Background. Tissue glues can minimize treatment invasiveness, mitigate the risk of infection, and reduce surgery time; ergo, they have been developed and used in surgical procedures as wound closure devices beside sutures, staples, and metallic grafts. Regardless of their structure or function, tissue glues should show an acceptable microbial barrier function before being used in humans. This study proposes a novel in vitro method using Escherichia coli Lux and bioluminescence imaging technique to assess the microbial barrier function of tissue glues. Different volumes and concentrations of E. coli Lux were applied to precured or cured polyurethane-based tissue glue placed on agar plates. Plates were cultured for 1 h, 24 h, 48 h, and 72 h with bioluminescence signal measurement subsequently. Herein, protocol established a volume of 5 μL of a 1 : 100 dilution of E. coli Lux containing around 2 × 10 7  CFU/mL as optimal for testing polyurethane-based tissue glue. Measurement of OD600nm, determination of CFU/mL, and correlation with the bioluminescence measurement in p/s unit resulted in a good correlation between CFU/mL and p/s and demonstrated good reproducibility of our method. In addition, this in vitro method could show that the tested polyurethane-based tissue glue can provide a reasonable barrier against the microbial penetration and act as a bacterial barrier for up to 48 h with no penetration and up to 72 h with a low level of penetration through the material. Overall, we have established a novel, sensitive, and reproducible in vitro method using the bioluminescence imaging technique for testing the microbial barrier function of new tissue glues.

Purpose GLP-1 and GLP-2 (glucagon-like peptide-1/2) are gut hormones secreted in response to food. While GLP-1 controls glucose metabolism, GLP-2 is a local gut growth factor regulating intestinal nutrient absorption. GLP-2 has been found to be upregulated in patients with colitis. We hypothesize that beyond its local intestinal function GLP-2 might be involved in systemic immune responses. Methods and results To analyze whether GLP-2 secretion is modulated by the immune system, we measured circulating GLP-2 levels in 2 clinical cohorts. In the first cohort (n=34) GLP-2 levels increased over time following cardiac surgery as an inflammatory stimulus. In the second cohort 223 patients with sepsis had a 3.9 fold increase of GLP-2 plasma levels vs. 53 healthy controls (3.0 ng/mL vs. 11.4 ng/mL; p<0.001). High GLP-2 levels were associated with markers of inflammation (IL-6, PCT, CRP), septic cardiomyopathy (NT-proBNP) and independently predicted mortality in humans with sepsis. Induction of sepsis in mice by endotoxin or cecal ligation puncture strongly increased GLP-2 levels independent from food intake. By injecting various proinflammatory cytokines and inducing sepsis in IL1R−/− and IL6−/− mice we identified that inflammation upregulates GLP-2 secretion through IL-6. To identify the source of GLP-2 secretion under inflammation, we induced sepsis in Gcg−/− mice lacking endogenous GLP-2 production with a tissue-specific reactivation of Gcg in gut L-cells (GcgRAΔvilCre) or pancretic alpha cells (GcgRAΔPDX1-Cre). We observed sepsis-induced GLP-2 secretion to be derived from the pancreas and not from the gut. Additional in-vitro and ex-vivo approaches revealed that IL-6 directly activates GLP-2 secretion from pancreatic alpha cells. Gcg−/− mice lacking GLP-2 production and Glp2r−/− mice show aggravated sepsis indicating that endogenous upregulation of GLP-2 is protective. Finally, we analyzed whether inflammatory upregulation of GLP-2 has immunomodulatory relevance. We administered GLP-2 or saline as control per central jugular vein catheter mice who underwent CLP. GLP-2 treatment improved LV-contractility (dp/dtmax) in septic cardiomyopathy (control 7361 vs. GLP-2 9500 mmHg/s; p<0.01), inhibited sepsis-induced hypotension and reduced mortality (p=0.018). Mechanistically GLP-2 reduced myeloid immune cell infiltration into heart and liver tissue and decreased proinflammatory cytokine levels in various organs and the blood (TNF-α, IL-6 and IL-1β). After broad GLP-2 receptor profiling we found maximum mRNA expression in gut tissues with no expression on immune cells. By further mechanistic studies we found GLP-2 to protect against sepsis-induced gut barrier dysfunction. Conclusions Here we identified a counter-regulatory control system in which IL-6 derived upregulation of GLP-2 secretion limits excessive innate immune responses and protects against sepsis. These findings might open new avenues for the treatment of patients with inflammatory diseases. Funding Acknowledgement Type of funding sources: Foundation. Main funding source(s): European Foundation for the Study of Diabetes, European Research Area Network on Cardiovascular Diseases (ERA-CVD and BMBF), Deutsche Forschungsgemeinschaft (DFG)

Tissue adhesives as a physical barrier to microorganism penetration provide an alternative method with many advantages for wound closure in surgical settings compared to the clinical standard. This raises the need of developing and conducting in vitro methods that are sensitive and reproducible to assess their microbial barrier properties. In this study, three different polyurethane-based tissue adhesives with different physicochemical properties were evaluated in comparison to Dermabond® as a clinical gold standard for topical wound closure. Here, physicochemical properties varied in lactide concentration, viscosity, processing, and the full polymerization time. To evaluate the microbial barrier function, a 5 μl aliquot of E. coli Lux inoculum containing at least 1 × 10 9   CFU / ml was applied to the surface of each test adhesive and sterile filter paper as the control that was placed on an agar plate and incubated at 37°C. Plates were observed for bacterial growth (morphology), the adhesion of the adhesive/filter paper, and bioluminescence after 24, 48, and 72 hours. The data presented in this in vitro model indicated that polyurethane-based tissue adhesives with lactide concentration ≥ 5 % provided a suitable barrier against microbial penetration with 95% confidence of 99% efficacy for 72 h along with Dermabond®. Interestingly, the here described method was able to discriminate between the different physicochemical properties showing a better microbial barrier function with increasing lactide concentration of the adhesive. Overall, the results of this study showed the noninferiority between Dermabond® and the two abovementioned polyurethane-based tissue adhesives.