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Preparing a Mice Model of Severe Acute Pancreatitis via a Combination of Caerulein and Lipopolysaccharide Intraperitoneal Injection
* These authors contributed equally
In This Article
Summary
Intraperitoneal drug administration is a safe and effective non-invasive approach for inducing pancreatic injury. This study compared five distinct intraperitoneal injection protocols on mice to induce varying degrees of pancreatic injury and established a model of severe pancreatic injury to investigate the pathological changes and treatment strategies for severe acute pancreatitis (SAP).
Abstract
The treatment of severe acute pancreatitis (SAP), with high mortality rates, poses a significant clinical challenge. Investigating the pathological changes associated with SAP using animal models can aid in identifying potential therapeutic targets and exploring novel treatment approaches. Previous studies primarily induced pancreatic injury through retrograde bile duct injection of sodium taviaurocholate, but the impact of surgical damage on the quality of the animal model remains unclear. In this study, we employed various frequencies of intraperitoneal Caerulein injections combined with different doses of LPS to induce pancreatic injury in C57BL/6J mice and compared the extent of injury across five intraperitoneal injection protocols. Regarding inducing acute pancreatitis in mice, an intraperitoneal injection protocol is proposed that results in a mortality rate as high as 80% within 5 days. Specifically, mice received ten daily intraperitoneal injections of Caerulein (50 µg/kg), followed by an injection of LPS (15 mg/kg) one hour after the last Caerulein administration. By adjusting the frequency and dosage of injected medications, one can manipulate the severity of pancreatic injury effectively. This model exhibits strong controllability and has a short replication cycle, making it feasible for completion by a single researcher without requiring expensive equipment. It conveniently and accurately simulates key disease characteristics observed in human SAP while demonstrating a high degree of reproducibility.
Introduction
Severe acute pancreatitis is characterized by rapid onset, rapid progression, and high mortality rates within the digestive system disease domain1. Its high fatality rate has always been a prominent focus of clinical research. Due to unpredictable changes in clinical conditions, heterogeneity of disease manifestations, and limited availability of human specimens, establishing animal models has become increasingly crucial for disease research.
Retrograde injection of sodium taurocholate into the common bile duct is commonly used to create a rat model of SAP2. By simulating pancreaticobiliary ob....
Protocol
This protocol was reviewed and approved by the Ethics Committee at The First Affiliated Hospital of Anhui University of Science and Technology (Huainan, China) (Ethics Code: 2023-KY-905-001). The study followed the National Institutes of Health guidelines for the care and use of research rodents in all animal procedures. C57BL/6J adult mice weighing 20-30 g were used for the present study. The mice were housed in an animal laboratory for one week under controlled conditions (approximately 21 °C with a 12 h alternati.......
Representative Results
The process of experimental mouse modeling is illustrated in Figure 1. After 12 h of injection completion, an open-field video recorder was used to monitor the movement distance and immobility duration of different experimental groups of mice for 5 cycles (Figure 2A). During the 5 cycles, mice in the PI V group maintained a low level of movement distance within 3 min, while the immobility ratio within 3 min increased with each subsequent cycle (.......
Discussion
Currently, there is a lack of effective means to improve the high mortality rate in patients with severe acute pancreatitis. It is crucial to investigate the efficacy of drugs in enhancing immune stability mechanisms. An urgent need exists for an ideal animal model for severe acute pancreatitis. Mice with a C57BL/6J genetic background are widely used in biomedical research, including studies on SAP pathophysiology. Over 70 years of genetic differentiation in B6J mice have resulted in the spontaneous deletion of several e.......
Acknowledgements
This study was supported by Research Projects in Health and Medical Science in Huainan City (No. HNWJ2023005); Municipal Guiding Science and Technology Plan Program in Huainan City (No.2023151); Anhui Provincial College Students' Innovation and Entrepreneurship Training Program (No. S202310361254); the ninth batch of the "50·Stars of Science and Technology" innovation teams in Huainan City and Anhui Provincial Key Clinical Specialty Construction Project. We would like to express our gratitude to the Laboratory Department of the First Affiliated Hospital of Anhui University of Science and Technology for providing the relevant testing data.
....Materials
| Name | Company | Catalog Number | Comments |
| 20× Citric Acid Antigen Repair Solution (pH 6.0) | Wuhan servicebio Technology Co.,Ltd, China | G1202-250 ml | |
| Amylase | Mindray,China | ||
| Annexin V-FITC/PI | Wuhan servicebio Technology Co.,Ltd, China | G1511 | diluted at 1:20 |
| Anti-HMGB1 Rabbit pAB | Wuhan servicebio Technology Co.,Ltd, China | GB11103 | diluted at 1:1800 |
| BCA protein quantitative detection kit | Wuhan servicebio Technology Co.,Ltd, China | G2026-200T | |
| BD FACSCanto II Flow Cytometer | BD Life Sciences, San Jose, CA, 95131, USA | BD FACSCanto II | |
| BSA | Wuhan servicebio Technology Co.,Ltd, China | GC305010-100g | |
| C57BL/6J | Cavion Experimental Animal Co., Changzhou, China | license number SCXY (Su) 2011–0003 | |
| Ceruletide | MCE, New Jersey, USA | 17650-98-5 | 50 µg/kg |
| Chemiluminescence imager | Cytiva CO.,LTD.;USA | ||
| Citric acid antigen repair Solution (Dry powder pH 6.0) | Wuhan servicebio Technology Co.,Ltd, China | G1201-5 L | |
| Collagenase IV | Wuhan servicebio Technology Co.,Ltd, China | GC305014 | 0.5 mg/mL |
| DAB (SA-HRP) Tunel Cell Apoptosis Detection Kit | Wuhan servicebio Technology Co.,Ltd, China | G1507-100 T | |
| Dimension EXL with LM Integrated Chemistry System | Siemens Healthcare Diagnostics Inc.Brookfield,USA | YZB/USA 8311-2014 | |
| ECL developer | Wuhan servicebio Technology Co.,Ltd, China | ||
| Eosin dye (alcohol soluble) | Wuhan servicebio Technology Co.,Ltd, China | G1001-100 ml | |
| EthoVision XT | Noldus, Netherlands | ||
| FITC-labeled goat anti-rabbit IgG | Wuhan servicebio Technology Co.,Ltd, China | GB22303 | diluted at 1:50 |
| Fully automatic blood cell analyzer | Zybio Inc. China | Zybio-Z3 CRP | |
| GapDH | Wuhan servicebio Technology Co.,Ltd, China | GB11103 | diluted at 1:1500 |
| Hematoxylin blue return solution | Wuhan servicebio Technology Co.,Ltd, China | G1040-500 ml | |
| Hematoxylin differentiation solution | Wuhan servicebio Technology Co.,Ltd, China | G1039-500 ml | |
| Hematoxylin dye | Wuhan servicebio Technology Co.,Ltd, China | G1004-100 ml | |
| HMGB-1 ELISA kits | njjcbio Co., Ltd, China | ||
| HOMOGENIZER | Wuhan servicebio Technology Co.,Ltd, China | KZ-III-F;IC111150 100222 | |
| HRP-labeled goat anti-rabbit IgG | Wuhan servicebio Technology Co.,Ltd, China | GB23303 | diluted at 1:1500 |
| IL-6 ELISA kits | Wuhan servicebio Technology Co.,Ltd, China | GEM0001 | |
| Lipase | Mindray,China | ||
| Lipopolysaccharide | Wuhan servicebio Technology Co.,Ltd, China | GC205009 | 15 mg/kg |
| Low temperature high speed centrifuge | Changsha Pingfan Apparatus&Instrument Co.,Ltd.,China | TGL-20M | |
| Membrane breaking liquid | Wuhan servicebio Technology Co.,Ltd, China | G1204 | |
| microtome | Jinhua Craftek Instrument Co., Ltd.;China | CR-601ST | |
| Nylon mesh | Wuhan servicebio Technology Co.,Ltd, China | 200-mesh | |
| One-step TUNEL cell apoptosis detection kit (DAB staining method) | Wuhan servicebio Technology Co.,Ltd, China | G1507-100T | |
| Paraffin tissue embedding machine | PRECISION MEDICAL INSTRUMENTS CO.,LTD;Changzhou,China | PBM-A | |
| Pathological tissue drying apparatus | PRECISION MEDICAL INSTRUMENTS CO.,LTD;Changzhou,China | PHY-III | |
| Phosphate-buffered saline | Wuhan servicebio Technology Co.,Ltd, China | G4202-100ML | |
| PMSF | Wuhan servicebio Technology Co.,Ltd, China | G2008-1 ml | |
| Positive fluorescence microscope | Olympus Corporation,Tokyo, Japan | BX53 | |
| Pro Calcitonin | Mindray,China | ||
| PVDF membrane | Millipore, USA | 0.22 µm | |
| RIPA | Wuhan servicebio Technology Co.,Ltd, China | G2002-100 ml | |
| SDS-PAGE | Beyotime Biotechnology,China | P0012A | |
| TNF-αELISA kits | Wuhan servicebio Technology Co.,Ltd, China | GEM0004 | |
| Ultrasonic water bath | DONGGUAN KQAO ULTRASONIC EQUIPMENT CO.,LTD.;China | KQ-200KDE | |
| Western Blot | Bio-Rad Laboratories, Inc.,USA | ||
| Western blot imaging System | Global Life Sciences IP Holdco LLC, JAPAN | Amersham ImageQuant 800 | |
| Whirlpool mixer | SCILOGEX;USA |
References
- Gliem, N., Ammer-Herrmenau, C., Ellenrieder, V., Neesse, A. Management of severe acute pancreatitis: An update. Digestion. 102 (4), 503-507 (2021).
- Duan, F., et al.
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