Impact of antithrombin Ⅲ on hepatic and intestinal microcirculation in experimental liver cirrhosis and bowel inflammation: An in vivo analysis

AIM： To analyze the hepatic and intestinal microcirculation in an animal model of liver cirrhosis and inflammatory bowel disease （IBD） and to characterize bhe anti-inflammatory action of antithrombin Ⅲ （ATⅢ） on leukocyte kinetics and liver damage. METHODS： Hepatic and intestinal microcirculation was investigated by intravital videomicroscopy. Standardized models of experimental chronic liver cirrhosis and bowel inflammation were employed. Animals were divided into four groups （n = 6/group）： controls, animals with cirrhosis, animals with cirrhosis and IBD, animals with cirrhosis and IBD treated with ATIII. RESULTS： Cirrhosis facilitated leukocyte rolling and sticking in hepatic sinusoids （1.91±0.28 sticker/μm vs 0.5±0.5 sticker/μm in controls, P〈0.05）. The effect enhanced in animals with cirrhosis and IBD （5.4±1.65 sticker/μm）, but reversed after ATIII application （3.97±1.04 sticker/μm, P〈0.05）. Mucosal blood flow showed no differences in cirrhotic animals and controls （5.3±0.31 nL/min vs5.4±0.25 nL/min） and was attenuated in animals wibh cirrhosis and IBD significantly （3.49±0.6 nL/min）. This effect was normalized in the treatment group （5.13±0.4 nL/min, P〈0.05）. Enzyme values rose during development of cirrhosis and bowel inflammation, and reduced after ATIII application （P〈0.05）. CONCLUSION： Liver cirrhosis in the presence of IBD leads to a significant reduction in mucosal blood flow and an increase in hepatic leukocyte adherence with consecutive liver injury, which can be prevented by administration of ATⅢ.

Recent advances in basic and clinical aspects of inflammatory bowel disease: Which steps in the mucosal inflammation should we block for the treatment of inflammatory bowel disease?

There are four steps in the interaction between intestinal microbes and mucosal inflammation in genetically predisposed individuals from the viewpoints of basic and clinical aspects of inflammatory bowel disease （IBD）. The first step is an interaction between intestinal microbes or their components and intestinal epithelial cells via receptors, the second step an interaction between macrophages and dendritic cells and mucosal lymphocytes, the third step an interaction between lymphocytes and vascular endothelial cells, and the fourth step an interaction between lymphocytes and granulocytes producing proinflammatory cytokines or free radicals and mucosal damage and repair. Recent therapeutic approaches for IBD aim to block these four steps in the intestinal inflammation of patients with IBD.

Fecal calprotectin in coeliac disease

We would like to share with the readers the results of our experience in 50 celiac disease (CD) patients, enrolled between September 2012 and April 2013, who were referred to our third-level CD Unit. The fecal calprotectin (FC) concentration of 50 adults with newly diagnosed CD was compared to that of a control group of 50 healthy subjects. FC level was determined by enzyme linked immunosorbent assay with diagnostic cut-off of 75 μg/g. In addition, we tried to correlate the FC level with symptoms, histological severity of CD (Marsh grade) and level of tissue transglutaminase antibodies (aTg) in CD patients. Finally, FC level was increased in five CD patients and in four controls (10% vs 8%, P = NS); mean FC concentration of patients and controls were 57.7 (SD ± 29.1) and 45.1 (SD ± 38.4) respectively. Furthermore, no significant correlation was seen between FC levels and symptoms/Marsh grade/aTg. The five CD patients did not show inflammatory lesions (e.g., ulcers, erosions) at upper endoscopy. The four healthy controls with positive FC were followed-up for further six months; in this observational period they did not show clinical signs of any underlying disease. On these bases, we think that FC is not able to investigate the subclinical inflammatory changes of active CD and FC should be considered a useless tool in the diagnostic work-up of uncomplicated CD but it should be accompanied by aTg when ruling out organic disease in patients with irritable bowel syndrome.

Anti-inflammatory effects of Lacto-Wolfberry in a mouse model of experimental colitis

AIM: To investigate the anti-inflammatory properties of Lacto-Wolfberry (LWB), bothin vitro and using a mouse model of experimental colitis. METHODS: The effects of LWB on lipopolysaccharide (LPS)-induced reactive oxygen species (ROS) and interleukin (IL)-6 secretion were assessed in a murine macrophage cell line. in vitro assessment also included characterizing the effects of LWB on the activation of NF-E2 related 2 pathway and inhibition of tumor necrosis factor-α (TNF-α)-induced nuclear factor-κB (NFκB) activation, utilizing reporter cell lines. Following the in vitro assessment, the anti-inflammatory efficacy of an oral intervention with LWB was tested in vivo using a preclinical model of intestinal inflammation. Multiple outcomes including body weight, intestinal histology, colonic cytokine levels and anti-oxidative measures were investigated.RESULTS: LWB reduced the LPS-mediated inductionof ROS production [+LPS vs 1% LWB + LPS, 1590 ± 188.5 relative luminescence units (RLU) vs 389 ± 5.9 RLU, P < 0.001]. LWB was more effective than wolfberry alone in reducing LPS-induced IL-6 secretion in vitro (wolfberry vs 0.5% LWB, 15% ± 7.8% vs 64% ± 5%, P < 0.001). In addition, LWB increased reporter gene expression via the anti-oxidant response element activation (wolfberry vs LWB, 73% ± 6.9% vs 148% ± 28.3%, P < 0.001) and inhibited the TNF-α-induced activation of the NF-κB pathway (milk vs LWB, 10% ± 6.7% vs 35% ± 3.3%, P < 0.05). Furthermore, oral supplementation with LWB resulted in a reduction of macroscopic (-LWB vs +LWB, 5.39 ± 0.61 vs 3.66 ± 0.59, P = 0.0445) and histological scores (-LWB vs +LWB, 5.44 ± 0.32 vs 3.66 ± 0.59, P = 0.0087) in colitic mice. These effects were associated with a significant decrease in levels of inflammatory cytokines such as IL-1β (-LWB vs +LWB, 570 ± 245 μg/L vs 89 ± 38 μg/L, P = 0.0106), keratinocyte-derived chemokine/growth regulated protein-α (-LWB vs +LWB, 184 ± 49 μg/Lvs 75 ± 20 μg/L,P = 0.0244), IL-6 (-LWBvs +LWB, 318 ± 99 μg/L vs 117 ± 18 μg/L, P = 0.0315) and other pro-inflammatory proteins such as cyclooxygenase-2 (-LWB vs +LWB, 0.95 ± 0.12 AU vs 0.36 ± 0.11 AU, P = 0.0036) and phosphorylated signal transducer and activator of transcription-3 (-LWB vs +LWB, 0.51 ± 0.15 AU vs 0.1 ± 0.04 AU, P = 0.057). Moreover, antioxidant biomarkers, including expression of gene encoding for the glutathione peroxidase, in the colon and the plasma anti-oxidant capacity were significantly increased by supplementation with LWB (-LWB vs +LWB, 1.2 ± 0.21 mmol/L vs 2.1 ± 0.19 mmol/L, P = 0.0095).CONCLUSION: These results demonstrate the antiinflammatory properties of LWB and suggest that the underlying mechanism is at least in part due to NF-κB inhibition and improved anti-oxidative capacity.