Journal: bioRxiv

Article Title: Inhibition of the gut ceramidase Asah2 decelerates the vertebrate ageing rate

doi: 10.64898/2026.04.30.721799

Figure Lengend Snippet: a , Schematic representation of ceramide hydrolysis by membrane-tethered Asah2. Cer, ceramide; Sph, sphingosine. Image was created with BioRender.com. b , Loss of Asah2 activity is associated with a longer lifespan. Protein domain architectures of Asah2 are shown. Blue box, catalytic domain; orange box, immunoglobulin (Ig)-like domain; white box, other regions; grey box, N. furzeri MZM-specific region; red vertical lines in human ASAH2, essential amino acid residues required for enzymatic activity. c, e , asah2 is specifically expressed in the intestine and increases with age. Expression levels of asah2 mRNA in various tissues ( c ) and in the intestine ( e ) of 2- and 4-month-old wild-type (WT) and asah2 knockout (KO) N. furzeri are shown. In c and e , mRNA levels were analysed by reverse transcription quantitative PCR (RT–qPCR) and RNA sequencing (RNA-seq), respectively. CPM, counts per million; FDR, false discovery rate. FDR values were derived from quasi-likelihood methods in edgeR. d , Asah2 protein localises to the apical /luminal side of intestinal epithelium in short-lived GRZ fish but is absent in long-lived MZM fish and asah2 KO fish. Representative images of intestinal sections immunostained with anti-Asah2 antibody (green) and 4′,6-diamidino-2-phenylindole (DAPI; blue) are shown. Scale bar, 200 μm. f , Pooled censoring-aware parental survival analysis showing that higher participant plasma ASAH2 z-score was associated with higher pooled parental hazard, consistent with lower parental longevity. The displayed p-value tests the overall ASAH2-parental hazard association in the stratified cubic B-spline Cox model. Primary statistical inference was based on the cluster-robust linear Cox model. g , CRISPR/Cas9-mediated asah2 knockout. Top, schematic representation of the asah2 locus (black boxes, exons; ATG, start codon) and two sgRNA target sites (orange boxes). Bottom left, DNA sequences of WT and asah2 KO. The red horizontal line denotes the 4 bp deletion. Bottom right, the mutation was predicted to result in loss of the C-terminal domain of the Asah2 protein. Blue, catalytic domain; orange, Ig-like domain; white, other regions; grey, asah2 KO N. furzeri- specific region. h , Expression of Asah2 in the intestine of short-lived N. furzeri analysed by Western blotting. i, j , asah2 KO extends lifespan ( i ) and improves locomotor activity ( j ). In i , survival curves of male WT (blue) and asah2 KO (orange) fish are shown on the left; the corresponding statistical results, mean lifespan and maximum lifespan are shown on the right. Black dots indicate censored individuals. Graphs in j show exploratory behaviour in male WT (blue) and asah2 KO (orange) fish. The y-axis indicates the average velocity. The log-rank test, fixed-time survival test, and unpaired two-tailed t -test were used for i , and a Brown–Forsythe and Welch ANOVA followed by Dunnett’s T3 multiple-comparisons test were used for j . Bars and error bars represent the mean ± SD.

Article Snippet: Human tissue expression data were obtained from the Human Protein Atlas , .

Techniques: Membrane, Activity Assay, Expressing, Knock-Out, Reverse Transcription, Real-time Polymerase Chain Reaction, Quantitative RT-PCR, RNA Sequencing, Derivative Assay, Clinical Proteomics, CRISPR, Mutagenesis, Western Blot, Two Tailed Test

Journal: bioRxiv

Article Title: Inhibition of the gut ceramidase Asah2 decelerates the vertebrate ageing rate

doi: 10.64898/2026.04.30.721799

Figure Lengend Snippet: a , Asah2 expression increases with age in the human terminal ileum of the small intestine. Data were obtained from voyAGEr . CPM, counts per million. b , Sex-stratified pooled censoring-aware parental survival analysis of participant plasma ASAH2 z-score and pooled parental hazard in male and female participants. Displayed p-values denote the overall ASAH2-parental hazard association from stratified cubic B-spline Cox models. c, d , asah2 KO does not affect body size or body mass index ( c ) or reproductive capacity ( d ). Graphs in c show body weight, body length, and body mass index in WT (blue) and asah2 KO (orange) male fish. Animals measured at each time point were different individuals. In d , the graph shows the number of fertilized eggs in WT (blue) and asah2 KO (orange) fish. Bars and error bars ( c, d ) represent the mean ± SD. An unpaired two-tailed t -test was used in c , and a Brown–Forsythe and Welch ANOVA followed by Dunnett’s T3 multiple-comparisons test were used in d .

Article Snippet: Human tissue expression data were obtained from the Human Protein Atlas , .

Techniques: Expressing, Clinical Proteomics, Two Tailed Test