असुरक्षित चूहे में एक तरल डिम्बग्रंथि के कैंसर के Orthotopic के लिए मॉडल Vivo में ट्यूमर और ट्यूमर प्रतिरक्षा प्रतिक्रिया के इमेजिंग निगरानी
Summary
अध्ययन करने के लिए<em> Vivo में</em> ट्यूमर के विकास और ट्यूमर microenvironment, हम एक syngeneic और orthotopic असुरक्षित पशुओं में डिम्बग्रंथि के कैंसर के माउस मॉडल का इस्तेमाल किया. हम Katushka फ्लोरोसेंट प्रोटीन (MOV1 के साथ एक माउस ट्यूमर कोशिका लाइन (MOV1) transduced<sup> कैट</sup>) और यहाँ हम अपने अंडाशय में orthotopic आरोपण दिखाने के और<em> Vivo में</em> इमेजिंग.
Abstract
Background: Ovarian cancer is generally diagnosed at an advanced stage where the case/fatality ratio is high and thus remains the most lethal of all gynecologic malignancies among US women 1,2,3. Serous tumors are the most widespread forms of ovarian cancer and 4,5 the Tg-MISIIR-TAg transgenic represents the only mouse model that spontaneously develops this type of tumors. Tg-MISIIR-TAg mice express SV40 transforming region under control of the Mullerian Inhibitory Substance type II Receptor (MISIIR) gene promoter 6. Additional transgenic lines have been identified that express the SV40 TAg transgene, but do not develop ovarian tumors. Non-tumor prone mice exhibit typical lifespan for C57Bl/6 mice and are fertile. These mice can be used as syngeneic allograft recipients for tumor cells isolated from Tg-MISIIR-TAg-DR26 mice.
Objective: Although tumor imaging is possible 7, early detection of deep tumors is challenging in small living animals. To enable preclinical studies in an immunologically intact animal model for serous ovarian cancer, we describe a syngeneic mouse model for this type of ovarian cancer that permits in vivo imaging, studies of the tumor microenvironment and tumor immune responses.
Methods: We first derived a TAg+ mouse cancer cell line (MOV1) from a spontaneous ovarian tumor harvested in a 26 week-old DR26 Tg-MISIIR-TAg female. Then, we stably transduced MOV1 cells with TurboFP635 Lentivirus mammalian vector that encodes Katushka, a far-red mutant of the red fluorescent protein from sea anemone Entacmaea quadricolor with excitation/emission maxima at 588/635 nm 8,9,10. We orthotopically implanted MOV1Kat in the ovary 11,12,13,14 of non-tumor prone Tg-MISIIR-TAg female mice. Tumor progression was followed by in vivo optical imaging and tumor microenvironment was analyzed by immunohistochemistry.
Results: Orthotopically implanted MOV1Kat cells developed serous ovarian tumors. MOV1Kat tumors could be visualized by in vivo imaging up to three weeks after implantation (fig. 1) and were infiltrated with leukocytes, as observed in human ovarian cancers 15 (fig. 2).
Conclusions: We describe an orthotopic model of ovarian cancer suitable for in vivo imaging of early tumors due to the high pH-stability and photostability of Katushka in deep tissues. We propose the use of this novel syngeneic model of serous ovarian cancer for in vivo imaging studies and monitoring of tumor immune responses and immunotherapies.
Protocol
Discussion
सर्जरी और Orthotopic इंजेक्शन
डिम्बग्रंथि बर्सा में Orthotopic इंजेक्शन प्रशिक्षण और परिशुद्धता की मांग. इस प्रकार
- पहली शवों के साथ गरीब शल्य अनुभव व्यवहार के मामले में.
- Preferentially multiparous महिलाओं का उप?…
Disclosures
The authors have nothing to disclose.
Acknowledgements
इस काम NIH अनुदान P01 ऐ 068730 (एस एन सी, एन एस), NIH अनुदान CA016520 / TAPITMAT (एन एस), Claneil फाउंडेशन (एन एस) से निजी धन, और डिम्बग्रंथि बीजाणु FCCC और पेन्सिलवेनिया विश्वविद्यालय के लिए अनुदान द्वारा समर्थित किया गया था ( CA83638 P50) और फॉक्स चेज़ कैंसर केंद्र कोर अनुदान (p30 CA06927) (डीसीसी). लेखकों धन्यवाद ऑप्टिकल / bioluminescence कोर द्वारा निर्देशित स्टेम सेल और xenograft कोर कैंसर पेंसिल्वेनिया के विश्वविद्यालय में डा. जी Danet – Desnoyers द्वारा निर्देशित से पेंसिल्वेनिया, एंथोनी Secreto के विश्वविद्यालय में डॉ. EJ Delikatny सुविधा के उत्कृष्ट तकनीकी सहायता प्रशिक्षण orthotopic इंजेक्शन तकनीक और सर्जरी पर सहायता के लिए Denada Dangaj / पेंसिल्वेनिया OCRC के विश्वविद्यालय में दक्षिणी नौसेना कमान के लिए केंद्र.
Materials
| Material Name | Type | Company | Catalogue Number | Comment |
|---|---|---|---|---|
| DMEM-GLUTAMAX | Invitrogen | 10564-011 | ||
| PBS | Gibco | 14040 | ||
| Versene | Lonza | 17-711E | ||
| Heating pad | Deltaphase | 39 DP | ||
| Povidone pads | Dynarex | 1108 | ||
| Alcohol pads | Fisher | 06-669-62 | ||
| Artificial tears ointment | Phoenix Pharma., Inc. | 17845-153 | ||
| Ketoprofen | Fort Dodge laboratories | |||
| 3cc/insulin syringe | BD | 309301 | ||
| Polyg Polyglycolic Acid suture/needle (3/8 19mm) | Syneture | 9612-31 | ||
| Tissue adhesive | Vetbond | 3M | ||
| Vet Bactrim/ oral suspension | Hi-tech Pharmacal | 840823 | ||
| IVIS-Lumina | Caliper lifesciences | |||
| Isofluorane | Phoenix Pharma., Inc. | J108013 | ||
| Fetal Bovine Serum, Qualified | Invitrogen | 10437036 | ||
| Penicillin/streptomycin | Gibco | 15140 | ||
| TurboFP635 mammalian vector | Evrogen | FP721 | ||
| T175 flasks | cellstar | 660-190 |
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