Anastomotic leaks remain one of the most dreaded complications in gastrointestinal surgery causing significant morbidity, that negatively affect the patients' quality of life. Experimental studies play an important role in understanding the pathophysiological background of anastomotic healing and there are still many fields that require further investigation. Knowledge drawn from these studies can lead to interventions or techniques that can reduce the risk of anastomotic leak in patients with high-risk features. Despite the advances in experimental protocols and techniques, designing a high-quality study is still challenging for the investigators as there is a plethora of different models used.

To review current state of the art for experimental protocols in high-risk anastomosis in rats.

This systematic review was performed according to The Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines. To identify eligible studies, a comprehensive literature search was performed in the electronic databases PubMed (MEDLINE) and Scopus, covering the period from conception until 18 October 2023.

From our search strategy 102 studies were included and were categorized based on the mechanism used to create a high-risk anastomosis. Methods of assessing anastomotic healing were extracted and were individually appraised.

Anastomotic healing studies have evolved over the last decades, but the findings are yet to be translated into human studies. There is a need for high-quality, well-designed studies that will help to the better understanding of the pathophysiology of anastomotic healing and the effects of various interventions.

For several decades, scientific efforts have been taken to develop strategies and medical aids for the reduction of anastomotic complications after intestinal surgery. Still, anastomotic leakage (AL) represents a frequently occurring postoperative complication with serious consequences on health, quality of life, and economic aspects. Approaches using collagen and/or fibrin-based sealants to cover intestinal anastomoses have shown promising effects toward leak reduction; however, they have not reached routine use yet. To assess the effects of covering intestinal anastomoses with collagen and/or fibrin-based sealants on postoperative leakage, a systematic review and meta-analysis were conducted.

PubMed, Web of Science, Cochrane Library, and Scopus (01/01/1964 to 17/01/2022) were searched to identify studies investigating the effects of coating any intestinal anastomoses with collagen and/or fibrin-based sealants on postoperative AL, reoperation rates, Clavien-Dindo major complication, mortality, and hospitalization length. Pooled odds ratios (ORs) with 95% confidence intervals (CIs) were calculated.

Overall, 15 studies (five randomized controlled trials, three nonrandomized intervention studies, six observational cohort studies) examining 1,387 patients in the intervention group and 2,243 in the control group were included. Using fixed-effects meta-analysis (I 2 < 50%), patients with coated intestinal anastomoses presented significantly lower AL rates (OR = 0.37; 95% CI 0.27-0.52; p < 0.00001), reoperation rates (OR, 0.21; 95% CI, 0.10-0.47; p = 0.0001), and Clavien-Dindo major complication rates (OR, 0.54; 95% CI, 0.35-0.84; p = 0.006) in comparison to controls, with results remaining stable in sensitivity and subgroup analyses (stratified by study design, age group, intervention used, location of anastomoses, and indication for surgery). The length of hospitalization was significantly shorter in the intervention group (weighted mean difference (WMD), -1.96; 95% CI, -3.21, -0.71; p = 0.002) using random-effects meta-analysis (I 2 ≥ 50%), especially for patients with surgery of upper gastrointestinal malignancy (WMD, -4.94; 95% CI, -7.98, -1.90; p = 0.001).

The application of collagen-based laminar biomaterials or fibrin sealants on intestinal anastomoses can significantly reduce postoperative rates of AL and its sequelae. Coating of intestinal anastomoses could be a step toward effective and sustainable leak prevention. To assess the validity and robustness of these findings, further clinical studies need to be conducted.

Digestive tract resections are usually followed by an anastomosis. Anastomotic leakage, normally due to failed healing, is the most feared complication in digestive surgery because it is associated with high morbidity and mortality. Despite technical and technological advances and focused research, its rates have remained almost unchanged the last decades. In the last two decades, stem cells (SCs) have been shown to enhance healing in animal and human studies; hence, SCs have emerged since 2008 as an alternative to improve anastomoses outcomes.

To summarise the published knowledge of SC utilisation as a preventative tool for hollow digestive viscera anastomotic or suture leaks.

PubMed, Science Direct, Scopus and Cochrane searches were performed using the key words "anastomosis", "colorectal/colonic anastomoses", "anastomotic leak", "stem cells", "progenitor cells", "cellular therapy" and "cell therapy" in order to identify relevant articles published in English and Spanish during the years of 2000 to 2021. Studies employing SCs, performing digestive anastomoses in hollow viscera or digestive perforation sutures and monitoring healing were finally included. Reference lists from the selected articles were reviewed to identify additional pertinent articles.Given the great variability in the study designs, anastomotic models, interventions (SCs, doses and vehicles) and outcome measures, performing a reliable meta-analysis was considered impossible, so we present the studies, their results and limitations.

Eighteen preclinical studies and three review papers were identified; no clinical studies have been published and there are no registered clinical trials. Experimental studies, mainly in rat and porcine models and occasionally in very adverse conditions such as ischaemia or colitis, have been demonstrated SCs as safe and have shown some encouraging morphological, functional and even clinical results. Mesenchymal SCs are mostly employed, and delivery routes are mainly local injections and cell sheets followed by biosutures (sutures coated by SCs) or purely topical. As potential weaknesses, animal models need to be improved to make them more comparable and equivalent to clinical practice, and the SC isolation processes need to be standardised. There is notable heterogeneity in the studies, making them difficult to compare. Further investigations are needed to establish the indications, the administration system, potential adjuvants, the final efficacy and to confirm safety and exclude definitively oncological concerns.

The future role of SC therapy to induce healing processes in digestive anastomoses/sutures still needs to be determined and seems to be currently far from clinical use.

Haemostatic materials such as: gelatine sponges, oxygenated cellulose meshes, tissue sealants, collagen matrices with human thrombin and fibrinogen are gaining on popularity in gastrointestinal surgery, especially in colorectal surgery. We searched for available scientific publications in the Pubmed and Cochrane database on the use of individual hemostatic materials in the field of gastrointestinal surgery. The analysis focused on the assessment of the safety of the use of individual materials in terms of the rate of bleeding complications and the rate of anastomotic leakage cases. The use of haemostatic materials has for years been a recognized method of reducing the rate of intra- and postoperative complications, both in gastrointestinal surgery and in other surgical specialties. Based on the available studies, it can be concluded that the use of hemostatic materials such as matrices, sponges and adhesives in gastrointestinal surgery, even in patients at high risk of anastomotic leakage and bleeding complications, reduces the incidence of complications. The growing popularity of haemostatics and sealants in surgery means that they are currently used in a wide range of indications, and surgeons are more and more willing to use them even in case of standard surgical procedures, which is reflected in the available studies. Choosing a haemostat should be a conscious decision, taking into account the site and type of bleeding, mechanism of action, ease of use, efficacy, safety, and price, among others.

In most cases, traditional techniques to perform an anastomosis following gastrointestinal resections lead to successful healing. However, despite focused research in the field, in certain high-risk situations leakage rates remain almost unchanged. Here, additional techniques may help the surgeon to protect the anastomosis and prevent leakage. We give an overview of some of the latest developments on experimental and clinical techniques for induction of anastomotic healing.

We performed a review of the current literature on approaches to improve anastomotic healing.

Many promising approaches with a high clinical potential are in the developmental pipeline. Highly experimental approaches like inhibition of matrix metalloproteinases, stem cell therapy, hyperbaric oxygen therapy, induction of the hypoxic adaptive response, and the administration of growth factors are still in the preclinical phase. Other more clinical developments aim to strengthen the anastomotic suture line mechanically while shielding it from the influence of the microbiome. Among them are gluing, seaming the staple line, attachment of laminar biomaterials, and temporary intraluminal tubes. In addition, individualized bowel preparation, selectively reducing certain detrimental microbial populations could become the next stage of bowel preparation. Compression anastomoses are evolving as an equivalent technique additional to established hand-sewn and stapled anastomoses. Fluorescence angiography and flexible endoscopy could complement intraoperative quality control additionally to the air leak tests. Virtual ileostomy is a concept to prepare the bowel for the easy formation of a stoma in case of leakage.

A variety of promising diagnostic and prophylactic measures that may support the surgeon in identifying high-risk anastomoses and support them according to their potential deficits is currently in development.

Adhesion formation is a major problem when a mesh is exposed to intraabodminal viscera, with potential severe complications (bowel occlusion, fistulas or abscesses). New methods for preventing adhesions from a polypropylene mesh placed intra-abdominally or to solve difficult situations, such as when the peritoneum cannot be closed during a TAPP repair for an inguinal hernia, are still being seeked. This study mimics in an animal model a situation that can be found in clinical practice during laparoscopic inguinal hernioplasty. A polypropylene mesh could be exposed to the intra-abdominal cavity even when the peritoneum is closed due to different circumstances, with no options to guarantee the prosthetic material of being exposed to the intrabdominal viscera. Different options have been suggested to solve these situations, being proposed in this study to cover the visceral surface of the mesh with an absorbable sponge containing thrombin, fibrinogen, and clotting factors (Tachosil^®, Nycomed, Takeda, Osaka, Japan), to assess its use as a barrier to prevent postoperative adhesion formation.

Thirty Wistar white rats (300-450 mg) were included in this study as experimental animals, being randomized into three groups (A, B, and C). We performed a bilateral prosthetic repair with conventional polypropylene mesh (2 × 2 cm, 82 kD). Prosthesis fixation was performed as follows. Group A: absorbable suture; group B: metal staples; group C: metal tackers. A piece of insulating absorbable sponge (Tachosil^® 5 × 5 cm) was placed to cover the visceral surface of mesh placed at the right side of each animal. After 10 days, we performed a gross examination (by laparoscopy and laparotomy), measuring the quantity and the quality of the adhesions. Samples were taken for histopathological analysis.

Tachosil^®-treated prostheses showed a statistically significant decrease in the quality of the adhesion found (p < 0.05). In addition, a smaller quantity of adhesions was identified in barrier-treated animals, although this lacked statistical significance. The histologic analysis showed no significant differences: more edema with the untreated mesh and increased angiogenesis and a lower degree of necrosis in mesh covered with Tachosil^®.

The use of Tachosil^® as a barrier material led to the absence of strong adhesions as it prevented direct contact between the mesh and the internal organs, preventing major problems associated with strong adhesions.

Anastomotic leakage is a disastrous complication after colorectal surgery. Although current methods for leakage prevention have different levels of clinical efficacy, they are until now imperfect solutions. Stem cell therapy using ASC sheets could provide a solution to this problem. ASCs are considered as promising candidates for promoting tissue healing because of their trophic and immunomodulatory properties. Here, we provide methods to produce high-density ASC sheets, that are transplanted onto a colorectal anastomosis in a rat model to reduce the leakage. ASCs formed cell sheets in thermo-responsive culture dishes that could be easily detached. On the day of the transplantation, a partial colectomy with a 5-suture colorectal anastomosis was performed. Animals were immediately transplanted with 1 ASC sheet per rat. ASC sheets adhered spontaneously to the anastomosis without any glue, suture, or any biomaterial. Animal groups were sacrificed 3 and 7 days postoperatively. Compared to transplanted animals, the incidence of anastomotic abscesses and leakage was higher in control animals. In our model, the transplantation of ASC sheets after colorectal anastomosis was successful and associated with a lower leakage rate.

The intestinal anastomotic failure is one of the most severe complications in gastrointestinal surgery. Despite the great surgical improvements during the last decade, anastomotic leak rates remain practically the same, with a dramatically high grade of morbidity for patients. Leakages are usually the final consequence of ischemia in the anastomosis, leading to tissue hypoxia. In response to hypoxia, the cell orchestrates a variety of coordinated responses in order to restore oxygen homeostasis. The molecular mechanism of hypoxia sensitivity involves oxygen sensing hydroxylases, prolyl-hydroxylases, orchestrating two main transcription factors related to induction of inflammation and angiogenesis, namely nuclear factor-κB and hypoxia-inducible factors. The immunohistochemical expression of two transcription factors hypoxia-inducible factors-1α and nuclear factor-κB p65 has already been described in several disorders, including wound healing, asthma and chronic obstructive lung disease, rheumatoid arthritis, cancer, inflammatory bowel disease, and acute colitis. In the surgical field, fibrin sealants have been widely used to prevent leaks in lung surgery and they might also be useful as a reinforcement of sutures in intestinal anastomosis. The commercial fibrin sealant patches are hemostatic and adhesive surgical agents mainly derived from human plasma. We herein report the results of a prospective randomized experimental study on pigs. We performed a high-risk leakage model of bowel anastomosis, causing a significant devascularization of 10-15 cm of the bowel wall before performing a conventional colo-ileal anastomosis. We randomized the animals to receive a covering of the anastomosis with a fibrin patch (case group) or not (control group). We report the changes in the immunohistochemical expression of the proteins involved in tissue response to hypoxia in the experimental model. Our results indicate that the fibrin patch delays the healing response, promoting a longer lasting inflammation in the surgical bed. Nevertheless, the fibrin patches effectiveness to reduce dehiscence shown in clinical practice suggests that this delay does not negatively affect patients' outcome. Impact statement The consequences of the anastomotic failure are dramatic for patients. Understanding how the ever-increasing use of fibrin sealant, that seems to have a beneficial effect on the anastomoses, interacts with the tissue and the healing process can help to justify its use and encourage research on how to improve this effect even more. We feel that the present work shows that the patch can improve healing by complex mechanisms other than the mere contention and physical support of the intestine. Furthermore, research is needed to confirm our preliminary findings.

One of the most severe complications after esophaguectomy is anastomotic dehiscence. The use of collagen sponges could be an effective way to resolve this complication. Our objective was to perform an experimental model of esophageal anastomosis in rats to study these mechanisms.

A total of 50 Sprague-Dawley rats were used divided into 2 groups, Tachosil^® group (n=25) and control group (n=25). After the section of the abdominal esophagus a single-layer esophago-gastric anastomosis was performed reinforced with 1cm of Tachosil^® wrapping the anastomosis in group 1. A functional study was performed using manometry as well as histopathological and immunohistochemical studies for angiogenic, fibrogenic and growth factors.

The mortality in our series was 8% in the collagen dressing group, compared to 36% in the control group. When esophageal manometry was performed, the dehiscence pressure was higher in the reinforced anastomosis, On microscopical analysis, in the collagen dressing group a profuse inflammatory reaction with abundant neutrophils and macrophages surrounded by a connective matrix with fibroblasts and blood vessels was observed, The expression of VEGF, FGF1 and FGF2 was noticeably higher in the collagen dressing group.

These results show that the application of collagen dressing facilitates tissue reparation phenomena, and therefore could be very useful as a reinforcement of esophago-gastric anastomosis to prevent dehiscence.

The most dreaded complication of colorectal surgery is anastomotic leakage. Adipose tissue-derived stem cell sheets (ASC sheets) prepared from temperature-responsive culture surfaces can be easily transplanted onto tissues. These sheets are proposed to improve cell transplant efficiency and enhance wound healing. The aim of this study was to investigate whether application of ASC sheets could prevent leakage of sutured colorectal anastomoses. Insufficient suturing of colorectal anastomoses was performed in Wistar rats to create a colorectal anastomotic leakage model. Rats were randomized to ASC sheet application or control group. Leakage, abscess formation, adhesion formation, anastomotic bursting pressure (ABP), and histology were evaluated on postoperative day 3 or 7. ASC sheet application significantly reduced anastomotic leakage compared to controls, without increased adhesion formation. ASC sheet transplantation resulted in more CD3+ T-cells and CD163+ anti-inflammatory macrophages at the anastomotic site than the control group. ABP, vessel density and collagen deposition were not different between groups. Using cell sheet technology, we generated ASC sheets that prevented disruption of sutured colorectal anastomoses as shown by reduced leakage. Increased numbers of anti-inflammatory macrophages and T-cells might have contributed to this positive effect.

Anastomotic leak in the gastrointestinal tract is one of the most important complications of resection. They are the main cause of reoperation, their occurrence worsens the prognosis of the patient, increasing the proportion of direct mortality, as well as being a significant risk factor for recurrence of cancer. The risk of leaks within the gastrointestinal tract is greatly varied, depending on the location and extent of the resection, but also on patient, disease or a surgical procedure, including surgeon. To determine the potential risk of leakage can be significant for introduction some prophylactic actions. Some of them have the character of general recommendations, as proper nutrition of the patient in the perioperative period, while another part is directly connected to the surgical procedure. The second group includes protective stoma, the use of tissue glues, insertion transrectal drain for rectal anastomosis decompression, the use of stents or the use of collagen matrix coated with fibrinogen and thrombin. Important to reduce the proportion of leaks can be more precise and targeted prophylactic recommendations, based on the individualized determination of risk factors leaks. Further research for this purpose are necessary for this purpose, the big hope can be associated with data obtained through mobile applications.