Development of an Orthotopic Human Pancreatic Cancer Xenograft Model Using Ultrasound Guided Injection of Cells

Mice have been employed as models of cancer for over a century, providing significant advances in our understanding of this multifaceted family of diseases. In particular, orthotopic tumor xenograft mouse models are emerging as the preference for cancer research due to increased clinical relevance over subcutaneous mouse models. In the current study, we developed orthotopic pancreatic cancer xenograft models in mice by a minimally invasive method, ultrasound guided injection (USGI) comparable to highly invasive surgical orthotopic injection (SOI) methods. This optimized method prevented injection complications such as recoil of cells through the injection canal or leakage of cells out of the pancreas into the peritoneal cavity. Tumor growth was monitored in vivo and quantified by ultrasound imaging weekly, tumors were also detected by in vivo fluorescence imaging using a tumor targeted molecular probe. The mean tumor volumes for the USGI and SOI models after 2 weeks of tumor growth were 205 mm3 and 178 mm3 respectively. By USGI of human pancreatic cancer cell lines, human orthotopic pancreatic cancer xenografts were established. Based on ultrasound imaging, the orthotopic human pancreatic cancer xenograft take rate was 100% for both human pancreatic cancer cell lines used, MiaPaCa-2 and Su86.86, with mean tumor volumes of 28 mm3and 30 mm3. We demonstrated that this USGI method is feasible, reproducible, facile, minimally invasive and improved compared to the highly-invasive SOI method for establishing orthotopic pancreatic tumor xenograft models suitable for molecular imaging

Development of an Orthotopic Human Pancreatic Cancer Xenograft Model Using Ultrasound Guided Injection of Cells

Mice have been employed as models of cancer for over a century, providing significant advances in our understanding of this multifaceted family of diseases. In particular, orthotopic tumor xenograft mouse models are emerging as the preference for cancer research due to increased clinical relevance over subcutaneous mouse models. In the current study, we developed orthotopic pancreatic cancer xenograft models in mice by a minimally invasive method, ultrasound guided injection (USGI) comparable to highly invasive surgical orthotopic injection (SOI) methods. This optimized method prevented injection complications such as recoil of cells through the injection canal or leakage of cells out of the pancreas into the peritoneal cavity. Tumor growth was monitored in vivo and quantified by ultrasound imaging weekly, tumors were also detected by in vivo fluorescence imaging using a tumor targeted molecular probe. The mean tumor volumes for the USGI and SOI models after 2 weeks of tumor growth were 205 mm3 and 178 mm3 respectively. By USGI of human pancreatic cancer cell lines, human orthotopic pancreatic cancer xenografts were established. Based on ultrasound imaging, the orthotopic human pancreatic cancer xenograft take rate was 100% for both human pancreatic cancer cell lines used, MiaPaCa-2 and Su86.86, with mean tumor volumes of 28 mm3and 30 mm3. We demonstrated that this USGI method is feasible, reproducible, facile, minimally invasive and improved compared to the highly-invasive SOI method for establishing orthotopic pancreatic tumor xenograft models suitable for molecular imaging

Soluble Immune Complexes Shift the TLR-Induced Cytokine Production of Distinct Polarized Human Macrophage Subsets towards IL-10

Contains fulltext : 109563.pdf (publisher's version ) (Open Access)BACKGROUND: Costimulation of murine macrophages with immune complexes (ICs) and TLR ligands leads to alternative activation. Studies on human myeloid cells, however, indicate that ICs induce an increased pro-inflammatory cytokine production. This study aimed to clarify the effect of ICs on the pro- versus anti-inflammatory profile of human polarized macrophages. MATERIALS AND METHODS: Monocytes isolated from peripheral blood of healthy donors were polarized for four days with IFN-gamma, IL-4, IL-10, GM-CSF, M-CSF, or LPS, in the presence or absence of heat aggregated gamma-globulins (HAGGs). Phenotypic polarization markers were measured by flow cytometry. Polarized macrophages were stimulated with HAGGs or immobilized IgG alone or in combination with TLR ligands. TNF, IL-6, IL-10, IL-12, and IL-23 were measured by Luminex and/or RT-qPCR. RESULTS: HAGGs did not modulate the phenotypic polarization and the cytokine production of macrophages. However, HAGGs significantly altered the TLR-induced cytokine production of all polarized macrophage subsets, with the exception of MPhi(IL-4). In particular, HAGGs consistently enhanced the TLR-induced IL-10 production in both classically and alternatively polarized macrophages (M1 and M2). The effect of HAGGs on TNF and IL-6 production was less pronounced and depended on the polarization status, while IL-23p19 and IL-12p35 expression was not affected. In contrast with HAGGs, immobilized IgG induced a strong upregulation of not only IL-10, but also TNF and IL-6. CONCLUSION: HAGGs alone do not alter the phenotype and cytokine production of in vitro polarized human macrophages. In combination with TLR-ligands, however, HAGGs but not immobilized IgG shift the cytokine production of distinct macrophage subsets toward IL-10

Mean 3D tumor volume measurements (n = 5) from the USGI and SOI models of orthotopic pancreatic cancer xenografts growth over time.

<p>Mean 3D tumor volume measurements (n = 5) from the USGI and SOI models of orthotopic pancreatic cancer xenografts growth over time.</p

Mean 3D tumor volume measurements (n = 5) from the orthotopic human pancreatic cancer xenografts using pancreatic cancer cell lines, MiaPaca-2 and SU86.86, growth over time.

<p>Mean 3D tumor volume measurements (n = 5) from the orthotopic human pancreatic cancer xenografts using pancreatic cancer cell lines, MiaPaca-2 and SU86.86, growth over time.</p

Mean <i>ex vivo</i> fluorescence signal from mouse organs (pancreas, spleen, heart, lungs, liver, kidneys, and GI tract) obtained 24 h post-administration of tumor targeted fluorescent-probe and 2 weeks post SOI or USGI, n = 5.

<p>Mean <i>ex vivo</i> fluorescence signal from mouse organs (pancreas, spleen, heart, lungs, liver, kidneys, and GI tract) obtained 24 h post-administration of tumor targeted fluorescent-probe and 2 weeks post SOI or USGI, n = 5.</p

Comparison of representative images of SOI and USGI orthotopic pancreatic cancer xenografts models 2 weeks post-injection of HCT116/δOR+ cells.

<p>The blue shaded area represents the tumor generated by performing 3D tumor volume measurements. A) ultrasound image of USGI model, B) <i>in vivo</i> fluorescence image of USGI model, C) <i>ex vivo</i> fluorescence image of the mouse pancreas from the USGI model, D) ultrasound image of SOI model, E) <i>in vivo</i> fluorescence image of SOI model, and F) <i>ex vivo</i> fluorescence image of the mouse pancreas from the SOI model.</p

Representative ultrasound acquired images of orthotopic human pancreatic cancer xenografts 2 weeks after USGI of: A) MiaPaCa-2 cells B) SU86.86 cells.

<p>The blue shaded area represents the tumor generated by performing 3D tumor volume measurements.</p

Representative H&E staining of mouse pancreata containing xenografts of the following cell lines: A) HCT116/δOR+, B) MiaPaca-2 and C) SU86.86.

<p>The 20X magnification view shows normal pancreas (indicated by black arrows) adjacent to tumor cells (indicated by red arrows).</p