Multimodal 3-D/2-D human islet and duct imaging in exocrine and endocrine lesion environment: associated pancreas tissue remodeling

Supplemental Materials

Supplemental Figures

Suppl. Fig. S1 (related to Fig. 1). Multimodal imaging of islet aggregation with duct-islet cell cluster in PDAC surgical margin. (A-F) and (G-L) are two examples of the coexistence of islet aggregation, PanIN, acinar atrophy, and stromal accumulation in the surgical margin distal to PDAC (case No. 1628870 and 4442070). In F and L, the high-resolution images identify the duct (CK7⁺) -islet (insulin⁺ or glucagon⁺) cell cluster. Green, CK7; blue, insulin; magenta, glucagon; white, nuclei.

Suppl. Fig. S2 (related to Fig. 2). Detection and confirmation of duct lesion in human donor pancreas (gallery display).

Suppl. Fig. S3-1 (related to Fig. 4S). Detection and characterization of an endocrine-exocrine lesion mixture in human donor pancreas (CgA⁺, PGP9.5⁺, glucagon⁺ microadenoma with CK7⁺ epithelium in a stroma-rich environment). (A-E) Detection of glucagon⁺ microadenoma. A-C show microadenoma (arrow) in vibratome section. Fluorescence tissue map (D) identifies the intra-microadenoma ducts (CK7⁺, green; enlarged in inset i and ii, asterisk). Magenta, glucagon; blue, insulin; white, nuclei. PGP9.5 staining (E, cyan) confirms the islet/neuroendocrine lesion. Red, CD31. (F-M) Confirmation of microadenoma-duct lesion mixture with multiplex CgA, H&E, CD45, and Ki-67 signals. The endocrine-exocrine lesion mixture is associated with stromal accumulation (F-I; asterisk, enlarged area), leukocyte infiltration (J, K; asterisk/arrow, enlarged area), and prominent cell proliferation (L, M; box, enlarged area; blue arrows, Ki-67⁺ nuclei). An adjacent PanIN (within 2 mm) is presented in the next page. Black arrows in F, G, J, L indicate the same microenvironment.

Suppl. Fig. S3-2 (related to Fig. 5). Detection of a PanIN lesion adjacent to the microadenoma in panel A-M (Suppl. Fig. S3-1). (N-P) Microadenoma and its microenvironment visualized via stereomicroscopy. N-P and A-M examine the same microenvironment (arrow). Vibratome section P is further processed for H&E histology to confirm the PanIN lesion. (Q, R) Gross view and enlarged H&E images of PanIN. The gold-standard H&E histology (cyan arrows from P to Q to R, same environment) confirms the low-grade PanIN lesion and peri-lesional stroma. Box in Q is magnified in R to specify the lesion.

Suppl. Fig. S4 (related to Fig. 4O-R). False positive result in fluorescence imaging of islet cell microadenoma in human pancreas. (A-C) Gross view and side-by-side display of fluorescence and transmitted light signals of microadenoma. The three images were derived from an optically cleared microadenoma (refractive index: 1.52). A: microadenoma alongside pancreatic lobules (red, blood vessels, CD31; green, lymphatic vessels, D2-40; white, nuclei, DAPI). B: enlarged fluorescence image of vasculature (blood and lymphatic vessels). C: overlay of transmitted light and nuclear signals. Adipocytes and blood clots (black spots) are prominently seen in C via in-depth transmitted light signals. Number 1-6 indicate the adipocytes (#1-2) and blood clots (#3-6) shown in both B and C. Alphabet a-f indicate the adipocytes and blood clots appear only in C. CD31 signals of B is presented in D. B-D examine the same microenvironment. (D, E) False positive result revealed by analysis of CD31 fluorescence signals of microadenoma. Relative signals intensity along the central line of D (0-2,000 pixels) is presented in E. Both CD31-labeled blood vessels and the blood clot-induced autofluorescence create local increases in the signal intensity (E). Number 4 in E indicates the autofluorescence caused by the blood clot #4 in panel B-D.

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Supplemental Tables

Suppl. Table S1 (Related to Materials and Methods). Clinical data of pancreatic ductal adenocarcinoma cases.

Suppl. Table S2 (Related to Materials and Methods). Pancreas organ donor information and lesion summary.

Suppl. Table S3 (Related to Materials and Methods). Summary of primary antibodies used in illustrations.

Suppl. Table S4 (Related to Materials and Methods). Summary of color codes presented in illustrations.

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Supplemental Video

Suppl. Video S1 (related to Fig. 3L-N). Tile scan and in-depth recording of PanIN microenvironment in human pancreas. This video shows a survey of PanIN microenvironment to identify the associated exocrine-endocrine tissue remodeling. Duct lesion and islet a- and b-cells form a complex in the peri-lobular space. Green, CK7; magenta, glucagon; blue, insulin; white, nuclei. Overlay of fluorescence and transmitted light (gray) signals is used to identify the boundaries of pancreatic lobules and CK7⁺ epithelium.