Journal: Cell reports

Article Title: Identification of the global miR-130a targetome reveals a role for TBL1XR1 in hematopoietic stem cell self-renewal and t(8;21) AML.

doi: 10.1016/j.celrep.2022.110481

Figure Lengend Snippet: Figure 2. Enforced expression of miR-130a expands HSCs by forcing them into cell cycle (A) Representative flow plots of HSPC populations from control and miR-130a OE xenografts at 12 weeks. (B) Proportion of CD34+CD38 compartment and frequency of LT-HSC, ST-HSC, and MLP (multi-lymphoid progenitor) cell populations (n = 4, each replicate contains pooled RF and BM from 2 to 4 individual mice). (C) Proportion of CD34+CD38+ compartment and frequency of CMP (common myeloid progenitor), MEP (megakaryocyte-erythroid progenitor), and GMP (granulocyte-macrophage progenitor) cell populations (n = 4, each replicate contains pooled RF and BM from 2 to 4 individual mice). (D) Secondary transplantation of CD45+mO+ from 12 week NSG mice at limiting dilution doses. Frequency of HSCs was evaluated from the engraftment in secondary mice (>0.05% CD45+mO+ cells in BM, n = 19–31 mice from two independent experiments). (E) Normalized enrichment score (NES) of the gene sets significantly different between miR-130a OE and control-transduced HSCPs (n = 3). (F) GSEA plots of HSC cell-cycle-primed gene sets and MYC targets in the transcriptome profile following miR-130a OE in CD34+ HSPCs. (G) Cell-cycle analysis of LT-HSCs, ST-HSCs, and CD34+CD38+ cells transduced with miR-130a or control lentiviruses (n = 3). (B and C) Unpaired t test, all error bars indicate ±SEM, *p < 0.05.

Article Snippet: Quantitative RT-PCR for expression levels of miR-130a in CBF AML Peripheral blood samples from t(8;21) and inv(16) patients (Table S5) were thawed as described and CD34+ blasts were enriched with Direct CD34+ Progenitor Isolation Kit (Miltenyi) or sorted by FACS.

Techniques: Expressing, Control, Transplantation Assay, Cell Cycle Assay, Transduction

Journal: Cell reports

Article Title: Identification of the global miR-130a targetome reveals a role for TBL1XR1 in hematopoietic stem cell self-renewal and t(8;21) AML.

doi: 10.1016/j.celrep.2022.110481

Figure Lengend Snippet: Figure 3. Mass spectrometry and chimeric AGO2 eCLIP identify miR-130a targetome in human HSPCs (A) Heatmap of downregulated pathways and proteins following miR-130a OE in CD34+ CB cells; Wilcoxon one-sided test, n = 3, p < 0.05. (B) List of miR-130a downregulated proteins that are predicted TarBase and mirDIP targets, limma t test. (C) Western blot showing repression of miR-130a targets. (D) Enrichment map showing gene sets containing miR-130a targets listed in (B); node size is proportional to NES; Wilcoxon one-sided test, p < 0.05. (E) GSEA plots of miR-130a targets from chimeric AGO2 eCLIP in proteome profile following miR-130a OE in CD34+ HSPCs. (F) Genome browser tracks from chimeric AGO2 eCLIP-seq in CD34+ HSPCs. (G) Enrichment map of miR-130a targets from chimeric AGO2 eCLIP in downregulated protein sets following miR-130a OE; false discovery rate (FDR) < 0.05, node size is proportional to NES, Wilcoxon one-sided test; p < 0.05.

Article Snippet: Quantitative RT-PCR for expression levels of miR-130a in CBF AML Peripheral blood samples from t(8;21) and inv(16) patients (Table S5) were thawed as described and CD34+ blasts were enriched with Direct CD34+ Progenitor Isolation Kit (Miltenyi) or sorted by FACS.

Techniques: Mass Spectrometry, Western Blot

Journal: Cell reports

Article Title: Identification of the global miR-130a targetome reveals a role for TBL1XR1 in hematopoietic stem cell self-renewal and t(8;21) AML.

doi: 10.1016/j.celrep.2022.110481

Figure Lengend Snippet: Figure 4. Repression of TBL1XR1 impairs B lymphoid differentiation and expands LT-HSC (A) Western blot showing TBL1XR1 protein levels in xenografts. (B) Human CD45+ chimerism in the right femur (RF) and bone marrow (BM) at 24 weeks post-transplantation (n = 2 biological experiments, 6–8 mice/experimental group). (C) Fold change in BFP+CD45+ cells in RF and BM at 24 weeks post-transplantation compared with input levels. (D) Lineage distribution of BFP+ xenografts. (E) Flow cytometry plots of HSPC populations, flow plots are representative of 3 samples overlaid together. (F) Distribution of BFP+CD34+CD38 cell populations from 24 week xenografts (n = 5, each replicate contains pooled RF from 2 to 4 individual mice), unpaired t test, *p < 0.05. (G) Secondary transplantation of CD45+BFP+ from 24 week NSG mice at limiting dilution doses. Frequency of HSC was evaluated from the engraftment in NSG- GF secondary mice (>0.05% CD45+BFP+cell in BM, n = 11–16 mice/experimental group). (H) Enrichment map of upregulated (red nodes) and downregulated (blue nodes) pathways following TBL1XR1 KD intersected with miR-130a targets from eCLIP, hypergeometric t test, p < 0.05. (I) GSEA plots of proliferative and quiescence genes in transcriptome profile following TBL1XR1 KD. (J) Western blot showing protein levels of NCoR1 and TBL1XR1 following TBL1XR1 KD in CD34+ CB cells. (K) GSEA plots of genes upregulated and downregulated by RA in the transcriptome profile following TBL1XR1 KD. (B–D) Unpaired Mann-Whitney U test, all error bars indicate ±SEM, *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001.

Article Snippet: Quantitative RT-PCR for expression levels of miR-130a in CBF AML Peripheral blood samples from t(8;21) and inv(16) patients (Table S5) were thawed as described and CD34+ blasts were enriched with Direct CD34+ Progenitor Isolation Kit (Miltenyi) or sorted by FACS.

Techniques: Western Blot, Transplantation Assay, Flow Cytometry, MANN-WHITNEY

Journal: Cell reports

Article Title: Identification of the global miR-130a targetome reveals a role for TBL1XR1 in hematopoietic stem cell self-renewal and t(8;21) AML.

doi: 10.1016/j.celrep.2022.110481

Figure Lengend Snippet: Figure 5. Loss of function of miR-130a and TBL1XR1 OE induces differentiation of t(8;21) AML (A) Clustering of TCGA AMLs into four groups based on scRNA-seq data (n = 173). (B) Expression of miR-130a in AML subtypes from the TCGA dataset (n = 170). (C) Kaplan-Meier curve showing correlation between miR-130a expression and CBF AML patient survival (n = 48). (D) qRT-PCR showing expression levels of miR-130a in CD34+ blasts from t(8;21) AML compared with healthy controls (n = 20) and CD34 cells from the same patient samples (n = 24).

Article Snippet: Quantitative RT-PCR for expression levels of miR-130a in CBF AML Peripheral blood samples from t(8;21) and inv(16) patients (Table S5) were thawed as described and CD34+ blasts were enriched with Direct CD34+ Progenitor Isolation Kit (Miltenyi) or sorted by FACS.

Techniques: Expressing, Quantitative RT-PCR

Journal: Journal of cancer research and therapeutics

Article Title: Syk expression in non-small-cell lung cancer and its relation with angiogenesis.

doi: 10.4103/0973-1482.154082

Figure Lengend Snippet: Figure 2: CD34 immunostaining locates in microvascular endothelial cells with brown granular in tumor tissue SABC × 200

Article Snippet: Streptavidin‐biotin complex (SABC) kit, CD34 antibody, and DAB chromogenic reagent were from Wuhan Boster Biotechnology Company.

Techniques: Immunostaining

Journal: Frontiers in Immunology

Article Title: Mice with humanized immune system as novel models to study HIV-associated pulmonary hypertension

doi: 10.3389/fimmu.2022.936164

Figure Lengend Snippet: (A) Human spleen and liver organoids in the mouse kidney capsule. NSG BLT mice were engrafted with fetal liver, thymus and hematopoietic cells; human engraftments were confirmed by gross inspections of the mouse kidney capsules at euthanasia, where organoids develop in humanized mice. (B) Experimental timeline. NSG mice were engrafted with either human hematopoietic CD34+ cells (cord-blood derived) or with human liver, thymus, and hematopoietic stem cells (fetal-derived). After confirmation of human engraftments by flow cytometry, humanized mice were infected with HIV and either placed in hypoxia or injected with SU5416 to induce PAH. Animals were monitored for any signs of disease including graft-vs-host disease, labored breathing or wasting throughout the course of the study. Mice were subjected to open-chest right heart catheterizations under anesthesia; specimens were collected for further analyses.

Article Snippet: Mice humanized with CD34 + stem cells were purchased from The Jackson Laboratory and showed a 31-48% level of human CD45 + cell engraftment.

Techniques: Capsules, Derivative Assay, Flow Cytometry, Infection, Injection