Design of chitosan- and dextran-based hydrogels for preventing postoperative adhesions [English translation]

Postoperative adhesions (POA) are a common complication following surgical procedures, particularly abdominal and pelvic surgeries. POA can lead to chronic pain, bowel obstruction, infertility, and organ dysfunction. These adhesions result from an excessive inflammatory response and a disorganized healing process, leading to the formation of abnormal fibrous bands between organs and tissues. Among the various strategies explored to prevent POA, hydrogels have emerged as promising biomaterials. Due to their water-rich three-dimensional (3D) structure, hydrogels act as physical barriers between surgical surfaces, preventing direct contact between nearby organs at the surgical site and thus reducing the formation of fibrous bridges responsible for adhesions. Hybrid hydrogels composed of chitosan and dextran have shown significant potential in this application. Chitosan (CHI) is selected for its natural origin, biocompatibility, biodegradability, as well as its hemostatic and antimicrobial properties, while dextran is favored for its anti-adhesive and low fouling properties, non-immunogenic nature, biocompatibility, and biodegradability. Until now, these two polymers have typically been crosslinked by chemically modifying dextran through oxidation to introduce aldehyde groups that can react with the amine groups of chitosan, forming a stable 3D structure. However, the preparation of aldehyde-modified dextran via oxidation raises concerns regarding biocompatibility, as the oxidation process, often performed using sodium periodate, can lead to partial degradation of the polymer and generate potentially cytotoxic by-products. This research project aims to change the crosslinking technique; its original contribution lies in the design of hybrid hydrogels based on chitosan and dextran without degradation of the biopolymers, without generating by-products, and using chemical crosslinking. Specifically, we modified dextran by carboxymethylation. The idea is to react the carboxylic acid groups of carboxymethylated dextran (CMD) with the native amine groups of chitosan to form covalent bonds. The selected coupling agent is NHS/EDC, widely used for protein grafting, as it forms stable amide bonds between activated carboxyl groups and free amine groups at neutral pH. 1-Ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) activates the carboxylic acids of CMD, while N-hydroxysuccinimide (NHS) stabilizes the reactive intermediate by forming an NHS ester, which is more durable and more reactive with amines. This coupling method is carried out in aqueous media, at room temperature, and without generating toxic by-products, making it an ideal strategy for designing biomaterials intended for medical use. However, chitosan (pKa ≃ 6.5) is only soluble under acidic conditions due to the protonation of its amine groups (-NH3+), and at the optimal pH for the NHS/EDC reaction (pH ≃ 7–8), chitosan alone tends to precipitate. In our research group, it was recently demonstrated that the addition of sodium bicarbonate increases the pH of the solution beyond the pKa of chitosan without causing precipitation. This approach will be used to couple, for the first time, chitosan and CMD via NHS/EDC chemistry, leading to the formation of a stable, biocompatible hydrogel that is potentially effective for the prevention of postoperative adhesions. The second part of the project focused on the characterization of the hydrogels. Their mechanical properties were evaluated through unconfined compression tests for various CHI:CMD volume ratios (ranging from 30:70 to 80:20 %v/v), using stock solutions at 20 mg/mL. All samples exhibited a modulus in the kPa range, similar to that of soft tissues. Gelation, assessed by rheometry, was found to be highly temperature-dependent: it occurred almost instantaneously at 37 °C but took approximately 30 minutes at 4 °C. This temperature-sensitive behavior enables control over the shape of the hydrogels (through molding for example), paving the way for biomedical applications, particularly in injectable systems. Finally, the last phase of the project aimed to evaluate the cell non-adhesiveness of the hydrogels. Human umbilical vein endothelial cells (HUVECs), commonly used in vitro to study cell-material interactions due to their key role in inflammation, tissue repair, and vascular formation, processes associated with postoperative adhesions, were selected for this purpose. HUVECs showed no adhesion to the CHI-CMD hydrogels, suggesting a low risk of cell colonization and fibrous tissue formation between organs. Unlike 2D surfaces that promote cell attachment, the 3D hydrogels prevented cell adhesion even in the presence of adhesion peptides. These results further support their potential in preventing postoperative adhesions. Thus, we present a simple and original method, both versatile and innovative, for the design of hydrogels intended for the prevention of postoperative adhesions. This approach avoids the formation of by-products, while opening new opportunities in hydrogel production processes and their application across various other biomedical fields.