Use of blends of bioabsorbable poly(L-lactic acid)/poly(hydroxybutyrate- co-hydroxyvalerate) as surfaces for Vero cell culture

Vero cells, a cell line established from the kidney of the African green monkey (Cercopithecus aethiops), were cultured in F-10 Ham medium supplemented with 10% fetal calf serum at 37°C on membranes of poly(L-lactic acid) (PLLA), poly(hydroxybutyrate-co-hydroxyvalerate) (PHBV) and their blends in different proportions (100/0, 60/40, 50/50, 40/60, and 0/100). The present study evaluated morphology of cells grown on different polymeric substrates after 24 h of culture by scanning electron microscopy. Cell adhesion was also analyzed after 2 h of inoculation. For cell growth evaluation, the cells were maintained in culture for 48, 120, 240, and 360 h. For cytochemical study, the cells were cultured for 120 or 240 h, fixed, processed for histological analysis, and stained with Toluidine blue, pH 4.0, and Xylidine ponceau, pH 2.5. Our results showed that cell adhesion was better when 60/40 and 50/50 blends were used although cells were able to grow and proliferate on all blends tested. When using PLLA/PHBV (50/50) slightly flattened cells were observed on porous and smooth areas. PLLA/PHBV (40/60) blends presented flattened cells on smooth areas. PLLA/PHBV (0/100), which presented no pores, also supported spreading cells interconnected by thin filaments. Histological sections showed that cells grew as a confluent monolayer on different substrates. Cytochemical analysis showed basophilic cells, indicating a large amount of RNA and proteins. Hence, we detected changes in cell morphology induced by alterations in blend proportions. This suggests that the cells changed their differentiation pattern when on various PLLA/PHBV blend surfaces.

Application of ftir in the determination of acrylate content in poly(sodium acrylate-co-acrylamide) superabsorbent hydrogels

Hydrogels have been prepared by free-radical solution copolymerization of acrylamide and sodium acrylate (NaAc), with molar ratio ranging from 25/75 to 80/20, respectively, using methylene bisacrylamide as the crosslinking agent. A FTIR spectroscopy procedure to determine the acrylate/acrylamide ratio in these hydrogels was proposed based on absorbance at 1410 cm-1 (nCOO-) and 2940 cm-1 (nCH and nCH2). A straight line with a good linear correlation coefficient (0.998) was obtained by plotting the acrylate content (Ac%) versus relative absorbance (Arel = A1410/A2940). Results were confirmed by the amount of sodium cation released in acid medium determined by atomic absorption spectrometry.

Starch/poly (butylene adipate-co-terephthalate)/montmorillonite films produced by blow extrusion

This study aims to prepare biodegradable films from cassava starch, poly (butylene adipate-co-terephthalate) (PBAT), and montmorillonite (MMT) using blow-extrusion process and analyze the effects of different types and concentrations of MMT on the microstructure, physicochemical, and mechanical properties of the resulting films. The films were produced by blending 30% of PBAT with glycerol (17.5%), starch (49.0-52.5%), and four different types of montmorillonite (Cloisite® Na+, 10A, 15A, and 30B) at two different concentrations (1.75% and 3.5%). All the films prepared in this study showed an increase in the basal spacing of MMT layers. In particular, the films with 10A and 30B showed the highest increase in intercalation basal spacing, suggesting the formation of intercalated composites. The addition of nanoclays decreased the elongation of films. The addition of Cloisite® 10A resulted in films with the lowest WVP values and the highest stability to water adsorption under different RH conditions.

Poly-DL-lactide-co-glycolide microspheres as a controlled release antigen delivery system

Successful vaccine application means maximum protection with minimal number of administrations. A rational development of vaccines involves studies of the nature of the antigen as well as of the adjuvant to be used to improve the immune responses. This has provided the impetus for studies to design the degradable devices and for different approaches to antigen delivery by different routes of administration. The development of controlled release systems based on polymeric devices that permit a sustained or pulsed release of encapsulated antigens has attracted much interest. Polymeric delivery systems consist of polymers that release their content continuously in a controlled manner over a period of time. The development of a biocompatible delivery system for parenteral administration offers several advantages in terms of immunoadjuvanticity over other compounds. It was found that, in contrast to other carriers, microspheres are more stable, thus permitting administration by the oral or parenteral route. In the present study, we describe the main characteristics and potentialities of this new immunoadjuvant for oral and parenteral administration.

Poly(3-hydroxybutyrate-co-3-hydroxyhexanoate)-based scaffolds for tissue engineering

Development and selection of an ideal scaffold is of importance for tissue engineering. Poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) (PHBHHx) is a biocompatible bioresorbable copolymer that belongs to the polyhydroxyalkanoate family. Because of its good biocompatibility, PHBHHx has been widely used as a cell scaffold for tissue engineering. This review focuses on the utilization of PHBHHx-based scaffolds in tissue engineering. Advances in the preparation, modification, and application of PHBHHx scaffolds are discussed.

Evaluation of thermotolerant capacity of lactic acid bacteria isolated from commercial sausages and the effects of their addition on the quality of cooked sausages

The thermotolerant capacity of several lactic acid bacteria strains isolated from cooked commercial sausages was determined. Four strains were positively identified as Lactobacillus plantarum, Lactobacillus curvatus, Pediococcus pentosaceus and Pediococcus acidilacti, after surviving thermal treatment (70 °C during 60 minutes). Thermotolerant strains were inoculated in sausage batters before cooking in order to determine their effect on color, texture, acceptance and inhibition of Enterobacteria during 12 days at 8 °C. No significant effect of the inoculated strains was detected on color parameters. Textural profile parameters, cohesiveness and resilience, were not affected by the inoculation of thermotolerant lactic acid bacteria, but L. curvatus sausages resulted softer than the rest of the treatments. Samples inoculated with L. curvatus also obtained the lowest scores for the sensory attributes evaluated, with the remaining treatments causing no unfavorable effects on sausage acceptance. There was a reduction in enterobacterial counts after 12 days of cold storage in inoculated samples. The performance of inoculated lactic acid bacteria strains can be explained in a similar way as that of starter cultures in dry-fermented sausages, where the growth in nests impairs other pathogenic microorganisms present in the rest of the sausage, since environmental conditions and the early inoculation of these thermotolerant strains favor them to become the dominant flora.

An alternative method for screening lactic acid bacteria for the production of exopolysaccharides with rapid confirmation

The accumulation of exopolysaccharides (EPS) produced by microorganisms occurs in the presence of excess substrate and limiting conditions of elements that are essential to growth, such as nitrogen, phosphorus, sulfur, and magnesium. The presence of EPS produced by bacterial cells contributes to slime colonies formation in solid medium and increased viscosity in liquid medium. This paper proposes an alternative method for screening EPS-producing lactic acid bacteria using solid medium-containing discs of filter paper that are saturated with active cultures. The screening was carried out under different culture conditions varying the type of sugar, pH, and temperature. EPS production was visualized by the presence of mucoid colonies on the discs, which was confirmed by the formation of a precipitate when part of this colony was mixed with absolute alcohol. The established conditions for obtaining a high number of isolates producing EPS were 10% sucrose, pH 7.5 and 28 ºC. This method proved to be effective and economical because several strains could be tested on the same plate, with immediate confirmation.

Antimicrobial properties of lactic acid bacteria isolated from uruguayan artisan cheese

Uruguayan artisan cheese is elaborated with raw milk and non-commercial starters. The associated native microbiota may include lactic acid bacteria and also potentially pathogenic bacteria. Lactic acid bacteria were isolated from artisan cheese, raw milk, and non-commercial starter cultures, and their potential bacteriocin production was assessed. A culture collection of 509 isolates was obtained, and five isolates were bacteriocin-producers and were identified as Enterococcus durans,Lactobacillus casei, and Lactococcus lactis. No evidence of potential virulence factors were found in E. durans strains. These are promising results in terms of using these native strains for cheese manufacture and to obtain safe products.

Screening and kinetics of glutaminase and glutamate decarboxylase producing lactic acid bacteria from fermented Thai foods

L-glutaminase and glutamic acid decarboxylase (GAD) catalyzes the hydrolysis of L-glutamine and glutamate, respectively. L-glutaminase widely used in cancer therapy along with a combination of other enzymes and most importantly these enzymes were used in food industries, as a major catalyst of bioconversion. The current investigation was aimed to screen and select L-glutaminase, and GAD producing lactic acid bacteria (LAB). A total of 338 LAB were isolated from fermented meat, fermented fish, fermented soya bean, fermented vegetables and fruits. Among 338 isolates, 22 and 237 LAB has been found to be positive for L-glutaminase and GAD, respectively. We found that 30 days of incubation at 35 ºC and pH 6.0 was the optimum condition for glutaminase activity by G507/1. G254/2 was found to be the best for GAD activity with the optimum condition of pH 6.5, temperature 40 ºC and ten days of incubation. These LAB strains, G507/1 and G254/2, were identified as close relative of Lactobacillus brevis ATCC 14869 and Lactobacillus fermentum NBRC 3956, respectively by 16S rRNA sequencing. Further, improvements in up-stream of the fermentation process with these LAB strains are currently under development.

Production of probiotic fresh white cheese using co-culture with Streptococcus thermophilus

In this research, the probiotic Streptococcus thermophilus was inoculated into milk as co-culture to produce probiotic cheese. The effects of using Streptococcus thermophilus with other probiotic bacteria on cheese composition, and microbiological viability during 28 days of storage were investigated. Sensorial properties were determined only at 1st and 28th days of storage. The results showed that the use of Streptococcus thermophilus as co-culture in probiotic cheese production did not affect negatively the cheese components. Fat and dry matter properties of cheese weren't influenced by added probiotic bacteria. However, different level of pH, salt and lactic acid were detected. All probiotic bacteria were present in high levels throughout storage of cheeses, above 7 Log cfu.g- 1, threshold required for probiotic activity. Sensory panel showed that the highest average sensory evaluation points were recorded in cheeses made with Streptococcus thermophilus plus Lactobacillus casei, whereas other probiotic bacteria combinations had been affected less in regard to taste or appearance.

Estratégias de cultivo para produção dos plásticos biodegradáveis poli(3-Hidroxibutirato) e poli(3-hidroxibutirato-co-3-hidroxivalerato) por bactérias

Polyhydroxyalkanoates (PHAs) are carbon and energy storage materials that are accumulated as intracellular granules in a variety of microorganisms during unbalanced growth. PHAs have drawn attention due to their properties similar to conventional plastics and complete biodegradability. They can be used for food and cosmetics packaging, and in medicine and agriculture. However, their applicability is reduced because of their high production cost compared to conventional plastics. An overview on production strategies of poly(3-hydroxybutyrate) and poly(3-hydroxybutyrate-co-3-hydroxyvalerate) aiming at reducing the production costs is presented.

CARACTERIZAÇÃO E AVALIAÇÃO IN VITRO DE NANOCOMPÓSITOS DE POLI (L-ÁCIDO LÁTICO) E NANOTUBOS DE CARBONO DE PAREDES MÚLTIPLAS PURIFICADOS

Carbon nanotubes (CNT) have been studied for biomedical applications due to their unique properties. However, pristine CNT have structural features and impurities that can cause toxicity to biological systems. In this work, we describe a method to purify multiwalled carbon nanotubes (MWCNT) by chemical modification and subsequent attachment of hydroxyl and carboxyl groups to improve dispersion and to decrease toxic effects. Nanocomposites from poly (L-lactic acid) (PLLA) and nanotubes were produced by the solvent casting method and characterized and evaluated for cytocompatibility with Vero cells. The nanocomposite interactions with Vero cells demonstrated that the cells were able to adhere and sustain proliferation and showed favorable cytocompatibility. In vitro studies also revealed an increase in fibroblast cell viability in the nanocomposites, compared with neat PLLA.

Biomechanical and histological evaluation of hydrogel implants in articular cartilage

We evaluated the mechanical behavior of the repaired surfaces of defective articular cartilage in the intercondylar region of the rat femur after a hydrogel graft implant. The results were compared to those for the adjacent normal articular cartilage and for control surfaces where the defects remained empty. Hydrogel synthesized by blending poly(2-hydroxyethyl methacrylate) and poly(methyl methacrylate-co-acrylic acid) was implanted in male Wistar rats. The animals were divided into five groups with postoperative follow-up periods of 3, 5, 8, 12 and 16 weeks. Indentation tests were performed on the neoformed surfaces in the knee joint (with or without a hydrogel implant) and on adjacent articular cartilage in order to assess the mechanical properties of the newly formed surface. Kruskal-Wallis analysis indicated that the mechanical behavior of the neoformed surfaces was significantly different from that of normal cartilage. Histological analysis of the repaired defects showed that the hydrogel implant filled the defect with no signs of inflammation as it was well anchored to the surrounding tissues, resulting in a newly formed articular surface. In the case of empty control defects, osseous tissue grew inside the defects and fibrous tissue formed on the articular surface of the defects. The repaired surface of the hydrogel implant was more compliant than normal articular cartilage throughout the 16 weeks following the operation, whereas the fibrous tissue that formed postoperatively over the empty defect was stiffer than normal articular cartilage after 5 weeks. This stiffness started to decrease 16 weeks after the operation, probably due to tissue degeneration. Thus, from the biomechanical and histological point of view, the hydrogel implant improved the articular surface repair.

An acid-sensing ion channel that detects ischemic pain

Ischemic pain occurs when there is insufficient blood flow for the metabolic needs of an organ. The pain of a heart attack is the prototypical example. Multiple compounds released from ischemic muscle likely contribute to this pain by acting on sensory neurons that innervate muscle. One such compound is lactic acid. Here, we show that ASIC3 (acid-sensing ion channel #3) has the appropriate expression pattern and physical properties to be the detector of this lactic acid. In rats, it is expressed only in sensory neurons and then only on a minority (~40%) of these. Nevertheless, it is expressed at extremely high levels on virtually all dorsal root ganglion sensory neurons that innervate the heart. It is extraordinarily sensitive to protons (Hill slope 4, half-activating pH 6.7), allowing it to readily respond to the small changes in extracellular pH (from 7.4 to 7.0) that occur during muscle ischemia. Moreover, both extracellular lactate and extracellular ATP increase the sensitivity of ASIC3 to protons. This final property makes ASIC3 a "coincidence detector" of three molecules that appear during ischemia, thereby allowing it to better detect acidosis caused by ischemia than other forms of systemic acidosis such as hypercapnia.

Antioxidant effect of mogrosides against oxidative stress induced by palmitic acid in mouse insulinoma NIT-1 cells

Excessive oxidative stress in pancreatic β cells, caused by glucose and fatty acids, is associated with the pathogenesis of type 2 diabetes. Mogrosides have shown antioxidant and antidiabetic activities in animal models of diabetes, but the underlying mechanisms remain unclear. This study evaluated the antioxidant effect of mogrosides on insulinoma cells under oxidative stress caused by palmitic acid, and investigated the underlying molecular mechanisms. Mouse insulinoma NIT-1 cells were cultured in medium containing 0.75 mM palmitic acid, mimicking oxidative stress. The effects of 1 mM mogrosides were determined with the dichlorodihydrofluorescein diacetate assay for intracellular reactive oxygen species (ROS) and FITC-Annexin V/PI assay for cell apoptosis. Expression of glucose transporter-2 (GLUT2) and pyruvate kinase was determined by semi-quantitative reverse-transcription polymerase chain reaction. Palmitic acid significantly increased intracellular ROS concentration 2-fold (P<0.05), and decreased expression of GLUT2 (by 60%, P<0.05) and pyruvate kinase (by 80%, P<0.05) mRNAs in NIT-1 cells. Compared with palmitic acid, co-treatment with 1 mM mogrosides for 48 h significantly reduced intracellular ROS concentration and restored mRNA expression levels of GLUT2 and pyruvate kinase. However, mogrosides did not reverse palmitic acid-induced apoptosis in NIT-1 cells. Our results indicate that mogrosides might exert their antioxidant effect by reducing intracellular ROS and regulating expression of genes involved in glucose metabolism. Further research is needed to achieve a better understanding of the signaling pathway involved in the antioxidant effect of mogrosides.