The prevention of peritoneal adhesions by in situ cross-linking hydrogels of hyaluronic acid and cellulose derivatives

Abstract

Post-operative peritoneal adhesions can cause pelvic pain, infertility, and potentially lethal bowel obstruction. We have designed and synthesized injectable hydrogels that are formed by mixing hydrazide-modified hyaluronic acid (HA) with aldehyde-modified versions of cellulose derivatives such as carboxymethylcellulose (CMC), hydroxypropylmethylcellulose (HPMC), and methylcellulose (MC). Gelation of these hydrogels occurred in less than 1 min, and had higher shear moduli than that of HA-HA gel (HAX). Hydrogels degraded in the presence of hyaluronidase in vitro, with HA-MC and HA-HPMC degrading more slowly than HAX and HA-CMC. The aldehyde-modified cellulose derivatives showed dose-dependent mild-to-moderate cytotoxicity to mesothelial cells and macrophages in vitro, but all were biocompatible in the murine peritoneum, causing no adhesions for 3 weeks. All the cellulose-derived gels showed efficacy in reducing the area of adhesion formation in a rabbit sidewall defect-bowel abrasion model.

Fig. 1

Chemical structure of polymers. Regarding the cellulose derivatives: for CMC-CHO: R is H or CH₂COOH. For HPMC-CHO: R is H or (CH₂CH(CH₃)O)_nH. For MC-CHO: R is H or CH₃.

Fig. 2

Degradation kinetics of the hydrogels in 10 unit/ml hyaluronidase in PBS at 37 ºC. Volume of the hydrogel (%) is the ratio of the volume of hydrogel at each time point to the initial volume, expressed as a percentage. Data are averages ± standard deviations (n=4).

Fig. 3

Effect of aldehyde polymers on cell viability measured by the MTT assay. (A) Mesothelial cells after 3 days incubation with polymers. (B) Macrophages (J774.A1 cell line), after 2 days incubation with polymers. Data are averages ± standard deviations (n=4).

Fig. 4

Peritoneums of mice 1 week after injection of hydrogels. (A) HAX: no residue. (B) HA-CMC: note the thin coating of gel-like material. (C) HA-MC: note the increased amount of residual material, demonstrated the forceps submerged beneath it.

Fig. 5

Prevention of peritoneal adhesions in a rabbit abrasion model. A. Induction of adhesions. Note the abdominal wall defect (arrow), and the bleeding surface of the cecum. B. Adhesions seen on dissection after 1 week in an animal treated with saline. C. Absence of adhesions after 1 week in an animal treated with HA-MC.

Fig. 6

Photomicrographs of tissues recovered 1 week after injury in the rabbit sidewall defect-bowel abrasion model. (A) Cross-section of an abrasion in a saline-treated animal. The cecal lumen (CE) is in the left upper corner of the picture. The cecal smooth muscle is fused to the striated muscle of abdominal musculature (AM). Magnification 100 X. (B) Hydrogel recovered from an animal treated with HA-MC, with inflammatory cells (predominantly macrophages and lymphocytes). Magnification 100 X. (C) Site of abdominal wall defect in an animal treated with HA-MC. The defect has been re-epithelialized (arrows), with a subjacent layer of healing tissue (predominantly fibroblasts). Magnification 400 X. (D) Normal untreated parietal peritoneum. The mesothelium (arrows) overlies connective tissue (CT) and abdominal muscle.