Deformable microlaser force sensing.

Mechanical forces are key regulators of cellular behavior and function, affecting many fundamental biological processes such as cell migration, embryogenesis, immunological responses, and pathological states. Specialized force sensors and imaging techniques have been developed to quantify these otherwise invisible forces in single cells and in vivo. However, current techniques rely heavily on high-resolution microscopy and do not allow interrogation of optically dense tissue, reducing their application to 2D cell cultures and highly transparent biological tissue. Here, we introduce DEFORM, deformable microlaser force sensing, a spectroscopic technique that detects sub-nanonewton forces with unprecedented spatio-temporal resolution. DEFORM is based on the spectral analysis of laser emission from dye-doped oil microdroplets and uses the force-induced lifting of laser mode degeneracy in these droplets to detect nanometer deformations. Following validation by atomic force microscopy and development of a model that links changes in laser spectrum to applied force, DEFORM is used to measure forces in 3D and at depths of hundreds of microns within tumor spheroids and late-stage Drosophila larva. We furthermore show continuous force sensing with single-cell spatial and millisecond temporal resolution, thus paving the way for non-invasive studies of biomechanical forces in advanced stages of embryogenesis, tissue remodeling, and tumor invasion.

In vitro evaluation of the effect of yogurt acid whey fractions on iron bioavailability.

A side effect of the raised consumption of Greek yogurt is the generation of massive amounts of yogurt acid whey (YAW). The dairy industry has tried several methods for handling these quantities, which constitute an environmental problem. Although the protein content of YAW is relatively low, given the huge amounts of produced YAW, the final protein amount in the produced YAW should not be underestimated. Taking into consideration the increased interest for bioactive peptides and the increased demand for dietary proteins, combined with protein and peptides content of YAW, efforts should be made toward reintroducing the latter in the food supply chain. In this context and in view of the prevalent dietary iron deficiency problem, the objective of the present study was the investigation of YAW fractions' effect on Fe bioavailability. With this purpose, an in vitro digest approach, following the INFOGEST protocol, was coupled with the Caco2 cell model. To evaluate whether YAW digest fractions exert positive, negative or neutral effect on Fe bioavailability, they were compared with the ones derived from milk, a well-studied food in this context. Milk and YAW showed the same effectiveness on both Fe bioavailability and the expression of relative genes (DCYTB, DMT1, FPN1, and HEPH). Focusing further on YAW fractions, by comparison with their blank digest control counterparts, it resulted that YAW 3- to 10-kDa digests fraction had a superior effect over the 0- to 3-kDa fraction on Fe-uptake, which was accompanied by a similar effect on the expression of Fe metabolism-related genes (DCYTB, FPN1, and HEPH). Finally, although the 3- to 10-kDa fraction of bovine YAW digests resulted in a nonsignificant increased Fe uptake, compared with the ovine and caprine YAW, the expression of DCYTB and FPN1 genes underlined this difference by showing a similar pattern with statistically significant higher expression of bovine compared with ovine and bovine compared with both ovine and caprine, respectively. The present study deals with the novel concept that YAW may contain factors affecting Fe bioavailability. The results show that it does not exert any negative effect and support the extensive investigation for specific peptides with positive effect as well as that YAW proteins should be further assessed on the prospect that they can be used in human nutrition.

Effect of Milk Origin and Seasonality of Yogurt Acid Whey on Antioxidant Activity before and after In Vitro Gastrointestinal Digestion.

Yogurt acid whey (YAW) is a by-product of Greek strained yogurt production. The disposal of YAW constitutes an environmental problem, and given the increasing demand of Greek yogurt worldwide, its handling is a challenge. However, whey-derived peptides, resulting from microbial fermentation as well as those resulting from further hydrolysis during the digestion process, have been linked to enhanced biological activities. In this study, the antioxidant capacity of 33 samples of YAW obtained from Greek dairy companies of bovine, ovine or caprine origin was investigated using both cell-free and cell-based assays. The YAW samples, their in vitro digestion products (YAW-Ds) and a fraction of the digests (less than 3 kDa; YAW-D-P3) were assessed using four biochemical assays, namely ORAC, ABTS, FRAP and P-FRAP. Our data revealed a higher antioxidant capacity for digested samples compared with undigested samples, with all four methods. ORAC values after in vitro digestion were higher for the ovine samples compared to their bovine (YAW-D and YAW-D-P3) and caprine (YAW-D-P3) counterparts. Furthermore, the YAW-D-P3 fraction derived from samples collected in the summer months exhibited higher ORAC values when compared to the respective fraction from the winter months' samples. The cellular antioxidant activity of ovine YAW-D-P3 was improved in H2O2-treated HT29 cells compared to the control H2O2-treated cells. However, YAW-D-P3 could not trigger either the pathways involving the transcription factors NF-κB or NFE2L2 or the gene expression of SOD1, CAT and HMOX1 in LPS-challenged THP-1-derived macrophages. These results suggest that YAW, and particularly YAW from ovine origin, could be used as a natural source for its antioxidant potential in human and animal nutrition.

Antioxidant Activity of Sweet Whey Derived from Bovine, Ovine and Caprine Milk Obtained from Various Small-Scale Cheese Plants in Greece before and after In Vitro Simulated Gastrointestinal Digestion.

Whey-derived peptides have been associated with different biological properties, but most peptides are usually further hydrolyzed during the digestive process. In the present study, the antioxidant capacity of 48 samples of sweet whey (SW) derived from cheeses obtained from small-scale cheese plants made with bovine, ovine, caprine or a mixture of ovine/caprine milk was assessed using both cell-free and cell-based assays. SW digestates (SW-Ds) and a fraction (<3 kDa; SW-D-P3) thereof were obtained after in vitro digestion and subsequent ultrafiltration. Antioxidant properties using four different assays were evaluated before and after digestion. Our data showed higher values (p < 0.05) for ORAC, ABTS, FRAP and P-FRAP after in vitro digestion (SW-Ds and SW-D-P3) when compared with the corresponding values before digestion. In the non-digested SW, ORAC values were higher (p < 0.05) for the bovine SW compared with all the other samples. In contrast, the ABTS assay indicated a higher antioxidant activity for the ovine SW both before digestion and for SW-D-P3 compared with the bovine SW. The fraction SW-D-P3 of the ovine SW, using HT29 cells and H2O2 as an oxidizing agent, increased (p < 0.05) the cellular antioxidant activity. Furthermore, the same fraction of the ovine/caprine mixed SW increased, through the NF-κB pathway, the expression of SOD1 and CAT, genes implicated in the oxidative response in macrophage-like THP-1 cells. These findings indicate that SW, and particularly bovine and ovine SW, could be a candidate source for physical antioxidants in human and animal nutrition.

Real-time imaging of cellular forces using optical interference.

Important dynamic processes in mechanobiology remain elusive due to a lack of tools to image the small cellular forces at play with sufficient speed and throughput. Here, we introduce a fast, interference-based force imaging method that uses the illumination of an elastic deformable microcavity with two rapidly alternating wavelengths to map forces. We show real-time acquisition and processing of data, obtain images of mechanical activity while scanning across a cell culture, and investigate sub-second fluctuations of the piconewton forces exerted by macrophage podosomes. We also demonstrate force imaging of beating neonatal cardiomyocytes at 100 fps which reveals mechanical aspects of spontaneous oscillatory contraction waves in between the main contraction cycles. These examples illustrate the wider potential of our technique for monitoring cellular forces with high throughput and excellent temporal resolution.

Feed supplementation of Lactobacillus plantarum PCA 236 modulates gut microbiota and milk fatty acid composition in dairy goats--a preliminary study.

This study aimed to evaluate the potential of a promising Lactobacillus plantarum isolate (PCA 236) from cheese as a probiotic feed supplement in lactating goats. The ability of L. plantarum to survive transit through the goat gastrointestinal tract and to modulate selected constituents of the gut microbiota composition, monitored at faecal level was assessed. In addition, L. plantarum effects on plasma immunoglobulins and antioxidant capacity of the animals as well as on the milk fatty acid composition were determined. For the purpose of the experiment a field study was designed, involving 24 dairy goats of the Damascus breed, kept in a sheep and goat dairy farm. The goats were divided in terms of body weight in two treatments of 12 goats each, namely: control (CON) without addition of L. plantarum and probiotic (PRO) treatment with in feed administration of L. plantarum so that the goats would intake 12 log CFU/day. The experiment lasted 5 weeks and at weekly time intervals individual faecal, blood and milk samples were collected and analysed. All faecal samples were examined for the presence of L. plantarum PCA 236. In addition, the culturable population levels of mesophilic aerobes, coliforms lactic acid bacteria (LAB), Streptococcus, Enterococcus, mesophilic anaerobes, Clostridium and Bacteroides in faeces were also determined by enumeration on specific culture media. In parallel, plasma IgA, IgM and IgG and antioxidant capacity of plasma and milk were determined. No adverse effects were observed in the animals receiving the lactobacillus during the experiment. Lactobacillus plantarum PCA 236 was recovered in the faeces of all animals in the PRO treatment. In addition, PRO treatment resulted in a significant (P