In vivo study of the biocompatibility of a novel compressed collagen hydrogel scaffold for artificial corneas

The experiments were designed to evaluate the biocompatibility of a plastically compressed collagen scaffold (PCCS). The ultrastructure of the PCCS was observed via scanning electron microscopy. Twenty New Zealand white rabbits were randomly divided into experimental and control groups that received corneal pocket transplantation with PCCS and an amniotic membrane, respectively. And the contralateral eye of the implanted rabbit served as the normal group. On the 1st, 7th, 14th, 21st, 30th, 60th, 90th, and 120th postoperative day, the eyes were observed via a slit lamp. On the 120th postoperative day, the rabbit eyes were enucleated to examine the tissue compatibility of the implanted stroma. The PCCS was white and translucent. The scanning electron microscopy results showed that fibers within the PCCS were densely packed and evenly arranged. No edema, inflammation, or neovascularization was observed on ocular surface under a slit lamp and few lymphocytes were observed in the stroma of rabbit cornea after histological study. In conclusion, the PCCS has extremely high biocompatibility and is a promising corneal scaffold for an artificial cornea. (c) 2013 Wiley Periodicals, Inc. J Biomed Mater Res Part A, 2013.

Aspects of in vitro and in vivo research approaches directed towards identifying probiotics and prebiotics for human use

The microbiota of the human gastrointestinal tract plays a key role in nutrition and health. Through the process of fermentation, gut bacteria metabolize various substrates (principally dietary components) to end products such as short-chain fatty acids and gases. This anaerobic metabolism is thought to contribute positively toward host daily energy requirements. However, under certain circumstances, the fermentative process may produce undesirable metabolites. This may cause the onset of gut disorders that can be manifest through both acute and chronic conditions. Moreover, the gut flora may become contaminated by transient pathogens that serve further to upset the normal community structure. There has been a recent increase in the use of dietary components that help to maintain, or even improve, the gut microflora "balance." Probiotics are live microbial feed supplements added to appropriate food vehicles (usually fermented milks), whereas prebiotics are dietary carbohydrates that have a selective metabolism in the colon and serve to increase numbers of bacteria seen as desirable. Because of their purported health-promoting properties, lactic acid-producing bacteria, including bifidobacteria, are the usual target organisms. The market value and biological potential of both approaches are enormous. This article will summarize how efficacious types can be identified.

Greater impact of dietary fat manipulation than apolipoprotein E genotype on ex-vivo cytokine production – insights from the SATgenε study

Apolipoprotein E (APOE) genotype is believed to play an important role in cardiovascular risk. APOE4 carriers have been associated with higher blood lipid levels and a more pro-inflammatory state compared with APOE3/E3 individuals. Although dietary fat composition has been considered to modulate the inflammatory state in humans, very little is known about how APOE genotype can impact on this response. In a follow-up to the main SATgene study, we aimed to explore the effects of APOE genotype, as well as, dietary fat manipulation on ex vivo cytokine production. Blood samples were collected from a subset of SATgene participants (n = 52/88), prospectively recruited according to APOE genotype (n = 26 E3/E3 and n = 26 E3/E4) after low-fat (LF), high saturated fat (HSF) and HSF with 3.45 g docosahexaenoic acid (DHA) dietary periods (each diet eight weeks in duration assigned in the same order) for the measurement of ex vivo cytokine production using whole blood culture (WBC). Concentrations of IL-1beta, IL-6, IL-8, IL-10 and TNF-alpha were measured in WBC supernatant samples after stimulation for 24 h with either 0.05 or 1 lg/ml of bacterial lipopolysaccharide (LPS). Cytokine levels were not influenced by genotype, whereas, dietary fat manipulation had a significant impact on TNF-a and IL-10 production; TNF-a concentration was higher after consumption of the HSF diet compared with baseline and the LF diet (P < 0.05), whereas, IL-10 concentration was higher after the LF diet compared with baseline (P < 0.05). In conclusion, our study has revealed the amount and type of dietary fat can significantly modulate the production of TNF-a and IL-10 by ex vivo LPS-stimulated WBC samples obtained from normolipidaemic subjects.

In vitro and in vivo models of colorectal cancer: antigenotoxic activity of berries.

The etiology of colorectal cancer (CRC), a common cause of cancer-related mortality globally, has strong associations with diet. There is considerable epidemiological evidence that fruits and vegetables are associated with reduced risk of CRC. This paper reviews the extensive evidence, both from in vitro studies and animal models, that components of berry fruits can modulate biomarkers of DNA damage and that these effects may be potentially chemoprotective, given the likely role that oxidative damage plays in mutation rate and cancer risk. Human intervention trials with berries are generally consistent in indicating a capacity to significantly decrease oxidative damage to DNA, but represent limited evidence for anticarcinogenicity, relying as they do on surrogate risk markers. To understand the effects of berry consumption on colorectal cancer risk, future studies will need to be well controlled, with defined berry extracts, using suitable and clinically relevant end points and considering the importance of the gut microbiota.

Virgin olive oil phenolics extract inhibit invasion of HT115 human colon cancer cells in vitro and in vivo

The decreased cancer risk associated with consumption of olive oil may be due to the presence of phenolics which can modulate pathways including apoptosis and invasion that are relevant to carcinogenesis. We have previously shown that a virgin olive oil phenolics extract (OVP) inhibited invasion of HT115 colon cancer cells in vitro. In the current study we assessed the in vitro effects of OVP (25 μg mL(-1)) on HT115 cell migration, spreading and integrin expression. Furthermore, the anti-metastatic activity of OVP - at a dose equivalent to 25 mg per kg per day for 2, 8 or 10 weeks - was assessed in a Severe Combined ImmunoDeficiency (SCID) Balb-c mouse model. After 24 h OVP did not inhibit cell migration but significantly reduced cell spreading on fibronectin (65% of control; p < 0.05) and expression of a range of α and β integrins was modulated. In vivo, OVP by gavage significantly (p < 0.05) decreased not only tumour volume but also the number of metastases in SCID Balb-c mice. Collectively, the data suggest that - possibly through modulation of integrin expression - OVP decreases invasion in vitro and also inhibits metastasis in vivo.

Comparison of in vivo and in vitro digestion on polyphenol composition in lingonberries: potential impact on colonic health

The composition of polyphenols in ileal fluid samples obtained from an ileostomy subject after lingonberry intake was compared with lingonberry extracts obtained after simulated in vitro digestion (IVDL) and subsequent faecal fermentation (IVFL). HPLC-PDA-MS/MS analysis confirmed similar patterns of lingonberry (poly)phenolic metabolism after the in vivo and in vitro digestion, with reduced recovery of anthocyanins and a similar pattern of recovery for proanthocyanidins observed for both methods of digestion. On the other hand, the IVFL sample contained none of the original (poly)phenolic components but was enriched in simple aromatic components. Digested and fermented extracts exhibited significant (P < 0.05) anti-genotoxic (Comet assay), anti-mutagenic (Mutation Frequency assay), and anti-invasive (Matrigel Invasion assay) effects in human cell culture models of colorectal cancer at physiologically-relevant doses (0-50 μg/mL gallic acid equivalents). The ileal fluid induced significant anti-genotoxic activity (P < 0.05), but at a higher concentration (200 μg/mL gallic acid equivalents) than the IVDL. Despite extensive structural modification following digestion and fermentation, lingonberry extracts retained their bioactivity in vitro. This reinforces the need for studies to consider the impact of digestion when investigating bioactivity of dietary phytochemicals.

Efficacy of condensed tannins against larval Hymenolepis diminuta (Cestoda) in vitro and in the intermediate host Tenebrio molitor (Coleoptera) in vivo

Natural anti-parasitic compounds in plants such as condensed tannins (CT) have anthelmintic properties against a range of gastrointestinal nematodes, but for other helminths such effects are unexplored. The aim of this study was to assess the effects of CT from three different plant extracts in a model system employing the rat tapeworm, Hymenolepis diminuta, in its intermediate host, Tenebrio molitor. An in vitro study examined infectivity of H. diminuta cysticercoids (excystation success) isolated from infected beetles exposed to different concentrations of CT extracts from pine bark (PB) (Pinus sps), hazelnut pericarp (HN) (Corylus avellana) or white clover flowers (WC) (Trifolium repens), in comparison with the anthelmintic drug praziquantel (positive control). In the in vitro study, praziquantel and CT from all three plant extracts had dose-dependent inhibitory effects on cysticercoid excystation. The HN extract was most effective at inhibiting excystation, followed by PB and WC. An in vivo study was carried out on infected beetles (measured as cysticercoid establishment) fed different doses of PB, HN and praziquantel. There was a highly significant inhibitory effect of HN on cysticercoid development (p = 0.0002). Overall, CT showed a promising anti-cestodal effect against the metacestode stage of H. diminuta.

Nitrosopersulfide (SSNO-) targets soluble guanylyl cyclase and induces vasodilation in vivo

Background Recent experimental evidence suggests that nitric oxide (NO) and hydrogen sulfide signaling pathways are intimately intertwined particularly in the vasculature, with mutual attenuation or potentiation of biological responses under control of the soluble guanylyl cyclase (sGC) / phopshodiesterase (PDE) pathway. There is now compelling evidence that part of the NO/sulfide cross talk has a chemical foundation via the formation of S/N-hybrid molecules including thionitrous acid (HSNO) and nitrosopersulfde (SSNO-). The aim of this study was to characterize the bioactive products of the interaction between sulfide and NO metabolites targeting sGC that may potentially regulate vasodilation. Results We found that the chemical interaction of sulfide with NO or nitrosothiols leads to formation of S/N-hybrid metabolites including SSNO- via intermediate formation of HSNO. Contrary to a recent report in the literature but consistent with the transient nature of HSNO, its formation was not detectable by high-resolution mass spectrometry under physiologically relevant conditions. SSNO- is also formed in non-aqueous media by the reaction of nitrite with oxidized sulfur species including colloidal sulfur and polysulfides. SSNO- is stable in the presence of high concentrations of thiols, release NO, and activates sGC in RFL-6 cells in an NO-dependent fashion. Moreover, SSNO- is a potent vasodilator in aortic rings in vitro and lowers blood pressure in rats in vivo. The presence of high concentrations of SOD or thiols does not affect SSNO- mediated sGC activation, while it potentiates and inhibits the effects of the nitroxyl (HNO) donor Angeli's salt, suggesting that HNO release from SSNO- is not involved in sGC activation. Conclusion The reaction between NO and sulfide leads to fomation of S/N-hybrid molecules including SSNO-, releasing NO, activating sGC and inducing vasodilation. SSNO- is considerably more stable than HSNO at pH 7.4 and thus a more likely biological mediator that can account for the chemical cross-talk between NO and sulfide.

1,8-cineol potentiates IRF3-mediated antiviral response in human stem cells and an ex vivo model of rhinosinusitis

Common cold is one of the most frequent human inflammatory diseases caused by viruses and can facilitate bacterial super-infections resulting in sinusitis or pneumonia. The active ingredient of the drug Soledum, 1,8-cineole, is commonly applied for treating inflammatory diseases of the respiratory tract. However, the potential of 1,8-cineole for treating primary viral infections of the respiratory tract remains unclear. In the present study, we demonstrate for the first time that 1,8-cineole potentiates Poly(I:C)-induced activity of the anti-viral transcription factor Interferon Regulatory Factor 3, while simultaneously reducing pro-inflammatory NF-κB-activity in human cell lines, inferior turbinate stem cells (ITSCs) and ex vivo cultivated human nasal mucosa. Co-treatment of cell lines with Poly(I:C) and 1,8-cineole resulted in significantly increased IRF3 reporter gene activity compared to Poly(I:C) alone, whereas NF-κB-activity was reduced. Accordingly, 1,8-cineole- and Poly(I:C)-treatment led to increased nuclear translocation of IRF3 in ITSCs and a human ex vivo model of rhinosinusitis compared to the Poly(I:C)-treated approach. Nuclear translocation of IRF3 was significantly increased in ITSCs and slice cultures treated with LPS and 1,8-cineole compared to the LPS-treated cells mimicking bacterial infection. Our findings strongly suggest that 1,8-cineole potentiates the antiviral activity of IRF3 in addition to its inhibitory effect on pro-inflammatory NF-κB-signalling and may thus broaden its field of application.

Review of current in vivo measurement techniques for quantifying enteric methane emission from ruminants

Ruminant husbandry is a major source of anthropogenic greenhouse gases (GHG). Filling knowledge gaps and providing expert recommendation are important for defining future research priorities, improving methodologies and establishing science-based GHG mitigation solutions to government and non-governmental organisations, advisory/extension networks, and the ruminant livestock sector. The objectives of this review is to summarize published literature to provide a detailed assessment of the methodologies currently in use for measuring enteric methane (CH4) emission from individual animals under specific conditions, and give recommendations regarding their application. The methods described include respiration chambers and enclosures, sulphur hexafluoride tracer (SF6) technique, and techniques based on short-term measurements of gas concentrations in samples of exhaled air. This includes automated head chambers (e.g. the GreenFeed system), the use of carbon dioxide (CO2) as a marker, and (handheld) laser CH4 detection. Each of the techniques are compared and assessed on their capability and limitations, followed by methodology recommendations. It is concluded that there is no ‘one size fits all’ method for measuring CH4 emission by individual animals. Ultimately, the decision as to which method to use should be based on the experimental objectives and resources available. However, the need for high throughput methodology e.g. for screening large numbers of animals for genomic studies, does not justify the use of methods that are inaccurate. All CH4 measurement techniques are subject to experimental variation and random errors. Many sources of variation must be considered when measuring CH4 concentration in exhaled air samples without a quantitative or at least regular collection rate, or use of a marker to indicate (or adjust) for the proportion of exhaled CH4 sampled. Consideration of the number and timing of measurements relative to diurnal patterns of CH4 emission and respiratory exchange are important, as well as consideration of feeding patterns and associated patterns of rumen fermentation rate and other aspects of animal behaviour. Regardless of the method chosen, appropriate calibrations and recovery tests are required for both method establishment and routine operation. Successful and correct use of methods requires careful attention to detail, rigour, and routine self-assessment of the quality of the data they provide.