserum/SDH-WB ratios at each level were 0.14, 1.41, and 0.68, whereas the SDC-WB/SDH-WB ratios were 0.35, 2.00, and 0.73, respectively. CONCLUSIONS: The ABL90 FLEX PLUS provided Cr and BUN results comparable with the four widely used analyzers. Among the candidates, the serum was suitable for Cr testing using the ABL90 FLEX PLUS, while the C-WB did not satisfy the acceptance criteria.

Transcriptome Analysis of Particulate Matter 2.5-Induced Abnormal Effects on Human Sebocytes.

Particulate matter 2.5 (PM2.5), an atmospheric pollutant with an aerodynamic diameter of <2.5 µm, can cause serious human health problems, including skin damage. Since sebocytes are involved in the regulation of skin homeostasis, it is necessary to study the effects of PM2.5 on sebocytes. We examined the role of PM2.5 via the identification of differentially expressed genes, functional enrichment and canonical pathway analysis, upstream regulator analysis, and disease and biological function analysis through mRNA sequencing. Xenobiotic and lipid metabolism, inflammation, oxidative stress, and cell barrier damage-related pathways were enriched; additionally, PM2.5 altered steroid hormone biosynthesis and retinol metabolism-related pathways. Consequently, PM2.5 increased lipid synthesis, lipid peroxidation, inflammatory cytokine expression, and oxidative stress and altered the lipid composition and expression of factors that affect cell barriers. Furthermore, PM2.5 altered the activity of sterol regulatory element binding proteins, mitogen-activated protein kinases, transforming growth factor beta-SMAD, and forkhead box O3-mediated pathways. We also suggest that the alterations in retinol and estrogen metabolism by PM2.5 are related to the damage. These results were validated using the HairSkin® model. Thus, our results provide evidence of the harmful effects of PM2.5 on sebocytes as well as new targets for alleviating the skin damage it causes.

Neutrophil Recruitment in Arterial Thrombus and Characteristics of Stroke Patients with Neutrophil-Rich Thrombus.

PURPOSE: Neutrophils contribute to thrombosis. However, there is limited information on the temporal course of neutrophil recruitment in thrombosis, the contribution of neutrophils to thrombus growth, and the characteristics of stroke patients with neutrophil-rich thrombi. MATERIALS AND METHODS: After inducing carotid artery thrombosis in Institute of Cancer Research mice using ferric chloride, aged thrombi were produced by ligating the distal portion of the carotid artery in mice for 0.5, 1, 2, 3, 6, or 24 h. For thrombus analysis in stroke patients, we used registry data and thrombi that were obtained during intra-arterial thrombectomy. Immunohistochemistry was performed to determine thrombus composition. RESULTS: In the thrombi of 70 mice, Ly6G positive cell counts (neutrophils) and histone H3-positive cell counts increased in a time-dependent manner (both p<0.001). Ly6G-positive cell count was strongly correlated with histone H3-positive cell counts (r=0.910, p<0.001), but not with thrombus size (p=0.320). In 75 stroke patients, atrial fibrillation and cardioembolism were more frequent in the higher neutrophil group (32/37, 86.5%) than in the lower neutrophil group (19/38, 50%) (p=0.002). The median erythrocyte fraction was higher [52.0 (interquartile range 39.9-57.8)] in the higher neutrophil group than in the lower neutrophil group [40.3 (interquartile range 23.5-53.2)]. The fraction of neutrophils was positively correlated with that of erythrocytes (R=0.35, p=0.002). CONCLUSION: Neutrophils were recruited and increased in arterial thrombosis in a time-dependent manner; however, they were not associated with the growth of formed thrombi. Neutrophil fractions in the thrombi of stroke patients appeared to be associated with atrial fibrillation and erythrocyte fraction.

Genomic Approach to the Assessment of Adverse Effects of Particulate Matters on Skin Cancer and Other Disorders and Underlying Molecular Mechanisms.

Air pollutants are in the spotlight because the human body can easily be exposed to them. Among air pollutants, the particulate matter (PM) represents one of the most serious toxicants that can enter the human body through various exposure routes. PMs have various adverse effects and classified as severe carcinogen by International Agency for Research on Cancer. Their physical and chemical characteristics are distinguished by their size. In this review, we summarized the published information on the physicochemical characteristics and adverse effects of PMs on the skin, including carcinogenicity. Through comparisons of biological networks constructed from relationships discussed in the previous scientific publications, we show it is possible to predict skin cancers and other disorders from particle-size-specific signaling alterations of PM-responsive genes. Our review not only helps to grasp the biological association between ambient PMs and skin diseases including cancer, but also provides new approaches to interpret chemical-gene-disease associations regarding the adverse effects of these heterogeneous particles.

Primary Ciliogenesis by 2-Isopropylmalic Acid Prevents PM2.5-Induced Inflammatory Response and MMP-1 Activation in Human Dermal Fibroblasts and a 3-D-Skin Model.

Particulate matters (PMs) increase oxidative stress and inflammatory response in different tissues. PMs disrupt the formation of primary cilia in various skin cells, including keratinocytes and melanocytes. In this study, we found that 2-isopropylmalic acid (2-IPMA) promoted primary ciliogenesis and restored the PM2.5-induced dysgenesis of primary cilia in dermal fibroblasts. Moreover, 2-IPMA inhibited the generation of excessive reactive oxygen species and the activation of stress kinase in PM2.5-treated dermal fibroblasts. Further, 2-IPMA inhibited the production of pro-inflammatory cytokines, including IL-6 and TNF-α, which were upregulated by PM2.5. However, the inhibition of primary ciliogenesis by IFT88 depletion reversed the downregulated cytokines by 2-IPMA. Moreover, we found that PM2.5 treatment increased the MMP-1 expression in dermal fibroblasts and a human 3-D-skin model. The reduced MMP-1 expression by 2-IPMA was further reversed by IFT88 depletion in PM2.5-treated dermal fibroblasts. These findings suggest that 2-IPMA ameliorates PM2.5-induced inflammation by promoting primary ciliogenesis in dermal fibroblasts.

Effect of leukopenia induced by cyclophosphamide on the initial stage of arterial thrombosis in mice.

INTRODUCTION: Leukocytes are found in organizing thrombi and are associated with thrombus growth. However, their role in the initial stage of thrombus formation is not well known. We investigated the role of leukocytes in the early stage of arterial thrombosis by inducing leukopenia. METHODS: In this double-blind, randomized, placebo-controlled study, 72 Institute of Cancer Research mice were randomly treated with intraperitoneal 100 mg/kg cyclophosphamide or normal saline. The primary outcome was time to occlusion after FeCl3 treatment. We also compared thrombus size, histological composition, and association with peripheral blood cell counts between cyclophosphamide and control groups. RESULTS: Cyclophosphamide treatment significantly decreased leukocyte counts by 82.8% compared to placebo (P < 0.001). The time to occlusion was significantly longer in the cyclophosphamide group (3.31 ± 1.59 min) than in the control group (2.30 ± 1.14 min; P = 0.003). The immunoreactivity for Ly6G-positive cells, intracellular histone H3, and released histone H3 in thrombi was significantly reduced in the cyclophosphamide group by 92.8%, 50.2%, and 34.3%, respectively. Time to occlusion had a moderate negative correlation with leukocyte count in peripheral blood (r = -0.326, P = 0.022) in the entire group. CONCLUSIONS: Cyclophosphamide-induced leukopenia attenuated thrombus formation during the early stage of arterial thrombosis. Our findings suggest the potential role of leukocytes in the initial stage of arterial thrombosis.

Diagnostic Validation of a Clinical Laboratory-Oriented Targeted RNA Sequencing System for Detecting Gene Fusions in Hematologic Malignancies.

Targeted RNA sequencing (RNA-seq) is a highly accurate method for sequencing transcripts of interest with a high resolution and throughput. However, RNA-seq has not been widely performed in clinical molecular laboratories because of the complexity of data processing and interpretation. We developed and validated a customized RNA-seq panel and data processing protocol for fusion detection using 4 analytical validation samples and 51 clinical samples, covering seven types of hematologic malignancies. Analytical validation showed that the results for target gene coverage and between- and within-run precision and linearity tests were reliable. Using clinical samples, RNA-seq based on filtering and prioritization strategies detected all 25 known fusions previously found by multiplex reverse transcriptase-PCR and fluorescence in situ hybridization. It also detected nine novel fusions. Known fusions detected by RNA-seq included two IGH rearrangements supported by expression analysis. Novel fusions included six that targeted just one partner gene. In addition, 18 disease- and drug resistance-associated transcript variants in ABL1, GATA2, IKZF1, JAK2, RUNX1, and WT1 were designated simultaneously. Expression analysis showed distinct clustering according to subtype and lineage. In conclusion, this study showed that our customized RNA-seq system had a reliable and stable performance for fusion detection, with enhanced diagnostic yield for hematologic malignancies in a clinical diagnostic setting.

Intense Pulsed Light Attenuates UV-Induced Hyperimmune Response and Pigmentation in Human Skin Cells.

The skin of an organism is affected by various environmental factors and fights against aging stress via mechanical and biochemical responses. Photoaging induced by ultraviolet B (UVB) irradiation is common and is the most vital factor in the senescence phenotype of skin, and so, suppression of UVB stress-induced damage is critical. To lessen the UVB-induced hyperimmune response and hyperpigmentation, we investigated the ameliorative effects of intense pulsed light (IPL) treatment on the photoaged phenotype of skin cells. Normal human epidermal keratinocytes and human epidermal melanocytes were exposed to 20 mJ/cm2 of UVB. After UVB irradiation, the cells were treated with green (525-530 nm) and yellow (585-592 nm) IPL at various time points prior to the harvest step. Subsequently, various signs of excessive immune response, including expression of proinflammatory and melanogenic genes and proteins, cellular oxidative stress level, and antioxidative enzyme activity, were examined. We found that IPL treatment reduced excessive cutaneous immune reactions by suppressing UVB-induced proinflammatory cytokine expression. IPL treatment prevented hyperpigmentation, and combined treatment with green and yellow IPL synergistically attenuated both processes. IPL treatment may exert protective effects against UVB injury in skin cells by attenuating inflammatory cytokine and melanogenic gene overexpression, possibly by reducing intracellular oxidative stress. IPL treatment also preserves antioxidative enzyme activity under UVB irradiation. This study suggests that IPL treatment is a useful strategy against photoaging, and provides evidence supporting clinical approaches with non-invasive light therapy.

The HLA-A*02:01:175 allele newly identified in a Korean hematopoietic stem cell donor by next-generation sequencing.

HLA-A*02:01:175 has a single synonymous nucleotide polymorphism when compared with HLA-A*02:01:01:01 [c165.G>C].

Changing the frequency and spectra of chromosomal aberrations in Korean patients with acute leukemia in a tertiary care hospital.

BACKGROUND: Chromosomal analysis is essential for the diagnosis and risk stratification of all leukemia patients. Not surprisingly, racial differences in chromosomal aberrations (CA) in hematological malignancies could be found, and CA incidence in leukemia might change over time, possibly due to environmental and lifestyle changes. Thus, we compared the frequency and range of CA in patients with acute leukemia (AL) during two time periods (2006‒2009 vs. 2010‒2015) and compared them with other prior studies. METHODS: We enrolled 717 patients with AL during a six-year period (2010‒2015). We compared the results to those of our earlier study (2006‒2009) [1]. Conventional cytogenetics, a multiplex reverse transcriptase (RT)-PCR system, and fluorescence in situ hybridization were employed to assess bone marrow specimens or peripheral blood at the diagnostic stage in AL patients to detect CA. RESULTS: The incidence of CA changed in the leukemia subgroups during the two time periods. Notably, the most frequent CA of childhood acute myeloid leukemia (AML) was PML/RARA, and was followed by RUNX1/RUNX1T1 in the current study. In contrast, the most common CA was RUNX1/RUNX1T1 in a previous study [1] and was followed by PML/RARA. In this study, the most frequent CA of the mixed phenotype AL was BCR/ABL1, which was followed by KMT2A/MLLT3. In a previous report, [1] the most frequent CA was BCR/ABL1, which was followed by KMT2A/ELL. CONCLUSION: The distribution of CA in Korean AL patients changed over time in a single institute. This change might be due to environmental and lifestyle changes.

Melasolv induces melanosome autophagy to inhibit pigmentation in B16F1 cells.

The melanosome is a specialized membrane-bound organelle that is involved in melanin synthesis, storage, and transportation. In contrast to melanosome biogenesis, the processes underlying melanosome degradation remain largely unknown. Autophagy is a process that promotes degradation of intracellular components' cooperative process between autophagosomes and lysosomes, and its role for process of melanosome degradation remains unclear. Here, we assessed the regulation of autophagy and its contributions to depigmentation associated with Melasolv (3,4,5-trimethoxycinnamate thymol ester). B16F1 cells-treated with Melasolv suppressed the α-MSH-stimulated increase of melanin content and resulted in the activation of autophagy. However, introduction of bafilomycin A1 strongly suppressed melanosome degradation in Melasolv-treated cells. Furthermore, inhibition of autophagy by ATG5 resulted in significant suppression of Melasolv-mediated depigmentation in α-MSH-treated cells. Taken together, our results suggest that treatment with Melasolv inhibits skin pigmentation by promoting melanosome degradation via autophagy activation.

ATP Production Relies on Fatty Acid Oxidation Rather than Glycolysis in Pancreatic Ductal Adenocarcinoma.

Glycolysis is known as the main pathway for ATP production in cancer cells. However, in cancer cells, glucose deprivation for 24 h does not reduce ATP levels, whereas it does suppress lactate production. In this study, metabolic pathways were blocked to identify the main pathway of ATP production in pancreatic ductal adenocarcinoma (PDAC). Blocking fatty acid oxidation (FAO) decreased ATP production by 40% in cancer cells with no effect on normal cells. The effects of calorie balanced high- or low-fat diets were tested to determine whether cancer growth is modulated by fatty acids instead of calories. A low-fat diet caused a 70% decrease in pancreatic preneoplastic lesions compared with the control, whereas a high-fat diet caused a two-fold increase in preneoplastic lesions accompanied with increase of ATP production in the Kras (G12D)/Pdx1-cre PDAC model. The present results suggest that ATP production in cancer cells is dependent on FAO rather than on glycolysis, which can be a therapeutic approach by targeting cancer energy metabolism.

Transglutaminase 2-Mediated p53 Depletion Promotes Angiogenesis by Increasing HIF-1α-p300 Binding in Renal Cell Carcinoma.

Angiogenesis and the expression of vascular endothelial growth factor (VEGF) are increased in renal cell carcinoma (RCC). Transglutaminase 2 (TGase 2), which promotes angiogenesis in endothelial cells during wound healing, is upregulated in RCC. Tumor angiogenesis involves three domains: cancer cells, the extracellular matrix, and endothelial cells. TGase 2 stabilizes VEGF in the extracellular matrix and promotes VEGFR-2 nuclear translocation in endothelial cells. However, the role of TGase 2 in angiogenesis in the cancer cell domain remains unclear. Hypoxia-inducible factor (HIF)-1α-mediated VEGF production underlies the induction of angiogenesis in cancer cells. In this study, we show that p53 downregulated HIF-1α in RCC, and p53 overexpression decreased VEGF production. Increased TGase 2 promoted angiogenesis by inducing p53 degradation, leading to the activation of HIF-1α. The interaction of HIF-1α and p53 with the cofactor p300 is required for stable transcriptional activation. We found that TGase 2-mediated p53 depletion increased the availability of p300 for HIF-1α-p300 binding. A preclinical xenograft model suggested that TGase 2 inhibition can reverse angiogenesis in RCC.

Inhibition of Transglutaminase 2 but Not of MDM2 Has a Significant Therapeutic Effect on Renal Cell Carcinoma.

More than 50% of human cancers harbor TP53 mutations and increased expression of Mouse double minute 2 homolog(MDM2), which contribute to cancer progression and drug resistance. Renal cell carcinoma (RCC) has an unusually high incidence of wild-type p53, with a mutation rate of less than 4%. MDM2 is master regulator of apoptosis in cancer cells, which is triggered through proteasomal degradation of wild-type p53. Recently, we found that p53 protein levels in RCC are regulated by autophagic degradation. Transglutaminase 2 (TGase 2) was responsible for p53 degradation through this pathway. Knocking down TGase 2 increased p53-mediated apoptosis in RCC. Therefore, we asked whether depleting p53 from RCC cells occurs via MDM2-mediated proteasomal degradation or via TGase 2-mediated autophagic degradation. In vitro gene knockdown experiments revealed that stability of p53 in RCC was inversely related to levels of both MDM2 and TGase 2 protein. Therefore, we examined the therapeutic efficacy of inhibitors of TGase 2 and MDM2 in an in vivo model of RCC. The results showed that inhibiting TGase 2 but not MDM2 had efficient anticancer effects.

Fc-saxatilin inhibits VEGF-induced permeability by regulating claudin-5 expression in human brain microvascular endothelial cells.

The disruption of the blood-brain barrier influences the degree of brain damage and prognosis in cerebral ischemia or other brain diseases accompanied by inflammation. Vascular endothelial growth factor (VEGF) released during brain ischemia or inflammation has been implicated in the breakdown of the blood-brain barrier by increasing endothelial permeability. Saxatilin, a disintegrin-containing RGD motif, has been reported to disaggregate platelets via interactions with platelet integrins and to have a thrombolysis effect. Additionally, the Fc-saxatilin fusion protein reduces vascular leakage in cerebral ischemia in mice. In this study, we show that Fc-saxatilin prevents VEGF-induced permeability in human brain microvascular endothelial cells (HBMECs). The activation of Src and Fak, downstream signaling proteins of VEGF in the induction of endothelial permeability, was inhibited by Fc-saxatilin in HBMECs. The downregulation of a tight junction protein, claudin-5, at the protein and mRNA levels by VEGF was recovered by Fc-saxatilin. Our findings suggest that Fc-saxatilin attenuates VEGF-induced endothelial permeability via the regulation of downstream signaling, and this may contribute to its protective effect against vascular leakage in the ischemic brain.

Transfer of a healthy microbiota reduces amyloid and tau pathology in an Alzheimer's disease animal model.

OBJECTIVE: Cerebral amyloidosis and severe tauopathy in the brain are key pathological features of Alzheimer's disease (AD). Despite a strong influence of the intestinal microbiota on AD, the causal relationship between the gut microbiota and AD pathophysiology is still elusive. DESIGN: Using a recently developed AD-like pathology with amyloid and neurofibrillary tangles (ADLPAPT) transgenic mouse model of AD, which shows amyloid plaques, neurofibrillary tangles and reactive gliosis in their brains along with memory deficits, we examined the impact of the gut microbiota on AD pathogenesis. RESULTS: Composition of the gut microbiota in ADLPAPT mice differed from that of healthy wild-type (WT) mice. Besides, ADLPAPT mice showed a loss of epithelial barrier integrity and chronic intestinal and systemic inflammation. Both frequent transfer and transplantation of the faecal microbiota from WT mice into ADLPAPT mice ameliorated the formation of amyloid ß plaques and neurofibrillary tangles, glial reactivity and cognitive impairment. Additionally, the faecal microbiota transfer reversed abnormalities in the colonic expression of genes related to intestinal macrophage activity and the circulating blood inflammatory monocytes in the ADLPAPT recipient mice. CONCLUSION: These results indicate that microbiota-mediated intestinal and systemic immune aberrations contribute to the pathogenesis of AD in ADLPAPT mice, providing new insights into the relationship between the gut (colonic gene expression, gut permeability), blood (blood immune cell population) and brain (pathology) axis and AD (memory deficits). Thus, restoring gut microbial homeostasis may have beneficial effects on AD treatment.

The natural phytochemical trans-communic acid inhibits cellular senescence and pigmentation through FoxO3a activation.

Ageing is characterized by the accumulation of chronic and irreversible oxidative damage, chronic inflammation and organ dysfunction. To attenuate these ageing-related changes, various natural phytochemicals are often applied. Trans-communic acid (TCA), an active component of brown pine leaf extract, has antimicrobial and cancer chemopreventive activity and inhibits ultraviolet B (UVB)-induced MMP-1 expression. To determine whether the phytochemical TCA could affect the lifespan of an ageing model, Caenorhabditis elegans prevent ageing-related phenotypes of the skin. Caenorhabditis elegans (C. elegans) wild-type N2 and mutant strains were used in this study to explore the lifespan extension effect of TCA and its mechanism. We estimated lipofuscin accumulation and melanin levels, which are closely associated with skin senescence. Moreover, we explored the mechanism of action associated with ageing attenuation. We performed oxidative stress resistance and thermotolerance assays in C. elegans and surface plasmon resonance analysis of TCA binding with the forkhead box-O3a (FoxO3a) protein. TCA, which is the active component in Korean red pine (Pinus densiflora), attenuated ageing-related changes in skin cells. TCA lowered lipofuscin accumulation in fibroblasts and decreased melanin levels in melanocytes. These protective effects were mediated by activation of the representative longevity gene FoxO3a, which was induced by direct binding with TCA. Interestingly, TCA extended the lifespan of C. elegans, although it did not affect stress resistance, oxidative stress or thermotolerance. These results strongly suggest that TCA prevents the senescent phenotype of model organisms and exhibits beneficial effects on ageing-related skin phenotypes through direct FoxO3a activation.

Histological features of intracranial thrombi in stroke patients with cancer.

The histological features of thrombus in stroke patients with cancer are not well known. Using immunohistochemical staining of thrombi retrieved during mechanical thrombectomy in stroke patients, thrombus compositions were compared between 16 patients with active cancer, 16 patients with inactive cancer, and 16 patients without any history of cancer. The active cancer group showed higher platelet and lower erythrocyte fractions than the inactive cancer or the control group. Four patients with vegetation showed very high platelet and low erythrocyte fractions. Patients with cryptogenic etiology in the active cancer group showed a similar pattern to those with vegetation. These findings may aid the determination of treatment strategies in cancer-associated stroke. ANN NEUROL 2019.