The following describes the performance of vertical sleeve gastrectomy in mice. This is a type of weight-loss surgery that involves removal of approximately 70% of the stomach.
Bariatric surgery, such as vertical sleeve gastrectomy (VSG), is a surgery of the gastrointestinal tract that is performed for the purpose of weight loss. Bariatric surgery is currently the most effective long-term treatment for obesity. In addition to weight loss, bariatric surgery produces additional health benefits such as remission of type 2 diabetes, remission of hypertension, and decreased risk of developing certain types of cancer. The mechanisms beyond weight loss for these benefits remain incompletely defined. Therefore, animal models of bariatric surgery are being developed and validated to identify the mechanisms leading to these benefits, with the goal of improving understanding of gastrointestinal physiology and identifying new therapeutic targets. VSG has become the most commonly performed bariatric procedure in the clinic in the United States because it is highly effective at producing weight loss and metabolic improvement, and is simpler to perform than other bariatric procedures. Therefore, we have developed and validated a murine model of VSG. This murine VSG model recapitulates many of the effects of VSG seen in humans, including improved glucose and blood pressure regulation. The method is based on isolation of the stomach, ligation of gastric vessels, and removal of 70% of the stomach by transecting along the greater curvature of the stomach. We have successfully applied this surgical protocol to various genetically modified mouse lines to define the mechanistic contributors to the benefits of VSG. Furthermore, this murine VSG model has been combined with other surgical techniques, to achieve deeper mechanistic insight. Therefore, this is a simple and versatile model for studying gastrointestinal physiology and the health benefits of bariatric surgery.
As the obesity epidemic continues to grow worldwide bariatric surgery has gained popularity as it is the most effective long-term treatment for obesity1. Unfortunately, weight loss by diet and exercise is difficult to achieve and relatively ineffective over the long-term2,3. Bariatric surgery, such as vertical sleeve gastrectomy (VSG), is defined as the manipulation of the gastrointestinal tract for the purpose of weight loss1,4. Although weight loss is a prominent outcome of bariatric surgery, bariatric surgery provides other health benefits such as improving obesity comorbidities and extending lifespan5. For example, bariatric surgery results in high rates of remission of type 2 diabetes and hypertension, and reductions in the lifetime risk of the development of certain types of cancer1,6,7. Of note, the effect of bariatric surgery causing remission of type 2 diabetes and hypertension is often observed soon after surgery and prior to weight loss8,9. This highlights the concept that there are mechanisms independent of body weight contributing to the health benefits observed after surgery. Animal models of bariatric surgery have been developed and are utilized to study the mechanisms by which these health benefits occur10,11,12.
We have validated a mouse model of VSG, which we have applied to various genetically modified mouse models to study the mechanisms by which bariatric surgery improves obesity comorbidities such as type 2 diabetes, hypertension, and colorectal cancer10,11,12. Rodent models allow more experimental control and the ability to perform genetic or pharmaceutical manipulation to define the role of specific genes or signaling pathways of interest. We are focusing primarily on VSG because VSG is the most commonly performed bariatric procedure in the clinic in the United States13. Additionally, VSG is a simple surgical model with fewer anatomic modifications compared to other procedures such as Roux-en-Y gastric bypass or biliopancreatic diversion.
Our mouse model of VSG recapitulates the following effects of bariatric surgery observed in humans: weight loss, reduced food intake, improved glucose regulation, improved islet function, increased post-prandial glucagon-like peptide-1 (GLP-1) secretion, reduced arterial blood pressure, and increased circulating bile acid concentrations10,11,12,13,14,15. Therefore, this is an ideal model to study the body weight dependent and independent mechanisms by which VSG improves or resolves obesity comorbidities. In addition, it is a reliable model that can be combined with other surgical procedures, allowing for the investigation of the impact of VSG under various disease conditions with greater mechanistic insight12.
All experimental protocols have been approved by the Cornell University Institutional Animal Care and Use Committee.
1. Pre-surgical Preparation
NOTE: Study mice are typically on a C57BL diet-induced obese background to make studies translationally relevant to human obesity and insulin resistance. Male and female mice may be studied as described in the subsequent steps.
2. Vertical Sleeve Gastrectomy and Sham Procedures
3. Post-operative Mouse Care and Measurements
The sham and VSG procedures are depicted in Figure 1. Figure 1A shows where the suture line is placed along the gastric walls during the sham procedure. This same area is where the stomach is cut during VSG surgery. Figure 1B shows the tubular remnant of stomach left after performance of VSG.
Statistics and Data Analysis
Data are presented as mean ± SEM. Data were analyzed by ANOVA with Tukey's post-test or by Student's t-test as indicated. Differences were considered significant at P<0.05.
VSG decreases energy intake and body weight and improves glucose tolerance
High fat diet fed male C57BL mice were operated on and studied as described above. VSG-operated mice exhibited reduced energy intake and body weight compared to ad libitum fed sham-operated mice (Cumulative energy intake: Sham = 474 ± 19, VSG = 385 ± 14 kcal; Final body weight: Sham = 34.5 ± 2.1, VSG = 29.9 ± 1 g; Figure 2A-B, *P<0.05). Three weeks after surgery an oral glucose tolerance test was performed. Glucose measurements were made using a glucometer (see Table of Materials). Serum insulin and GLP-1 concentrations were measured by multiplex sandwich electrochemiluminescence immunoassay (see Table of Materials). VSG improved glucose tolerance (Glucose AUC0-120: Sham =1,467 ± 76, VSG = 1,061 ± 72 mmol/L x 120 minutes; Figure 3A-B, P <0.01), increased glucose-stimulated insulin secretion (Percent increase in insulin from baseline to 15 minutes after the glucose gavage: Sham = 92 ± 54, VSG = 2,720 ± 1,241%; Figure 3C, P <0.05), and increased post-prandial GLP-1 secretion compared with sham-operated ad libitum fed control mice (GLP-1 AUC0-120: Sham = -124 ± 45, VSG = 111 ± 48 pmol/L x 120 minutes; Figure 3D, *P <0.01). These results are consistent with what is seen in humans and other rodent models after bariatric surgery9,17,18,19.
Figure 1: Diagram of VSG. Depiction of line of transection during VSG. (A) This same area is where a simple continuous line of suture is placed along the gastric walls during the sham procedure. (B) Depiction of the tubular remnant left after completion of the VSG procedure. Please click here to view a larger version of this figure.
Figure 2: VSG lowers energy intake and body weight. Cumulative energy intake (A) and body weight (B). **P <0.05 compared with Sham by Student's t-test and *P <0.05 Sham compared with VSG by two-factor ANOVA. Results shown as mean ±SEM. n = 6-7 per group. Please click here to view a larger version of this figure.
Figure 3: VSG improves glucose tolerance, increases glucose-stimulated insulin secretion and increases post-prandial GLP-1 secretion during an OGTT. (A) Blood glucose concentration, (B) glucose area under the curve (AUC), (C) percent increase in serum insulin concentrations from baseline to 15 minutes after glucose gavage, and (D) serum total GLP-1 concentrations during an OGTT. *P <0.05, **P <0.01, ***P <0.001 VSG compared with Sham by Student's t-test of the AUC or percent change in insulin. Results shown as mean ± SEM. n = 6-7 per group. Please click here to view a larger version of this figure.
Bariatric surgery is the most effective long-term treatment for obesity and results in other health benefits such as high rates of type 2 diabetes and hypertension remission1,9,15. Murine models of bariatric surgery provide a powerful tool with which to identify the mechanisms by which bariatric surgery causes rapid and pronounced improvements in obesity comorbidities. Furthermore, murine models of bariatric surgery provide a novel paradigm for studying the basic biology by which the gut regulates various processes, such as metabolism, cardiovascular function, and carcinogenesis10,11,12.
We have developed a mouse model of vertical sleeve gastrectomy to study the mechanisms by which bariatric surgery improves obesity comorbidities such as type 2 diabetes and hypertension10,11,12. This model of VSG consists of ligation of stomach vessels followed by removal of 70% of the stomach. The control condition is a sham surgery in which suture is placed along the gastric wall in the same location where the stomach would be transected in a VSG procedure. Herein, we present data where sham-operated mice were fed ad libitum after surgery. In order to investigate body weight-independent effects, a sham-operated control group that is food restricted in order to match their body weight to VSG-operated mice should be studied, as we have previously described11,12. Our data demonstrate that this VSG model achieves both reductions in body weight and food intake compared to ad libitum fed sham-operated controls. Oral glucose tolerance tests are done at least two weeks after surgery to allow animals enough time to fully recover from surgery, so that surgical recovery is not a confounding factor. Consistent with the decrease in body weight, the presented model exhibits improved glucose tolerance. Similar to what is observed in human patients after bariatric surgery, VSG-operated mice exhibit remarkable increases in glucose-stimulated insulin secretion and postprandial GLP-1 secretion14,17.
Lab members are thoroughly trained on sham surgery and VSG and practice for several months to master this technique and achieve a survival rate of greater than 95%; however, this will vary based on prior experience. Prior to initiating a full study, it is recommended that a new surgeon validate his/her VSG and sham surgeries by measuring body weight, food intake, and glucose tolerance in practice sham and VSG-operated mice to ensure that the appropriate phenotype is being achieved. Furthermore, it is important to closely monitor mice after surgery for signs of surgical failure. The most common post-operative complication is leakage of stomach contents. Post-operative signs of this complication include lack of food consumption, lack of fecal production, and a palpable upper abdominal mass. Animals that exhibit signs of surgical complication should be promptly euthanized. If post-operative complications are a reoccurring problem, a reassessment of surgical technique must be performed. The most common problem is inadequate closure of the gastric wall leading to leakage of gastric contents. Closure of the gastric wall is the most important step in the protocol. Once suture placement is complete, there should be no visible gaps between knots. It is advised that CTAs are used to verify that there is no leakage of stomach contents prior to closing the abdomen. Another less common issue is induction of gastrointestinal stasis by inadvertent manipulation of the small intestine when initially removing the stomach from the abdominal cavity. Care must be taken to not manipulate the small intestines because the mouse small intestine is very fragile and excessive manipulation of the small intestine can lead to stasis.
Our mouse model of VSG has similar surgical outcomes to that of other groups and survival rates greater than 95%. Similar to human patients and other mouse models of VSG, the presented mouse VSG models exhibited weight loss, reduced food intake, improved glucose tolerance, and increased post-prandial GLP-1 secretion4,18,19. This sham procedure differs from other models as it involves placement of suture while other models only apply pressure to the stomach with blunt forceps19,20. Placing sutures allows control of the effect of placing foreign material in the stomach while also providing surgical manipulation of the gastric tissue. Like any mouse model, the limitations of this technique are that the translation of data generated from this mouse model to humans is limited by species differences. However, mice allow for greater control and experimental manipulation, providing deeper mechanistic insight than can be achieved in human patients. Although the majority of murine bariatric studies have primarily focused on the glucoregulatory benefits of these procedures, we use the presented murine VSG model to define the mechanisms driving the impact of bariatric surgery on hypertension and cancer12. This highlights important future applications of this model.
Murine models of bariatric surgery provide an important tool to study the mechanisms by which bariatric surgery produces weight loss and confers health benefits. Furthermore, bariatric models provide a novel paradigm with which to study how the gut interfaces with other physiologic processes in the body. Described here is a model of VSG which has been previously validated and recapitulates many of the effects seen in humans after VSG10,11,12. Furthermore, this is a versatile model that can be successfully combined with other surgical procedures for assessment of different disease processes and/or more detailed mechanistic assessment12.
The authors have nothing to disclose.
This research was supported by NIH/NCI R21CA195002-01A1, The President's Council of Cornell Women and the SUNY Graduate Diversity Fellowship. Dr. Cummings' laboratory also received funding during the project period from the Cornell Comparative Cancer Biology Training Program and Eli Lilly and Company.
45% high fat diet | Research Diets | D12451 | |
60% high fat diet | Research Diets | D12492 | |
Boost | Nestle | 160-67538 | rich chocolate flavor |
6-0 Suture | Ethicon | Z432 | monofilament absorbable/taper |
7-0 Suture | Covidien | 8866127-01 | monofilament absorbable/taper |
Cotton Swabs | Fisherbrand | 23-400-118 | small |
Cotton Swabs | Fisherbrand | 233-400-101 | large |
Gauze | Various | 4×4 4 ply and 2×2 4 ply | |
Foil | Various | ||
Surgery drape | Various | ||
0.9% saline solution | Various | ||
LRS | Hospira | 170RX | |
Betadine | Various | ||
Alcohol | Various | ||
Eye Ointment | Paralube® Vet Ointment | 17033-211-38 | |
Tissue Adhesive | Vetbond | 1469SB | |
Meloxicam (Metacam) | Boehringer Ingelheim | 141-213 | 5 mg/ml |
Enrofloxacin | Baytril | 08713254-186599 | 22.7 mg/ml |
Thin tipped hemostats | Fine Science Tools | 13021-12 | |
Metzenbaum Scissor | Fine Science Tools | 14018-18 | |
Iris Scissors | Fine Science Tools | 14058-09 | |
Dumont Forcep | Fine Science Tools | 11251-20 | |
Serrated Forcep | Fine Science Tools | 11020-12 | |
Gavage needle | Fine Science Tools | 18060-20 | |
Microneedle driver | Fine Science Tools | 12075-14 | |
Spring Scissor | Fine Science Tools | 15396-00 | |
Insulin syringe | Various | ||
1mL syringe | Various | ||
20mL syringe | Various | ||
Glucometer (one touch ultra mini) | Lifescan | 70021208 | |
Multiplex insulin and GLP-1 kit | Meso Scale Discovery | K15171C-1 | |
GraphPad Prism 6.00 | GraphPad Software | ||
Nestlets | Ancare | NES3600 |