Fig 1.
KPIC mice formed a local invasive tumor with 3-months mean survival.
(A) The gross anatomy of PC in a KPIC mouse showed cancer in the tail and body of pancreas, and the swelling of small intestine. The KPIC tumor locally invaded the neighboring organs (In, intestine; ki, kidney; yellow circle, tumor region). (B) Kaplan-Merrier curves of KPIC and KIC mice. Log-rank test. (C) H&E and Mason Trichrome staining of KPIC and KIC tumors. Both KPIC and KIC tumors showed a character of typical adenocarcinoma that tumor cells grow in a ductal manner. (D) Mason Trichrome staining showed the dense desmoplastic stroma of KPIC and KIC tumors. (E) Sox-9 immuostaining of KPIC and KIC tumors. Scale bar, 100μm.
Fig 2.
The molecular profiles and EMT of neoplastic cells of KPIC tumors resemble human PC.
(A) E-cadherin and CK immunostaining showed that KPIC tumors are epithelial. (B-D) Co-immuostaining p53, pMAPK and Her2 antibodies with E-cadherin antibody in KPIC tumors showed that their molecular profiles are similar to that in human PC. (E, F) Triple immunofluorescent staining of E-cadherin, vimentin and cytokeratin or Her2 staining showed that KPIC tumors harbor a significant number of EMT cells. Her2 expression in EMT cells was upregulated. (circled region, EMT region; white yellow and white arrows, EMT cells). Scale bar, 50μm.
Fig 3.
The proliferative and glucose uptake ability and microvasculature characteristics of KPIC tumors.
(A) Immuostaining of KPIC tumor and typical human PC with Ki67 antibody showed that the neoplastic cells of KPIC tumor are highly proliferative, and similar to the highly proliferative neoplastic cells in human PC. Scale bar, 50μm. n, sample size. (B) CD34 immunofluorescent staining showed that KPIC tumor has a “hairy” microvasculature with basal microvilli that is rarely branched and similar to the typical “hairy” microvasculature in human PC (Human PC, Age 69, Male, tumor in head of pancreas). Scale bar, 20μm. (C) Accumulations of 2-NBDG-Alex 488 and Lectin-Alex-633 in KPIC tumors. First panel shows a panIN lesion; Second panel shows a typical adenocarcinoma. Scale bar, 20μm. (D) Accumulations of 2-NBDG-Alex 633 and Lectin-Alex 488 in the neoplastic cells of KIC tumors. Scale bar, 20μm.
Fig 4.
The isolation and characterization of the neoplastic cells of KPIC tumor.
(A) The morphology of KPIC cells showed that KPIC cells have the characteristics of epithelial cells, and formed an adhesive, dense and compact monolayer. Scale bar, 100μm. (B-F) Co-immunostainig of KPIC tumor cells with Sox-9, CK, p53p, pMAPK, and pAkt (ser 473) antibodies and E-cadherin antibody. Scale bar, 50μm (B); 20μm (C-F). (G) Genotyping of KPIC cells showed that KPIC cells carried four mutant alleles. (H) Ki67 staining showed that the proliferation of KPIC cells are faster than that of KPC cells. **, P<0.01 using student t-test. Scale bar, 50μm. (I) Comparing aneuploid cells in KPIC with KPC cells. N.S, not statistically significant by student t-test. Scale bar, 50μm.
Fig 5.
KPIC cells are easy to EMT under drug stress.
(A) The morphological changes of KPIC and KPC cells under gemcitabine and Abraxane stresses. (B) Vimentin and E-cadherins staining of gemcitabine and Abraxane treated KPIC and KPC cells. Gemcitabine and Abraxane treatment induced EMT in both cells. Scale bar, 50μm.
Fig 6.
KPIC cells are tumorigenic in immunocompetent mouse.
(A) The gross pathology of an orthotopic KPIC cell tumor showed that the orthotopic tumor invaded the spleen and liver (white circle, orthotopic tumor; green line, liver invasion; yellow line, spleen invasion). (B) H&E staining, Mason Trichrome and Ki67 immunostaining of orthotopic KPIC tumors showed that the orthotopic tumors are invasive, and contain desmoplastic stroma and highly proliferative cells. (C) H&E staining, p53 and CK antibodies immunohistochemistry of the orthotopic tumor of KPIC cells showed the invasion of orthotopic KPIC tumor to liver. These KPIC cells are ductal epithelial cells and accumulate p53. (D, E) H&E staining showed the orthotopic tumor invasion to spleen and lymph node. White dotted lines (B-D), the boundary between tumor and normal tissue. Ca, cancer; Pan, pancreas. (F) Histology of original spontaneous tumor of KPIC. Scale bar, 100μm.
Fig 7.
KPIC cells are metastatic in immunocompetent mouse.
(A) Orbital intravenous injection of KPIC cells to immunocompetent mice formed multiple metastatic lesions in the lung (19 days). Metastatic KPIC cells are proliferative and Sox-9- and p53-positive. Scale bars, 500μm and 100μm. (B) KPIC cells formed an in situ tumor at the injection site which also invaded to the muscles of eye. KPIC tumor cells are proliferative and Sox-9- and p53- positive. (yellow arrow, tumor in eye; green arrows, muscle; white arrows, tumor cells). Scale bar, 100μm.