Journal: bioRxiv

Article Title: Primary cilia regulate GLP-1 signaling in pancreatic β cells

doi: 10.64898/2026.02.22.707280

Figure Lengend Snippet: ( A ) Dynamic insulin secretion from perifused mouse islets. Wild-type (WT, black) and β-cell-specific cilia knockout (βCKO, red) islets were sequentially exposed to 2 mM glucose (2G), 16 mM glucose (16G), 20 nM liraglutide (Lira), and 30 mM KCl as indicated. N = 6 replicates of 50 islets per genotype from 5 mice. *p<0.05, two-way ANOVA with Sidak’s multiple comparisons test. ( B–D ) Quantitative analysis of secretion traces. ( B ) Area under the curve (AUC) during liraglutide stimulation (minutes 30–50). ( C ) Total AUC of the entire perifusion (minutes 0-70). ( D ) AUC during KCl depolarization (minutes 60–70). βCKO islets secreted significantly less insulin in response to liraglutide and overall but showed no defect in KCl-induced secretion. Data are mean ± SEM. **p<0.01; ns, not significant, unpaired student’s t-test. ( E–J ) IFT88 knockdown impairs GLP-1-augmented secretion in human islets. Static glucose-stimulated insulin secretion (GSIS) assays from islets of male ( E–G ) and female ( H–J ) donors. Islets transduced with control (Ctl, white) or IFT88 -targeting shRNA (IFT88 KD, red) were incubated at 1 mM glucose (1G), 11 mM glucose (11G), and 11G + 100 nM liraglutide (Lira). Donor ages are indicated. IFT88 KD significantly reduced liraglutide-potentiated insulin secretion. Data are mean ± SEM of triplicate samples per donor. *p<0.05, **p<0.01, ***p<0.001, ****p<0.0001; ns, not significant; one-way ANOVA with Tukey’s multiple comparisons test.

Article Snippet: IFT88 knockdown in human islets: Healthy non-diabetic human islets were transduced with adenoviral vectors encoding either GFP-tagged shRNA targeting human IFT88 (Ad-GFP-h-IFT88-shRNA) or scrambled control (Ad-GFP-U6-scrmb-shRNA, #1122N; Vector Biolabs).

Techniques: Knock-Out, Knockdown, Transduction, Control, shRNA, Incubation

Journal: bioRxiv

Article Title: Primary cilia regulate GLP-1 signaling in pancreatic β cells

doi: 10.64898/2026.02.22.707280

Figure Lengend Snippet: ( A ) Representative z-projected confocal images of human islets (28-year-old female donor) transduced with scrambled (Scr) or IFT88 -targeting shRNA adenovirus. GFP (green) marks transduced cells; ARL13B (red) labels primary cilia; DAPI (blue) stains nuclei. Scale bar = 20 µm. Cilia length was quantified from both conditions, showing significantly reduced mean length following IFT88 knockdown. n=132 cilia from 6 islets (Scr shRNA) and 467 cilia from 8 islets (IFT88 shRNA). ( B–C ) IFT88 knockdown efficiency was assessed across multiple donors to demonstrate reproducibility. ( B ) qPCR analysis of IFT88 mRNA levels in islets from two male donors (ages 27 and 44). ( C ) Immunoblot analysis of IFT88 protein levels in islets from three male donors (ages 36, 49 and 45), with corresponding quantifications. Bars represent scrambled control (gray) and IFT88 shRNA (pink). Data are mean ± SEM of duplicate or triplicate samples per donor. **p < 0.01, ****p < 0.0001, ns = not significant, by student’s unpaired t-test.

Article Snippet: IFT88 knockdown in human islets: Healthy non-diabetic human islets were transduced with adenoviral vectors encoding either GFP-tagged shRNA targeting human IFT88 (Ad-GFP-h-IFT88-shRNA) or scrambled control (Ad-GFP-U6-scrmb-shRNA, #1122N; Vector Biolabs).

Techniques: Transduction, shRNA, Knockdown, Western Blot, Control

Journal: Frontiers in Cellular Neuroscience

Article Title: Chemogenetic Schwann cell activation impairs early myelination and triggers adult demyelination in the peripheral nervous system

doi: 10.3389/fncel.2026.1771951

Figure Lengend Snippet: Expression and functional validation of hM3Dq in SCs within DRG co-culture. (A) CNO stimulation of hM3Dq-expressing SCs in DRG co-culture visualized using GCaMP fluorescence. Representative images of basal and CNO-stimulated cells are shown. Scale bar = 80 μm. (B) Immunocytochemical confirmation of hM3Dq expression in SCs using HA-tag and Sox10 antibodies. Representative images are shown. Scale bars = 60 μm, 40 μm, and 20 μm from left to right. (C) Functional validation of hM3Dq activation in SCs, demonstrated by an increase in GCaMP dF/F₀ following initial CNO stimulation. (D) Quantification of basal Ca 2+ levels, peak frequency, and peak amplitude in SCs after 3 days of CNO treatment. Representative Ca 2+ traces from individual vehicle- and CNO-treated SCs are shown. Data were collected from at least four independent biological replicates per condition, with approximately 100 cells analyzed per replicate. (E,F) SCs pre-treated with CNO for 3 days were subsequently stimulated with potassium (50 mM), glutamate (100 μM), ATP (100 μM), or acetylcholine (100 μM) in either Ca 2+ -free or Ca 2+ -containing media. (E) Representative Ca 2+ traces of vehicle- and CNO-treated SCs stimulated with glutamate, ATP, and acetylcholine in the presence of extracellular Ca 2+ are shown. Each trace represents a single cell; dF/F₀ measurements were recorded over a 480-s period from approximately 300 individual cells. (F) Bar graphs display changes in GCaMP fluorescence (dF/F₀) during stimulation, quantified as area under the curve (AUC). Dots represent values from independent biological replicates, each analyzing approximately 100 cells. Data are presented as mean ± SEM. Statistical significance: * p < 0.05; ** p < 0.01; *** p < 0.001 versus corresponding vehicle control.

Article Snippet: On day 7, cells were transduced with an adenoviral vector encoding the Ca 2+ indicator GCaMP6m (Ad-GCaMP6m; Vector Biolabs) at a multiplicity of infection (MOI) of 25 for 24 h. Following transduction, cultures were treated for 3 days with either vehicle (0.9% saline solution) or 10 μM CNO to activate hM3Dq signaling.

Techniques: Expressing, Functional Assay, Biomarker Discovery, Co-Culture Assay, Fluorescence, Activation Assay, Single Cell, Control

Journal: bioRxiv

Article Title: TACE reprograms RANKL-mediated differentiation of macrophages by activating the non-canonical pathway of IRF3

doi: 10.64898/2026.02.11.705073

Figure Lengend Snippet: TACE f/f mice were crossed with myeloid-specific LysM-Cre mice to generate mice lacking TACE in myeloid cells (TACE ΔM ). ( a and b ) BMDMs from TACE con and TACE ΔM mice were stimulated with RANKL (50 ng/ml) for the indicated times. ( a ) Immunoblot analysis using antibodies against phosphorylated ERK (p-ERK), JNK (p-JNK), or p38. ( b ) Nuclear p65 activity was assessed by immunoblot analysis. Lamin B1 was used as a control for nuclear fractions. ( c ) Immunoblot analysis of NFATc1 and α-tubulin during osteoclastogenesis. ( d - i ) BMDMs from TACE and TACE mice were treated with M-CSF and RANKL for two days. Total RNA was analyzed by bulk RNA-seq. ( d ) Schematic diagram of the experimental conditions representing the early phase of RANKL stimulation. ( e ) Volcano plot showing 39 genes upregulated and 118 genes downregulated in TACE cells (|log2 FC| ≥ 2, p < 0.05). ( f ) Gene Ontology (GO) enrichment analysis of the upregulated genes in TACE cells. ( g ) Hallmark pathway analysis of the upregulated genes in TACE cells. ( h ) Expression of IFN-dependent genes CXCL10 and IRF7 was measured by RT-qPCR. ( i ) Upstream regulator analysis was performed using Ingenuity Pathway Analysis (IPA). ( j ) BMDMs from WT mice were stimulated with RANKL (50 ng/ml) for the indicated times. Nuclear and cytosolic proteins were extracted and analyzed by immunoblotting with IRF3, Lamin B1, or α-tubulin antibodies. ( k ) BMDMs from TACE mice were stimulated with RANKL (50 ng/ml) for the indicated times. Nuclear fractions were analyzed by immunoblotting with IRF3 and Lamin B1 antibodies. All data are presented as mean ± SEM. *p < 0.05; **p < 0.01 by one-way ANOVA with post hoc Tukey’s test ( h ). Data represent at least three independent experiments.

Article Snippet: For adenoviral transduction, recombinant adenoviral particles encoding human TACE (ADV-200349) or IRF3 (ADV-212405) and control (1060) adenoviral particles encoding green fluorescent protein (Ad-CMV-GFP) were purchased from Vector Biolabs (Malvern, PA, USA).

Techniques: Western Blot, Activity Assay, Control, RNA Sequencing, Expressing, Quantitative RT-PCR

Journal: bioRxiv

Article Title: TACE reprograms RANKL-mediated differentiation of macrophages by activating the non-canonical pathway of IRF3

doi: 10.64898/2026.02.11.705073

Figure Lengend Snippet: ( a and b ) BMDMs from WT and IRF3 KO mice were cultured with M-CSF and RANKL for 3 days(n=3). ( a ) Efficiency of IRF3 knockout (KO). IRF3 mRNA expression was measured by RT-qPCR and normalized to HPRT. ( b ) Osteoclastogenesis assay. Left: representative images of TRAP-stained cells. Right: quantification of TRAP-positive multinucleated cells as a percentage relative to WT cells. ( c and d ) BMDM from WT mice were transduced with either control-GFP or hIRF3-expressing adenovirus and cultured with M-CSF (20 ng/ml) and RANKL (40 ng/ml) for 3 days(n=3). ( c ) Efficiency of hIRF3. hIRF3 mRNA expression was measured by RT-qPCR and normalized to HPRT ( d ) Osteoclastogenesis assay. Left: TRAP staining of representative images. Scale bar: 100 µm. Right: Quantification of TRAP-positive multinucleated cells. ( e and f ) BMDMs from WT and IRF3 KO mice were nucleofected with either control (CTL) or TACE-targeting siRNAs and cultured with M-CSF. ( e ) Knockdown (KD) efficiency. TACE mRNA levels were measured by qPCR and normalized to HPRT. ( f ) Osteoclastogenesis assay. Left: representative images of TRAP-stained cells. Right: quantification of TRAP-positive multinucleated cells as a percentage relative to WT-NC cells. All data are presented as mean ± SEM. n.s., not significant. Scale bar: 100 µm. *p < 0.05; **p < 0.01 by two-tailed, unpaired t-test ( b , c , d ) or one-way ANOVA with post hoc Tukey’s test ( a , e , f ). Data represent at least three independent experiments.

Article Snippet: For adenoviral transduction, recombinant adenoviral particles encoding human TACE (ADV-200349) or IRF3 (ADV-212405) and control (1060) adenoviral particles encoding green fluorescent protein (Ad-CMV-GFP) were purchased from Vector Biolabs (Malvern, PA, USA).

Techniques: Cell Culture, Knock-Out, Expressing, Quantitative RT-PCR, Staining, Transduction, Control, Knockdown, Two Tailed Test

Journal: bioRxiv

Article Title: TACE reprograms RANKL-mediated differentiation of macrophages by activating the non-canonical pathway of IRF3

doi: 10.64898/2026.02.11.705073

Figure Lengend Snippet: ( a - c ) BMDMs from WT and IRF3 KO mice were treated with M-CSF and RANKL for one day. Total RNA was analyzed by bulk RNA-seq with two biological replicates. ( a ) Volcano plot showing 692 genes upregulated and 262 genes downregulated in IRF3-deficient cells (|log2 FC| ≥ 2, p < 0.05). ( b ) Gene Ontology (GO) enrichment analysis of the upregulated genes in IRF3 KO cells. ( c ) Hallmark pathway analysis of the upregulated genes in IRF3 KO cells. ( d and e ) BMDMs from WT and IRF3 KO mice were stimulated with RANKL (50 ng/ml) for the indicated times(n=3). ( d ) Expression of IFN-dependent genes CXCL10, IRF7, and Mx1 was measured by RT-qPCR. ( e ) Expression of NFATc1 was measured by RT-qPCR as a marker of osteoclastogenesis. All data are presented as mean ± SEM. *p < 0.05; **p < 0.01 by one-way ANOVA with post hoc Tukey’s test ( d , e ).

Article Snippet: For adenoviral transduction, recombinant adenoviral particles encoding human TACE (ADV-200349) or IRF3 (ADV-212405) and control (1060) adenoviral particles encoding green fluorescent protein (Ad-CMV-GFP) were purchased from Vector Biolabs (Malvern, PA, USA).

Techniques: RNA Sequencing, Expressing, Quantitative RT-PCR, Marker

Journal: bioRxiv

Article Title: TACE reprograms RANKL-mediated differentiation of macrophages by activating the non-canonical pathway of IRF3

doi: 10.64898/2026.02.11.705073

Figure Lengend Snippet: ( a – c ) BMDMs from WT and IRF3 KO mice were treated with M-CSF and RANKL for one day, and total RNA was analyzed by RNA sequencing. ( a ) Differential gene expression (DEG) analysis was performed to compare IRF3-dependent genes with those induced by RANKL. ( b ) GSEA analysis of the upregulated genes by IRF3 deficiency in Group C. ( c ) Heatmap of genes in the Hallmark_TNFA signaling pathway. (d) Expression of the Hbegf gene upon RANKL stimulation in WT and IRF3 KO BMDMs was measured by RT-qPCR. ( e ) Expression of the Hbegf gene upon RANKL stimulation in WT and IFNAR KO BMDMs was measured by RT-qPCR. ( f – h ) BMDMs from WT mice were cultured with M-CSF and RANKL in the presence or absence of recombinant HB-EGF (100ng/ml). ( g ) BMDMs from WT mice were stimulated with RANKL (50 ng/ml) for 24 hours. Immunoblotting was performed using NFATc1 and β-actin antibodies. ( h ) Nuclear fractions were analyzed by immunoblotting with IRF3 and Lamin B1 antibodies. ( i ) Schematic representation of TACE-dependent cross-regulation between IRF3 and HB-EGF, which fine-tunes osteoclast differentiation and arthritic bone erosion. All data are presented as mean ± SEM. Scale bar: 100 µm. *p < 0.05; ****p < 0.001 by one-way ANOVA with post hoc Tukey’s test ( d , e ) or two-tailed, unpaired t-test ( f ). Data represent at least three independent experiments.

Article Snippet: For adenoviral transduction, recombinant adenoviral particles encoding human TACE (ADV-200349) or IRF3 (ADV-212405) and control (1060) adenoviral particles encoding green fluorescent protein (Ad-CMV-GFP) were purchased from Vector Biolabs (Malvern, PA, USA).

Techniques: RNA Sequencing, Gene Expression, Expressing, Quantitative RT-PCR, Cell Culture, Recombinant, Western Blot, Two Tailed Test