Bioaccessibility and Digestibility of Proteins in Plant-Based Drinks and Cow's Milk: Antioxidant Potential of the Bioaccessible Fraction.

During the last years, a strong increase in the sales volume and consumption of plant-based drinks was observed, which were partly used as an alternative to cow's milk. As milk is a relevant protein source in many countries, we have investigated the protein bioaccessibility and digestibility of soy, almond, and oat drinks in comparison to milk using the tiny-TIMsg gastrointestinal model. The relative protein digestibility of all products was between 81% (soy drink) and 90% (milk). The digestible indispensable amino acid score (DIAAS) in vitro method was used to estimate the protein nutritional quality. The highest DIAAS values were obtained for milk in tryptophan (117%) and soy drink in sulfur containing amino acids (100%). Oat drink was limited in lysine (73%), almond drink in lysine (34%) and the sulfur containing amino acids (56%). Additionally, the antioxidant activity of the bioaccessible fractions was analyzed using Trolox equivalent antioxidative capacity and oxygen radical absorbance capacity assays, revealing a higher antioxidative potential of milk and soy drink compared to oat and almond drink.

Pomegranate (Punica granatum L.) Extract and Its Anthocyanin and Copigment Fractions-Free Radical Scavenging Activity and Influence on Cellular Oxidative Stress.

Secondary plant metabolites, e.g., polyphenols, are widely known as health-improving compounds that occur in natural functional foods such as pomegranates. While extracts generated from these fruits inhibit oxidative stress, the allocation of these effects to the different subgroups of substances, e.g., anthocyanins, "copigments" (polyphenols without anthocyanins), or polymeric compounds, is still unknown. Therefore, in the present study, polyphenols from pomegranate juice were extracted and separated into an anthocyanin and copigment fraction using adsorptive membrane chromatography. Phenolic compounds were determined by high performance liquid chromatography with photodiode array (HPLC-PDA) detection and HPLC-PDA electrospray ionization tandem mass spectrometry (HPLC-PDA-ESI-MS/MS), while the free radical scavenging activity of the pomegranate XAD­7 extract and its fractions was evaluated by the Trolox equivalent antioxidant capacity (TEAC) assay and electron spin resonance (ESR) spectroscopy. Compared to juice, the total phenolic content and free radical scavenging potential was significantly higher in the pomegranate XAD-7 extract and its fractions. In comparison to the anthocyanin and copigment fraction, pomegranate XAD-7 extract showed the highest radical scavenging activity against galvinoxyl and DPPH radicals. Moreover, the enriched XAD-7 extract and its fractions were able to protect human hepatocellular HepG2 cells against oxidative stress induced by hydrogen peroxide. Overall, these results indicated that anthocyanins and copigments act together in reducing oxidative stress.

Synthesis and in vitro characterization of the genotoxic, mutagenic and cell-transforming potential of nitrosylated heme.

Data from epidemiological studies suggest that consumption of red and processed meat is a factor contributing to colorectal carcinogenesis. Red meat contains high amounts of heme, which in turn can be converted to its nitrosylated form, NO-heme, when adding nitrite-containing curing salt to meat. NO-heme might contribute to colorectal cancer formation by causing gene mutations and could thereby be responsible for the association of (processed) red meat consumption with intestinal cancer. Up to now, neither in vitro nor in vivo studies characterizing the mutagenic and cell transforming potential of NO-heme have been published due to the fact that the pure compound is not readily available. Therefore, in the present study, an already existing synthesis protocol was modified to yield, for the first time, purified NO-heme. Thereafter, newly synthesized NO-heme was chemically characterized and used in various in vitro approaches at dietary concentrations to determine whether it can lead to DNA damage and malignant cell transformation. While NO-heme led to a significant dose-dependent increase in the number of DNA strand breaks in the comet assay and was mutagenic in the HPRT assay, this compound tested negative in the Ames test and failed to induce malignant cell transformation in the BALB/c 3T3 cell transformation assay. Interestingly, the non-nitrosylated heme control showed similar effects, but was additionally able to induce malignant transformation in BALB/c 3T3 murine fibroblasts. Taken together, these results suggest that it is the heme molecule rather than the NO moiety which is involved in driving red meat-associated carcinogenesis.

Impact of defined thermomechanical treatment on the structure and content of dietary fiber and the stability and bioaccessibility of polyphenols of chokeberry (Aronia melanocarpa) pomace.

Dietary fiber is a potential replacement for other ingredients such as starch in reformulated extruded breakfast cereals. Analysis of chokeberry pomace powder revealed a total dietary fiber content of 57.8 ± 2 g/100 g with 76% being insoluble, 20% high molecular soluble and 4% low molecular soluble dietary fiber. The fiber polysaccharide composition was analyzed in detail by using a variety of analytical approaches. Extrusion-like processing conditions were studies in a Closed Cavity Rheometer enabling the application of defined thermal (temperature range 100-160 °C) and mechanical treatments (shear rates between 0.1 s-1 and 50 s-1) to chokeberry pomace powder. Application of temperatures up to 140 °C irrespective of the mechanical treatment does not remarkably alter dietary fiber structure or content, but reduces the initial content of total polyphenols by about 40% to a final content of 3.3 ± 0.5 g/100 g including 0.63 ± 0.1 g/100 g of anthocyanins, 0.18 ± 0.02 g/100 g of phenolic acids and 0.090 ± 0.007 g/100 g of flavonols, respectively. The retained polyphenols are fully bioaccessible after in vitro digestion, and antioxidant capacity remains unchanged as compared to the untreated pomace powder. Glucose bioaccessibility remains unaffected, whereas glucose content is reduced. It is concluded that chokeberry pomace powder is a good source of dietary fiber with the potential to partially substitute starch in extruded breakfast cereals.

Photoautotrophically Grown Chlorella vulgaris Shows Genotoxic Potential but No Apoptotic Effect in Epithelial Cells.

This study investigated the effect of Chlorella vulgaris (C. vulgaris) on genotoxicity, cytotoxicity, and apoptosis in Caco-2 and HT-29 cells. C. vulgaris significantly induced DNA damage in both cell lines at a concentration of 200 µg dry matter/mL (comet tail intensity CTI: 24.6 ± 4.7% for Caco-2, 16.6 ± 0.9% for HT-29). The application of processing (sonication, ball-milling) did not affect the genotoxicity negatively and lowered the lipid peroxidation in C. vulgaris preparations. C. vulgaris-induced intracellular formation of reactive oxygen species in human cell lines and might be responsible for the genotoxic effect. A solid fraction mainly triggered the observed DNA damage (CTI: 41.5 ± 1.9%), whereas a hydrophilic (CTI: 7.9 ± 1.7%) and lipophilic (CTI: 10.2 ± 2.1%) fraction revealed a significantly lower tail intensity. C. vulgaris significantly induced DNA damage in both cell lines possibly through intracellular formation of reactive oxygen species; however, it was repaired after a 2 h recovery time or was even avoided at lower concentrations. In addition, none of the preparations indicated an adverse effect on cell proliferation or revealed apoptotic activity.

A Lipophilic Fucoxanthin-Rich Phaeodactylum tricornutum Extract Ameliorates Effects of Diet-Induced Obesity in C57BL/6J Mice.

Phaeodactylum tricornutum (P. tricornutum) comprise several lipophilic constituents with proposed anti-obesity and anti-diabetic properties. We investigated the effect of an ethanolic P. tricornutum extract (PTE) on energy metabolism in obesity-prone mice fed a high fat diet (HFD). Six- to eight-week-old male C57BL/6J mice were switched to HFD and, at the same time, received orally placebo or PTE (100 mg or 300 mg/kg body weight/day). Body weight, body composition, and food intake were monitored. After 26 days, blood and tissue samples were collected for biochemical, morphological, and gene expression analyses. PTE-supplemented mice accumulated fucoxanthin metabolites in adipose tissues and attained lower body weight gain, body fat content, weight of white adipose tissue (WAT) depots, and inguinal WAT adipocyte size than controls, independent of decreased food intake. PTE supplementation was associated with lower expression of Mest (a marker of fat tissue expandability) in WAT depots, lower gene expression related to lipid uptake and turnover in visceral WAT, increased expression of genes key to fatty acid oxidation and thermogenesis (Cpt1, Ucp1) in subcutaneous WAT, and signs of thermogenic activation including enhanced UCP1 protein in interscapular brown adipose tissue. In conclusion, these data show the potential of PTE to ameliorate HFD-induced obesity in vivo.

Particle size of milled chokeberry pomace did not influence in vitro cellular absorption and transport efficiencies of anthocyanins, phenolic acids and flavonols.

Industrial chokeberry pomace is very rich in polyphenols. The main focus here lies on the possible relationship between the particle size of chokeberry milled pomace and an enhanced absorption and transport of polyphenols by Caco-2 cells. Wet milling was used to produce materials with particle size distributions in the micrometre and in the sub-micrometre to nanometre ranges starting from chokeberry pomace. Milled materials with about 50% of the particles with a mean size (x50,3) of 223 ± 13 µm (coarse milling) and about 90% of the particles with x50,3 of 160 ± 40 nm (fine milling, sonication) were obtained. None of the milled materials exhibited cytotoxic effects within the tested concentration-ranges. The polyphenol absorption and the transport efficiencies from the fine and the coarse milled materials were similar. Thus, no effect of the particle size upon cellular uptake and transport could be established, but agglomeration of particle during incubation cannot be excluded as the cause. Furthermore, based on polyphenol stability we postulate that direct milling may be applied to valorise the processing by-product from commercial fruit juice production.

Effect of sonication on bioaccessibility and cellular uptake of carotenoids from preparations of photoautotrophic Phaeodactylum tricornutum.

With regard to its cost-effective cultivation and the composition of high-value nutrients, the diatom Phaeodactylum tricornutum (P. tricornutum) attracts interest for the use in human nutrition. Besides a number of important nutrients, it is rich in carotenoids. Therefore, this study aimed to investigate the potential of P. tricornutum as a carotenoid source for human nutrition. In photoautotrophically produced P. tricornutum biomass the carotenoid constitution, bioaccessibility (in vitro digestion model) and cellular uptake in differentiated Caco-2 cells (Transwell model system) was determined. Furthermore, the influence of sonication on these parameters was investigated. The results indicate that ß-carotene, zeaxanthin and fucoxanthin were the main carotenoids found in P. tricornutum. Moreover, these carotenoids showed a good bioaccessibility (ß-carotene: 25%, zeaxanthin: 27%, fucoxanthin: 57%), which is further improved by sonication for ß-carotene and fucoxanthin. In line with the good bioaccessibility, fucoxanthin was the most abundant carotenoid in Caco-2 cells followed by zeaxanthin. In contrast, ß-carotene could not be detected in the cells. The present study demonstrated that P. tricornutum represents a good source of carotenoids, particularly fucoxanthin. Thus, this diatom can contribute to the intake of bioaccessible carotenoids, even without processing. In addition, sonication might be a useful tool to improve the carotenoid bioaccessibility.

Isolation and Characterization of Lactic Acid Bacteria from Fermented Goat Milk in Tajikistan.

The lactobacilli associated with a fermented goat milk product from Tajikistan were isolated to characterize their technological properties and antibiotic resistances in order to assess their suitability for development as starter cultures. In this study, twenty three strains were identified by 16S rRNA sequencing as typical dairy-associated lactic acid bacterial strains, i.e. L. plantarum, L. pentosus, L. delbrueckii, L. helveticus and L. paracasei. These strains were generally susceptible to most antibiotics tested in this study and this allowed a selection of strains as safe starters. The draft genomes of four representative strains were sequenced and the number of contigs of the four assembled genomes ranged from 51 to 245 and the genome sizes ranged from 1.75 to 3.24 Mbp. These representative strains showed differences in their growth behavior and pH-reducing abilities in in vitro studies. The co-inoculation of these Lactobacillus spp. strains together with a yeast Kluyveromyces marxianus MBT-5698, or together with the yeast and an additional Streptococcus thermophilus MBT-2, led to a pH reduction to 3.4 after 48 h. Only in the case of fermentation inoculated with the co-culture, the viscosity of the milk increased noticeably. In contrast, fermentations with single strains did not lead to gelation of the milk or to a decrease in the pH after 24h. The results of this study provide a comprehensive understanding of the predominant lactobacilli related to Tajikistani fermented milk products.

Bioavailability and Safety of Nutrients from the Microalgae Chlorella vulgaris, Nannochloropsis oceanica and Phaeodactylum tricornutum in C57BL/6 Mice.

Microalgae are rich in macronutrients and therefore, they have been proposed as a potential future food source preserving natural resources. Here, we studied safety and bioavailability of algae nutrients in mice. Three microalgae species, Chlorella vulgaris, Nannochloropsis oceanica and Phaeodactylum tricornutum, were studied after ball mill disruption at different doses (5%, 15% and 25% dry weight) for 14 days. In response to all three algae diets, we observed a weight gain similar or superior to that in response to the control diet. No substantial differences in organ weights nor gut length occurred. Protein bioavailability from the algae diets did not differ from the control diet ranging from 58% to 77% apparent biological value. Fat absorption was lower for microalgae compared to soy oil in control diets, albeit still substantial. High liver eicosapentaenoic acid levels were measured following feeding with N. oceanica, the algae richest in omega-3 fatty acids. Neither histological nor serum analyses revealed any heart, kidney or liver toxicity induced by any of the algae diets. Algae-rich diets were thus well accepted, well tolerated and suitable for the maintenance of body weight and normal organ function. No toxicological effects were observed.

Dephosphorylation of myo-inositol phosphates in the in vitro intestinal Caco-2 cell model.

Plant and microbial phytases present in raw materials can cause a dephosphorylation of phytate (myo-inositol hexakisphosphate) (InsP6)) during food processing resulting in a broad range of different myo-inositol phosphates such as pentakisphosphate (InsP5) and tetrakisphosphate (InsP4) in foods. Here, we investigated whether the human intestinal epithelium is able to dephosphorylate myo-inositol phosphates (InsP6, InsP5-, InsP4-, InsP3-isomers) using an in vitro model with differentiated human Caco-2 cells cultured on semipermeable inserts. Incubation of InsP6 and an InsP5-isomer with cells for 3 h showed no dephosphorylation of both InsPs. Treatment of cells with a mixture of different InsP4-isomers, however, caused a formation of about 3.5% of an InsP3-isomer (Ins(1,5,6)P3) and treatment with a mixture of different InsP3-isomers caused about 20% formation of InsP2-isomers, respectively. Thus, human intestinal cells can contribute to the dephosphorylation of myo-inositol phosphates of partly dephosphorylated forms such as InsP3 and InsP4.

Quantification of Slackia and Eggerthella spp. in Human Feces and Adhesion of Representatives Strains to Caco-2 Cells.

Eggerthella and Slackia spp. are gut associated bacteria that have been suggested to play roles in host lipid and xenobiotic metabolism. A quantitative PCR method for the selective enumeration of bacteria belonging to either the genus Eggerthella or Slackia was developed in order to establish the numbers of these bacteria occurring in human feces. The primers developed for selective amplification of these genera were tested first in conventional PCR to test for their specificity. Representative species of Eggerthella and Slackia, as well as closely related genera of the Coriobacteriia, were included in the investigation. The selected primers were shown to be capable of specific amplification of species of the genera Eggerthella and Slackia, but not all species of the genera may be amplified by the respective primers. Their use in qPCR experiments to assess the levels of Slackia equolifaciens and Eggerthella lenta in the feces of 19 human volunteers showed they occurred at mean counts of 7 × 10(5) and 3.1 × 10(5) CFU/g for Eggerthella spp. and Slackia spp., respectively. Electron microscopy investigations showed that while E. lenta cells exhibited slender and very regular shaped rods, Slackia cells showed a remarkably pleomorphic phenotype. Both species did not appear to have fimbriae or pili. Some S. equolifaciens cells showed a characteristic "ribbon" of presumably extracellular material around the cells, particularly at the areas of cell division. The two species also differed markedly in their adhesion behavior to Caco-2 cells in cell culture, as E. lenta DSMZ 15644 showed a high adhesion capacity of 74.2% adherence of the bacterial cells added to Caco-2 cells, while S. equolifaciens DSM 24851(T) on the other hand showed only low adhesion capability, as 6.1% of bacterial cells remained bound. Speculatively, this may imply that the ecological compartments where these bacteria reside in the gut may be different, i.e., E. lenta may be associated more with the gut wall, while Slackia may be free living in the lumen.

Ultra high pressure homogenization of almond milk: Physico-chemical and physiological effects.

Ultra high pressure homogenization (UHPH) of food is a processing technology to improve food safety and shelf life. However, despite very short treatment duration UHPH may lead to changes in chemical and physico-chemical properties including formation of submicro-/nano-particles. This may affect the physiological or toxicological properties of the treated food. Here, we treated raw almond milk (AMr) with UHPH at 350 MPa and 85 °C (AMuhph), known able to inactivate food relevant microorganisms. UHPH-treatment led to about a threefold increase of the mean particle size. There was a nearly complete loss of antigenicity investigated by ELISA for determination of traces of almond proteins. The content of vitamins B1 and B2 remained unchanged, while free exposed sulfhydryl groups decreased. Despite of observed modifications, UHPH-treatment of almond milk did not cause any changes in cyto- or genotoxic effects and antigenotoxic capability of protecting intestinal cells against iron induced DNA damage in vitro.

Bioaccessibility of carotenoids from Chlorella vulgaris and Chlamydomonas reinhardtii.

Microalgae can contribute to a balanced diet because of their composition. Beside numerous essential nutrients, carotenoids are in the focus for food applications. The bioavailability of carotenoids from photoautotrophic-cultivated Chlorella vulgaris (C. vulgaris) and Chlamydomonas reinhardtii (C. reinhardtii) was compared. An in vitro digestion model was used to investigate carotenoid bioaccessibility. Furthermore, the effect of sonication on bioaccessibility was assessed. Lutein was the main carotenoid in both species. C. reinhardtii showed higher amounts of lutein and ß-carotene than C. vulgaris. In contrast to C. reinhardtii, no ß-carotene and only 7% of lutein were bioaccessible in nonsonicated C. vulgaris. Sonication increased the bioaccessibility of carotenoids from C. vulgaris to a level comparable with C. reinhardtii (ß-carotene: ≥ 10%; lutein: ≥ 15%). Thus, C. reinhardtii represents a good carotenoid source for potential use in foods without processing, while the application of processing methods, like sonication, is necessary for C. vulgaris.

Cyto-genotoxic and oxidative effects of a continuous UV-C treatment of liquid egg products.

UV-C treatment of food is a promising non-thermal processing technology to improve food safety and preservation. Most of the chemical constituents of food absorb UV-C light that can lead to chemical modifications and quality changes. This work investigated the effects of UV-C treatment of liquid egg products on lipid, protein oxidations and potential cyto- and genotoxic effects on intestinal epithelial cells in vitro. Egg preparations (egg white, yolk, liquid whole egg) were treated with UV-C (254 nm, volumetric doses between 0 and 115,619 J L(-1)) using a commercial UV-C processing unit equipped with a Dean Flow reactor. UV-C treatment at high doses (from 32,181 J L(-1), about 2 times higher than that needed to inactivate 5 log of relevant microorganisms) showed an increased lipid oxidation in egg yolk and slight effects in liquid whole eggs; this was confirmed by slightly but not statistically significant increased peroxide values. UV-C induced also slight protein damage, characterised by the total sulfhydryl group reduction. These UV-C-induced oxidative modifications in egg preparations however did not cause any increase in the cyto- or genotoxic (DNA strand breaks) effects in intestinal Caco-2 cells.

Anthocyanins suppress the cleavable complex formation by irinotecan and diminish its DNA-strand-breaking activity in the colon of Wistar rats.

In the present study, the question was addressed whether anthocyanins interfere with the topoisomerase I poison irinotecan in vivo. In vivo complexes of enzyme to DNA bioassay was used to detect irinotecan-induced stabilization of topoisomerase I/DNA complexes and single cell gel electrophoresis to determine DNA-strand-break induction in the colon of male Wistar rats. Furthermore, analysis of anthocyanin concentrations in rat plasma and rat colon was included in the testing, demonstrating that anthocyanins reach the colon and the concentrations do not differ between rats that only received anthocyanins and the anthocyanin/irinotecan group. Blackberry extract was found to significantly reduce irinotecan-mediated topoisomerase I/DNA cleavable complex formation. Overall, anthocyanins did not notably increase cleavable complex formation. However, a significant increase of DNA damage was shown after a single dose of irinotecan as well as the single compounds cyanidin (cy) and cyanidin-3-glucoside (cy-3-g). Furthermore, a significant reduction of irinotecan-induced DNA-strand breaks after a pretreatment with cy, cy-3-g and blackberry extract was observed. Thus, the question arises whether anthocyanin-rich preparations might interfere with chemotherapy or whether, due to low systemic bioavailability, the preparations might provide protective potential in the gastrointestinal tract.

Accelerated aging phenotype in mice with conditional deficiency for mitochondrial superoxide dismutase in the connective tissue.

The free radical theory of aging postulates that the production of mitochondrial reactive oxygen species is the major determinant of aging and lifespan. Its role in aging of the connective tissue has not yet been established, even though the incidence of aging-related disorders in connective tissue-rich organs is high, causing major disability in the elderly. We have now addressed this question experimentally by creating mice with conditional deficiency of the mitochondrial manganese superoxide dismutase in fibroblasts and other mesenchyme-derived cells of connective tissues in all organs. Here, we have shown for the first time that the connective tissue-specific lack of superoxide anion detoxification in the mitochondria results in reduced lifespan and premature onset of aging-related phenotypes such as weight loss, skin atrophy, kyphosis (curvature of the spine), osteoporosis and muscle degeneration in mutant mice. Increase in p16(INK4a) , a robust in vivo marker for fibroblast aging, may contribute to the observed phenotype. This novel model is particularly suited to decipher the underlying mechanisms and to develop hopefully novel connective tissue-specific anti-aging strategies.

In vivo bioassay to detect irinotecan-stabilized DNA/topoisomerase I complexes in rats.

Irinotecan is an anticancer agent that stabilizes topoisomerase I/DNA complexes. So far, no test system has been reported for directly determining irinotecan-induced stabilization of topoisomerase I/DNA complexes in organs in vivo. We adapted an 'in vivo complexes of enzyme to DNA' (ICE) bioassay to assess irinotecan activity in the stomach, duodenum, colon and liver of male Wistar rats after a single treatment with irinotecan (100 mg/kg body weight, intraperitoneally). This was compared to the control group receiving 0.9% sodium chloride intraperitoneally. In addition, the DNA strand breaking properties of irinotecan were measured in mucosal cells from the distal colon by single-cell gel electrophoresis (comet assay) to investigate the association of topoisomerase poisoning and DNA damage in vivo. A single dose of irinotecan significantly increased amounts of topoisomerase I covalently bound to DNA in stomach, duodenum, colon and liver. Concomitantly, the irinotecan-treated group showed significantly higher amounts of DNA strand breaks in colon mucosa cells compared to the control group. The ICE bioassay and the comet assay represent two test systems for investigating the impact of topoisomerase I poisons on DNA integrity in colon tissues of Wistar rats.

Prevention of colon carcinogenesis by apple juice in vivo: impact of juice constituents and obesity.

It is estimated that 75-85% of all chronic diseases are linked to lifestyle-related and environmental factors. The development of colon cancer is positively associated with obesity and inversely associated with the intake of dietary fibre, fruit and vegetable. Apple juice is the most widely consumed fruit beverage in Germany. It contains a specific spectrum of polyphenols and other components that may reduce the risk of colon cancer. Epidemiologic studies suggest an inverse correlation between apple consumption and colon cancer risk, although the mechanisms for these observations are not clear. The present review summarizes the preventive potential of apple juices and different apple constituents on biomarkers related to colon carcinogenesis with special focus on the in vivo evidence and the cancer promoting condition of obesity. However, under the cancer promoting condition of obesity, apple juice did not show cancer-preventive bioactivity. In our experiments a cancer-preventive bioactivity of apple juice is lacking in rats under the cancer-promoting condition of obesity. To further investigate, whether this lack of efficacy observed in obese rats might be representative for obese individuals human intervention studies on high risk groups such as obese or diabetic individuals are of interest and will be conducted.

Overexpression of manganese superoxide dismutase in human dermal fibroblasts enhances the contraction of free floating collagen lattice: implications for ageing and hyperplastic scar formation.

Cell-matrix interactions are of significant importance for tissue homeostasis of the skin and, if disturbed, may lead to ageing and hyperplastic scar formation. We have studied fibroblasts stably overexpressing manganese superoxide dismutase (MnSOD) with a defined capacity for the removal of superoxide anions and concomitant accumulation of hydrogen peroxide to evaluate the role of enhanced MnSOD activity on the dynamics of cell-matrix interactions in the three-dimensional collagen lattice contraction assay. MnSOD overexpressing fibroblast populated collagen lattices revealed a significantly enhanced contraction compared to collagen lattices populated with vector control cells. The enhanced collagen lattice contraction was in part due to an increase in active TGF-beta1 and the accumulation of H2O2 in MnSOD overexpressing fibroblasts populated collagen lattices. Inhibition of TGF-beta1 signalling by the ALK4,5,7 kinases' inhibitor SB431542 at least partly inhibited the enhanced collagen lattice contraction of MnSOD overexpressing fibroblasts populated lattices. In addition, supplementation of vector control fibroblast populated collagen lattices with recombinant TGF-beta1 concentration dependently enhanced the collagen lattice contraction. In the presence of the antioxidant Ebselen, a mimic of H2O2 and other hydroperoxides/peroxynitrite-detoxifying glutathione peroxidase, collagen lattice contraction and the activation of TGF-beta1 were significantly reduced in collagen lattices populated with MnSOD overexpressing fibroblasts. Collectively, these data suggest that H2O2 or other hydroperoxides or peroxynitrite or a combination thereof may function as important second messengers in collagen lattice contraction and act at least in part via TGF-beta1 activation.