Lymphocyte Reaction Research Articles

Features of subset composition and functional activity of blood lymphocytes in tick-borne infections of different etiologies

Aim. To perform a comparative assessment of subset composition and functional activity of peripheral blood lymphocytes in patients with tick-borne encephalitis (TBE) and ixodid tick-borne borreliosis (ITBB) in the acute phase of the disease.Materials and methods. The study involved 22 patients with febrile and meningeal TBE, 15 patients with ITBB with and without erythema, and 11 healthy controls. Subset composition of blood lymphocytes was determined by flow cytometry. The blast transformation assay was applied to assess lymphocyte proliferation. Cytokine-producing activity of cells was studied in 24-hour incubated mononuclear cell cultures. Cytokine concentrations (interleukin (IL)-2, IL-4, IL-10, interferon (IFN)γ) were determined in the supernatants by the enzyme-linked immunosorbent assay (ELISA).Results. Patients with TBE demonstrated an increase in the proportion of helper – inducer T-cells, a pronounced decrease in the proportion and absolute count of cytotoxic T cells, and low T lymphocyte count compared to the control values. The study in ITBB patients revealed an increase in the helper – inducer T-cell count and the proportion of NK-cells, a decrease in the cytotoxic T cell count, and the T lymphocyte count comparable to normal values. The most significant decrease in the levels of phytohemagglutinin-induced lymphocyte proliferation was found in patients with TBE. Patients of both groups showed a decrease in IL-2 secretion in the mononuclear cell culture, a rise in IL-4 and IL-10 production, and IFNγ production levels comparable to control values.Conclusion. The study of TBE patients revealed relative lymphocytopenia with changes in the subset composition of lymphocytes characterized by an increase in the proportion of helper – inducer T-cells and a decrease in the absolute cytotoxic T lymphocyte count. Patients with ITBB demonstrated an increase in the proportion of NK-cells and a more pronounced imbalance in the T-helper / cytotoxic T lymphocyte ratio. Changes in the functional phenotype of lymphocytes, regardless of the etiology of tick-borne infection, were characterized by reduced proliferative reserve, low IL-2 secretion, increased IL-4 and IL-10 production, and depressed reactivity of lymphocytes with respect to IFNγ secretion.

A-156 Comparison of blast and variant lymphocyte flagging between the Alinity hq from Abbott and XN-1000 from Sysmex

Abstract Background Blast cells are immature white blood cells (WBC). The presence of blasts in the peripheral blood may be associated with certain diseases such as myelodysplastic syndrome and acute myelogenous leukemia. Variant lymphocytes (i.e., reactive lymphocytes, atypical lymphocytes) are WBCs produced by the immune system in reaction to infections caused by pathogens such as Epstein-Barr virus, cytomegalovirus, etc. It is clinically useful that the automated hematology analyzer is capable of flagging these cells. In this study, the blast (shown as ‘BLAST’ on the Alinity hq and ‘Blast/Abn Lympho?’ or ‘Blast?’ on XN-1000) and variant lymphocyte (shown as ‘VAR LYM’ on the Alinity hq, and ‘Atypical Lympho?’ on XN-1000) flagging was compared between the Alinity hq from Abbott and XN-1000 from Sysmex. Alinity hq has different sensitivity settings available for BLAST and VAR LYM flagging. In this study, the default setting for Alinity hq was used. For XN-1000, the Q-Flag default setting is 100. In this study, the setting was 120 for ‘Blast/Abn Lympho’ and 100 for “Atypical lympho?’. Methods Blood specimens were tested on the Alinity hq and XN-1000 analyzers. Two smears per specimen were made manually or by the Alinity hs slide maker stainer and read by two independent readers. The specimens showing any of the following flagging were included in the analysis: ‘BLAST’ on Alinity hq, ‘VAR LYM’ on Alinity hq, ‘Blast/Abn Lympho?’ or ‘Blast?’ on XN-1000, ‘Atypical lympho?’ on XN-1000. The flagging reports of these samples from both analyzers were compared to the smear review to determine if each analyzer's blast and variant lymphocyte flagging were true positive, false positive, true negative, or false negative. Results A total of 520 specimens were tested, yielding 41 specimens for blast flagging analysis and 80 specimens for variant lymphocyte flagging analysis. Due to the scarce availability of true positive samples from the manual review, true positive and false negative rate were not compared. For blast flagging using smear results of >=1% or >=5% as the positive criterion, Alinity hq yielded a 9.8% (=4/41) false positive rate and 87.8% (=36/41) true negative rate. XN-1000 yielded a 75.6% (=31/41) false positive rate and 22.0% (=9/41) true negative rate. For variant lymphocyte flagging using smear results of >=5% as the positive criterion, Alinity hq yielded a 6.3% (=5/80) false positive rate and 92.5% (=74/80) true negative rate. XN-1000 yielded a 91.3% (=73/80) false positive rate and 7.5% (=6/80) true negative rate. Conclusions This study demonstrated that Alinity hq achieved a much lower false positive rate and much higher true negative rate for both blast and variant lymphocyte flagging when compared to the XN-1000. Manual blast and variant lymphocyte morphology classification is laboratory- and technician-dependent. Both analyzers allow different sensitivity/specificity levels to flag blasts and variant lymphocytes. Each individual laboratory should evaluate and choose the setting that is most suitable for their specific patient populations to obtain the desired flagging rate.

Serum Procalcitonin, Hematology Parameters, and Cell Morphology in Multiple Clinical Conditions and Sepsis.

The clinical value of procalcitonin (PCT) in infection diagnosis and antibiotic stewardship is still unclear. This study aimed to investigate the association between serum PCT and different clinical conditions as well as other infectious/inflammatory parameters in different septic patients in order to elucidate the value of PCT detection in infection management. Chemiluminescence immunoassay was used for serum PCT analysis. Hematology analysis was used for complete blood cell count. Digital automated cell morphology analysis was used for blood cell morphology examination. Blood, urine, and stool cultures were performed according to routine clinical laboratory standard operating procedures. C-reactive protein (CRP) was analyzed by immunoturbidimetry. Erythrocyte sedimentation rate test was performed using natural sedimentation methods. Outpatients, ICU patients, and patients under 2 years of age with respiratory infections had higher serum PCT levels. Septic patients had the highest-serum PCT levels and other infection indexes. PCT levels in the blood, urine, and stool culture-positive patients were significantly higher than in culture-negative patients. The neutrophil granulation and reactive lymphocytes were observed together with the PCT-level increments in different septic patients, and these alterations were lessened after treatment. There was no significant change in monocyte morphology between pre- and posttreatment septic patients. Serum PCT is associated with neutrophil cytotoxicity and lymphocyte morphology changes in sepsis; thus, the combination of neutrophil and lymphocyte digital cell morphology evaluations with PCT detection may be a useful examination for guiding the clinical management of sepsis.

Adversarial training collaborating hybrid convolution-transformer network for automatic identification of reactive lymphocytes in peripheral blood.

Reactive lymphocytes may indicate diseases such as viral infections. Identifying these abnormal lymphocytes is crucial for disease diagnosis. Currently, reactive lymphocytes are mainly manually identified by pathological experts with microscopes and morphological knowledge, which is time-consuming and laborious. Some studies have used convolutional neural networks (CNNs) to identify peripheral blood leukocytes, but there are limitations in the small receptive field of the model. Our model introduces a transformer based on CNN, expands the receptive field of the model, and enables it to extract global features more efficiently. We also enhance the generalization ability of the model through virtual adversarial training (VAT) without changing the parameters of the model. Finally, our model achieves an overall accuracy of 93.66% on the test set, and the accuracy of reactive lymphocytes also reaches 88.03%. This work takes another step toward the efficient identification of reactive lymphocytes.

Priming lymphocyte responsiveness and differential T cell signaling in pediatric IBD patients with Cannabis use.

The prevalence of inflammatory bowel disease (IBD) has increased dramatically in recent years, particularly in pediatric populations. Successful remission with current therapies is limited and often transient, leading patients to seek alternative therapies for symptom relief, including the use of medical marijuana (Cannabis sativa). However, chronic cannabis use among IBD patients is associated with increased risk for surgical interventions. Therefore, determining the direct impact of cannabis use on immune modulation in IBD patients is of critical importance. Peripheral blood mononuclear cells of cannabis using and non-using pediatric IBD patients were phenotyped by flow cytometry and functionally assessed for their cytokine production profile. A phospho-kinase array was also performed to better understand changes in immune responses. Results were then compared with serum phytocannabinoid profiles of each patient to identify cannabinoid-correlated changes in immune responses. Results demonstrated elevated levels of a myriad of pro-inflammatory cytokines in users versus non-users. Differences in signaling cascades of activated T cells between users and non-users were also observed. A number of anti-inflammatory cytokines were inversely correlated with serum phytocannabinoids. These results suggest that cannabis exposure, which can desensitize cannabinoid receptors, may prime pro-inflammatory pathways in pediatric IBD patients.

Blood smears analysis: a valuable tool for profiling circulating erythrocytes and leukocytes in mouse model of leishmaniasis

Although blood smear analysis can yield valuable insights into the health status of laboratory animals, it is not frequently performed in studies involving mouse models of leishmaniasis. Thus, to underscore its importance, we conducted microscopic evaluation of blood smears prepared from 10 µL of blood from L. (L.) amazonensis-infected BALB/c mice. Subsequently, we explored potential correlations between the identified hematological alterations, clinical evaluation data, and serum levels of total proteins (TP), albumin (Alb), globulins (Glob), IL-17A, and IL-10. Our findings revealed that infected mice exhibited an increased erythrocyte count, alongside a higher prevalence of microcytic and hypochromic erythrocytes. Moreover, it was observed elevated relative and absolute numbers of neutrophils and monocytes and reduced relative and absolute numbers of lymphocytes. A significant proportion of monocytes and lymphocytes in infected animals displayed reactive features. The relative number of lymphocytes was negatively correlated with the serum levels of Glob. Furthermore, the frequency of reactive lymphocytes was positively correlated with the serum levels of Glob and IL-17A. There was also a positive correlation between the absolute number of neutrophils and the serum levels of IL-17A and the frequency of reactive monocytes and the serum levels of Glob. Therefore, these findings, collectively, underscore the importance of blood smear analysis for elucidating both the quantitative and qualitative hematological changes experimentally induced by Leishmania infection. Integration of this technique into research protocols involving different mouse models of leishmaniasis holds immense potential for advancing our knowledge of this infectious disease and enhancing clinical management strategies.

Evaluation of Biochemical and Hematological Dynamics as Markers for Severe Dengue Forms

Dengue is a viral infection that has become a serious problem in recent years. It is a major cause of mortality and morbidity. The present study is a prospective, hospital-based, observational study done from 2015 to October 2022. The objective of our study was to consider whether dengue infection can be suspected based on hematological and biochemical findings. The study included 100 patients positive for dengue infection. Complete haemograms, transaminases for liver injury, blood urea, and serum creatinine levels for renal assessment were performed for these patients. The most common hematological findings were thrombocytopenia, leucopenia, an increase in the mixed cell fraction of the leucocytes, and the presence of reactive lymphocytes. Biochemical parameters like aminotransferases, blood urea, and serum creatinine levels were significantly raised. Hence, a platelet count of <100,000 cells/μl, leucopenia of < 4000 cells/μl, and aspartate aminotransferase levels of >82.2 U/L can be considered predictors of dengue infection

Cytomorphological findings in confirmed cases of tubercular lymphadenitis.

Tuberculosis (TB) remains a major health problem, especially in the developing countries. Fine-needle aspiration cytology is the first line of investigation for tubercular lymphadenitis as it is easy to perform, less invasive, quick, and economical. The typical cytopathological features of tuberculosis TB include epithelioid cell granulomas with Langhans giant cells and caseous necrosis. The present study aimed to evaluate the cytomorphological features of newly diagnosed cases of tubercular lymphadenitis confirmed by GeneXpert. This was a retrospective study in which all fine-needle aspirates from newly diagnosed cases of tubercular lymphadenitis confirmed by GeneXpert over a 1-year period from July 2022 to July 2023 were included in the study. The May-Grunwald-Giemsa stained smears from these aspirates were categorized into three patterns-epithelioid cell granulomas with necrosis, epithelioid granulomas without necrosis, and necrosis only. The granulomas were further categorized into well-formed, ill--formed, and splintered. The background of the aspirate which included a reactive lymphoid background, lymphocytes, eosinophils, and neutrophils was tabulated for all the cases. Out of the three cytomorphological patterns, epithelioid granulomas with necrosis were the most predominant (67.5%), followed by necrosis only (20.8%) and granulomas without necrosis (11.6%). An acid-fast bacilli (AFB) positivity of 53.3% (64 cases) was seen on the Ziehl-Neelsen stain. Well-composed, poorly formed, and splintered granulomas were seen in 55 (57.9%), 42 (44.2%), and 21 (22.1%) cases, respectively. Neutrophils were the most common background population (42, 35%) followed by lymphocytes (36, 30%). Reactive lymphoid cells and eosinophils were seen in 12 (10%) and 3 (2.5%) cases, respectively. Fine-needle aspiration cytology is a rapid inexpensive minimally invasive test for tubercular lymphadenitis as epithelioid cell granulomas along with caseous necrosis are highly suggestive of TB. However, manual acid-fast AFB detection has a low sensitivity as is illustrated in the present study where only AFBwas demonstrated in only 53.3% of cases.

Abstract 4149: Real-time intravital trafficking of tumor infiltrating lymphocytes in breast cancer models

Abstract Cancer immunotherapy utilizing the immunoreactive efficiency of immune cells via adoptive cell therapy (ACT) has been a novel method of cancer treatment. In tumor microenvironment, immune cell reactivity of tumor-infiltrating lymphocytes (TILs) can facilitate cancer cell attack, which finally derives efficient suppression of tumor growth. However, the conventional evaluation for cancer immunotherapy has been generally performed by ex vivo methods, providing restricted information on lymphocyte motility and immune cell dynamics within the tumor microenvironment. In vivo data with the dynamic behavior of TILs in breast cancer can provide new insight to investigate novel targets of immune checkpoint inhibitors. In this research, longitudinal intravital imaging of motility trafficking of TILs in breast cancer animal models has been performed by utilizing intravital microscopy to unveil the immunoreactive dynamics of T cell activity in early stage. In vivo analysis of TIL motility within different breast cell lines, including 4T1 (mouse breast cancer cell), MDA-MB-231 (human triple-negative breast cancer cell), and MCF7 (human breast cancer cell), respectively, has been investigated for immune responses to cancer. We’ve monitored spatiotemporal dynamics of TIL over 11 days in breast cancer xenograft mouse model generated by tumor implantation into the dorsal skinfold imaging chamber and adoptive T cell transfer by blood circulation. In the results, the adoptive immunoreactivity of TIL has been differentiated by the timepoint of adoptive T cell transfer, distance proximity of T cells in tumors, and breast cancer types. Furthermore, anti-cancer effect coordinated with dendritic cells behaviors showed induced immune responses with accumulated dendritic cells with increased TIL count over time in breast cancer model treated with various cancer therapy such as immunotherapy, radiation therapy and anti-cancer drug (anti-PD-L1), which implied a relationship between programmed death-ligand 1 expression levels and TIL/dendritic cell motility. The result demonstrates that intravital TIL motility trafficking analysis in cancer model can be an invaluable assessment method to explain the detailed cellular/molecular mechanism of immune response to cancer involved with T lymphocytes, dendritic cells, and immune checkpoint inhibitors to accelerate the efficiency of cancer immunotherapy. Citation Format: Soyeon Ahn, Kubra Akyildiz, Hyunseok Kim, Pilhan Kim. Real-time intravital trafficking of tumor infiltrating lymphocytes in breast cancer models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4149.

Haematological Profile and its Clinical Implications in Dengue Viral Infections

Introduction: Dengue fever is the most common arboviral disease in the world. Transmission to humans of dengue virus initiates a spectrum of illness, from inapparent to severe and sometimes lethal infections. Complete blood count (CBC) is an important part of diagnostic workup of patients. Comparison of various findings in CBC and peripheral blood film can help the physician in diagnosis, management and assessment of prognosis of the patients. Objectives: To find out the pattern of haematological findings in dengue infection. Methods: This cross sectional study was carried out on 62 dengue cases in CMH Savar from May to October 2019. Diagnosis of dengue was based on clinical features, haematological and biochemical findings and confirmed by serological tests. Haematological parameters analyzed include haemoglobin (Hb), haematocrit, total leucocyte count (TLC), differential leucocyte count (DLC), platelet count and peripheral blood film. Results: Important findings in haematological profile were haemoconcentration, raised haematocrit, low WBC count, early neutrophilia and late lymphocytosis, reactive lymphocytes in peripheral blood and thrombocytopenia. Haemoglobin ranged from 10.0-18.5gm/dl. Haematocrit was raised in 13(20.96%) cases. Thrombocytopenia (platelet count <1.5 Lac/mm3) was observed in 59(95.16%) cases. Platelet count 1-1.5 Lac was seen in 4(6.45%) cases, 50,000-1 Lac in 25(40.32%) cases, 20,000-50,000 in 26(41.93%) cases and < 20,000 in 4(6.45%) cases. Leucopenia (TLC<4000/mm3) was seen in 46(74.19%) cases and >11000mm3 in 2(3.22%) cases. On peripheral blood smear reactive lymphocytes including plasmacytoid lymphocytes were seen in almost all the cases. Conclusion: CBC examination gives enough clue for early diagnosis of dengue infection which can be confirmed by dengue serology. Serial CBC examination is also important in disease monitoring, treatment of the patient and prediction of prognosis. JAFMC Bangladesh. Vol 19, No 1 (June) 2023:11-14

From periphery immunity to central domain through clinical interview as a new insight on schizophrenia

Identifying disease predictors through advanced statistical models enables the discovery of treatment targets for schizophrenia. In this study, a multifaceted clinical and laboratory analysis was conducted, incorporating magnetic resonance spectroscopy with immunology markers, psychiatric scores, and biochemical data, on a cohort of 45 patients diagnosed with schizophrenia and 51 healthy controls. The aim was to delineate predictive markers for diagnosing schizophrenia. A logistic regression model was used, as utilized to analyze the impact of multivariate variables on the prevalence of schizophrenia. Utilization of a stepwise algorithm yielded a final model, optimized using Akaike’s information criterion and a logit link function, which incorporated eight predictors (White Blood Cells, Reactive Lymphocytes, Red Blood Cells, Glucose, Insulin, Beck Depression score, Brain Taurine, Creatine and Phosphocreatine concentration). No single factor can reliably differentiate between healthy patients and those with schizophrenia. Therefore, it is valuable to simultaneously consider the values of multiple factors and classify patients using a multivariate model.

Automatic classification of acute lymphoblastic leukemia cells and lymphocyte subtypes based on a novel convolutional neural network.

Acute lymphoblastic leukemia (ALL) is a life-threatening disease that commonly affects children and is classified into three subtypes: L1, L2, and L3. Traditionally, ALL is diagnosed through morphological analysis, involving the examination of blood and bone marrow smears by pathologists. However, this manual process is time-consuming, laborious, and prone to errors. Moreover, the significant morphological similarity between ALL and various lymphocyte subtypes, such as normal, atypic, and reactive lymphocytes, further complicates the feature extraction and detection process. The aim of this study is to develop an accurate and efficient automatic system to distinguish ALL cells from these similar lymphocyte subtypes without the need for direct feature extraction. First, the contrast of microscopic images is enhanced using histogram equalization, which improves the visibility of important features. Next, a fuzzy C-means clustering algorithm is employed to segment cell nuclei, as they play a crucial role in ALL diagnosis. Finally, a novel convolutional neural network (CNN) with three convolutional layers is utilized to classify the segmented nuclei into six distinct classes. The CNN is trained on a labeled dataset, allowing it to learn the distinguishing features of each class. To evaluate the performance of the proposed model, quantitative metrics are employed, and a comparison is made with three well-known deep networks: VGG-16, DenseNet, and Xception. The results demonstrate that the proposed model outperforms these networks, achieving an approximate accuracy of 97%. Moreover, the model's performance surpasses that of other studies focused on 6-class classification in the context of ALL diagnosis. RESEARCH HIGHLIGHTS: Deep neural networks eliminate the requirement for feature extraction in ALL classification The proposed convolutional neural network achieves an impressive accuracy of approximately 97% in classifying six ALL and lymphocyte subtypes.

Association of Blood Metabolomics Biomarkers with Brain Metabolites and Patient-Reported Outcomes as a New Approach in Individualized Diagnosis of Schizophrenia.

Given its polygenic nature, there is a need for a personalized approach to schizophrenia. The aim of the study was to select laboratory biomarkers from blood, brain imaging, and clinical assessment, with an emphasis on patients' self-report questionnaires. Metabolomics studies of serum samples from 51 patients and 45 healthy volunteers, based on the liquid chromatography-electrospray ionization-mass spectrometry (LC-ESI-MS/MS), led to the identification of 3 biochemical indicators (cortisol, glutamate, lactate) of schizophrenia. These metabolites were sequentially correlated with laboratory tests results, imaging results, and clinical assessment outcomes, including patient self-report outcomes. The hierarchical cluster analysis on the principal components (HCPC) was performed to identify the most homogeneous clinical groups. Significant correlations were noted between blood lactates and 11 clinical and 10 neuroimaging parameters. The increase in lactate and cortisol were significantly associated with a decrease in immunological parameters, especially with the level of reactive lymphocytes. The strongest correlations with the level of blood lactate and cortisol were demonstrated by brain glutamate, N-acetylaspartate and the concentrations of glutamate and glutamine, creatine and phosphocreatine in the prefrontal cortex. Metabolomics studies and the search for associations with brain parameters and self-reported outcomes may provide new diagnostic evidence to specific schizophrenia phenotypes.

Characterization of CD4+ and CD8+ T cells responses in the mixed lymphocyte reaction by flow cytometry and single cell RNA sequencing.

The Mixed Lymphocyte Reaction (MLR) consists in the allogeneic co-culture of monocytes derived dendritic cells (MoDCs) with T cells from another donor. This in vitro assay is largely used for the assessment of immunotherapy compounds. Nevertheless, the phenotypic changes associated with lymphocyte responsiveness under MLR have never been thoroughly evaluated. Here, we used multiplex cytokine and chemokine assays, multiparametric flow cytometry and single cell RNA sequencing to deeply characterize T cells activation and function in the context of CD4+- and CD8+-specific MLR kinetics. We showed that CD4+ and CD8+ T cells in MLR share common classical markers of response such as polyfunctionality, increased proliferation and CD25 expression but differ in their kinetics and amplitude of activation as well as their patterns of cytokines secretion and immune checkpoints expression. The analysis of immunoreactive Ki-67+CD25+ T cells identified PBK, LRR1 and MYO1G as new potential markers of MLR response. Using cell-cell communication network inference and pathway analysis on single cell RNA sequencing data, we also highlighted key components of the immunological synapse occurring between T cells and the stimulatory MoDCs together with downstream signaling pathways involved in CD4+ and CD8+ T cells activation. These results provide a deep understanding of the kinetics of the MLR assay for CD4+ or CD8+ T cells and may allow to better characterize compounds impacting MLR and eventually identify new strategies for immunotherapy in cancer.

Performance comparison of two automated digital morphology analyzers for leukocyte differential in patients with malignant hematological diseases: Mindray MC-80 and Sysmex DI-60.

The MC-80 (Mindray, Shenzhen, China), a newly available artificial intelligence (AI)-based digital morphology analyzer, is the focus of this study. We aim to compare the leukocyte differential performance of the Mindray MC-80 with that of the Sysmex DI-60 and the gold standard, manual microscopy. A total of 100 abnormal peripheral blood (PB) smears were compared across the MC-80, DI-60, and manual microscopy. Sensitivity, specificity, predictive value, and efficiency were calculated according to the Clinical and Laboratory Standards Institute (CLSI) EP12-A2 guidelines. Comparisons were made using Bland-Altman analysis and Passing-Bablok regression analysis. Additionally, within-run imprecision was evaluated using five samples, each with varying percentages of mature leukocytes and blasts, in accordance with CLSI EP05-A3 guidelines. The within-run coefficient of variation (%CV) of the MC-80 for most cell classes in the five samples was lower than that of the DI-60. Sensitivities for the MC-80 ranged from 98.2% for nucleated red blood cells (NRBC) to 28.6% for reactive lymphocytes. The DI-60's sensitivities varied between 100% for basophils and reactive lymphocytes, and 11.1% for metamyelocytes. Both analyzers demonstrated high specificity, negative predictive value, and efficiency, with over 90% for most cell classes. However, the DI-60 showed relatively lower specificity for lymphocytes (73.2%) and lower efficiency for blasts and lymphocytes (80.1% and 78.6%, respectively) compared with the MC-80. Bland-Altman analysis indicated that the absolute mean differences (%) ranged from 0.01 to 4.57 in MC-80 versus manual differential and 0.01 to 3.39 in DI-60 versus manual differential. After verification by technicians, both analyzers exhibited a very high correlation (r = 0.90-1.00) with the manual differential results in neutrophils, lymphocytes, and blasts. The Mindray MC-80 demonstrated good performance for leukocyte differential in PB smears, notably exhibiting higher sensitivity for blasts identification than the DI-60.

Extended Inflammation Parameters (EIP) as Markers of Inflammation in Systemic Sclerosis.

Systemic sclerosis (SSc) is an autoimmune disease characterized by inflammation, progressive vasculopathy, and fibrosis of skin and internal organs. The aim of the study was to evaluate extended inflammatory parameters (EIP) in patients with SSc in comparison to the control group of healthy subjects. A total of 28 patients with SSc and 29 healthy controls (HCs) were included in the study. The following EIP parameters were analyzed: neutrophil reactive intensity (NEUT-RI), neutrophil granularity intensity (NEUT-GI), antibody-synthesizing lymphocytes (AS-LYMP), and reactive lymphocytes (RE-LYMP). Patients with SSc showed significantly higher values of parameters determining neutrophil reactivity and neutrophil granularity when compared to HCs (respectively, 49.16 FI vs. 44.33 FI, p < 0.001, and 152.01 SI vs. 147.51 SI, p < 0.001). Moreover, patients with SSc had higher absolute numbers of RE-LYMP than HCs (0.69 × 103/µl vs. 0.04 × 103/µl, p < 0.001). Importantly, significant correlations between the RE-LYMP and either IL-6 (R = 0.447, p < 0.001) or ESR (R = 0.532, p < 0.001) were found among patients with SSc. Changes in NEUT-RI, NEUT-GI, and RE-LYMP levels positively correlate with inflammation in SSc and, thus, could potentially be used as an additional reliable inflammatory biomarker to assess inflammation in this disease.

КЛИНИЧЕСКИЕ ОСОБЕННОСТИ ОСТРОЙ ЦИТОМЕГАЛОВИРУСНОЙ ИНФЕКЦИИ, СОПРОВОЖДАЮЩЕЙСЯ ТРОМБОТИЧЕСКИМИ ОСЛОЖНЕНИЯМИ, У ИММУНОКОМПЕТЕНТНЫХ ПАЦИЕНТОВ

Objective of the study. To identify and analyse clinical and laboratory features in immunocompetent patients with thrombotic complications in acute cytomegalovirus infection. Development of the algorithm of prognosis and early diagnosis of thrombotic complications in order to reduce the risk of severe course of the disease and lethal outcome. Materials and Methods. The study included 100 immunocompetent adult patients undergoing inpatient observation and treatment with laboratory-proven acute cytomegalovirus infection. All patients had a detailed clinical and epidemiological history for the possible presence of modifiable and/or non-modifiable prothrombotic risk factors; a multifaceted laboratory and instrumental examination was performed. Results. Out of one hundred patients with acute cytomegalovirus infection selected in the study, thrombotic diseases were observed in 7% of cases (in 7 patients). The mean age of the patients was 41 (38; 42) years. Patients were hospitalised on average on 12 (10; 14) days from the onset of the disease. The main complaints on admission were: prolonged fever up to 38°C and marked general weakness. The patients had elevated ALT and AST in 88% and 80% of cases, respectively. The presence of reactive lymphocytes was registered in 57% of cases. Enlargement of liver and spleen was observed in 67% and 70% of cases, respectively. Compared to patients with uncomplicated course of acute cytomegalovirus infection, statistically significant laboratory differences were observed in SRB (p&lt;0.001) and D-dimers (p&lt;0.05) in patients who had thrombotic complications. All patients who had genetic material (DNA) to CMV in the blood had positive tests for rr65 antigenemia. The polymorphism of factor V gene G1691A (FVLeiden) was detected in only one patient, who was a carrier of heterozygous GA allele. Conclusion. Acute cytomegalovirus infection in immunocompetent patients includes a symptom complex consisting of a typical clinical picture (prolonged febrile intoxication, hepatosplenomegaly, elevated hepatic transaminases, CRP and D-dimer levels) and obligatory laboratory detection of the virus (positive PCR and/or Ag pp65 tests for CMV). Cytomegalovirus is an independent and reliable prothrombotic risk factor, with thrombotic complications occurring in 7% of cases. Patients with risk factors for thrombotic complications with verified acute cytomegalovirus infection are recommended to be prescribed anticoagulant therapy at prophylactic doses. Also, patients with a proven case of thrombotic process on the background of acute cytomegalovirus infection are recommended a course of etiotropic antiviral therapy.

Анализ клинических и лабораторных предикторов сочетанной инфекции безэритемной формы иксодового клещевого боррелиоза и клещевого энцефалита

Laboratory confirmation of the diagnosis of tick-borne infections is often hampered due to delayed seroconversion. Objective. Assessment of clinical and laboratory predictors to develop a differential diagnosis model between the mixed infection of Lyme borreliosis with tick-borne encephalitis and the monoinfection of Lyme borreliosis non-erythemal form at the onset of the disease. Patients and methods. In the first week of the disease, 54 clinical and laboratory parameters were studied in 27 patients with the mixed infection of Lyme borreliosis with tick-borne encephalitis (mean age: 47.2 years) and 65 patients with borreliosis monoinfection (mean age: 45.3 years). The logistic regression model was developed using Statistica 12.0. Results. Predictive factors for diagnosis of the mixed infection included painful eye movement, photophobia, nausea, meningism, intoxication syndrome intensity (AUC = 0.69) fever intensity (AUC = 0.79), neutrophil to lymphocyte ratio (AUC = 0.75), neutrophil to monocyte ratio (AUC = 0.73), neutrophils count (AUC = 0.76), immature granulocytes count (AUC = 0.80), lymphocytes count (AUC = 0.73) and reactive lymphocytes count (AUC = 0.70). A logistic regression model was developed with very good prognosis of mixed infection (AUC = 0.88). Conclusion. The model is allowed to assess the probability of a preliminary diagnosis of the mixed infection of Lyme borreliosis and tick-borne encephalitis with good sensitivity and specificity values. Key words: Lyme borreliosis, tick-borne encephalitis, mixed infection, non-erythemal form, clinical predictors, neutrophil to monocyte ratio, hemogram parameters, logistic regression model

Current Landscape of Immune Checkpoint Inhibitors for Metastatic Urothelial Carcinoma: Is There a Role for Additional T-Cell Blockade?

Urothelial carcinoma (UC) is the most common form of bladder cancer (BC) and is the variant with the most immunogenic response. This makes urothelial carcinoma an ideal candidate for immunotherapy with immune checkpoint inhibitors. Key immune checkpoint proteins PD-1 and CTLA-4 are frequently expressed on T-cells in urothelial carcinoma. The blockade of this immune checkpoint can lead to the reactivation of lymphocytes and augment the anti-tumor immune response. The only immune checkpoint inhibitors that are FDA-approved for metastatic urothelial carcinoma target the programmed death-1 receptor and its ligand (PD-1/PD-L1) axis. However, the overall response rate and progression-free survival rates of these agents are limited in this patient population. Therefore, there is a need to find further immune-bolstering treatment combinations that may positively impact survival for patients with advanced UC. In this review, the current immune checkpoint inhibition treatment landscape is explored with an emphasis on combination therapy in the form of PD-1/PD-L1 with CTLA-4 blockade. The investigation of the current literature on immune checkpoint inhibition found that preclinical data show a decrease in tumor volumes and size when PD-1/PD-L1 is blocked, and similar results were observed with CTLA-4 blockade. However, there are limited investigations evaluating the combination of CTLA-4 and PD-1/PD-L1 blockade. We anticipate this review to provide a foundation for a deeper experimental investigation into combination immune checkpoint inhibition therapy in metastatic urothelial carcinoma.

CytogeneticsPathBrowser: an educational tool to convert textbook images and captions into flashcards

Abstract Background Competence in reaching the correct diagnosis from analysis of clinical pathology specimens requires exposure to a large volume and breadth of cases. Images from textbooks are critical resources that supplement actual cases with well curated information. However, learning from textbooks has several setbacks. Images from individual textbooks are limited in number and often do not adequately capture variation of the disease entity. Textbooks also do not usually provide a platform for learners to assess their diagnostic abilities. This burdens the learner with creating flashcards for self-assessment, which is manually intensive. Methods To address these limitations, we developed PathBrowser, a web browser-accessible program that leverages the PyMuPDF Python package (Artifex) to extract images and captions from textbooks in PDF format. Captions are written to the image files using ExifTool (Phil Harvey), making the images searchable by keywords. The program then displays the image while hiding the caption, challenging the user to enter the appropriate diagnosis. To minimize work by the user to prepare flashcard questions and answers, the program can automatically generate quiz questions by using natural language processing tools and a Naïve Bayes (NB) classifier (NLTK and SpaCy packages) to identify important terms from the caption as either diagnoses or cytologic features. Results As a proof of principle, we used PathBrowser to build an image repository of myeloblasts and common mimickers, including reactive lymphocytes and chronic lymphoid leukemias, towards training junior pathology residents to identify blasts in peripheral blood and body fluids. In total, PathBrowser extracted 173 images (63 myeloblasts and 110 non-blasts) from 2 textbook sources (Atlas of Diagnostic Hematology by Salama et al., and Hematopathology by Jaffe et al.). We trained a NB classifier to label images as “myeloblast” or “non-myeloblast,” based on keywords in the caption. We manually ascertained that PathBrowser extracted the appropriate caption from 30/30 (100%) images of myeloblasts. Of these, the NB classifier correctly extracted the term “myeloblast” in 28 (93%) images. A survey of pathology residents showed that 89% of respondents agreed the program is a useful pathology learning tool. Conclusions We developed a program that makes reading from textbooks an interactive learning experience. By automating the manual-intensive steps of extracting images and captions and preparing flashcards with quiz questions using caption terms classified by machine learning tools, learners can focus on recognizing the diagnostic features represented by the images. Finally, our program enables the rapid generation of a repository of images from diverse sources, so that learners can browse a wide spectrum of disease entities and their variants.