Journal: eLife

Article Title: Lipolysis of bone marrow adipocytes is required to fuel bone and the marrow niche during energy deficits

doi: 10.7554/eLife.78496

Figure Lengend Snippet:

Article Snippet: Commercial assay or kit , BCA Protein Assay Kit , Thermo Fisher Scientific , 23225 , .

Techniques: Generated, Transgenic Assay, Animal Model, Bicinchoninic Acid Protein Assay, Enzyme-linked Immunosorbent Assay, TRAP Assay, Electron Microscopy, Reverse Transcription, Software, Flow Cytometry, Microscopy, Imaging, Micro-CT, Real-time Polymerase Chain Reaction

Journal: Heliyon

Article Title: Transplantation of IGF-1-induced BMSC-derived NPCs promotes tissue repair and motor recovery in a rat spinal cord injury model

doi: 10.1016/j.heliyon.2022.e10384

Figure Lengend Snippet: Supplementation of EGF, bFGF, and IGF-1 enhanced the cellular proliferation of BMSC-derived NPCs via downregulation of miR-22-3p. (A) Representative photomicrograph of BMSCs in culture. Immunocytochemical staining indicated that BMSCs expressed CD90, CD44, fibronectin, vimentin, and nestin. Nuclei were counterstained with Sytox Blue. Images were viewed under a confocal microscope. Scale bar: 100 μm. (B) Photomicrograph of free-floating neurospheres generated in neurobasal media with and without growth factor supplementation. Scale bar: 100 μm. The size of each neurosphere was measured using ImageJ based on the longest diameter. (C) Immunofluorescence staining of NPCs with Sox2 (scale bar: 50 μm) and fibronectin (scale bar: 20 μm). (D) Proliferation analysis of NPCs under different growth factors. Cells were incubated with an MTS reagent for 4 h, and changes in proliferation were studied at different time intervals. Data are represented as the mean optical density (OD) at 540 nm. (E) Expression level of miR-22-3p in BMSC-derived NPCs relative to the control (without growth factor). (F) Relative expression of the Pten , Akt1 , and Tp53 genes in NPCs derived from different growth factor combinations compared with the control. The indicated fold-change values were normalized to the Actb and Gapdh controls. All experiments were repeated in three biological replicates. Data are presented as mean ± SD. Statistical analysis was performed using one-way ANOVA. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, and ∗∗∗∗ p < 0.0001.

Article Snippet: All reactions were run in triplicate using master mix and pre-designed primers as follows (all products were purchased from Applied Biosystem, CA, USA): for microRNA detection assay and endogenous controls: TaqMan Universal PCR Master Mix II, miR-22-3p (Assay ID: 000398), snoRNA (Assay ID: 001718), and small nucleolar RNA U87 (Assay ID: 001712); for gene expression assay and endogenous controls: TaqMan Fast Advanced Master Mix, Akt-1 (Assay ID: Rn00583646_m1, RefSeq ID: NM_033230.2, Amplicon length: 87), Tp53 (Assay ID: Rn00755717_m1, RefSeq ID: NM_030989.3, Amplicon length: 94), Pten (Assay ID: Rn00477208_m1, RefSeq ID: NM_031606.1, Amplicon length: 73), Actb (Assay ID: Rn00667869_m1, RefSeq ID: NM_031144.3, Amplicon length: 91), and Gapdh (Assay ID: Rn01775763_g1, RefSeq ID: NM_017008.4, Amplicon length: 174).

Techniques: Derivative Assay, Staining, Microscopy, Generated, Immunofluorescence, Incubation, Expressing, Control

Journal: Heliyon

Article Title: Transplantation of IGF-1-induced BMSC-derived NPCs promotes tissue repair and motor recovery in a rat spinal cord injury model

doi: 10.1016/j.heliyon.2022.e10384

Figure Lengend Snippet: Transplantation of BMSC-derived NPCs in a spinal cord injury animal model promoted locomotor and sensory recoveries (N = 3 per group). (A) A 4-mm-long longitudinal cut along the midline of the spinal cord for lateral hemisection at the T9-T10 level. (B) Timeline of the entire in vivo transplantation study. (C) Open-field locomotor assessment. The hind limb function of all rats was assessed using the Basso, Beattie, and Bresnahan (BBB) locomotor scale. (D) Mechanical sensory assessment as conducted with the Von Frey filament test. Two-way ANOVA was performed, followed by a Tukey test to compare the mean difference among the groups and times. An asterisk (∗) indicates a significant difference of the treatment groups and the control group. A hashtag (#) indicates a significant difference between treatment days. (E) Relative expression of Map2, Gfap, Olig2, Mbp, Sox2 , and nestin within the transplanted region of the spinal cord. (F) Western blot analysis and relative quantification of TUJ1, GFAP, APC, and MBP. Each protein band was normalized to GAPDH. Data are presented as mean ± SD. Statistical analysis was performed using one-way ANOVA to compare within groups. ∗ p < 0.05, # p < 0.05, ∗∗∗ p < 0.001, and ∗∗∗∗ p < 0.0001.

Article Snippet: All reactions were run in triplicate using master mix and pre-designed primers as follows (all products were purchased from Applied Biosystem, CA, USA): for microRNA detection assay and endogenous controls: TaqMan Universal PCR Master Mix II, miR-22-3p (Assay ID: 000398), snoRNA (Assay ID: 001718), and small nucleolar RNA U87 (Assay ID: 001712); for gene expression assay and endogenous controls: TaqMan Fast Advanced Master Mix, Akt-1 (Assay ID: Rn00583646_m1, RefSeq ID: NM_033230.2, Amplicon length: 87), Tp53 (Assay ID: Rn00755717_m1, RefSeq ID: NM_030989.3, Amplicon length: 94), Pten (Assay ID: Rn00477208_m1, RefSeq ID: NM_031606.1, Amplicon length: 73), Actb (Assay ID: Rn00667869_m1, RefSeq ID: NM_031144.3, Amplicon length: 91), and Gapdh (Assay ID: Rn01775763_g1, RefSeq ID: NM_017008.4, Amplicon length: 174).

Techniques: Transplantation Assay, Derivative Assay, Animal Model, In Vivo, Control, Expressing, Western Blot, Quantitative Proteomics

Journal: Nature Metabolism

Article Title: Cell-specific DNA methylation in human alpha and beta cells regulates gene expression in type 2 diabetes

doi: 10.1038/s42255-026-01498-9

Figure Lengend Snippet: a , b , Volcano plots showing T2D-associated DEGs in human alpha ( a ) and beta ( b ) cells, P < 0.05 based on DESeq2 (ref. ). c , mRNA expression of T2D - associated beta cell DEGs selected for functional follow-up based on q < 0.1 and DESeq2 (ref. ). Data from 15 non-diabetic donors and 7 donors with T2D. Women and men are represented by filled and open circles, respectively. Data are presented as the mean ± s.e.m. d , siRNA-mediated knockdown of ADCY9 , FBLL1 and ZSWIM9 in human islets from five donors. e , Silencing of Adcy9 , Fbll1 and Zswim9 using siRNA in rat INS-1 832/13 beta cells. n = 4 ( Fbll1 ) or 6 ( Adcy9 and Zswim9 ) biological replicates. f , Insulin secretion from human islets stimulated with 1 mM and 20 mM glucose. Measurements were performed on islets from the same five donors as in d . Data are presented as the mean ± s.e.m. g , Insulin secretion from INS-1 832/13 beta cells during 1 h of stimulation with low (2.8 mM) or high (16.7 mM) glucose. n = 6 biological replicates. h , Depolarization-induced insulin secretion (35 mM K + ) from INS-1 832/13 beta cells. n = 6 biological replicates from the same experiments as e . i , Overexpression of GNAL (OE- GNAL ), ONECUT2 (OE- ONECUT2 ), SNX1 (OE- SNX1 ) or TRAPPC13 (OE- TRAPPC13 ) in human islets from five ( SNX1 ) or six ( GNAL , ONECUT2 and TRAPPC13 ) donors. j , OE- GNAL , OE- ONECUT2 , OE- SNX1 and OE- TRAPPC13 in INS-1 832/13 beta cells. n = 5 ( ONECUT2 ) or 6 ( GNAL , SNX1 and TRAPPC13 ) biological replicates. k , Insulin secretion from human islets stimulated with 1 mM or 20 mM glucose for 1 h. Measurements were performed on islets from the same donors as in i . l , m , Insulin secretion from INS-1 832/13 beta cells during 1 h of stimulation with 2.8 mM or 16.7 mM glucose in absolute values ( l ) and as fold change, that is, secretion at 16.7 mM divided by secretion at 2.8 mM glucose ( m ). n = 6 biological replicates. n , Depolarization-induced insulin secretion by stimulation with 35 mM K + in INS-1 832/13 beta cells. n = 3 biological replicates. In d – n , data are presented as the mean ± s.e.m. and * P < 0.05, ** P < 0.01, *** P < 0.001 versus siNC or GFP, based on two-tailed ( d – n ) paired Student’s t -tests. The same colour represents paired samples included in the same experiment, for human islets from the same donor and for the INS-1 832/13 beta cells from the same cell passage. The q values for b , c were corrected for multiple testing, while the other P values throughout were not.

Article Snippet: TaqMan assays were used to measure expression levels of human ADCY9 (Hs00181599_m1), human FBLL1 (Hs01584505_s1), ZNF431 (Hs00819968_m1), human ZSWIM9 (Hs00736775_m1), human ONECUT2 (Hs00191477_m1), human RHOT1 (Hs00430256_m1), human SNX1 (Hs00162052_m1), human TRAPPC13 (Hs01556088_m1), human GNAL (Hs00181836_m1), rat Adcy9 (Rn01424753_m1), rat Fbll1 (Rn06322963_s1), rat Zswim9 (Rn01753793_m1), rat Onecut2 (Rn01265320_m1), rat Snx1 (Rn01418446_m1), rat Trappc13 (Rn01493919_m1), rat Gnal (Rn01491898_m1) and rat Ins1 (Rn02121433_g1).

Techniques: Expressing, Functional Assay, Knockdown, Over Expression, Two Tailed Test

Journal: Nature Metabolism

Article Title: Cell-specific DNA methylation in human alpha and beta cells regulates gene expression in type 2 diabetes

doi: 10.1038/s42255-026-01498-9

Figure Lengend Snippet: a , Venn diagram showing T2D-associated differentially expressed genes (DEGs) in human α- and β-cells overlapping with DEGs identified in whole human islets by Bacos et al. b , c , Examples of T2D-associated DEGs found in β-cells (n = 15 non-diabetic donors and 7 donors with T2D) that overlap with T2D-associated DEGs found in human islets by Bacos et al. : GABRA2 , SOCS1 , SYT1 , PAX5 , CABLES1 and TBC1D4 ( *p < 0.05, **p < 0.01 based on DESeq2 ). d-j pre-T2D/T2D-associated differences in the absolute DNA methylation level (%) of individual CpG sites annotated to ADCY9 ( d ), ZNF431 ( e ) , ZSWIM9 ( f ) , GNAL ( g ), ONECUT2 ( h ), SNX1 ( i ), and TRAPPC13 ( j ), respectively in β-cells. *p < 0.05, **p < 0.01 pre-T2D/T2D (n = 3 donors) versus non-diabetic controls (n = 11 donors) based on two-tailed Wilcoxon rank-sum test. k , GNAL, NTN1 , and PRKCH are T2D-associated DEGs in both sorted α- and β-cells, as well as in whole islets from Bacos et al. . Data shown is for β-cells from pre-T2D/T2D (n = 7 donors) versus non-diabetic controls (n = 15 donors), *p < 0.05, ***p < 0.001. In b-k , women and men are represented by filled and open circles, respectively. Data are presented as mean ± SEM. For exact p-values, see Supp . Table (panels b , c , and k ) and Source data files (panels d-j ).

Article Snippet: TaqMan assays were used to measure expression levels of human ADCY9 (Hs00181599_m1), human FBLL1 (Hs01584505_s1), ZNF431 (Hs00819968_m1), human ZSWIM9 (Hs00736775_m1), human ONECUT2 (Hs00191477_m1), human RHOT1 (Hs00430256_m1), human SNX1 (Hs00162052_m1), human TRAPPC13 (Hs01556088_m1), human GNAL (Hs00181836_m1), rat Adcy9 (Rn01424753_m1), rat Fbll1 (Rn06322963_s1), rat Zswim9 (Rn01753793_m1), rat Onecut2 (Rn01265320_m1), rat Snx1 (Rn01418446_m1), rat Trappc13 (Rn01493919_m1), rat Gnal (Rn01491898_m1) and rat Ins1 (Rn02121433_g1).

Techniques: DNA Methylation Assay, Two Tailed Test

Journal: Nature Metabolism

Article Title: Cell-specific DNA methylation in human alpha and beta cells regulates gene expression in type 2 diabetes

doi: 10.1038/s42255-026-01498-9

Figure Lengend Snippet: a , Candidate gene selection for functional follow-up from RNA-seq data in sorted α- and β-cells from donors with pre-T2D/T2D versus controls. Flowchart shows the differentially expressed genes (DEGs) in pre-T2D/T2D versus non-diabetic controls in α- and β-cells, respectively, and selection of eight DEGs in β-cells for follow-up. b , Gene manipulation in EndoC-βH1 β-cells often gives different phenotypes compared to human islets and INS-1 β-cells. Figure shows that RHOT1 knockdown (left, ***p = 0.0002) resulted in increased glucose-stimulated insulin secretion (right, **p = 0.0041) in EndoC-βH1 β-cells, which should be compared to reduced insulin secretion seen in human islets and INS-1 β-cells treated similarly (Figs. f and in previous publication ). n = 6 biological replicates and paired t-tests. c , Reduced ADCY9 protein levels in INS-1 β-cells after siRNA transfection, n = 4 biological replicates, *p = 0.037 versus negative control (siNC) based on one-tailed paired Student’s t-test. d , Increased ONECUT2 protein levels in INS-1 β-cells after lentiviral transduction, n = 6 biological replicates, **p = 0.0079 versus GFP (control) based on paired Student’s t-test. e , No significant effect on oxygen consumption rate in siAdcy9 INS-1 β-cells, versus siNC, n = 4 biological replicates, p > 0.05 based on paired Student’s t-test. f , Reduced ATP5A protein levels in OE- ONECUT2 INS-1 β-cells, n = 6 biological replicates, *p = 0.049 versus GFP based on paired Student’s t-test. g , Microarray showed decreased expression of NDUFA1 (**p = 0.0022), NDUFA7 (**p = 0.0022), and COX10 (**p = 0.0043) in OE- ONECUT2 islets (n = 6 biological replicates/donors) based on Mann-Whitney U-tests, presenting unlogged data. h , Expression of NRID1 (**p = 0.0022), PTPRN (**p = 0.0022), and SLC8B1 (**p = 0.0087), contributing to the enrichment of the down-regulated gene set ‘regulation of insulin secretion’ in GSEA of OE- ONECUT2 human islets versus GFP (Mann-Whitney U-tests, presenting unlogged data). n = 6 biological replicates/donors. i , emapplot visualizing GSEA results of transcriptome data in human islets transfected with siADCY9 versus siNC. n = 5 biological replicates and GSEA was corrected for multiple testing. For full data, see Supplementary Table . j , ONECUT2 expression, and k , insulin secretion, presented as fold change (secretion at 16.7 mM glucose divided by secretion at 2.8 mM glucose), when silencing ONECUT2 using siRNA (siONECUT2) versus siNC in EndoC-βH1 β-cells exposed to high glucose (19 mM glucose) plus palmitate (1 mM palmitate, HGP) or control media for 72 h. n = 6 biological replicates, data analyzed by one-way ANOVA ( j , p = 0.0293, and k , p = 0.0108) with paired Student’s t-tests as post hoc. j , *p = 0.023: control+siNC versus HGP+siONECUT2, ***p = 4.38E-05: HGP+siNC versus HGP+siONECUT2, and k , *p = 0.017: control+siNC versus HGP+siNC, *p = 0.049: HGP+siNC versus HGP+siONECUT2. n.s.: Not significant. l , Cartoon summarizing key results from this study; created in BioRender; Ofori, J. https://BioRender.com/1w54ey4 (2026). Data in b-h and i-j are presented as mean ± SEM. Tests were two-tailed.

Article Snippet: TaqMan assays were used to measure expression levels of human ADCY9 (Hs00181599_m1), human FBLL1 (Hs01584505_s1), ZNF431 (Hs00819968_m1), human ZSWIM9 (Hs00736775_m1), human ONECUT2 (Hs00191477_m1), human RHOT1 (Hs00430256_m1), human SNX1 (Hs00162052_m1), human TRAPPC13 (Hs01556088_m1), human GNAL (Hs00181836_m1), rat Adcy9 (Rn01424753_m1), rat Fbll1 (Rn06322963_s1), rat Zswim9 (Rn01753793_m1), rat Onecut2 (Rn01265320_m1), rat Snx1 (Rn01418446_m1), rat Trappc13 (Rn01493919_m1), rat Gnal (Rn01491898_m1) and rat Ins1 (Rn02121433_g1).

Techniques: Selection, Functional Assay, RNA Sequencing, Knockdown, Transfection, Negative Control, One-tailed Test, Transduction, Control, Microarray, Expressing, MANN-WHITNEY, Two Tailed Test

Journal: Nature Metabolism

Article Title: Cell-specific DNA methylation in human alpha and beta cells regulates gene expression in type 2 diabetes

doi: 10.1038/s42255-026-01498-9

Figure Lengend Snippet: a , Reduced OCR in INS-1 832/13 beta cells overexpressing ONECUT2 (OE- ONECUT2 ). The left graph shows the oxygen consumption trace, and the right graph the quantification of derived respiratory parameters. n = 4 independent experiments. * P < 0.05 based on two-tailed paired Student’s t -tests. b , Reduced ATP production in OE- ONECUT2 INS-1 832/13 beta cells, n = 73 (GFP) and 116 (OE- ONECUT2 ) cells from six different experiments. *** P = 4.61 × 10 −14 based on a two-tailed Student’s t -test (right). c , emapplot visualizing results of the GSEA of the transcriptome data generated in OE- ONECUT2 versus control (GFP) human islets of six non-diabetic donors presented in Supplementary Table . The R packages clusterProfiler and enrichplot were used to generate the emapplot, and 23 gene sets are preselected. GSEA was corrected for multiple testing. d , The genes contributing to the enrichment for the downregulated gene set ‘regulation of insulin secretion’ in the GSEA of OE- ONECUT2 human islets, as well as selected individual downregulated genes in the gene set: FOXO1 (* P = 0.026), G6PC2 (* P = 0.0152), GCK (** P = 0.0087), GIPR (** P = 0.0022), GPR119 (* P = 0.0441), PDX1 (* P = 0.0441) and SLC2A2 (* P = 0.0441). P values are based on two-tailed Mann–Whitney U- tests and presenting unlogged data. n = 6 biological replicates of cells expressing GFP or ONECUT2 . e , WGCNA based on weighted correlations among OE- ONECUT2 DEGs with q < 0.1 showed that ONECUT2 is part of an expression cluster containing 114 DEGs (in network) and 108 are shown in the figure (due to Cytoscape cut-off 0.55), with direct connection to 30 DEGs. f – h , Male GK rats, a T2D animal model, showed higher Onecut2 (* P = 0.0458; f ), and nominally lower Ins ( P = 0.12; g ) expression in islets, as well as higher blood glucose levels (*** P = 6.11 × 10 −7 ; h ) compared with male control Wistar rats ( n = 5) based on two-tailed Student’s t -tests. Data in a , b , d and f – h are presented as the mean ± s.e.m. P values presented in a , b , d and f – h were not corrected for multiple testing.

Article Snippet: TaqMan assays were used to measure expression levels of human ADCY9 (Hs00181599_m1), human FBLL1 (Hs01584505_s1), ZNF431 (Hs00819968_m1), human ZSWIM9 (Hs00736775_m1), human ONECUT2 (Hs00191477_m1), human RHOT1 (Hs00430256_m1), human SNX1 (Hs00162052_m1), human TRAPPC13 (Hs01556088_m1), human GNAL (Hs00181836_m1), rat Adcy9 (Rn01424753_m1), rat Fbll1 (Rn06322963_s1), rat Zswim9 (Rn01753793_m1), rat Onecut2 (Rn01265320_m1), rat Snx1 (Rn01418446_m1), rat Trappc13 (Rn01493919_m1), rat Gnal (Rn01491898_m1) and rat Ins1 (Rn02121433_g1).

Techniques: Derivative Assay, Two Tailed Test, Generated, Control, MANN-WHITNEY, Expressing, Animal Model