Type 2 diabetes alters quiescent pancreatic stellate cells to tumor-prone state
Yutaro Hara, Hiroki Mizukami, Takahiro Yamada, Shuji Shimoyama, Keisuke Yamazaki, Takanori Sasaki, Zhenchao Wang, Hanae Kushibiki, Masaki Ryuzaki, Saori Ogasawara, Hiroaki Tamba, Akiko Itaya, Norihisa Kimura, Keinosuke Ishido, Shinya Ueno, Kenichi Hakamada
Yutaro Hara, Hiroki Mizukami, Takahiro Yamada, Shuji Shimoyama, Keisuke Yamazaki, Takanori Sasaki, Zhenchao Wang, Hanae Kushibiki, Masaki Ryuzaki, Saori Ogasawara, Hiroaki Tamba, Akiko Itaya, Norihisa Kimura, Keinosuke Ishido, Shinya Ueno, Kenichi Hakamada
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Research Article Endocrinology Gastroenterology

Type 2 diabetes alters quiescent pancreatic stellate cells to tumor-prone state

Abstract

Pancreatic stellate cells (PSCs) are the origin of cancer-associated fibroblasts. Type 2 diabetes mellitus (T2D) may promote pancreatic ductal adenocarcinoma (PDAC), eliciting changes in the quiescent PSC (qPSC) population from the precancerous stage. However, the details are unknown. We evaluated the subpopulations of qPSCs and the impact of T2D. PSCs isolated from 8-week-old C57BL/6J mice and diabetic db/db mice were analyzed by single-cell RNA-seq. Sorted qPSCs and PDAC cells were transplanted into allogenic mice. The isolated qPSCs were broadly classified into mesothelial cell and pancreatic fibroblast (Paf) populations by single-cell RNA-seq. Pafs were subclassified into inflammatory Pafs, myofibroblastic Pafs (myPafs) and a small population named tumor immunity- and angiogenesis-promoting Pafs (tapPafs), expressing Cxcl13. In the subcutaneous transplantation model, the tumors transplanted with myPafs were significantly larger than the tumors transplanted with tapPafs. An increase in myPafs and a decrease in tapPafs were observed from the precancerous stage in human T2D, indicating the effects of tumor progression. This study revealed the subpopulation changes in qPSCs in T2D. A therapy that increases the number of tapPafs could be a therapeutic option for patients with PDAC and T2D and even those in a precancerous stage of T2D.

Authors

Yutaro Hara, Hiroki Mizukami, Takahiro Yamada, Shuji Shimoyama, Keisuke Yamazaki, Takanori Sasaki, Zhenchao Wang, Hanae Kushibiki, Masaki Ryuzaki, Saori Ogasawara, Hiroaki Tamba, Akiko Itaya, Norihisa Kimura, Keinosuke Ishido, Shinya Ueno, Kenichi Hakamada

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Figure 5

Tumor immunity and angiogenesis elicited by tapPafs in the allogeneic transplantation model mice with normal immunity.

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Tumor immunity and angiogenesis elicited by tapPafs in the allogeneic tr...
(A) The tumor growth rate was evaluated after the subcutaneous injection of 1.0×105 KPCY tumor cells cotransplanted with 1.0×105 myPafs or 2.0×105 KPCY tumor cells cotransplanted with 1.0×105 tapPafs or 1.0×105 total PSCs in syngeneic C57BL/6J mice (n = 4 mice per condition). (B) Representative photographs showing dissected transplanted PDAC tumors from C57BL/6J mice. (C) Representative images of pathological H&E staining and immunohistochemical analysis of CD3-, CD8- and B220-positive immune cell infiltration in tumors transplanted with myPafs and tapPafs (original magnification, ×20). (D) The densities of total cells and CD3-, CD8- and B220-positive cells quantitatively evaluated in immunostained sections of the transplanted tumors (n = 4 per each group). (E) Immunofluorescence for CD31 revealing microvessel channels in the tumors. The full size of the tumor is shown in the inset (original magnification, ×20). (F) MVD quantified in the CD31-positive area using Otsu’s method for image thresholding (n = 4 per each group). (G) PKH26-labeled Pafs (red fluorescence) retained in tumors 4 weeks after transplantation (original magnification, ×20). (H) Comparison of mRNA fold changes measured by microarray. Statistical analysis was performed by 2-way ANOVA with post hoc multiple-comparison tests. MVD, microvascular density; Pafs, pancreatic fibroblasts; myPafs, myofibroblastic Pafs; PSCs, pancreatic stellate cells; tapPafs, tumor immunity- and angiogenesis-promoting Pafs; Fd, field. The data are presented as the means ± SDs. *P < 0.05, ** P < 0.01, *** P < 0.001. Scale bars: 100 μm (C and G) and 300 μm (E).