Formulation engineering of food systems for 3D-printing applications - A review.

The efficient development of extrusion-based 3D-printing requires flexibility in both formulation- and process design. This task requires a fundamental understanding of the influence of material rheological properties on the extrusion process. Within this review, a qualitative toolbox for food extrusion is presented which provides guidelines for the formulation and engineering of extrusion processes in general and 3D-printing in particular. The toolbox is based on current knowledge of highly viscous food systems and the influence of individual components on the overall rheology. It includes the efficiency of particle packing, microstructure and the influence of shear rate, as well as the formation of self-supporting structures by gelation of the liquid phase and crowding of particles. Physical laws and semi-empirical equations are discussed to describe the rheology and relate relevant theory to the extrusion process. Practical information is presented, including examples of extrusion and 3D-printing of food and non-food systems. The qualitative extrusion toolbox provides a general framework for the emerging field of extrusion-based 3D-printing of food products. It can be used to identify which specific material and process parameters can be changed and how they may be altered to optimize the 3D-printing process. The general framework will assist researchers, as well as industry.

Synthesis of N-Acetyllactosamine and N-Acetyllactosamine-Based Bioactives.

N-Acetyllactosamine (LacNAc) or more specifically ß-d-galactopyranosyl-1,4-N-acetyl-d-glucosamine is a unique acyl-amino sugar and a key structural unit in human milk oligosaccharides, an antigen component of many glycoproteins, and an antiviral active component for the development of effective drugs against viruses. LacNAc is useful itself and as a basic building block for producing various bioactive oligosaccharides, notably because this synthesis may be used to add value to dairy lactose. Despite a significant amount of information in the literature on the benefits, structures, and types of different LacNAc-derived oligosaccharides, knowledge about their effective synthesis for large-scale production is still in its infancy. This work provides a comprehensive analysis of existing production strategies for LacNAc and important LacNAc-based structures, including sialylated LacNAc as well as poly- and oligo-LacNAc. We conclude that direct extraction from milk is too complex, while chemical synthesis is also impractical at an industrial scale. Microbial routes have application when multiple step reactions are needed, but the major route to large-scale biochemical production will likely lie with enzymatic routes, particularly those using ß-galactosidases (for LacNAc synthesis), sialidases (for sialylated LacNAc synthesis), and ß-N-acetylhexosaminidases (for oligo-LacNAc synthesis). Glycosyltransferases, especially for the biosynthesis of extended complex LacNAc structures, could also play a major role in the future. In these cases, immobilization of the enzyme can increase stability and reduce cost. Processing parameters, such as substrate concentration and purity, acceptor/donor ratio, water activity, and temperature, can affect product selectivity and yield. More work is needed to optimize these reaction parameters and in the development of robust, thermally stable enzymes to facilitate commercial production of these important bioactive substances.

Challenging immunodominance of influenza-specific CD8+ T cell responses restricted by the risk-associated HLA-A*68:01 allomorph.

Although influenza viruses lead to severe illness in high-risk populations, host genetic factors associated with severe disease are largely unknown. As the HLA-A*68:01 allele can be linked to severe pandemic 2009-H1N1 disease, we investigate a potential impairment of HLA-A*68:01-restricted CD8+ T cells to mount robust responses. We elucidate the HLA-A*68:01+CD8+ T cell response directed toward an extended influenza-derived nucleoprotein (NP) peptide and show that only ~35% individuals have immunodominant A68/NP145+CD8+ T cell responses. Dissecting A68/NP145+CD8+ T cells in low vs. medium/high responders reveals that high responding donors have A68/NP145+CD8+ memory T cells with clonally expanded TCRαßs, while low-responders display A68/NP145+CD8+ T cells with predominantly naïve phenotypes and non-expanded TCRαßs. Single-cell index sorting and TCRαß analyses link expansion of A68/NP145+CD8+ T cells to their memory potential. Our study demonstrates the immunodominance potential of influenza-specific CD8+ T cells presented by a risk HLA-A*68:01 molecule and advocates for priming CD8+ T cell compartments in HLA-A*68:01-expressing individuals for establishment of pre-existing protective memory T cell pools.

A review of salty waste stream management in the Australian dairy industry.

Saline wastewater is a by-product of cheese manufacturing and whey processing that can have serious environmental and economic consequences. Salty streams originating from dairy processing operations include chromatography wastes, clean-in-place wastewater, acid whey, salty whey and waste generated from whey demineralization processes such as nanofiltration, electrodialysis and ion exchange. With the participation of the major dairy companies in Australia, an industry wide survey was conducted to acquire a comprehensive understanding of the management strategies for these salty waste streams. High salinity waste streams are commonly directed to evaporation ponds. However, environmental impacts from land degradation, odour and dust have prevented the construction of further evaporation ponds in some areas of Australia. The survey results also show that disposal to municipal trade waste is not always effective, as the current levels of some salinity-related parameters are significantly higher than the levels allowed by the local water/environmental authorities. For high salinity streams, salt removal can lead to more substantial savings in trade waste charges compared to wastewater volume reduction. Thus, salt removal and recovery from salty waste streams has become a major focus of the sustainability agenda of the Australian dairy industry.

A fluorescence in situ staining method for investigating spores and vegetative cells of Clostridia by confocal laser scanning microscopy and structured illuminated microscopy.

Non-pathogenic spore-forming Clostridia are of increasing interest due to their application in biogas production and their capability to spoil different food products. The life cycle for Clostridium includes a spore stage that can assist in survival under environmentally stressful conditions, such as extremes of temperature or pH. Due to their size, spores can be investigated by a range of microscopic techniques, many of which involve sample pre-treatment. We have developed a quick, simple and non-destructive fluorescent staining procedure that allows a clear differentiation between spores and vegetative cells and effectively stains spores, allowing recovery and tracking in subsequent experiments. Hoechst 34580, Propidium iodide and wheat germ agglutinin WGA 488 were used in combination to stain four strains of Clostridia at different life cycle stages. Staining was conducted without drying the sample, preventing changes induced by dehydration and cells observed by confocal laser scanner microscopy or using a super-resolution microscope equipped with a 3D-structured illumination module. Dual staining with Hoechst/Propidium iodide differentiated spores from vegetative cells, provided information on the viability of cells and was successfully applied to follow spore production induced by heating. Super-resolution microscopy of spores probed by Hoechst 34580 also allowed chromatin to be visualised. Direct staining of a cheese specimen using Nile Red and Fast Green allowed in situ observation of spores within the cheese and their position within the cheese matrix. The proposed staining method has broad applicability and can potentially be applied to follow Clostridium spore behaviour in a range of different environments.

Effect of temperature on the microstructure of fat globules and the immunoglobulin-mediated interactions between fat and bacteria in natural raw milk creaming.

Natural creaming of raw milk is the first step in production of Grana Padano and Parmigiano Reggiano Protected Denomination of Origin cheeses. This process decreases the fat content and plays an important role in the removal of clostridia species that may cause late-blowing defects in ripened cheeses. Partial coalescence of fat globules-that may influence fat behavior in cheese making and affect the microstructure of fat in the final cheese product-was observed at creaming temperatures higher than 22°C by confocal laser scanning microscopy. The widespread practice of heating of milk at 37°C before creaming at 8°C resulted in important changes in the size distribution of fat globules in raw milk, potentially altering the ability of fat to entrap clostridia spores. We investigated the role of immunoglobulin classes in both the clustering of fat globules and the agglutination of Clostridium tyrobutyricum to fat globules during creaming. Immunogold labeling and transmission electron microscopy showed that IgA and IgM but not IgG were involved in both clustering and agglutination. Both vegetative cells and spores were clearly shown to agglutinate to fat droplets, a process that was suppressed by thermal denaturation of the immunoglobulins. The debacterization of raw milk through natural creaming was improved by the addition of purified immunoglobulins. Overall, these findings provide not only a better understanding of the phenomena occurring during the natural creaming but also practical insights into how the process of creaming may be optimized in cheese production plants.

Solubility of Calcium Phosphate in Concentrated Dairy Effluent Brines.

The solubility of calcium phosphate in concentrated dairy brine streams is important in understanding mineral scaling on equipment, such as membrane modules, evaporators, and heat exchangers, and in brine pond operation. In this study, the solubility of calcium phosphate has been assessed in the presence of up to 300 g/L sodium chloride as well as lactose, organic acids, and anions at 10, 30, and 50 °C. As a neutral molecule, lactose has a marginal but still detectable effect upon calcium solubility. However, additions of sodium chloride up to 100 g/L result in a much greater increase in calcium solubility. Beyond this point, the concentrations of ions in the solution decrease significantly. These changes in calcium solubility can readily be explained through changes in the activity coefficients. There is little difference in calcium phosphate speciation between 10 and 30 °C. However, at 50 °C, the ratio of calcium to phosphate in the solution is lower than at the other temperatures and varies less with ionic strength. While the addition of sodium lactate has less effect upon calcium solubility than sodium citrate, it still has a greater effect than sodium chloride at an equivalent ionic strength. Conversely, when these organic anions are present in the solution in the acid form, the effect of pH dominates and results in much higher solubility and a calcium/phosphate ratio close to one, indicative of dicalcium phosphate dihydrate as the dominant solid phase.

Crystallisation of minerals from concentrated saline dairy effluent.

An understanding of crystallisation within saline effluents is important for the design of both brine crystallisers and brine disposal ponds. In this work, crystallisation of a saline effluent concentrate from the Australian dairy industry has been examined at 22 wt% and 30 wt% total solids and at temperatures between 10 and 70 °C. The precipitation occurs more rapidly at higher temperatures. This trend is dictated by precipitation of calcium phosphate salts, albeit the major constituents of the mixture are NaCl and lactose. The crystallisation induction time can be shortened by introducing cavitation induced by ultrasound. In particular, the use of two short acoustic pulses between 3.7 J/g and 16 J/g at 20 kHz spaced ten minutes apart has maximum impact upon both induction time and crystal size. It is believed that the first ultrasound pulse either generates new nuclei or enhances the mass transfer of solute toward the surface of sub-micron growing crystals. Conversely, the second pulse disrupts the growing crystals and forms secondary nuclei. The ultrasound cannot shift the solution equilibrium and so is not able to improve the low crystal yield. To increase this total yield, further evaporation is necessary. The work provides direction to personnel in the dairy industry of the feasibility of brine crystallisation with respect to energy demand and solid recovery.

An enzyme-responsive controlled release system based on a dual-functional peptide.

A new controlled release system was developed by loading a dual-functional peptide (DFP) on a mesoporous silica material. One-pot synthesis produced a DFP that was stimuli responsive, releasing a therapeutic peptide by protease cleavage. The design provides new steps towards smart biomaterials.

[Critical haematuria after prostate biopsies with RIVAROXABAN. Case report]. / Hématurie critique post-biopsies de prostate sous RIVAROXABAN. Cas clinique.

Managing patients with new oral anticoagulants in perioperative period is not yet well protocolized. We report a clinical case of a critical haematuria after prostate biopsies to a patient treated with RIVAROXABAN. Monitoring and treatment of the haematuria have been difficult due to the lack of biological control and antidote for this treatment.

Tissue engineering as a potential alternative or adjunct to surgical reconstruction in treating pelvic organ prolapse.

INTRODUCTION AND HYPOTHESIS: Cell-based tissue engineering strategies could potentially provide attractive alternatives to surgical reconstruction of native tissue or the use of surgical implants in treating pelvic organ prolapse (POP). METHODS: Based on a search in PubMed, this review focuses on candidate cell types, scaffolds, and trophic factors used in studies examining cell-based tissue engineering strategies to treat POP, stress urinary incontinence (SUI), and the closely related field of hernias. RESULTS: In contrast to the field of SUI, the use of cell-based tissue engineering strategies to treat POP are very sparsely explored, and only preclinical studies exist. CONCLUSION: The available evidence suggests that the use of autologous muscle-derived cells, fibroblasts, or mesenchymal stem cells seeded on biocompatible, degradable, and potentially growth-promoting scaffolds could be an alternative to surgical reconstruction of native tissue or the use of conventional implants in treating POP. However, the vagina is a complex organ with great demands of functionality, and the perfect match of scaffold, cell, and trophic factor has yet to be found and tested in preclinical studies. Important issues such as safety and economy must also be addressed before this approach is ready for clinical studies.

Snakebite on the hand: lessons from two clinical cases illustrating difficulties of surgical indication

Snakebite is a particularly important health problem in rural areas of tropical regions. A large number of victims survive with permanent physical sequelae due to local tissue necrosis. However, necrosis may be associated with compartment syndrome especially when the bite is on the hands or feet. Herein, we describe two cases reported at a rural district hospital in Central African Republic. The present study suggests that active multidisciplinary management may improve patient prognosis while evidencing how difficult it is to decide on surgical intervention.(AU)

Essential role of microphthalmia transcription factor for DNA replication, mitosis and genomic stability in melanoma.

Malignant melanoma is an aggressive cancer known for its notorious resistance to most current therapies. The basic helix-loop-helix microphthalmia transcription factor (MITF) is the master regulator determining the identity and properties of the melanocyte lineage, and is regarded as a lineage-specific 'oncogene' that has a critical role in the pathogenesis of melanoma. MITF promotes melanoma cell proliferation, whereas sustained supression of MITF expression leads to senescence. By combining chromatin immunoprecipitation coupled to high throughput sequencing (ChIP-seq) and RNA sequencing analyses, we show that MITF directly regulates a set of genes required for DNA replication, repair and mitosis. Our results reveal how loss of MITF regulates mitotic fidelity, and through defective replication and repair induces DNA damage, ultimately ending in cellular senescence. These findings reveal a lineage-specific control of DNA replication and mitosis by MITF, providing new avenues for therapeutic intervention in melanoma. The identification of MITF-binding sites and gene-regulatory networks establish a framework for understanding oncogenic basic helix-loop-helix factors such as N-myc or TFE3 in other cancers.

The effect of milk processing on the microstructure of the milk fat globule and rennet induced gel observed using confocal laser scanning microscopy.

Confocal laser scanning microscopy (CLSM) was successfully used to observe the effect of milk processing on the size and the morphology of the milk fat globule in raw milk, raw ultrafiltered milk, and standardized and pasteurized milk prepared for cheese manufacture (cheese-milk) and commercial pasteurized and homogenized milk. Fat globule size distributions for the milk preparations were analyzed using both image analysis and light scattering and both measurements produced similar data trends. Changes to the native milk fat globule membrane (MFGM) were tracked using a MFGM specific fluorescent stain that allowed MFGM proteins and adsorbed proteins to be differentiated on the fat globule surface. Sodium dodecyl sulfate polyacrylamide gel electrophoresis confirmed the identity of native MFGM proteins isolated from the surface of fat globules within raw, UF retentate, and cheese-milk preparations, whereas only casein was detected on the surface of fat globules in homogenized milk. The microstructure, porosity, and gel strength of the rennet induced gel made from raw milk and cheese-milk was also found to be comparable and significantly different to that made from homogenized milk. Our results highlight the potential use of CLSM as a tool to observe the structural details of the fat globule and associated membrane close to its native environment.

Parametric instabilities and their control in advanced interferometer gravitational-wave detectors.

A detailed simulation of Advanced LIGO test mass optical cavities shows that parametric instabilities will excite 7 acoustic modes in each fused silica test mass, with parametric gain R up to 7 and only 1 acoustic mode with R approximately 2 for alternative sapphire test masses. Fine-tuning of the test mass radii of curvature causes the instabilities to sweep through various modes with R as high as approximately 2000. Sapphire test mass cavities can be tuned to completely eliminate instabilities using thermal g-factor tuning. In the case of fused silica test mass, instabilities can be minimized but not eliminated.

Expression in yeast and purification of functional recombinant human poly(ADP-ribose)polymerase (PARP). Comparative pharmacological profile with that of the rat enzyme.

Human poly(ADP-ribose)polymerase (PARP) was expressed in the yeast line JEL1 under the control of a GAL promoter. Proteins were extracted and human recombinant PARP purified to apparent homogeneity. The pharmacological profile of this human enzyme was characterised in terms of the effects of known inhibitors of PARP belonging to various chemical families and this was compared with that of the rat enzyme purified from rat testes, using the same purification protocol. The rat and the human enzymes appeared very similar in terms of their sensitivities to those selected inhibitors.

Pituitary adenylate cyclase-activating peptide stimulates acute progesterone production in rat granulosa/Lutein cells via two receptor subtypes.

Pituitary adenylate cyclase-activating peptide (PACAP) is transiently expressed in ovarian granulosa/lutein cells from eCG/hCG-treated rats, and in vitro immunoneutralization of endogenously released PACAP inhibits acute progesterone secretion and subsequent luteinization in such cells. This suggests that PACAP mediates locally some of the effects of the LH surge, but the putative PACAP receptor(s) involved in such an auto or paracrine activity is presently unknown. Reverse-transcription polymerase chain reaction with specific primers to the three cloned PACAP-binding receptors called PAC(1), VPAC(1), and VPAC(2) demonstrated both PAC(1) and VPAC(2) mRNA in extracts from preovulatory follicular cells. Radioligand-binding assays revealed the presence of high-affinity binding sites with characteristics of these two receptors on the intact cells, and autoradiography demonstrated that the binding was restricted to a minor proportion of the follicular cells as well as the oocytes. Pituitary adenylate cyclase-activating peptide and vasoactive intestinal peptide (VIP) dose-dependently stimulated cAMP accumulation and acute progesterone accumulation. Forskolin and db-cAMP also stimulated acute progesterone accumulation, and the protein kinase A inhibitor H89 dose-dependently inhibited peptide induced acute progesterone accumulation, suggesting involvement of cAMP and the protein kinase A pathway in the process. In conclusion, two of the three PACAP binding receptors are present on preovulatory follicular cells and are involved in the effects of PACAP on acute progesterone production. The data provide further evidence to establish PACAP as an auto- or paracrine regulator of LH-induced acute progesterone production in rat preovulatory follicles.

Pituitary adenylate cyclase-activating polypeptide is an auto/paracrine stimulator of acute progesterone accumulation and subsequent luteinization in cultured periovulatory granulosa/lutein cells.

Recently, we have demonstrated that pituitary adenylate cyclase-activating polypeptide (PACAP) is transiently expressed in steroidogenic ovarian cells during the periovulatory period. This prompted us to establish an in vitro system in which the potential local regulatory role of PACAP during periovulatory progesterone production could be examined. Granulosa/lutein cells from PMSG- and human CG (hCG)-stimulated immature rats were used. The cells were isolated from preovulatory follicles 4-6 h after the hCG injection, at which time the transient ovarian PACAP expression begins in vivo. By immunocytochemistry on intact cells and RIA on cell extracts and culture medium, granulosa/lutein cells were found to accumulate and secrete PACAP during incubation. Furthermore, the cells responded to exogenous PACAP 38 with a rapid (10(-7) M induced a peak value 20-fold higher than controls at 2 h) and dose-dependent accumulation of progesterone. PACAP 38 (5 x 10(-9) M), in combination with an approximately half-maximal dose of hCG (1 ng/ml), showed an additive effect on progesterone accumulation. Immunoneutralization of endogenously released PACAP was performed using the IgG fraction from a specific PACAP antiserum that dose-dependently inhibits the progesterone accumulating effect of exogenous PACAP 38. The acute effects of endogenously released PACAP were studied during 8 h of incubation of granulosa/lutein cells with anti-PACAP IgG (100 microg/ml). A significant reduction in progesterone accumulation was observed after 4, 6, and 8 h [38.7% (P < 0.05), 41.2% (P < 0.02), and 50% (P < 0.002), respectively], compared with nonimmune IgG (100 microg/ml) treated cultures. The long-term effects on luteinization induced by endogenously released PACAP were studied after incubation of the cells with anti-PACAP IgG or nonimmune IgG for 24 h, followed by incubation for 9 days in serum-containing medium. Under these conditions, nonimmune IgG-treated cells assumed a luteal phenotype, accumulating large and stable amounts of progesterone and acquiring hypertrophic cell bodies with numerous lipid droplets and distinct nucleoli in the large nuclei. Anti-PACAP IgG-treated cells displayed morphological and functional signs of impaired luteinization being smaller and more irregular and with progesterone accumulation being significantly lower throughout the incubation period [56.4% (P < 0.02), 69.2% (P < 0.05), 43.8% (P < 0.02), and 52.2% (P < 0.02) at 1, 4, 7, and 10 days, respectively]. Together, these findings support an auto- or paracrine role for PACAP during gonadotropin-induced acute periovulatory progesterone production and subsequent luteinization in granulosa/lutein cells.