Zwitterionic Chitosan Derivative, a New Biocompatible Pharmaceutical Excipient, Prevents Endotoxin-Mediated Cytokine Release

Chitosan is a cationic polymer of natural origin and has been widely explored as a pharmaceutical excipient for a broad range of biomedical applications. While generally considered safe and biocompatible, chitosan has the ability to induce inflammatory reactions, which varies with the physical and chemical properties. We hypothesized that the previously reported zwitterionic chitosan (ZWC) derivative had relatively low pro-inflammatory potential because of the aqueous solubility and reduced amine content. To test this, we compared various chitosans with different aqueous solubilities or primary amine contents with respect to the intraperitoneal (IP) biocompatibility and the propensity to induce pro-inflammatory cytokine production from macrophages. ZWC was relatively well tolerated in ICR mice after IP administration and had no pro-inflammatory effect on naïve macrophages. Comparison with other chitosans indicates that these properties are mainly due to the aqueous solubility at neutral pH and relatively low molecular weight of ZWC. Interestingly, ZWC had a unique ability to suppress cytokine/chemokine production in macrophages challenged with lipopolysaccharide (LPS). This effect is likely due to the strong affinity of ZWC to LPS, which inactivates the pro-inflammatory function of LPS, and appears to be related to the reduced amine content. Our finding warrants further investigation of ZWC as a functional biomaterial

Novel patch biomaterial treatment for colon diverticulosis in swine model

Current leading managements for diverticular disease cannot prevent the recurrence of diverticulitis, bleeding and/or other complications. There is an immediate need for developing new minimal invasive therapeutic strategies to prevent and treat this disease. Through a biomechanical analysis of porcine colon with diverticular lesions, we proposed a novel adhesive patch concept aiming at mechanical reconstruction of the diseased colon wall. This study aims to evaluate the surgical feasibility (safety and efficacy) of pulmonary visceral pleura (PVP) patch therapy using a pig model of diverticulosis. Six female Yucatan miniature pigs underwent collagenase injection (CI) for the development of diverticular lesions. The lesions in each animal either received patch implantation (treated group, n = 40 for 6 pigs) or left intact (untreated group, n = 44 for 6 pigs). The normal colonic wall in each animal received patch implantation at two spots to serve as control (n = 12 for 6 pigs). After 3 months of observation, the performance and safety of the patch treatment were evaluated through macroscopic and histological examination. We found that 95% of pouch-like herniation of the mucosa was prevented from the colon wall with the treatment. The pouch diameter was significantly reduced in the treated group as compared to the untreated group (p < 0.001). The patch application caused a significant increase in the levels of collagen of the colon tissue as compared to the untreated and control groups (p < 0.001). No difference was found in the lymphocyte and macrophage inflammatory infiltrate between the groups. Our results suggest that patch treatment efficiently inhibits the diverticular pouch deformation and promotes the healing of the colon wall with a normal inflammatory response, which may minimize the risk of diverticulosis reoccurrence and complications over time

Effect of chitosan treatment (all in 2 mg/mL) on the levels of proinflammatory cytokines released from (A) naïve mouse peritoneal macrophages and (B) LPS-challenged macrophages.

<p>Cytokine levels are determined by Milliplex Multi-Analyte Profiling cytokine/chemokine panel. Media of the LPS-challenged macrophages were 10 times diluted prior to analysis. Graphs on the right are displayed in narrow y-scales. ZWC (An/Am = 0.7); C-Gt: chitosan glutamate; Gly-C: glycol chitosan. Data are expressed as averages with standard deviations of three repeated measurements. *: p<0.05; **: p<0.01; ***: p<0.001 vs PBS.</p

Viability of mouse peritoneal macrophages in the presence of ZWC (An/Am ratio = 0.7), chitosan glutamate (C-Gt) and glycol chitosan (Gly-C).

<p>Data are expressed as averages with standard deviations of three repeated measurements. *: p<0.05; **: p<0.01 vs PBS.</p

MIP-2 production from macrophages incubated with ZWC (2 mg/mL), LPS (1 µg/mL), a mixture of LPS (1 µg/mL) and ZWC (2 mg/mL) (LPS co-inc w ZWC), or LPS pre-incubated with ZWC (equivalent to 1 µg/mL LPS under an assumption that all the LPS remained in the supernatant; LPS pre-inc w ZWC).

<p>ELISA was performed on 10 times-diluted culture media. Data are expressed as averages with standard deviations of three repeated measurements. ***: p<0.001.</p

Chitosan precipitates (arrows) in the peritoneal cavity.

<p>Mice injected with chitosan glutamate intraperitoneally were examined 7 days after injection. (A) Chitosan precipitates found between the liver and the stomach. (B) Chitosan precipitates stuck on the spleen (top) and the liver (bottom). (C) Lobes of the liver were connected via chitosan residue.</p