Ортотопическая Модель серозного рака яичников у иммунокомпетентных мышей для В естественных условиях Опухоль изображений и мониторинг иммунного ответа опухоли
Summary
Для изучения<em> В естественных условиях</em> Рост опухоли и микроокружения, мы использовали сингенных и ортотопической мышиной модели рака яичников у иммунокомпетентных животных. Мы трансдуцированных клетка опухоли мышей линии (MOV1) с Katushka флуоресцентный белок (MOV1<sup> KAT</sup>), И здесь мы показываем свою ортотопической имплантации в яичнике и<em> В естественных условиях</em> Изображений.
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
Хирургии и Ортотопическая инъекций
Ортотопическая инъекций в яичниках сумки требует подготовки и точности. Таким образом
- В случае плохой хирургический опыт, практика с трупов в первую очередь.
- Использование преимущественно повторнородящих женщин (один или…
Divulgazioni
The authors have nothing to disclose.
Acknowledgements
Эта работа была поддержана NIH грант P01 А.И. 068730 (SNC, Н. С.), грант NIH CA016520 / TAPITMAT (NS), частное финансирование из Фонда Claneil (NS) и яичников SPORE предоставить РКИК и Университета Пенсильвании ( P50 CA83638) и Fox Chase Cancer Center Основные Грант (P30 CA06927) (ДКК). Авторы выражают благодарность отличном техническом содействии оптический / Биолюминесценция Основные фонда направлено доктором Э. Delikatny в Университете Пенсильвании, Энтони Secreto из стволовых клеток и ксенотрансплантата Основные режиссера д-р Г. Дане-Денуайе в Университете Пенсильвании рака Центр подготовки к SNC ортотопической технике инъекций и Denada Dangaj в Университете штата Пенсильвания / ОЦРК для оказания помощи по хирургии.
Materials
| Material Name | Tipo | 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 |
Riferimenti
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