HUMAN CYTOPENIA VARIANTS AT DIVERSE HIV INFECTION STAGES

Over decades, HIV infection and its complications have been one of the most debated problems in the world. The human immunodeficiency virus leads not only to weakened immune system, but also disrupts normal hematopoiesis manifested as cytopenia (anemia, thrombocytopenia and neutropenia). Materials and methods. A retrospective analysis of cases of combined HIV infection and inhibited hematopoiesis was carried out according to hemogram data of patients admitted for treatment at the IKB No. 1 named after D. Dalmatov, Omsk. The inclusion criteria were cytopenia during hospitalization detected in detailed blood test (by calculating hemoglobin level, counts of erythrocytes, leukocytes, platelets). The age of the patients included in the study differed: from 20 to 29 years - 27 patients (24.6%), from 30 to 39 years - 69 subjects (62.7%), from 40 to 49 years - 13 patients (11.8%), over 50 years old 1 patient (0.9%). All patients had suppression of at least one hematopoietic cell lineage. Anemia was considered as decreased hemoglobin level below than 130 g / l in men and 120 g / l in women. Erythrocytopenia was considered as decreased erythrocyte count below 4.76x10 * 12 / l. Leukopenia was defined as decreased total count of leukocytes below 4.0x10 * 9 / L, while a decrease in the absolute count of neutrophils below 1000 cells / μL was considered as neutropenia. Thrombocytopenia was determined as decreased platelet count below 150x10 * 9 \ l. Results. All patients had suppression of at least one hematopoietic cell lineage. 6 patients with stage 2 had one-cell lineage cytopenias, 7 – two- cell lineages. While analyzing the data obtained, it can be concluded that in patients with stage 2 HIV, inhibition of erythroid and platelet cell lineage predominates, whereas thrombocytopenia reached grade IV. At stage 3 HIV, all 7 patients had inhibition of only one cell lineage. In this group, the inhibition of hematopoiesis had a lighter degree in all hematopoietic cell lineages. In 46 patients with stage 4, there were various oppression of one of the hematopoietic cell lineages, in 44 patients there were two-cell lineage cytopenias. For patients with a more advanced stage of HIV, a decrease in the number of all cellular elements of the blood in the hemogram is characteristic; these disorders are more severe and persistent.

Transcriptomic landscape of early age onset of colorectal cancer identifies novel genes and pathways in Indian CRC patients

Past decades of the current millennium have witnessed an unprecedented rise in Early age Onset of Colo Rectal Cancer (EOCRC) cases in India as well as across the globe. Unfortunately, EOCRCs are diagnosed at a more advanced stage of cancer. Moreover, the aetiology of EOCRC is not fully explored and still remains obscure. This study is aimed towards the identification of genes and pathways implicated in the EOCRC. In the present study, we performed high throughput RNA sequencing of colorectal tumor tissues for four EOCRC (median age 43.5 years) samples with adjacent mucosa and performed subsequent bioinformatics analysis to identify novel deregulated pathways and genes. Our integrated analysis identifies 17 hub genes (INSR, TNS1, IL1RAP, CD22, FCRLA, CXCL3, HGF, MS4A1, CD79B, CXCR2, IL1A, PTPN11, IRS1, IL1B, MET, TCL1A, and IL1R1). Pathway analysis of identified genes revealed that they were involved in the MAPK signaling pathway, hematopoietic cell lineage, cytokine–cytokine receptor pathway and PI3K-Akt signaling pathway. Survival and stage plot analysis identified four genes CXCL3, IL1B, MET and TNS1 genes (p = 0.015, 0.038, 0.049 and 0.011 respectively), significantly associated with overall survival. Further, differential expression of TNS1 and MET were confirmed on the validation cohort of the 5 EOCRCs (median age < 50 years and sporadic origin). This is the first approach to find early age onset biomarkers in Indian CRC patients. Among these TNS1 and MET are novel for EOCRC and may serve as potential biomarkers and novel therapeutic targets in future.

OGP46 Induces Differentiation of Acute Myeloid Leukemia Cells via Different Optimal Signaling Pathways

Acute myelogenous leukemia (AML) is characterized by blockage of cell differentiation leading to the accumulation of immature cells, which is the most prevalent form of acute leukemia in adults. It is well known that all-trans retinoic acid (ATRA) and arsenic trioxide (ATO) are the preferred drugs for acute promyelocytic leukemia (APL). However, they can lead to irreversible resistance which may be responsible for clinical failure after complete remission (CR). Moreover, the differentiation therapy of ATRA-based treatment has not been effective against AML with t(8;21) translocation. Here we aimed to identify the differentiation effect of OGP46 on AML cell lines (HL-60, NB4, and Kasumi-1) and explore its possible mechanisms. We found that OGP46 has significant inhibitory activity against these cells by triggering cell differentiation with cell-cycle exit at G1/G0 and inhibited the colony-formation capacity of the AML cells. It was shown that OGP46 induced the differentiation of NB4 cells via the transcriptional misregulation in cancer signaling pathway by PML-RARα depletion, while it was attributed to the hematopoietic cell lineage and phagosome pathway in Kasumi-1 cells, which are all critical pathways in cell differentiation. These results highlight that OGP46 is an active agent not only in the APL cell line NB4 but also in AML-M2 cell lines, especially Kasumi-1 with t(8;21) translocation. Therefore, OGP46 may be a potential compound for surmounting the differentiation blockage in AML.

Identification of Potential Diagnostic Gene Targets for Pediatric Sepsis Based on Bioinformatics and Machine Learning

Purpose: To develop a comprehensive differential expression gene profile as well as a prediction model based on the expression analysis of pediatric sepsis specimens.Methods: In this study, compared with control specimens, a total of 708 differentially expressed genes in pediatric sepsis (case–control at a ratio of 1:3) were identified, including 507 up-regulated and 201 down-regulated ones. The Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis of differentially expressed genes indicated the close interaction between neutrophil activation, neutrophil degranulation, hematopoietic cell lineage, Staphylococcus aureus infection, and periodontitis. Meanwhile, the results also suggested a significant difference for 16 kinds of immune cell compositions between two sample sets. The two potential selected biomarkers (MMP and MPO) had been validated in septic children patients by the ELISA method.Conclusion: This study identified two potential hub gene biomarkers and established a differentially expressed genes-based prediction model for pediatric sepsis, which provided a valuable reference for future clinical research.

5-aminoimidazole-4-carboxamide ribonucleoside induces differentiation in a subset of primary acute myeloid leukemia blasts

Background All-trans retinoic acid (ATRA)-based treatment of acute promyelocytic leukemia (APL) is the most successful pharmacological treatment of acute myeloid leukemia (AML). Recent development of inhibitors of mutated isocitrate dehydrogenase and dihydroorotate dehydrogenase (DHODH) has revived interest in differentiation therapy of non-APL AML. Our previous studies demonstrated that 5-aminoimidazole-4-carboxamide ribonucleoside (AICAr) induced differentiation of monocytic cell lines by activating the ATR/Chk1 via pyrimidine depletion. In the present study, the effects of AICAr on the viability and differentiation of primary AML blasts isolated from bone marrow of patients with non-APL AML were tested and compared with the effects of DHODH inhibitor brequinar and ATRA. Methods Bone marrow samples were obtained from 35 patients and leukemia blasts were cultured ex vivo. The cell viability was assessed by MTT assay and AML cell differentiation was determined by flow cytometry and morphological analyses. RNA sequencing and partial data analysis were conducted using ClusterProfiler package. Statistical analysis was performed using GraphPad Prism 6.0. Results AICAr is capable of triggering differentiation in samples of bone marrow blasts cultured ex vivo that were resistant to ATRA. AICAr-induced differentiation correlates with proliferation and sensitivity to DHODH inhibition. RNA-seq data obtained in primary AML blasts confirmed that AICAr treatment induced downregulation of pyrimidine metabolism pathways together with an upregulation of gene set involved in hematopoietic cell lineage. Conclusion AICAr induces differentiation in a subset of primary non-APL AML blasts, and these effects correlate with sensitivity to a well-known, potent DHODH inhibitor.

Serum biomarkers discovery of steroid-induced femoral head osteonecrosis in Chinese female by comparative proteome

Background Steroid-induced osteonecrosis of the femoral head (SONFH) is a disabling, aseptic and ischemic disease due to excessive glucocorticoids (GCs) usage. Patients with SONFH are commonly asymptomatic, which makes its early diagnosis is challenge, the pathological mechanisms of SONFH are not well-known, the purpose of the present study was to screen diagnostic biomarkers for SONFH. Methods The differential expression of serum proteins from SONFH, traumatic osteonecrosis of the femoral head (TONFH) patients and healthy volunteers (CK) in Chinese females was compared using iTRAQ, and potential diagnostic biomarkers were verified by western blotting. Results Gene Ontology, Kyoto Encyclopedia of Genes and Genomes (KEGG), Domain and Clusters of Orthologous Groups (COG) analyses revealed key groups of proteins, pathways and domains differentially regulated among SONFH, TONFH and healthy volunteers in Chinese female, the results showed that peptidase S1, fibrinogen, transferrin, lipid transport domains and hematopoietic cell lineage, fat digestion, absorption, peroxisome proliferator-activated receptor (PPAR) pathways were associated with the development of SONFH. Finally, C-reactive protein (CRP), serum amyloid A protein (SAA1), alpha-1-acid glycoprotein 1 (ORM1) and dopamine beta-hydroxylase were selected for verification of differential expression using western blotting. Conclusions Our data suggest that dysfunction of hematopoietic cell lineage, adhesion, fat digestion and absorption, PPAR pathways may be involved in the pathogenesis of SONFH, serum proteins SAA1, ORM1 could be used as new potential diagnostic biomarkers for SONFH.

Infection in the Patient with Sickle Cell Anemia

This chapter provides a brief overview of the clinical manifestations of and management strategies for infectious complications in the immunocompromised sickle cell disease patient. The chapter discusses infections in various organ systems, including the respiratory tract, central nervous system, bone, hematopoietic cell lineage, and blood-borne infections. Differentiating infections from noninfectious processes that often have similar presentations in the sickle cell patient may at times be difficult, and clinicians managing sickle cell patients should be keenly aware of this fact. This chapter discusses the common bacterial pathogens associated with infection and a notable viral agent known to profoundly worsen anemia in the sickle cell host, parvovirus B19. Additionally, fundamental antimicrobial regimens and primary and secondary prophylactic strategies are included in this concise summary prepared for clinicians involved in the acute care management of the sickle cell patient.

DEK and WT1 Affect Alternative Splicing of Genes Involved in Hematopoietic Cell Lineage and Resistance to Chemotherapy in Acute Myeloid Leukemia Cells.

Abstract 2392 In humans, the majority of all protein-coding transcripts contain introns that are removed by mRNA splicing carried out by spliceosomes. Mutations in the spliceosome machinery have recently been identified using whole-exome/genome technologies in myelodysplastic syndromes (MDS) and in acute myeloid leukemia (AML). In MDS the frequency of somatic spliceosomal mutations (SSM) range from 1–3% for U2AF1 in RARS/RCMD-RS to more than 70% for SF3B1 in ARSI. These values are significantly lower in AML whereas AML cells cumulate numerous splicing defects. Beside SSMs, one can propose that alternative splicing (AS) might be disturbed by other processes such as abnormal protein-protein interactions. DEK and WT1 are 2 oncogenes overexpressed in most patients with AML. They physiologically influence AS through physical interactions with the heterodimer U2AF1/U2AF2 involved in the recognition of splice acceptor site by the splicing machinery. It is therefore possible that the leukemogenic overexpression of DEK or WT1 might deregulate AS in AML cells, even in the absence of SSM. Here we show that DEK and WT1 affect AS in AML cells. Exon expression profiling was performed in triplicate with MOLM13, KASUMI and KG1 AML cells stably knocked down or not for DEK and WT1 through shRNA. The efficiency of shRNA-mediated silencing was confirmed by western blot and total RNA was analyzed using the Exon microarray platform GeneChip Human Exon 1.0 ST (Affymetrix). Microarray data were cross-compared between cell lines and only statistically significant modifications (p<0.05) shared by MOLM13, KASUMI and KG1 cells were selected. DEK and WT1 knock-down induced the transcriptional deregulation of 1613 (813 up) and 3280 (998 up) genes in AML cell lines, respectively. AS events were selected and annotated with fasterDB database (http://fasterdb.com/faster/home.pl) for genes displaying either no or low (<2) differential transcription. With this approach, differential expression of DEK coincided with changes in 1049 AS events over 934 genes. Those were distributed in 4 alternative acceptor sites (ACC), 222 first exons (AFE), 257 last exons (ALE), 539 spliced exon (ASE), 6 deletions (DEL), 21 donor sites (DON). Differential expression of WT1 led to modifying 1371 alternative splicing entities over 1198 genes. Those were distributed in 6 ACC, 385 AFE, 343 ALE, 590 ASE, 13 DEL, and 34 DON. Genes with AS events were then sorted based on gene function with DAVID bioinformatics resources version 6.7 (http://david.abcc.ncifcrf.gov/). The results indicated that, in both DEK- and WT1-dependent assays, a large subset of genes were related to hematopoietic cell lineage followed by other functional categories such as calcium signaling, ATP-binding cassette (ABC) transporters, and focal adhesion pathways that have been previously reported to be affected in AML cells and involved in resistance to chemotherapy. Differential expression of WT1 modified AS of CD3E, CD9, CSF1R, CSF3R, CR2, GP1BA, ITGA1, ITGA3, ITGB3, IL1A and IL6 while that of DEK led to modulate AS of CD19, CD1d, CD36, CD3G, CSF1R, CR2, ITGA1, ITGA2B, ITGA4, ITGA6, IL7, HLA-DRB5, and MME. Microarray data were validated by exon specific RT-PCR. Exon expression profiling of fresh AML bone marrow samples with or without U2AFs mutations and various levels of DEK or WT1 expression is currently in progress and will be presented. In conclusion in AML cells, DEK and WT1 overexpression affects AS of numerous key genes involved in hematologic differentiation, leukemogenesis and resistance to chemotherapy. These posttranscriptional effects occur in the absence of transcriptional change and therefore highlight hitherto unknown phenotypic alterations having putative diagnostic, prognostic and therapeutic interests. DEK and WT1 are overexpressed in more than 80% of AML and target AS via U2AFs that are mutated in only 0–8% of cases. Accordingly present results strongly suggest that other factors than SSM divert AS in AML. Disclosures: No relevant conflicts of interest to declare.

Proper levels of c-Myb are discretely defined at distinct steps of hematopoietic cell development

The definitive hematopoietic cell lineages have been proposed to originate from hemogenic endothelial cells during mouse embryogenesis. c-Myb is a transcription factor that is essential for the development of definitive hematopoiesis. To investigate the functional role of c-Myb in hematopoietic cell development from endothelial cells, we introduced a c-myb transgene expressed under the control of a tetracycline-regulated promoter into the c-myb–/– embryonic stem (ES) cell line, with the aim of inducing c-Myb expression at any stage and at any level. Induction of c-Myb expression after replating c-myb–/– endothelial cells rescued the generation and proliferation of definitive hematopoietic progenitor cells, suggesting that c-Myb expression in developing endothelial cells is not a prerequisite for their hematogenic potential. Overexpression of c-Myb, however, prevented the terminal differentiation of erythrocytes and megakaryocytes and completely abolished B-lymphocyte development. Our results indicate that c-Myb is a major factor that controls differentiation as well as proliferation of hematopoietic progenitor cells derived from hemogenic endothelial cells, and that appropriate levels of c-Myb protein are strictly defined at distinct differentiation steps of each hematopoietic cell lineage.