Journal: Cell reports

Article Title: Latexin regulates sex dimorphism in hematopoiesis via gender-specific differential expression of microRNA 98-3p and thrombospondin 1

doi: 10.1016/j.celrep.2023.112274

Figure Lengend Snippet: (A) RNA-seq data of LSK cells from WT female, Lxn −/− female, WT male, and Lxn −/− male mice. (B and C) The mRNA (B) and protein (C) levels of Thbs1 in LSK cells of WT female, Lxn −/− female, WT male, and Lxn −/− male mice. (D) Representative flow cytometric analysis of active caspase-3 protein in male WT and Lxn −/− LSK cells. (E) Frequency of LSK cells positive for active caspase-3. (F) The protein level of cleaved caspase-3 protein in LSK cells of WT and Lxn −/− male mice by western blot. (G) Percentage of LSK cells in one femur of Thbs1 +/− and WT male mice. (H) Percentage of LSK cells in one femur of Thbs1 +/− and WT female mice. All data are shown as mean (n = 5–6) ± SD and were analyzed by two-tailed t test. *p ≤ 0.05 and ****p < 0.0001.

Article Snippet: Latexin antibody was bought from Proteintech, Thbs1 antibody was bought from Santa Cruz, actin antibody were bought from Sigma.

Techniques: RNA Sequencing, Western Blot, Two Tailed Test

Journal: Cell reports

Article Title: Latexin regulates sex dimorphism in hematopoiesis via gender-specific differential expression of microRNA 98-3p and thrombospondin 1

doi: 10.1016/j.celrep.2023.112274

Figure Lengend Snippet: (A) The expression of microRNA in LSK cells of WT female, Lxn −/− female, WT male, and Lxn −/− male mice by qPCR. (B) The expression of miR98-3p in 3T3 cells transfected with control or miR98-3p mimics. (C) The expression of Thbs1 in 3T3 cells transfected with control or miR98-3p mimics. (D) The expression of miR98-3p in 293T cells transfected with pcDNA3.1 control vector or pre-miR98. (E) The scheme for potential binding of Thbs1 -3′ UTR and miR98-3p. The predicted binding (seed) region for miR98-3p spans from 2,041 to 2,047 of the Thbs1 -3′ UTR. The mutant was generated as the negative control ( Thbs1 -3′ UTR-mut). (F) The luciferase activity in 293T cells co-transfected with Thbs1 -3′ UTR-WT and empty vector control (pcDNA3.1) or pre-miR98 plasmid or with Thbs1 -3′ UTR-mut and empty vector control (pcDNA3.1) or pre-miR98 plasmid. All data were derived from 2 independent experiments with 3 replicates from each experiment, are shown as mean ± SD, and were analyzed by two-tailed t test. **p < 0.01 and ****p < 0.0001.

Article Snippet: Latexin antibody was bought from Proteintech, Thbs1 antibody was bought from Santa Cruz, actin antibody were bought from Sigma.

Techniques: Expressing, Transfection, Control, Plasmid Preparation, Binding Assay, Mutagenesis, Generated, Negative Control, Luciferase, Activity Assay, Derivative Assay, Two Tailed Test

Journal: Cell reports

Article Title: Latexin regulates sex dimorphism in hematopoiesis via gender-specific differential expression of microRNA 98-3p and thrombospondin 1

doi: 10.1016/j.celrep.2023.112274

Figure Lengend Snippet: (A) The expression of miR98-3p in female LSK cells transfected with control or miR98-3p mimics was assessed by qPCR and quantified. (B) The mRNA level of Thbs1 in female LSK cells transfected with control or miR98-3p mimics was assessed by qPCR and quantified. (C) The protein level of Thbs1 in female LSK cells transfected with control or miR98-3p mimics was assessed by western blot. (D) The absolute number of clones, defined by the CAFC assay, observed at day 21. (E) Percentages of PB (CD45.2) chimerismin recipient mice (CD45.1, n = 5) that were transplanted with LSK cells treated with control or miR98-3p mimics at 4, 8, and 14 weeks after transplantation. (F and G) Lxn mRNA expression (F) and Lxn protein expression (G) (top panel) and quantification (bottom panel) in WBCs, BM Lin− cells, and HSPC-enriched LSK cells in male and female mice. (H and I) Thbs1 mRNA expression (H) and Thbs1 protein expression (I) in WBCs, BM Lin− cells, and HSPC-enriched LSK cells in male and female mice. Note: because Thbs1 protein expression is very low in LSK cells, we used different exposure times in order for the band to show. We thus could not do the band quantification. (J) miR-98-3p expression in male and female LSK cells treated with miR-98-3p antagonist or control. (K) Thbs1 protein expression in male and female LSK cells treated with miR-98-3p antagonist or control. (L) The model for sex-/gender-specific regulation of HSCs and hematopoiesis by the Lxn , mirR98-3p , and Thbs1 pathway. In female HSPCs, Lxn and low miR98-3p expression upregulates Thbs1 expression, which inhibits HSC repopulation and increases HSC apotosis, thus impairing HSC function. In contrast, miR98-3p expression is very high in male HSCs, leading to a very low level of Thbs1 , which abrogates the functional effect of Lxn on male HSCs. Differential expression of sex-chromosome-specific miR98-3p contributes to sex-dependent regulation of HSCs and hematopoiesis by Lxn-Thbs1 signaling. All the data are shown as mean ± SD and were analyzed by two-tailed t test. *p ≤ 0.05, **p < 0.01, and ****p < 0.0001.

Article Snippet: Latexin antibody was bought from Proteintech, Thbs1 antibody was bought from Santa Cruz, actin antibody were bought from Sigma.

Techniques: Expressing, Transfection, Control, Western Blot, Clone Assay, Transplantation Assay, Functional Assay, Quantitative Proteomics, Two Tailed Test