Associate Research Professor Johns Hopkins University, United States
Introduction: Cellular senescence plays a crucial role in regulating several physiological processes. However, understanding the distribution and abundance of senescent cells in human tissue remains limited. In this study, we developed a multiplex immunofluorescence (mxIF) workflow to simultaneously detect six markers of senescence, namely P53, P21, P16, HMGB1, 53BP1, and LaminB1, on formalin-fixed paraffin-embedded (FFPE) tissue sections of the human pancreas. By integrating H&E staining and mxIF imaging on serial sections, we generated large 3D volumetric maps of senescence markers in both diseased and adjacent healthy human pancreatic tissues, encompassing a substantial volume of 15mm × 15mm × 0.5mm (150 sections). Our analysis mapped approximately 18 million cells with expression level of senescence markers (across 30 sections). Furthermore, we employed deep learning models to extract tissue compartment information (10+ compartments) in the volume from H&E images and integrated this data with the mxIF volume to explore the relationships between senescence markers and tissue identities.
Materials and
Methods: Human healthy pancreas tissues are offered by Laura'd lab in Johns Hopkins Medicine and JAX laboratory.3D reconstruction of H&E images is performed by CODA platform developed by our group. The mxIF is performed by CLAP-mxIF methods. Low abundance proteins are detected with TSA signal amplification method, and high abundance proteins are detected by cycIF or IMC.
Results, Conclusions, and Discussions: Our 3D volumetric analysis indicated a strong association between senescence marker expression and specific tissue compartments within the pancreas. Specifically, we observed predominant p16 expression in the endocrine (i.e., islet) regions, while p21 and p53 were more enriched in exocrine (acinar) regions. The 3D volumetric visualization suggested the overexpression of acinar related senescence markers like p53 and p21 in pancreatitis region.
