This protocol is for the performance of one-anastomosis gastric bypass (OAGB) on rat. The operator performs a long and tubular-stapled gastric pouch followed by hand-sewn anastomosis. For this model, the operator reproduces the same ratio between biliopancreatic and common limb in humans; therefore, the biliopancreatic limb measures 35 cm.
The goal of this protocol is to set up a preclinical model of bariatric surgery and, more specifically, OAGB in obese rats. Based on this preclinical model, longitudinal studies can be carried out to provide an improved understanding of the mechanisms underlying the outcomes seen after bariatric surgery in humans. For this purpose, rats are operated on through a laparotomy under general anesthesia with isoflurane. First, the surgeon creates a long and tubular gastric pouch: after greater curve and hiatal dissection, the nonglandular stomach is stapled and removed. Then, the remaining stomach is also stapled in order to create a gastric tube and exclude the antrum of the stomach. After that, the surgeon performs a single end-to-side gastrojejunostomy 35 cm from the duodenojejunal angle. This limb length has been chosen in order to reproduce the same ratio between the biliopancreatic limb (BPL) and common limb (CL) length as in human bariatric surgery. The operation ends by aponeurotic and cutaneous closure. The early postoperative management consists of subcutaneous hydration, an intramuscular prophylactic antibiotic injection, a parietal injection of xylocaine, the administration of painkillers, and a progressive reintroduction of diet.
Morbid obesity is a major health issue in the western world, with increasing prevalence1. Bariatric surgery remains the only long-term effective therapy for morbid obesity2, with clear benefits on obesity-related comorbidities3. Roux-en-Y gastric bypass (RYGB) remains the gold standard as it is one of the oldest procedures and one of the most-performed worldwide with the longest follow-up4. OAGB is a more recent procedure first described in 2001 by Rutledge5. It has been described as an effective alternative to RYGB6 and has some advantages. Early data indicates a similar efficacy to RYGB with respect to weight-loss and the resolution of obesity-related comorbidities6,7. Furthermore, the risks of stenosis or leakage of the jejunojejunal anastomosis in RYGB are avoided, and the risk of an internal hernia may be lower than with RYGB8.
The physiological mechanisms underlying weight-loss after bariatric surgery remain controversial and can be difficult to study in humans9. Animal models are useful to understand how bariatric surgeries may impact glucose metabolism, neuroendocrine modifications, or satiety mechanisms. Comparative data between procedures are lacking and long-term effects of these surgeries need to be better understood10. It is not possible to fully standardize pre- and postoperative clinical care, and satisfactory follow-up can be difficult to achieve: some studies report loss-to-follow-up rates as high as 30% for the first year11.
The concept of gastric bypass is a combination of a restrictive procedure, due to the creation of a gastric pouch, and a moderate degree of malabsorption, resulting from the exclusion of the duodenum and the proximal jejunum from the gastrointestinal transit. Currently, limb length in bariatric surgery is still a matter of debate12,13. Available data show a high variability of procedures and no consensus has yet been reached on the optimal limb length for OAGB14,15. Nonetheless, many authors have reported a 200 cm BPL length as the standard for OAGB16,17,18. The mean length of the small bowel is about 700 cm19.
Several rodent models of RYGB have been developed worldwide20, with great variations in terms of pouch size, limb length, and the preservation of the vagal nerve. But very few authors reported a model of experimental OAGB; for this reason, theresearch team developed a model of OAGB in collaboration with bariatric surgeons. The rationale behind the development of the model was to mimic gastric bypass practiced in humans. The model incorporates the construction of a gastric pouch with the exclusion of the antrum and the body of the stomach. For it, the surgeon makes an ablation of the nonglandular stomach using a stapling device. Compared to a previous protocol20, the team decided to add a per-operative prophylactic antibiotic treatment in order to diminish the morbidity linked to gastrointestinal anastomosis, as is routine in human surgery21.
The rat limb length used in this model was chosen to reproduce the ratio between the BPL and the CL in humans. Preliminary measurements have shown the total small bowel length in rats to be between 90 and 120 cm22. Consequently, the length of the BPL chosen for OAGB was 35 cm. Furthermore, this model is practicable without making any vascular ligation, avoiding gastric pouch ischemia.
All animal use conformed to the European Community guidelines and was approved by the local ethics committee (Comité d'éthique en expérimentation animale N° 121) and the Ministry of Higher Education and Research (Ref #02285.03).
1. Animals and Diets
2. Material
3. Surgery
4. Postoperative Care
This protocol is reproducible and mimics OAGB as practiced on humans. The safety of this protocol is supported with published data23,24 with a low mortality rate between 12% and 25%. Different surgeons in the research team with a similar mortality rate have practiced this protocol.
Nonetheless, learning rodent surgery is complex, and the use of magnifying glasses is mandatory. Indeed, the critical step of the construction of a microscopic anastomosis requires a learning period from around 15 to 20 operations to reach an optimal dexterity. A surgical defect can lead to an anastomotic leakage of a gastric obstruction, which is the main death cause in our experience. Another key to success is the postoperative nutrition; indeed, an early postoperative solid nutrition can be dangerous. This protocol also provides a precise postoperative nutrition protocol, which has been demonstrated to be safe. The concept of this surgery on humans and on rats is illustrated in Figure 1. A result of the operation after sacrifice is provided in Figure 2.
Results obtained on weight loss after operation on obese rats were quite similar to those observed in humans, with a body weight loss of about 20%, stable after the operation. The results are detailed in Figure 3. Better insulin sensitivity and glucose tolerance were observed as soon as 2 weeks after the surgery, as assayed by an oral glucose test (OGTT) and an insulin tolerance test (ITT). These results are detailed in Figure 4. HFD-induced obesity has been described to promote insulin resistance and glucose intolerance in Wistar rats25; comparable data have been gathered in our unit (unpublished). Wistar rats are not a model of type 2 diabetes.
Figure 1: Illustration of OAGB on humans and on this rat model. (A) Scheme of an OAGB in humans; the gastric pouch is long and tubular and the BPL measures between 150 and 200 cm. (B) Rat model of an OAGB. The ratio of the BPL and the AL/CL reproduce the one practiced for human surgery; the BPL measures 35 cm. Abbreviations: GP = gastric pouch; Ana = anastomosis; AL = alimentary limb; CL = common limb. Please click here to view a larger version of this figure.
Figure 2: Illustration of OAGB on rats after sacrifice. Abbreviations: GP = gastric pouch; Ana = anastomosis; AL = alimentary limb; CL = common limb; Oes = Esophagus; Duo = duodenum Please click here to view a larger version of this figure.
Figure 3: Postoperative weight evolution (median ± SD) after OAGB and sham surgery. The sham surgery consisted of a laparotomy and a pinch on the great gastric curve with non-traumatic forceps. Please click here to view a larger version of this figure.
Figure 4: Glucose metabolism after OAGB and sham surgery; post-operative OGTT and ITT. (A) Post-operative OGTT. OAGB and sham rats were made to fast before being subjected to an oral glucose tolerance test (OGTT: 1 g/kg of body weight). (B) Post-operative ITT: an injection of 1 U insulin/kg of body weight. Rats were made to fast 4 h before receiving the insulin injection. Blood was harvested in the tail vein at t = 0 and at 10, 20, 30, 60, and 120 min after the insulin or glucose administration. Glucose was measured using a blood glucose meter. Glycemia was expressed in mg/dL ± standard deviation. Abbreviations:OGTT = oral glucose tolerance test; ITT = insulin tolerance test; BW = body weight. * p < 0.05, ** p < 0.01, *** p < 0.001; after two-way ANOVA statistical test. The sham surgery consisted of a laparotomy with a nontraumatic pinch of the great gastric curve using non-traumatic forceps. All data are presented as mean ± SD. Please click here to view a larger version of this figure.
This protocol has the advantage of mimicking OAGB practiced routinely in humans. Its safety and reproducibility in rats have been demonstrated in clinical and metabolic results.
The anastomosis in this protocol poses a technical difficulty and is the main critical step. For this, it is necessary to use specific microsurgical tools, such as a microsurgery needle-holder or magnifying glasses. In order to realize a hermetic anastomosis without any stenosis, it is necessary to perform at least seven to eight close sutures on each side of the anastomosis and to avoid taking any mucosa in any stitch in order to bury it inside the suture. The effective management of pain and animal welfare, as well as progressive postoperative nutrition as described previously, is mandatory.
However, this protocol has some limitations. First, the surgical approach is a laparotomy instead of a laparoscopy, due to the small size of the animals. There is no division on the right side of the gastric pouch. This choice has been made for technical reasons. Nonetheless, no repermeabilization of the gastric pouch has been observed after 15 postoperative weeks. Furthermore, mimicking the restrictive effects on rats is very difficult while maintaining ad libitum access to food. Animals may increase the frequency of oral intake in order to counterbalance the restrictive effect of the gastric pouch22,24. Additionally, there is no gastric resection in human surgery, whereas the rat's nonglandular stomach is known to be significantly expandable if not resected.
No protocol of OAGB in rodents mimicking the human surgery closely has ever been published by other teams. This protocol might offer a good balance between surgical feasibility, safety, and clinical relevance.
This method may also be used in the future to highlight physiological changes observed after RYGB and OAGB in humans, such as the theoretical increased risk of malignancy and the higher malabsorptive effect of OAGB, compared to RYGB.
The authors have nothing to disclose.
The authors would like to thank Jean Pierre Marmuse for his constant help and support; Eglantine Voitellier, Jean-Baptiste Cavin, and Houneyda El Jindi for setting up the models and the illustrations; Anne-Charlotte Jarry and Andreia Goncalves for their help with the videos; Sara J. Zaat for editing the text in English. Matthieu A. Siebert and Lara Ribeiro-Parenti were recipients of sponsorships from the Fondation pour la recherché médicale (FRM) and the Association française de chirurgie (AFC).
High-Fat Diet, 45 % fat | Genestil, Royaucourt, France | C 1090-45 | http://www.genestil.com/ |
Metzenbaum Scissors | World Precision instrument, Sarasota, US | 501739 | https://www.wpi-europe.com |
Dumont Tweezers | World Precision instrument, Sarasota, US | 14098 | https://www.wpi-europe.com |
Iris Forceps | World Precision instrument, Sarasota, US | 15915 | https://www.wpi-europe.com |
Iris Scissors 10cm | World Precision instrument, Sarasota, US | 14218 | https://www.wpi-europe.com |
Needle Holder | World Precision instrument, Sarasota, US | 14110 | https://www.wpi-europe.com |
Alm Self Retaining Retractor | World Precision instrument, Sarasota, US | 14240 | https://www.wpi-europe.com |
Baby Mixter Hemostatic Forceps, Right Angle | World Precision instrument, Sarasota, US | 501240 | https://www.wpi-europe.com |
Non woven sterile swabs | LCH Medical Products, Paris, France | SN40-0755 | www.lch-medical.com |
Galilean Binocular Loupe 2.5x | World Precision instrument, Sarasota, US | 504056 | https://www.wpi-europe.com |
Digestive thread: Isotactic polypropylene, monofil Prolene 7-0 | Ethicon, Issy les Moulineaux, France | EH7813E | www.ethicon.com |
Parietal thread : Coated Braided Polyester Ti-Cron 2-0 | Covidien, Mannsfield, MA, USA | 3003-52 | www.covidien.com |
Cutaneous thread : Polyglactin, VICRYL RAPIDE 4-0 | Ethicon, Issy les Moulineaux, France | VR3100 | www.ethicon.com |
ETS-Flex 35- mm staple gun | Ethicon, Issy les Moulineaux, France | ATS45 | www.ethicon.com |
Proximate Reloadable Staplers | Ethicon, Issy les Moulineaux, France | XR30V | www.ethicon.com |
TA-DST 30mm-3.5mm | Ethicon, Issy les Moulineaux, France | TX30B | www.ethicon.com |
Alcoholic 5% Betadine | MEDA Pharma, Merignac, France | 41085 | www.medapharma.fr |
LIDOCAINE AGUETTANT 20 mg/ml | LABORATOIRE AGUETTANT, Lyon, France | 3400940000000.0 | https://www.aguettant.fr |
Penicillin G 5MUI | Panpharma, Luitre, France | 3.40E+12 | https://www.panpharma.eu/fr |
Bionolyte G5 ; Sodium chloride 0.4%, glucose 5%, potassium chloride 0.2%) | Baxter, Maurepas, France | ||
Liquid diet | Genestil, Royaucourt, France | C-0200T | http://www.genestil.com/ |
Isoflurane 100% | Centravet, Plancoet, France | ISO007 | http://cooperative.centravet.net |