Intraperitoneal Adhesions After Open or Laparoscopic Abdominal Procedure: An Experimental Study in the Rat

Abstract

Background:

Adhesion formation is common after abdominal surgery. The incidence and severity of adhesion formation following open or laparoscopic surgery remain controversial. The role of CO₂ pneumoperitoneum is also widely discussed. This study aimed to compare adhesion formation following peritoneal injury by electrocoagulation performed through open or laparoscopic procedures in a rat model.

Materials and Methods:

Sixty male rats were randomized to undergo a 1.5-cm peritoneal injury with unipolar cautery under general anesthesia: open surgery (Group A, n=20), laparoscopic surgery with CO₂ pneumoperitoneum (Group B, n=20), and laparoscopic surgery with air pneumoperitoneum (Group C, n=20). Duration of the procedures was fixed at 90 minutes in all groups, and pneumoperitoneum pressure was kept at 10 mm Hg. Ten days later, the animals underwent a secondary laparotomy to score peritoneal adhesions using qualitative and quantitative parameters.

Results:

Forty-five rats developed at least one adhesion: 95% in Group A, 83% in Group B, and 55% in Group C (P<.01; Group C versus Group A, P<.01). According to number, thickness, tenacity, vascularization, extent, type, and grading according to the Zühkle classification, no significant difference was observed between Groups A and B. The distribution of adhesions after open surgery was significantly different than after laparoscopic surgery (P<.001). It is interesting that Group C rats developed significantly fewer adhesions at the traumatized site, and their adhesions had less severe qualitative scores compared with those after open surgery (P<.01).

Conclusions:

In this animal model, CO₂ laparoscopic surgery did not decrease the formation of postoperative adhesion, compared with open surgery. The difference with the animals operated on with air pneumoperitoneum emphasizes the role of CO₂ in peritoneal injury leading to adhesion formation.

Introduction

Adhesion formation is common after abdominal surgery.¹ The majority of patients who undergo laparotomy will later develop some degree of peritoneal adhesion.² Indeed, the prevalence of adhesion formation after major abdominal procedures has been estimated at 63%–97%.^2–4 These peritoneal adhesions may result in bowel obstruction, pelvic or abdominal pain, and infertility.⁵ In addition, from the surgical point of view, adhesions may prolong the time needed to gain access to the abdominal cavity and increase the risk of bowel injury during subsequent surgery.^6,7 Small bowel obstruction is the most common complication of peritoneal adhesion formation.^2,8,9 As a consequence, postoperative adhesions represent a real health problem and a significant economic impact.¹⁰

Laparoscopic surgery is considered to be less traumatic than open surgery. Some publications have suggested that laparoscopic surgery results in fewer adhesions than equivalent open surgery.^11–14 This, however, has been difficult to scientifically establish in a convincing manner. The response of the peritoneum to laparoscopic surgery and CO₂ pneumoperitoneum is not fully understood.¹⁵ Recent publications have demonstrated that the risk of intestinal occlusion does not seem to be reduced after laparoscopic surgery compared with open procedures.¹⁶

We aimed to develop a reproducible model of postoperative peritoneal adhesion formation in the rat, in order to later evaluate some prophylactic measures against this process. In this study, we compared the formation of peritoneal adhesions after the same peritoneal lesion was surgically induced during a 90-minute procedure performed through open laparotomy or laparoscopy with CO₂ pneumoperitoneum. In addition, a third group using laparoscopy with air pneumoperitoneum was evaluated, to identify the proper role of peritoneal CO₂ in the adhesion formation process.

Materials and Methods

This study was performed at the experimental research center of the Department of Surgery of the University of Liège, Liège, Belgium, following approval by the University Veterinary Ethical Committee. Care and handling of the animals were performed in accordance with international guidelines regarding animal research.

Animals and anesthesia

Inbred adult male Sprague–Dawley rats weighing 350–450 g were used for this study. They were housed in a climate-controlled (24°C) room under a 12-hour light/dark cycle. They were given tap water and commercial rat chow ad libitum before and after surgery. All surgical procedures and euthanasia were performed with the animals under general anesthesia induced by intraperitoneal injection of pentobarbital (Nembutal^®; Abbott Laboratories) (25–35 mg/kg of body weight) and with use of sterile surgical techniques. The anesthetized animals were placed under a heating lamp, and body core temperature was controlled by intrarectal monitoring and maintained between 36°C and 38°C.

Experimental design

Sixty male rats were randomized in three groups of 20 animals to undergo a primary surgical procedure aiming to induce peritoneal injury with unipolar cautery under three different surgical approaches: open surgery (Group A), laparoscopic surgery with CO₂ pneumoperitoneum (Group B), and laparoscopic surgery with air pneumoperitoneum (Group C). Ten days after this procedure, the animals underwent a secondary laparotomy to score intraperitoneal adhesions.

Primary surgical procedure

The rats were shaved and secured to the small-animal operating table in a supine position. In Group A, a xyphopubic midline incision was performed. In Groups B and C, a 10 mm Hg pneumoperitoneum was established using a Veress needle in the left hypochondrium (model 16L high flow insufflator; Stryker Endoscopy, San Jose, CA) and maintained throughout the operation (cold-dry insufflations for 90 minutes at a constant maximal flow of 100 mL/minute). Neither the air nor the CO₂ was humidified. Two 5-mm trocars (one optic and one working trocar) were introduced in the left flank. In all animals, a 1.5-cm peritoneal burn was performed with unipolar electrocautery in the right flank, through either open surgery (Group A) or laparoscopic technique (Groups B and C) using a clinical laparoscopic column (Stryker Endoscopy). After 90 minutes of procedure in all groups, the abdominal cavity was closed in two layers for all incisions, including the 5-mm port sites.

Secondary laparotomy

The animals were kept and observed under standard laboratory conditions. Ten days after the primary surgical procedure, all rats underwent a second surgery through xyphopubic midline laparotomy under general anesthesia. The whole abdominal cavity was explored to score peritoneal adhesions. The animals were then euthanized by exsanguination under general anesthesia.

Scoring of peritoneal adhesions

Peritoneal adhesions were scored at secondary laparotomy. Adhesions were defined as bonds (a thin film of connective tissue or a thick fibrous bridge) or as direct contact between two organ surfaces. The presence or the absence of peritoneal adhesions was recorded and scored using qualitative and quantitative parameters. The quantitative evaluation of adhesions was performed according to the scale described by Moreno et al.¹⁷ (Table 1). The density and the quality of adhesions were assessed according to qualitative parameters¹⁸ (Table 2) and the Zühlke classification¹⁹ (Table 3). All adhesions were considered for the quantitative scoring. The adhesions to the induced peritoneal lesion were evaluated as a percentage of the involved peritoneum expressed as extent in Table 2.

Table 1.

Quantification of Adhesion Formation (Adapted from Moreno et al.¹⁷)

Criteria	Comment
Number of adhesions	1 for each
Site of adhesions
Pelvic fat body–abdominal wall	Parietal
Omentum–abdominal wall	Parietal
Intestine–abdominal wall	Parietal
Omentum–liver/stomach	Visceral
Intestine–intestine	Visceral
Liver–stomach	Visceral
Thickness of adhesions
<3 mm
>3 mm
Tenacity
Type 0	No adhesion
Type I	Simple, without dissection
Type II	Dissection needed to separate adherent area
Type III	Dissection needed to cut the adhesions
Vascularization of adhesions
Yes	Vascularized
No	Avascularized

Table 2.

Qualitative Scoring System for Adhesion Formation (Adapted from Binda et al.¹⁸)

Category, description	Score
Extent
No adhesion	0
1%–25% of the injured of surface involved	1
26%–50% of the injured surface involved	2
51%–75% of the injured of surface involved	3
76%–100% of the injured of surface involved	4
Type
No adhesion	0
Filmy	1
Dense	2
Vascular	3
Tenacity
No adhesion	0
Easily fall apart	1
Require traction	2
Require sharp dissection	3
Total (extent+type+tenacity)	0–10

Table 3.

The Zühkle Grading Classification of Adhesion Density ¹⁹

Grade	Description
Grade 0	No adhesion
Grade I	Filmy adhesions that require gentle, blunt dissection to be divided
Grade II	Mild adhesions that require aggressive blunt dissection to be freed
Grade III	Moderate adhesions that require sharp dissection to be freed
Grade IV	Severe adhesions that are not dissectible without damaging adherent organs

Statistical analysis

Data are presented as absolute numbers or mean±SD values. Statistical analysis was performed using Epi Info™ version 3.3.2 (Centers for Disease Control and Prevention, Atlanta, GA) for Windows XP. Differences between groups were evaluated with the chi-squared test and Yates's corrected test. P<.05 was considered to be statistically significant.

Results

Follow-up and death

The overall mortality rate was 3.3% (n=2). Deaths occurred in rats from Group B (P>.05). One animal died during the initial anesthesia and CO₂ pneumoperitoneum, and one died at postoperative Day 1. Autopsy showed massive hemoperitoneum that caused the animal's death. These two rats that died early were excluded from subsequent analyses.

Adhesion formation

Forty-five rats developed at least one adhesion: 19 (95%) in Group A, 15 (83%) in Group B, and 11 (55%) in Group C (P<.01). Fewer rats from Group C than Group A developed adhesions (P<.01).

Quantification of adhesions

In total, 107 adhesions were observed in the 58 animals studied (Table 4). There was no statistical difference among the three groups (P>.05).

Table 4.

Adhesion Quantification According to the Modified Moreno's Scale

	Group A (n=20)	Group B (n=18)	Group C (n=20)	P
Number	49	36	22	.12
Site				<.001
Parietal	39	23	15
Visceral	10	13	7
Thickness				.57
<3 mm	14	11	9
>3 mm	35	25	13
Tenacity				.004
Type 0	1	3	9
Type I	9	10	6
Type II	19	9	5
Type III	21	17	11
Vascularization				.84
Yes	21	17	11
No	28	19	11

Site of adhesions

The sites of adhesions after open surgery (Group A) were significantly different from those after laparoscopic surgery (P<.001). A significant difference was observed between Group A and Group B (P<.001) and between Group A and Group C (P<.001). No significant difference was seen between the two laparoscopic groups (P>.05). Furthermore, there was no significant difference according to involved organs after laparoscopic surgery and open surgery (P>.05).

The rats predominantly developed parietal adhesions (n=77) (Table 4). Omentum was the internal organ mostly involved in adhesion formation (n=69). In laparoscopic groups (Groups B and C), adhesions predominantly developed between omentum (n=39) or the small intestine (n=15) and the right flank, rather than the scars of the optical (n=5) or the working (n=9) trocars. Animals from Group A also developed more parietal adhesions (n=39) between omentum, small bowel, or liver and the laparotomy scar (n=14) or the cauterized site (n=25). According to the development of adhesions at the surgical scars, 60% of the animals operated on by the open procedure developed adhesions to the laparotomy scars, compared with 31% of the rats operated on by laparoscopy that developed adhesions to the trocar sites (P>.05). In addition, animals that underwent open surgery and CO₂ laparoscopic surgery were more prone to develop several visceral-type adhesions (Table 4). In Group B, visceral-type adhesions developed, especially between the intestines (n=3), omentum and intestine (n=6), and omentum and liver (n=3). In addition, one bridle was formed between intestine loops.

Thickness

Animals mostly developed adhesions more than 3 mm thick (Table 4). The thickness of adhesions after laparoscopic or open surgery was not significantly different (P>.05).

Tenacity

Many adhesions needed sharp dissection for tissue separation (Table 4). There was a significant difference in adhesion tenacity between Groups A and C (P<.01) but no significant difference between Groups A and B (P>.05).

Vascularization

The majority of adhesions that developed in the three groups were not vascularized (Table 4), and no significant difference was observed among all the groups investigated (P>.05).

Extent

The extent of adhesion at the traumatized site is reported in Table 5. This score did not show a significantly difference after open and CO₂ laparoscopic surgery (P>.05). It is remarkable that rats operated on under air pneumoperitoneum demonstrated fewer postoperative adhesions at the traumatized site (difference between Groups A and C, P<.01; between Groups B and C, P<.05).

Table 5.

Extent of Adhesions at the Traumatized Site

Score	Group A (n=20)	Group B (n=18)	Group C (n=20)
0	3	5	14
1	2	0	0
2	2	2	0
3	6	6	4
4	7	5	2

Type

Type of adhesions (Table 6) after air laparoscopic surgery and open surgery was significantly different (P<.01). However, no significant difference was found between Groups A and B (P>.05).

Table 6.

Type of Adhesions

Score	Group A (n=20)	Group B (n=18)	Group C (n=20)
0	1	3	9
1	9	10	6
2	19	9	5
3	21	17	11

Adhesion score

The adhesions scores are shown in Table 7. The smallest score was observed in Group C, but the difference was not significant among the groups.

Table 7.

Adhesion Score

	Group A (n=20)	Group B (n=18)	Group C (n=20)	P
Extent	2.13±1.33	2.69±1.36	1.60±1.49	.06
Type	2.20±0.80	2.02±1.01	1.58±1.25	.16
Tenacity	2.08±0.75	2.00±1.00	1.54±1.23	.20
Total	6.41±2.88	6.71±3.37	4.72±3.97	.16

Adhesion grading

According to the Zühkle classification¹⁹ (Table 8), the difference observed between both laparoscopic groups and the open group was significant (P<.01); a difference was also observed between Groups A and C (P<.01). However, no significant difference in adhesions scores was found between Groups A and B (P>.05).

Table 8.

Adhesion Grading According to the Zühkle Classification ¹⁹

	Group A (n=20)	Group B (n=18)	Group C (n=20)
Grade 0	1	3	9
Grade I	9	10	6
Grade II	5	3	3
Grade III	20	7	3
Grade IV	15	16	10

Discussion

The development of peritoneal adhesions is an inevitable consequence of surgery in the abdominal cavity. These peritoneal adhesions may later cause intestinal obstruction as their most serious complication. The results of this study demonstrated that for the same 90-minute surgical procedure (i.e., 1.5-cm peritoneal burn), rats operated on with a CO₂ laparoscopic approach may not develop less severe peritoneal adhesions compared with rats that underwent open laparotomy procedure. In addition, the results of this study showed that compared with the clinically used CO₂ pneumoperitoneum, rats operated on under air pneumoperitoneum conditions may develop significantly fewer peritoneal adhesions.

When comparing adhesions induced by laparoscopic and open abdominal procedures, not only the overall adhesions should be evaluated, but, more importantly, three different adhesion sites should be separately analyzed: the adhesions at the surgical procedure site itself (in this study, the 1.5-cm peritoneal burn in the right flank), the adhesions developed where the abdominal cavity was opened for realization of the procedure (laparotomy or port site scars), and the adhesions developed elsewhere in the peritoneal cavity.

In this model, the extent of adhesions at the traumatized site was not significantly different in the laparotomy group compared with the CO₂ laparoscopic group, indicating that for the same surgical trauma, in our model at least, CO₂ laparoscopic surgery did not reduce the development of adhesions. It is interesting that this extent of adhesion was significantly reduced in rats operated on under air pneumoperitoneum compared with both other groups. Schippers et al.²⁰ compared laparoscopic and open cecal resection and a deserosation of the abdominal wall in a dog model and demonstrated fewer adhesions in the animals operated on laparoscopically. Gutt et al.¹¹ reviewed the influence of laparoscopy, comparative to laparotomy, in the process of adhesion formation. According to adhesions at the operative site, nine studies (two clinical and most of the experimental ones) demonstrated a reduced adhesion formation after laparoscopy, while five studies, all experimental, found an equal risk of adhesion formation. Moore et al.²¹ and Luciano et al.²² evaluated the incidence of adhesions after laparoscopic and open surgery in pig and rabbit models, respectively. They found absence or fewer adhesions at the operative sites after laparoscopy. However, Marana et al.²³ and Filmar et al.²⁴ found no statistically significant difference for one or another of the surgical methods according to adhesion at operative sites in animal models. Furthermore, Jacobi et al.²⁵ demonstrated that in conditions of peritonitis, adhesion formation is the same after laparoscopic or open cecal resection.

According to these controversial results, the improved result we observed with air pneumoperitoneum may implicate the negative impact of the CO₂ pneumoperitoneum in adhesion formation. Molinas and Koninckx²⁶ demonstrated that CO₂ pneumoperitoneum is a cofactor in adhesion formation and strongly suggested peritoneal hypoxia as the driving mechanism, with an increase in adhesions with increased pneumoperitoneum duration and pressure. Molinas and Koninckx²⁶ did not find a difference between CO₂ and helium pneumoperitoneum according to adhesion formation risk. However, Jacobi et al.²⁵ found that the overall adhesion score was significantly lower in the helium group than in the two other operation groups (laparotomy and CO₂ group). Furthermore, there was no difference between the laparotomy and CO₂ groups according to adhesion score.²⁵

Although the difference was not significant when evaluating the number of adhesions at the surgical scars in our study, animals operated on by the open procedure developed two times more adhesions to the laparotomy scars, compared with rats operated on by laparoscopy to the trocar sites. It is clear that, in this model, the total length of abdominal wall incision was reduced in animals from both laparoscopic groups (2×5 mm for the two trocars) compared with the full xyphopubic midline incision (approximately 5–6 cm) in the animals that underwent open surgery. This finding certainly reflects one of the major advantages of the laparoscopy approach: its minimal lesions to the abdominal wall, leading to less adhesion at the scars and also less risk of incisional hernia. Moore et al.²¹ found fewer adhesions in the trocar sites (12.5%), whereas in all the pigs in the open nephrectomy group, adhesions were present along the incision site. According to the literature search of Gutt et al.¹¹ on adhesion formation, only one experimental study favored laparotomy over laparoscopy. Indeed, Krähenbühl et al.²⁷ observed fewer adhesions in the laparotomy group, whereas in the laparoscopic group several adhesions were observed in the scars of the optical and working trocars. But among the clinical studies, Audebert and Gomel²⁸ found the best results in the laparoscopic group according to adhesions to the operative wound.

In addition, the animals of the different groups of this study also developed adhesions elsewhere in the abdominal cavity, but the sites and the quality of these adhesions were different. In total, the laparoscopic approaches seem to induce fewer adhesions with a lower severity score than the open surgery, with again a reduction of severity in the animals from the air pneumoperitoneum group. These results are in accordance with the review of Gutt et al.,¹¹ which described only four experimental animal studies evaluating distant intervisceral adhesions. Three studies favored laparoscopy, and one showed equal impact on adhesion formation for the two surgical techniques. Krähenbühl et al.,²⁷ Schippers et al.,²⁰ and Luciano et al.²² reported no or less adhesion at distant intervisceral adhesions in the laparoscopic groups, but for laparotomy, they found significantly more frequent intervisceral adhesions where no apparent injury had been inflicted.

Therefore, in our study, the animals that underwent air laparoscopy were the ones with little adhesion formation. This finding may correlate with the fact that cold-dry CO₂ laparoscopy induces peritoneal ischemia and acidosis. In this study, we deliberately chose to use cold-dry CO₂ to mimic the usual clinical conditions of laparoscopy and to perform air pneumoperitoneum in the same conditions. The fewer adhesions in the air laparoscopic group might be explained by the fact that the presence of oxygen in air pneumoperitoneum may allow a better peritoneal cell metabolism leading to less peritoneal trauma and less postoperative adhesions. This finding is corroborated by data from Molinas and Koninckx²⁶ and Molinas et al.²⁹ demonstrating that CO₂ pneumoperitoneum increases adhesion in a time-and pressure-dependent manner and that this process is reduced by the addition of 2%–4% oxygen, suggesting peritoneal hypoxia as an important mechanism in the adhesion formation during CO₂ pneumoperitoneum. Indeed, prolonged laparoscopic surgery requires long duration and large-volume gas insufflations, which raises concerns about the adverse effects of prolonged CO₂ insufflations.³⁰ The standard CO₂ used in current laparoscopic practice is cold-dry CO₂ that is not physiological to the normal condition of the peritoneal cavity. Many studies have shown that short-duration (<3 hours) laparoscopic procedures, with cold-dry CO₂ insufflations, can cause peritoneal alterations and result in numerous detrimental outcomes, including postoperative adhesion formation.^30–34 It is supposed that when fibrinolytic activity decreases, the process of adhesion formation does not depend on the type of surgery anymore, but evolves on its own account. The benefits of heated humidified CO₂ insufflations (37°C and 95% relative humidity, physiological condition) have been reported to include less hypothermia, less postoperative pains, shortened recovery room stay, better convalescence, less tumor spread and growth, and less adhesion formation.^30–34 The superiority of air pneumoperitoneum in this animal model should not be reproduced in the human setting, as the risks of gaseous embolism are more severe with air than CO₂. This should only be considered as additional information on the effects of the clinically used CO₂ on peritoneum, irrespective of the pressure and the length of the procedure.

This animal study may confirm the clinical findings that abdominal surgeons experience on an everyday basis: patients who undergo laparoscopic surgery develop less severe adhesions than patients operated on by laparotomy, particularly if adhesions to the abdominal wall are concerned. Even if this fact is difficult to scientifically prove in animal models or in prospective clinical trials, it is clinically evident that getting laparoscopic or open access to an abdominal cavity that was already operated by laparoscopy or laparotomy is a different challenge. However, in addition to this secondary technical issue, the main clinical question should be to determine if, for the same procedure, patients operated on by laparoscopy have less risks of developing clinical complications related to these adhesions, particularly intestinal obstruction or abdominal pain. Recent prospective studies did not observe such an advantage of laparoscopy over an open procedure,¹⁶ but this is still a matter of debate.

Footnotes

Acknowledgments

W.A. is a PhD student supported by Cooperation Technique Belge, the Belgian Development Agency. O.D. is helped by a grant from Wallonie Bruxelles International (Wallonie, Belgique), aiming to develop laparoscopic surgery in the Democratic Republic of Congo. This study was supported in part by an unrestricted grant from Roche, Belgium.

Disclosure Statement

No competing financial interests exist.

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