We started our experiments by testing different surgical instruments (ranging from 2 to 5 mm), different endoscopic cameras (ranging from 2.7 to 5 mm) and different operation settings for the laparoscopic procedures in rats. The use of 5-mm instruments turned out to be uncomfortable since the instruments were rather long and the required trocars were large and heavy. Slight movements of the instruments easily led to a dislocation of the trocars even when they were fixed with stay sutures and the barycentre was not on the same level as the abdominal wall. Also the graspers and scissors were too large for them to be comfortably handled intracorporeally. The operation was difficult to perform and appeared to be unsafe. The 2- and 3-mm endoscopic instruments were shorter, making them easier to handle intracorporeally and therefore they better fitted our needs while simultaneously keeping the procedures safe. One drawback we experienced were difficulties cutting the endoloop or caecum using the 2-mm endo-scissors. This problem was solved when we changed to 3-mm scissors. The cameras tested ranged in size from 5 mm, which provided a very good view but also a large access trauma, to 2 mm, providing a very poor view but less than half of the access trauma. An optimal compromise between both was provided by a 2.7-mm camera.
Finally, we were able to perform safely complete intracorporeally laparoscopic caecum resections. These were most effectively achieved using a 2.7-mm 30° Hopkins® optic (Karl Storz GmbH & Co KG) in combination with 2- and 3-mm endoscopic instruments (Karl Storz GmbH & Co KG) and a 5-mm LigaSure™ device (Covidien Deutschland GmbH).
Operation time and surgical trauma
The median operation times differed among the three groups. The longest operation time was for Group 2 (median: 39.5 min; range: 35 to 48 min) while the shortest operations were performed in Group 3 (median: 21 min; range: 15 to 31 min). ANOVA showed a significant difference in the median operation time with P = 0.0001. The post hoc t-test revealed a significant difference in the median operation time between Groups 1 and 2 (P = 0.0186), Groups 1 and 3 (P = 0.0024) and Groups 2 and 3 (P < 0.0001). The size of the caecum specimens resected did not differ significantly among the three series (P = 0.0863), showing that size variations did not bias the different findings among the groups.
For Group 1, five of the ten stumps were insufficient intraoperatively, resulting in stool release into the abdominal cavity. The relaparoscopy identified stump insufficiency for each of the five remaining rats, as demonstrated by microbiological analysis. For Group 2, intraoperatively as well as during relaparoscopy, no stool was detectable in the abdominal cavity and the microbiological analysis showed the complete absence of bacterial growth on McConkey agar plates. The results for all ten operations were satisfactory. For Group 3, intraoperatively, there was no indication of leakage or damage, and during relaparoscopy the stump closure was sufficient and no stool translocation was macroscopically observed. The latter was confirmed by microbiological analysis.
In a long-term follow-up group (data not shown), there was no macroscopic or microbiological evidence for contamination or leakage of the stump closure. These results indicate that the stump closure technique described in Group 2 was also sufficient after the critical postoperative time of 5 days.
LS is becoming more and more popular and is available in general and visceral surgery departments around the world. There are striking advantages over open surgery, such as faster recovery, less operative trauma and better cosmetic results. However, evidence is missing for whether the surgical trauma of LS leads to differences within the cellular and molecular responses, i.e. the innate immune response or wound healing. To address this question, appropriate animals model are needed. Unfortunately, LS models are rare and inhomogeneous and often of insufficient description. In the present study, we evaluated three different techniques for LA. We have provided detailed instructions and recommendations that are helpful for addressing further LS approaches within animal studies.
An initial literature search identified several studies that provided valuable information for the present research project [
11‐
24]. It can be assumed that a standard laparoscopic procedure was not used for these studies. Most used a hybrid operation, meaning that the investigators resected the caecum extracorporeally, instead of performing complete intracorporeal laparoscopic resections. Only in the study by Polat
et al. was it identified that an entire laparoscopic caecum resection was performed using two endoloops [
23]. Additionally, most studies do not list all operation parameters in detail (such as the way stumps are disinfected, the kinds of trocar or incision that were used and to what extent they enlarged the incision to exteriorize the resectate) (Table
2).
Table 2
Studies reporting on laparoscopic caecum resections in rats
Allendorf et al. 1996 [ 16] | 150 g | Ketamine | 4 to 6 mmHg CO2 | 3 ports | Yes (25 gauge) | 4 mm | 2 × 2 mm | 4 mm | Extracorporeal | Ligature |
Allendorf et al. 1997 [ 17] | 150 g | Ketamine | 4 to 6 mmHg CO2 | 3 ports | Yes (25 gauge) | 4 mm | 2 × 2 mm | 4 to 5 mm | Extracorporeal | Ligature |
Le Moine et al. 1998 [ 18] | unknown | Ketamine | 12 mmHg CO2 | 3 trocars | No | unknown | unknown | unknown | Extracorporeal | Ligature |
| unknown | Barbiturate | 8 mmHg CO2/Helium | 3 trocars | No | 5 mm (uncertain) | unknown | 1 cm | Extracorporeal | Suture |
| 250 to 300 g | Barbiturate | 8 mmHg CO2 | 1 trocar, 2 ports | No | 3.5 mm | 2 × 3.5 mm | unknown | Extracorporeal | Suture |
| unknown | Ketamine | 4 mmHg CO2 | 2 incisions | Yes (18 gauge) | 5 mm | 1 × 2 mm | unknown | Extracorporeal | Ligature |
| 250 to 300 g | Barbiturate | 8 mmHg CO2 | 3 trocars | No | 3.5 mm (uncertain) | unknown | unknown | Extracorporeal | Suture |
| 300 to 350 g | Ketamine | 4 to 6 mmHg CO2 | unknown (3 accesses) | Yes (25 gauge) | 4 mm | 2 × 2 mm | 4 mm (uncertain) | Intracorporeal/extracorporeal | Loop suture/ligature |
Schmelzer et al. 2008 [ 24] | unknown | Isoflurane | 4 mmHg CO2 | 1 trocar, 1 incision | No | 3 mm | 1 × 2 mm | 1 cm | Extracorporeal | Ligature |
Furthermore, the technical and procedural approach varied among the published studies. While most investigators used barbiturate [
19,
20,
22] or ketamine [
16‐
18,
21,
23] for narcosis, we used isoflurane (Forene®, Abbott GmbH & Co KG, Wiesbaden, Germany), which was also used by Schmelzer
et al. [
24]. We found that this narcosis was easy to handle, very reliable and the rats recovered quickly afterwards. Overdoses or even narcosis-related deaths were not observed.
Regarding the capnoperitoneum, several investigators used an angiocatheter (18 or 25 gauge) in a trocarless method to insufflate the peritoneal cavity [
16,
17,
21,
23], while others used proper trocars [
18‐
20,
22,
24]. As the usage of trocars appears to imitate LA more realistically in comparison with modern human laparoscopy, we consequently used trocars within the present study. Trocar usage was comfortable and ensured a constant capnoperitoneum without loss of pressure. Furthermore, different laparoscopes were used in the studies, ranging in size from 3 mm [
24] to 5 mm [
19,
21]. In our opinion, a 2.7-mm laparoscope is the ideal solution since it provides a compromise between the improved view from the 5-mm laparoscope and the lower access trauma of the 2-mm laparoscope. We suppose that the rapid and constant technological progress will give better vision even using small calibre laparoscopes in the future. Additionally, we found that instrument size is a crucial factor in rat LS, which was unfortunately not explicitly listed in a number of previous publications. Three studies used 2-mm forceps and scissors [
16,
17,
23] and another group 3.5-mm forceps and scissors [
20], while two other groups performed their operations with 2-mm forceps [
21,
24]. To extract the specimen, some investigators needed a 4-mm incision [
16,
17,
23] while others required a widening of 1 cm [
24]. We compared several instrument sizes and found that 2-mm and 3-mm laparoscopic instruments and trocars were the ideal size for high-precision interventions while keeping the operative trauma at the lowest possible level. Based on the results of this instrument comparison, we strongly recommend usage of a 2.7-mm laparoscope and 2- to 3-mm instruments for LA. Furthermore, the present instrument setting, particularly the choice of appropriate calibres of laparoscopic instruments, appears to be transferable to other experimental laparoscopic animal models.
After establishing an appropriate instrument setting, we compared three different kinds of appendectomy and stump closure techniques. The only group for which stump insufficiencies were observed was Group 1, in which we used bipolar coagulation. Of note, all animals in Group 1 demonstrated stump insufficiencies. Conflictingly, Aslan
et al. [
11] had sufficient stump closures using a bipolar coagulation device in a previous rat study. Although the bipolar coagulation devices differed between the studies, Take-apart® Manhes Bipolar Coagulation Forceps (Karl Storz GmbH & Co KG) versus Power Blade™ (Lina Tripol 5, Denmark) [
11], our results demonstrate severe failure in sufficient stump closure when using only bipolar coagulation for bowel resection surgery. This is confirmed by bursting pressure experiments in the study by Aslan
et al., which demonstrated a significant decrease in bursting pressure to levels as low as 11 cm H
2O for bipolar-coagulated stumps. In contrast, for Groups 2 and 3, all stumps were sufficiently closed and no peritoneal bacterial contamination was detected, indicating that the stumps were effectively sealed. The fastest and most comfortable operations were performed with the 5-mm Ligasure™ device for Group 3, although this is also the most expensive technique. Since for Group 2, all stump closures were sufficient and the procedure is the closest to reality, as it is comparable to the one most often used in children, we recommend the use of an endoloop plus bipolar coagulation. This procedure can be performed easily and appears to be safe and cost-effective, at least for research purposes.
One should note that our present study did not include long-time observation of anastomotic stump closure in all groups. We only pursued 3- and 5-day follow-up inspections for Group 2 as this procedure turned out to be the most reliable technique, particularly in terms of safety, simulating reality and cost-effectiveness. Additionally, we are currently not able to adapt the most frequently used appendiceal stump closure techniques for LA for adult humans, because the linear stapler tools used have diameters that are too large (10 mm) and would lead to inappropriate large access traumas in a rat model.
Appendectomy models have historically been useful in a variety of research fields. In CA models, for example, electrosurgical devices (bipolar cautery [
11,
14], LigaSure Precise [
12,
14]), endoclips [
12‐
14], endoloops [
13,
23] and staplers [
13,
15] have been compared, and all investigated methods have been shown to be feasible, safe and leakproof. While some of the LA rat models also investigated caecum resection and closure techniques, other groups focused on topics such as postoperative immune function [
16,
17,
21,
24], risk of neoplastic port site metastasis after LS [
18,
23], influence of taurolidine/heparin on local tumour growth after laparoscopy [
20,
22] and postoperative adhesion formation [
19]. With our standardized model, we hope to facilitate such research in the future to improve LS.
In the present study, we present evidence to show that endoloop ligation combined with bipolar coagulation is the most appropriate and easy-to-learn model for LA and we hope that this technique will be helpful and further improve research in the field of LA.