Integrating Magnetic Resonance Chemical Shift Imaging for Localized Prostate Cancer Risk Stratification on the Basis of the Impact of Periprostatic Brown Adipocytes Within Tumor Microenvironment

Figures

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  Fig. 1. A, B Collection method for preprostate fat during the operation; after developing the space of Retzius and anterior prostate dissection, direct visualization of the anterior prostate and periprostatic fat was dissected; C, D chemical shift MRI imaging of water-to-oil fraction incorporated into T2 series showing the orientation of periprostate tumor fat collected during the operation; E, F significantly higher water-to-oil ratio signal of periprostate tumor fat was shown in high-risk patient compared with that in low-risk patients; G receiver operating characteristic curve showing area under curve of 0.90 in determining high risk patient from iPSA and water-fat-ratio imaging

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  Fig. 2. A–C Hematoxylin and eosin stain of periprostatic fat, (A) mice interscapular BAT as positive control, (B) human adipocytes of periprostate tumor region, (C) human adipocytes of pre-prostate fat distant from tumor, and D apparent diffusion coefficient revealing the tumor located at right peripheral zone; E chemical shift MRI imaging showing orientation of periprostate tumor fat collected during surgery; F hematoxylin and eosin stain (H&E stain) in periprostate fat near tumor showing two different types of adipocyte; G qRT-PCR in periprostatic tumor fat and subcutaneous fat; increased expression of BAT-specific markers including UCP1, TBX, and LHX8 in periprostatic tumor fat compared with preprostate fat; increased expression of WAT-specific markers including HOXC8, HOXC9, and Leptin in preprostate fat compared with periprostatic tumor fat; ; H–I different signal intensities of UCP-1 in IHC reveals the distribution of BAT (red box) and WAT (blue box) within periprostatic fat near tumor region

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  Fig. 3. Immunofluorescence staining (IF) with primary antibody UCP-1 Rb mAb 1:50 dilution revealed positive uncoupling protein 1 (UCP-1) signal within periprostate tumor fat (A–D); (A) interscapular fat containing BAT collected in mice; (B–D) periprostate fat tissue collected in patient number 6: (B) human subcutaneous fat (400×), (C) periprostate tumor fat (400×), and (D) preprostate fat tissue (400×)

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  Fig. 4. UCP1-positive brown adipocyte within periprostate tumor fat; A chemical shift modified DIXON MRI image depicting the orientation of periprostate fat; B apparent diffusion coefficient sequence of image showing the location of prostate tumor; C–F cell culture at 1 week in periprostate fat near tumor and periprostate fat distant from tumor; (C,D) adiopose tissue distant from tumor after 1 week showing cluster of spherical adipocytes containing a large, single lipid droplet suggesting white adipocyte; (E,F) primary culture result of periprostate adipose tissue showing multilocular droplets, spherical and centrally located nuclei suggesting brown adipocyte; G qRT-PCR results reveal significantly higher expression of UCP1 within periprostate tumor fat compared with preprostate fat; H Kaplan–Meier survival analysis of UCP-1 expression and time to biochemical recurrence in patients with prostate cancer

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  Fig. 5. A–D Transwell invasion assay in different periprostate adipose culture medium groups; (A) representative images of the invasion assay in PC3 cells; PC3 cell invasion through collagen-coated inserts after a 24 h exposure to medium containing supernatant of peritumor adipocyte culture suggesting periprostate tumor fat affects prostate cancer cell invasion potential (n = 14); (B) LNCaP cell invasion from collagen-containing inserts after a 24-h exposure to medium containing supernatant of peritumor adipocyte culture (n = 15); all signals were normalized to 10.0% FBS (C–D) Representative images of the invasion assay from patient number 13 showing increased cancer cell migration potential attracted by peri-tumor fat culture medium. E–G Peri-tumor adipocyte co-culture with prostate cancer cell. Periprostate tumor fat affects prostate cancer cell invasion potential is partially contributed by epithelial mesenchymal transition. (E) RT-PCR showing periprostate tumor fat exhibit higher RNA transcription of N-cadherin, Slug, snail, ZEB1,ZEB2, vimentin, and ABCB1 and a lower E-cadherin. (F) Western blots of LNCaP cancer cells 72 hours after co-culture with human peri-prostatic adipocyte…