The Evaluation of CD3, CD 5, CD10, CD 19, and CD20 Markers in the Differential Diagnosis of the Lymphoma Subtypes in Sudanese Patients

The objective of this paper was to evaluate the use of CD3, CD5, CD10, CD19, and CD20 markers in the differential identification of lymphoma subtypes. Methods and Results: This was a retrospective cross-sectional study included 82 patients with palpable lymphadenopathies. The formalin-fixed paraffin block sections immunostained with the Dako flex were investigated. CD3, CD5, CD10, CD19, and CD20 staining was performed on sections. The current study found that the two main types of lymphoma, Hodgkin’s lymphoma and non-Hodgkin’s lymphoma, have a significant association with CD3, CD10, and CD19, and a highly significant association with CD20, implying that these CD markers are crucial for general classification and diagnosis of lymphoma. CD3 had a highly significant relationship with gender. CD3 and CD20 were demonstrated to have a significant relationship with the lymphoma subtypes. The CD20 marker is the most consistent and useful marker for differentiating lymphoma subtypes.

Responses of platelet CD markers and indices to resistance exercise with and without blood flow restriction in patients with type 2 diabetes

BACKGROUND: Diabetes mellitus is a common disorder with the risk of vascular injury. OBJECTIVE: The aim of this study was to compare the effects of low-intensity resistance exercise with blood flow restriction versus high-intensity resistance exercise on platelet CD markers and indices in patients with type 2 diabetes. METHODS: Fifteen female patients with type 2 diabetes (Mean±SD; age, 47.6±7.2 yrs) randomly completed two resistance exercise at an intensity corresponding to 20% and 80% of one-repetition maximum (1-RM), with and without blood flow restriction (REBFR and RE), respectively. We measured markers of platelet activation (P-selectin, GpIIb/IIIa, and CD42) and platelet indices before and immediately after exercise, and after 30 min recovery. RESULTS: Platelet count (PLT) and plateletcrit (PCT) increased in response to REBFR more than the RE (p < 0.05), though, no significant differences in PDW and MPV were observed (p < 0.05). Although P-selectin (CD62P), CD61, CD41, and CD42 were reduced following resistance exercise in both trials, these reductions were non-significant (p < 0.05). Besides, no significant between-group differences were found for platelet CD markers (p < 0.05). CONCLUSIONS: It is concluded that REBFR induces thrombocytosis, but responses of platelet CD markers in patients with type 2 diabetes are similar following low-intensity REBFR and high-intensity RE.

Targeting Chronic Myeloid Leukemia Stem/Progenitor Cells Using Venetoclax-Loaded Immunoliposome

CML is a hematopoietic stem-cell disorder emanating from breakpoint cluster region/Abelson murine leukemia 1 (BCR/ABL) translocation. Introduction of different TKIs revolutionized treatment outcome in CML patients, but CML LSCs seem insensitive to TKIs and are detectable in newly diagnosed and resistant CML patients and in patients who discontinued therapy. It has been reported that CML LSCs aberrantly express some CD markers such as CD26 that can be used for the diagnosis and for targeting. In this study, we confirmed the presence of CD26+ CML LSCs in newly diagnosed and resistant CML patients. To selectively target CML LSCs/progenitor cells that express CD26 and to spare normal HSCs/progenitor cells, we designed a venetoclax-loaded immunoliposome (IL-VX). Our results showed that by using this system we could selectively target CD26+ cells while sparing CD26− cells. The efficiency of venetoclax in targeting CML LSCs has been reported and our system demonstrated a higher potency in cell death induction in comparison to free venetoclax. Meanwhile, treatment of patient samples with IL-VX significantly reduced CD26+ cells in both stem cells and progenitor cells population. In conclusion, this approach showed that selective elimination of CD26+ CML LSCs/progenitor cells can be obtained in vitro, which might allow in vivo reduction of side effects and attainment of treatment-free, long-lasting remission in CML patients.

Deregulation of miR-34a, miR-221 and miR-222 after HT29 cells treatment with Silibinin encapsulated in polymersomes as an anti-cancer stem cell agent

Colorectal Cancer (CRC) has the most common malignant gastrointestinal cancer which representing about13% of all malignant tumor. CRC Cancer Stem Cell is the major reasons for recurrence of disease cause of solid tumor metastasis, relapse of cancer after treatment and drug resistance. Silibinin, an herbal extract from milk thistle plant, has been identified as a potential cancer medicine that can target the signaling pathway of CSCs and change their abilities. In our study, the results of CSC confirmation test such as specific surface CD markers and ability to form colonospheres was indicated the HT-29 cells as CSC-CRC. To increase the effectiveness of Silibinin, and also, to evaluate therapeutic intentions on HT-29 cancer stem-like cells, we encapsulated Silibinin in polymersome nanoparticle and validated the anti-proliferative and apoptotic activities of this new patent by MTT assay, AnnexinV/PI method, cell cycle analysis and DAPI staining. Furthermore, the efficacy of drug on Multicellular Tumor Spheroid (MCTS) and single cell suspension was showed that SPN had succeed to decrees the expression level of CSC CD markers compared with control group. Follow by using miRNAs as a novel and minus invasive expertise for prognostic, RT-qPCR confirmed that SPNs can repress oncogenic miRNAs such as miR-221 and miR-222. Silibinin encapsulated in Polymersome Nanoparticles (SPNs) can also enhance the expression of tumor suppressor miR-34a and some of its proapoptotic target genes such as P53, BAX, CASP9, CASP3, and CASP8. Our results suggested that SPNs can be recognized as a new stimulant factor to direct the HT-29 cancer cells toward apoptosis pathways thorough modify expression of some miRNAs and their apoptotic target genes directly and/or indirectly.

Tissue Treg Secretomes and Transcription Factors Shared With Stem Cells Contribute to a Treg Niche to Maintain Treg-Ness With 80% Innate Immune Pathways, and Functions of Immunosuppression and Tissue Repair

We used functional -omics angles and examined transcriptomic heterogeneity in CD4+Foxp3+ regulatory T cells (Treg) from spleen (s-Treg), lymph nodes (LN-Treg), intestine (int-Treg), and visceral adipose tissue (VAT-Treg), and made significant findings: 1) Five new shared Treg genes including NIBAN, TNFRSF1b, DUSP4,VAV2, and KLRG1, and 68 new signatures are identified. Among 27 signaling pathways shared in four tissue Treg, 22 pathways are innate immune pathways (81.5%); 2) s-Treg, LN-Treg, int-Treg, and VAT-Treg have zero, 49, 45, and 116 upregulated pathways, respectively; 3) 12, 7, and 15 out of 373 CD markers are identified as specific for LN-Treg, int-Treg, and VAT-Treg, respectively, which may initiate innate immune signaling; 4) 7, 49, 44, and 79 increased cytokines out of 1176 cytokines are identified for four Treg, respectively, suggesting that Treg have much more secretory proteins/cytokines than IL-10, TGF-β, and IL-35; 5) LN-Treg, int-Treg, and VAT-Treg have 13 additional secretory functions more than s-Treg, found by analyzing 1,706 secretomic genes; 6) 2, 20, 25, and 43 increased transcription factors (TFs) out of 1,496 TFs are identified four Treg, respectively; 7) LN-Treg and int-Treg have increased pyroptosis regulators but VAT-Treg have increased apoptosis regulators; 8) 1, 15, 19, and 31 increased kinases out of 661 kinome are identified for s-Treg, LN-Treg, int-Treg, and VAT-Treg, respectively; 9) comparing with that of s-Treg, LN-Treg, int-Treg, and VAT-Treg increase activated cluster (clusters 1–3) markers; and decrease resting cluster (clusters 4–6) markers; and 10) Treg promote tissue repair by sharing secretomes and TFs AHR, ETV5, EGR1, and KLF4 with stem cells, which partially promote upregulation of all the groups of Treg genes. These results suggest that stem cell-shared master genes make tissue Treg as the first T cell type using a Treg niche to maintain their Treg-ness with 80% innate immune pathways, and triple functions of immunosuppression, tissue repair, and homeostasis maintenance. Our results have provided novel insights on the roles of innate immune pathways on Treg heterogeneity and new therapeutic targets for immunosuppression, tissue repair, cardiovascular diseases, chronic kidney disease, autoimmune diseases, transplantation, and cancers.

Evaluation of Immunoglobulins, CD4/CD8 T Lymphocyte Ratio and Interleukin-6 in COVID-19 Patients

Introduction: The outbreak of novel coronavirus COVID-19 infections that started in China late 2019 has spread rapidly and cases have been recorded worldwide. So, in this study, we sought clarification of the clinical characteristics and importance of changing the lymphocyte group, antibodies, CD markers, and interleukin-6 in the serum of COVID-19 patients, which may help to clarify the pathogen and develop new biomarkers. Material and Methods: Venous blood samples had been accumulated from patients before taking any medications. Sera had been separated and saved at (-20°C) until analysis. Serum anti-SARS-CoV-2 immunoglobulins (IgG, IgA, and IgM) were determined in plasma samples using enzyme-linked immunosorbent assays (ELISA) and Serum IL-6 was assessed. Results: Median IgM (p=0.001), IgG (p<0.0001), and IgA (p<0.001), were decreased in patients comparing with control the control group. There is a significant decrease in CD3+ and CD4+ cells compared to healthy individuals in patients infected with COVID-19 (p<0.0001). CD19+ cell count decreased in COVID-19 patients compared to that of the control group (p<0.0001). After calculating CD4+/CD8+ cell ratio decreased in COVID-19 patients (p<0.0001). However, CD56+ cells were found to be increased (p<0.0001). Conclusions: IgM, IgG, IgA levels and CD19+, CD4+ cells, CD4+/CD8+ cell ratio were found to be decreased whereas CD8+, CD3+, CD4+ cells were detected to be increased in COVID-19 patients compared to those of healthy controls. Keywords: IL-6, COVID-19, IgG, IgA, IgM, CD markers

Best Practices for Validation of Measurable Residual Disease Assessments By Multiparameter Flow Cytometry in Emerging Clinical Trials of Acute Myeloid Leukemia

Background: Measurable Residual Disease (MRD) assessments are gaining increasing acceptance as a prognostic factor for tailoring treatment in hematological malignancies. Acute Myeloid Leukemia (AML) is a heterogeneous disease with high relapse rates and presents a high unmet need for effective treatment options. Measurement of residual disease after therapy reflects a combination of all resistance mechanisms and is currently used for guiding treatment options. Study Design: In this study, we aimed to validate an AML-MRD assay by multiparameter flow cytometry (MFC) methodology. This is a 4-tube, 8-parameter assay designed to incorporate cell differentiation (CD) markers for identification of a diverse group (covering roughly 90% of patients, Cloos et al, 2018) of Leukemia Associated Immunophenotypes (LAIPs) to accurately identify both native phenotypes and phenotype shifts after drug treatment. These CD markers were selected based on extensive investigation of many markers and in line with the consensus recommendations from European Leukemia Network AML working party (Schuurhuis et al, 2018), while specimen testing and interpretation principles were performed in accordance with Cloos et al, 2018. The assay validation focused on evaluation of sensitivity (MRD cut point and LOD), precision and accuracy as key criteria for evaluating assay performance utilizing primary patient specimens and AML cell lines representing different LAIPs. The results were orthogonally verified in a blinded manner by morphologic assessment at Navigate and by the MRD-team at VUMC Amsterdam. Results: Two experimental approaches were adopted to evaluate analytical and functional sensitivity (clinical applicability) of the assay. Results indicated analytical sensitivity (LOD) as low as 0.01% LAIPs of total WBC and functional sensitivity (LOQ) of 0.1% (MRD cut point). Excellent repeatability and reproducibility (less than 20% CV) was observed across instruments, operators and independent measurements (n = 75). The frequencies of AML blasts detected by MFC and morphological examination were highly concordant (Spearman r = 0.95, P value &lt; 0.001, n = 24). LAIPs deduced across nine patient specimens by the Navigate laboratory were independently confirmed by the MRD-team at VUMC Amsterdam. Conclusion: In summary, based on the use of consensus markers recommended by ELN for reliable capture of a broad group of LAIPs in AML patients and verification of key assay performance characteristics, we believe this comprehensive MFC based AML MRD assay is fit-for-purpose for accurately assessing measurable residual disease. Following clinical trial validation, MRD might be used as a surrogate endpoint for approval of emerging agents. Disclosures Marques Ramos: Novartis: Current Employment. Larson:BMS, Bioline, Celgene, Juno, Janssen: Research Funding; TORL Biotherapeutics: Current equity holder in private company. Sarikonda:Novartis: Current Employment.