Phosphocalcic metabolism and its potential association with biomarkers of kidney disease in dogs with spontaneous hyperadrenocorticism.

The pathogenesis of increased serum phosphate concentration and proteinuria in dogs with spontaneous hyperadrenocorticism (HAC) is unclear. A potential link between proteinuria and calcium/phosphate metabolism has never been studied in dogs with HAC. The aims of the study were: (1) To evaluate calcium/phosphate metabolism in dogs with spontaneous HAC and compare to healthy dogs as well as to dogs with non-HAC illness; (2) to look for associations between markers of calcium/phosphate metabolism and biomarkers of kidney disease in dogs with HAC. Fifty-four dogs were included in the study, classified as HAC (n=27), non-HAC disease (n=17), and healthy (n=10). Serum calcium, phosphate, 25(OH)Vitamin D, 1,25(OH)2Vitamin D, plasma intact parathyroid hormone concentration (iPTH), FGF23, and urinary fractional excretion of calcium and phosphate were evaluated in all dogs at diagnosis and compared between each group. The correlation between these variables and urine protein-to-creatinine ratio (UPC) and urinary N-acetylglucosaminidase-to-creatinine ratio (uNAG/C) was evaluated in the HAC group. Medians [range] of serum phosphate concentration, urinary fractional excretion of calcium (FE(Ca)), and iPTH were significantly higher in dogs with HAC than in dogs with non-HAC illness (P<0.01) and healthy dogs (P<0.01). Increased 1,25(OH)2Vitamin D/25(OH)Vitamin D was also observed (P<0.001). In HAC group, UPC was significantly negatively correlated with 25(OH)Vitamin D (r(s): -0.54; P<0.01). Urinary NAG/C was significantly positively correlated with serum phosphate (r(s): 0.46; P=0.019). Increased serum phosphate, urinary excretion of calcium, and hyperparathyroidism were observed in dogs with HAC. Vitamin D metabolism may be shifted towards increased 1-alpha hydroxylation.

Correlation of calpain sensitivity, Bradford assay instability, and electrophoretic mobility in phosphomimetic mutants of GlyT2 N-terminus.

The glycine transporter GlyT2 plays an important role in glycine-inhibitory neurotransmission of the hindbrain and spinal cord. Its special feature is the extended N-terminus, which contains a large number of potentially phosphorylated serine and threonine residues. Due to its unstructured nature, it is difficult to address the changes introduced by potential phosphorylation. Here, we used relatively simple methods such as calpain sensitivity, Bradford instability, and SDS electrophoretic mobility shift to investigate the effect of multiple phosphomimetic mutations versus neutral mutations on GlyT2N properties. The replacement of several serines or threonines with neutral alanines did not have a significant effect on the studied GlyT2N properties. Replacement of the same residues with phosphomimetic aspartate resulted in significant alterations in calpain cleavage patterns, Bradford instability, and SDS gel protein mobility. Interestingly, a correlation between the relative intensity of the measured effects was observed, indicating that they all reflect similar structural changes introduced by potential phosphorylation in vivo. Results indicate that a potential single or multiple phosphorylation significantly alters the proteomic properties of the glycine transporter GlyT2 N-terminus. Assays can be helpful in the first screening of structurally significant and possibly phosphorylated residues in the N-terminus of GlyT2.

Bioprospecting of serratiopeptidase-producing bacteria from different sources.

Anti-inflammatory enzymes have wide applications in the pharmaceutical industry. The objective of this study was to find new and efficient strains for the commercial production of serratiopeptidase enzyme. Vast number of samples were processed for the isolation of potent strains. The experimental treatment includes processing of twenty soil samples, silkworm gut, and sugarcane stem. The total protein and protease activity was estimated by Lowry's method and casein hydrolysis. The HRBC stabilization assay was performed for finding the anti-inflammatory potential of all strains. The serratiopeptidase production was confirmed by HPLC with the standard. Molecular characterization of selected potent strains was done by 16S rDNA and confirmed the taxonomy. The one step rapid purification of serratiopeptidase was performed by Ultra three phase partitioning method. The clot lysis potential of the Serratia marcescens VS56 was observed by modified Holmstorm method. The results of the study revealed that among the 60 strains, 12 strains were protease-positive on skim milk agar plates and showed significant protease activity. All 12 strains were screened for serratiopeptidase using high-performance liquid chromatography (HPLC) and VS56, VS10, VS12 and VS18 showed a similar retention time (4.66 ± 0.10 min) with standard. The selected potent strain, Serratia marcescens VS56 showed a proteolytic activity of 21.30 units/mL and produced a total protein of 102 mg/mL. The HRBC suspension results also showed a percentage of 94.6 ± 1.00 protection, which was compared to the standard diclofenac. The clot lysis potential of Serratia marcescens VS56 was 53% in 4 h. Furthermore, the molecular weight of the protein was identified to confirm the presence of serratiopeptidase. The study hence contributed successfully to isolating, screening, and identifying a potent producer for serratiopeptidase from an environmental source. This inherent advantage of the strain will undoubtedly contribute much to the coco comm commercial production of serratiopeptidase in the near future.

Shwachman-Diamond syndrome mimicking mitochondrial hepatopathy.

Shwachman-Diamond syndrome (SDS) is a genetic disorder caused by mutations in the Shwachman-Bodian-Diamond syndrome (SBDS) gene. The syndrome is characterized by multiorgan dysfunction primarily involving the bone marrow and exocrine pancreas. Frequently overlooked is the hepatic dysfunction seen in early childhood which tends to improve by adulthood. Here, we report a child who initially presented with failure to thrive and elevated transaminases, and was ultimately diagnosed with SDS. A liver biopsy electron micrograph revealed hepatocytes crowded with numerous small mitochondria, resembling the hepatic architecture from patients with inborn errors of metabolism, including mitochondrial diseases. To our knowledge, this is the first report of the mitochondrial phenotype in an SDS patient. These findings are compelling given the recent cellular and molecular research studies which have identified SBDS as an essential regulator of mitochondrial function and have also implicated SBDS in the maintenance of mitochondrial DNA.

A two-photon fluorescent probe for highly selective detection of Cys over GSH and Hcy based on the Michael addition and transcyclization mechanism and its application in bioimaging and protein straining in SDS-PAGE.

BACKGROUND: Cysteine (Cys), glutathione (GSH), and homocysteine (Hcy), as three major biothiols are involved in a variety of physiological processes and play a crucial role in plant growth. Abnormal levels of Cys can cause plants to fail to grow properly. To date, although a very large number of fluorescent probes have been reported for the detection of biothiols, very few of them can be used for the selective discrimination of Cys from GSH and Hcy due to their structural similarity, and only a few of them can be used for plant imaging. RESULTS: Here, three fluorescent probes (o-/m-/p-TMA) based on TMN fluorophore and the ortho-/meta-/para-substituted maleimide recognition groups were constructed to investigate the selective response effect of Cys. Compared to the o-/m-TMA, p-TMA can selectively detect Cys over GSH and Hcy with a rapid response time (10 min) and a low detection limit (0.26 µM). The theoretical calculation confirmed that the intermediate p-TMA-Cys-int has shorter interatomic reaction distances (3.827 Å) compared to o-/m-TMA-Cys (5.533/5.287 Å), making it more suitable for further transcyclization reactions. Additionally, p-TMA has been employed for selective tracking of exogenous and endogenous Cys in Arabidopsis thaliana using both single-/two-photon fluorescence imaging. Furthermore, single cell walls produced obvious two-photon fluorescence signals, indicating that p-TMA can be used for high-concentration Cys analysis in single cells. Surprisingly, p-TMA can be used as a fluorescent dye for protein staining in SDS-PAGE with higher sensitivity (7.49 µg/mL) than classical Coomassie brilliant blue (14.11 µg/mL). SIGNIFICANCE: The outstanding properties of p-TMA make it a promising multifunctional molecular tool for the highly selective detection of Cys over GSH and Hcy in various complex environments, including water solutions, zebrafish, and plants. Additionally, it has the potential to be developed as a fluorescent dye for a simple and fast SDS-PAGE fluorescence staining method.

Current phenotypic and genetic spectrum of syndromic deafness in Tunisia: paving the way for precision auditory health.

Hearing impairment (HI) is a prevalent neurosensory condition globally, impacting 5% of the population, with over 50% of congenital cases attributed to genetic etiologies. In Tunisia, HI underdiagnosis prevails, primarily due to limited access to comprehensive clinical tools, particularly for syndromic deafness (SD), characterized by clinical and genetic heterogeneity. This study aimed to uncover the SD spectrum through a 14-year investigation of a Tunisian cohort encompassing over 700 patients from four referral centers (2007-2021). Employing Sanger sequencing, Targeted Panel Gene Sequencing, and Whole Exome Sequencing, genetic analysis in 30 SD patients identified diagnoses such as Usher syndrome, Waardenburg syndrome, cranio-facial-hand-deafness syndrome, and H syndrome. This latter is a rare genodermatosis characterized by HI, hyperpigmentation, hypertrichosis, and systemic manifestations. A meta-analysis integrating our findings with existing data revealed that nearly 50% of Tunisian SD cases corresponded to rare inherited metabolic disorders. Distinguishing between non-syndromic and syndromic HI poses a challenge, where the age of onset and progression of features significantly impact accurate diagnoses. Despite advancements in local genetic characterization capabilities, certain ultra-rare forms of SD remain underdiagnosed. This research contributes critical insights to inform molecular diagnosis approaches for SD in Tunisia and the broader North-African region, thereby facilitating informed decision-making in clinical practice.

The association of symptomatic vitreous floaters with depression and anxiety.

PURPOSE: To evaluate the levels of anxiety and depression in patients with symptomatic vitreous floaters and to determine the possible correlations of psychological implications with the symptoms duration and possible improvement, the degree of posterior vitreous detachment, and the discomfort severity. METHODS: Ninety patients complaining for floaters and fifty-seven age- and gender-matched healthy-control subjects were recruited. Every participant underwent a complete ophthalmological examination, including funduscopy and optical coherence tomography scans, while clinical and demographic data were also gathered. The Patient Health Questionnaire-9 (PHQ-9), the Zung Depression Inventory-Self-Rating Depression Scale (Zung SDS), and the Hospital Anxiety and Depression Scale (HADS) were completed by everyone. RESULTS: Between the studied groups, no significant differences were detected regarding the clinical and demographic data (p > 0.05). The patients with floaters had significantly higher scores of PHQ-9, Zung SDS, HADS Anxiety, and HADS Depression (p < 0.001). After adjustment for several confounders, PHQ-9 (p = 0.041), Zung SDS (p = 0.003), and HADS Anxiety (p = 0.036) values remained significantly impaired. Among the patients, PHQ-9 and Zung SDS scores were significantly elevated in the patients with floaters duration less than 4 weeks (p < 0.05). Finally, anxiety and depression were significantly correlated with the symptoms duration and intensity, with the floater-associated discomfort, and with the stage of posterior vitreous detachment. CONCLUSION: Vitreous floaters have a negative impact on patients' psychological status, by the terms of enhanced depressive and anxiety levels. To the best of our knowledge, our study is the first in the literature to elaborate the aforementioned association, by assessing three different questionnaires simultaneously.

Biochemical Fractionation of Human α-Synuclein in a Drosophila Model of Synucleinopathies.

Synucleinopathies are a group of central nervous system pathologies that are characterized by the intracellular accumulation of misfolded and aggregated α-synuclein in proteinaceous depositions known as Lewy Bodies (LBs). The transition of α-synuclein from its physiological to pathological form has been associated with several post-translational modifications such as phosphorylation and an increasing degree of insolubility, which also correlate with disease progression in post-mortem specimens from human patients. Neuronal expression of α-synuclein in model organisms, including Drosophila melanogaster, has been a typical approach employed to study its physiological effects. Biochemical analysis of α-synuclein solubility via high-speed ultracentrifugation with buffers of increasing detergent strength offers a potent method for identification of α-synuclein biochemical properties and the associated pathology stage. Unfortunately, the development of a robust and reproducible method for the evaluation of human α-synuclein solubility isolated from Drosophila tissues has remained elusive. Here, we tested different detergents for their ability to solubilize human α-synuclein carrying the pathological mutation A53T from the brains of aged flies. We also assessed the effect of sonication on the solubility of human α-synuclein and optimized a protocol to discriminate the relative amounts of soluble/insoluble human α-synuclein from dopaminergic neurons of the Drosophila brain. Our data established that, using a 5% SDS buffer, the three-step protocol separates cytosolic soluble, detergent-soluble and insoluble proteins in three sequential fractions according to their chemical properties. This protocol shows that sonication breaks down α-synuclein insoluble complexes from the fly brain, making them soluble in the SDS buffer and thus enriching the detergent-soluble fraction of the protocol.

"Exotic" seeds from Southern Africa as potential Novel Foods? - Chemical composition of manketti nuts (Schinziophyton rautanenii) and ushivi beans (Guibourtia coleosperma).

Various species of Southern African plants and their edible seeds have gained huge importance due to positive health aspects, and there is increasing interest to introduce such seeds as Novel Food on the international market. Especially the seeds of Schinziophyton rautanenii (manketti) and Guibourtia coleosperma (ushivi) could have great potential as a food and food ingredient. Hence, extensive analyses on the chemical composition of manketti nuts and ushivi beans including the analysis of total solids, protein and fat content, soluble carbohydrates, ash, total and free amino acids, biogenic amines and polyamines, fatty acid profile as well as the content of certain B-vitamins and tocopherols were performed. Results obtained showed a valuable nutritional composition, e.g., a true protein content of 22.6% with a ratio of essential amino acids to total amino acid composition of 48% in manketti nuts, while ushivi beans had a true protein content of 8.2% with a similar ratio of essential to total amino acids (45%). Lipid content was 54.1% in manketti nuts, ushivi beans had a value of 7.7%. In both, linoleic acid was the most abundant. Furthermore, ushivi beans had high amounts of vitamin B1 and B2.

Characteristics of higher depressive tendency in the patients with olfactory disorder.

OBJECTIVE: Most patients with olfactory dysfunction experience stress and anxiety because of the inconvenience and changes caused by the loss of olfaction. However, psychological assessment is not performed routinely in patients with olfactory dysfunction, and the characteristics of these patients with psychological depression are unclear. METHODS: In this study, we used the Self-rating Depression Scale to evaluate the degree of depression in patients who visited our clinic with olfactory dysfunction and examine the characteristics of these patients with strong depressive tendencies. Patients who visited our clinic between April 2019 and March 2020 with complaints of olfactory dysfunction were included in the study. RESULTS: A total of 180 patients (79 male and 101 female) underwent olfactory examination and completed the Self-rating Depression Scale. Eighty-six and 94 patients were included in the low depression and high depression groups, respectively. Binomial logistic regression analysis showed significant positive associations of Self-rating Depression Scale scores with female sex and the presence of parosmia/phantosmia (p < 0.05). CONCLUSION: In our study, approximately half of the patients with olfactory dysfunction had depressive tendencies especially in female and parosmia/phantosmia patients. We believe that psychological assessments, such as that with the SDS, can help identify patients with olfactory dysfunction who may be at a greater risk of developing depression.

Whey protein dynamics in goat mammary secretions during colostrum and early lactation periods.

The protein composition in goat milk undergoes changes throughout the different lactation periods, displaying distinct characteristics that are influenced by the dynamic nature of protein composition and concentration during the transition from colostrum secretion to mature milk. To evaluate the dynamics of whey proteins of Saanen goats during the colostral phase and the first month of lactation, 110 milk samples from 11 healthy mammary halves of seven Saanen goats were selected through a clinical evaluation. Whey was obtained by rennet coagulation of the mammary secretion. The biuret method determined total protein concentration, and their fractions were identified by 12% dodecyl sulfate-polyacrylamide gel electrophoresis. Maximum concentrations of all protein fractions were observed in the first 12 h of lactation, reducing throughout the study. Modification of the protein predominance was also observed. The transition from colostrum secretion to milk occurred 5 or 7 d postpartum.

High-pressure microfluidisation positively impacts structural properties and improves functional characteristics of almond proteins obtained from almond meal.

Almond protein isolate (API) obtained from almond meal was processed using dynamic high-pressure microfluidisation (0, 40, 80, 120, and 160 MPa pressure; single pass). Microfluidisation caused significant reductions in the particle size and increased absolute zeta potential. SDS-PAGE analysis indicated reduction in band intensity and the complete disappearance of bands beyond 80 MPa. Structural analysis (by circular dichroism, UV-Vis, and intrinsic-fluorescence spectra) of the API revealed disaggregation (up to 80 MPa) and then re-aggregation beyond 80 MPa. Significant increments in protein digestibility (1.16-fold) and the protein digestibility corrected amino acid score (PDCAAS; 1.15-fold) were observed for the API (80 MPa) than control. Furthermore, significant improvements (P < 0.05) in the functional properties were observed, viz., the antioxidant activity, protein solubility, and emulsifying properties. Overall, the results revealed that moderate microfluidisation treatment (80 MPa) is an effective and sustainable technique for enhancing physico-chemical and functional attributes of API, thus potentially enabling its functional food/nutraceuticals application.

Human and camel cystic echinococcosis - a polyclonal antibody-based sandwich ELISA for its serodiagnosis with molecular identification.

Cystic echinococcosis (CE) is an emergent neglected disease affecting human and animals in Egypt with a wide distribution and incidence. This study aimed to evaluate the use of a polyclonal antibody-based sandwich ELISA in the detection of Echinococcus granulosus antigen in human and camel sera. Hydatid cyst protoscoleces antigen (PsAg) was isolated from hydatid cysts collected from naturally infected camel livers and lungs. PsAg was used for immunization of rabbits to raise IgG polyclonal antibodies (IgG PsAb). IgG PsAb were then precipitated, purified using Protein-A Sepharose gel and labeled with horseradish peroxidase enzyme. We assayed the purity of the IgG PsAb, and the two prepared E. granulosus antigens CPsAg from camel cysts and HPsAg from human cysts by Sodium dodecyl-sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The resulted protein bands of the prepared CPsAg appeared at different molecular weights: 180, 90, 68, 54, 42 and 22 kDa while, HPsAg shared with it in 4 common bands at 68, 54, 42, and 22 kDa. The purified IgG PsAb had been resolved at two bands at 52 kDa and at 32 kDa. Sandwich ELISA were performed for the detection of circulating E. granulosus antigens in sera of human (n = 183) and camels (n = 190). The purified IgG PsAb showed strong reactivity against E. granulosus infected human and camel samples and no cross reactivity neither with free-healthy negative sera nor with others parasitic diseases (Schistosomiasis, Fascioliasis, Toxoplasmosis, Ancylostomiasis for human samples and Fascioliasis, ticks' infestation, Eimeriosis, Cryptosporidiosis, Nasal myiasis, Toxoplasmosis for camel samples). The sensitivity of the assay was 98.25% (56/57) and 96.9% (31/32) against human and camel samples, respectively. Specificity was 100% in both human and camel samples. Sandwich ELISA detected CE in 33.3% (24/72) and 55.6% (50/90) random human and camel samples, respectively. Indirect ELISA, using CPsAg, was used for detection of antibodies in positive human and camels' sera and detected 96.5% (55/57) and 93.8% (30/32) of human and camel samples, respectively. In our study, Genomic DNA was extracted from protoscoleces fluid of human liver hydatid cysts to identify the Echinococcus sp. isolate based on NADH dehydrogenase subunit 1 (NAD1) gene by Polymerase Chain Reaction (PCR) and the isolate (GenBank: OP785689.1) were identified as E. granulosus sensu lato genotype. In conclusion, Sandwich ELISA technique was found to be a potent and sensitive assay for detection of hydatid antigen in both human and camel samples.

Two-Dimensional Gel Electrophoresis for Protein Separation of Plant Samples.

This chapter provides a description of the procedure for two-dimensional electrophoresis that can be performed for any given gel size and isoelectric focusing range. This will enable the operator to recognize critical steps and gain sufficient information to generate 2D images suitable for computer-assisted analysis of 2D-gel, as well as mass spectrometry analysis for protein identification and characterization.

Physio-biochemical responses and crop performance analysis in chickpea upon botanical priming.

Chickpea is a highly nutritious protein-rich source and one of the major crops to alleviate global malnutrition, but poor seed quality affects its productivity. Seed quality is essential for better crop establishment and higher yields, particularly in the uncertain climate change. The present study investigated the impact of botanical priming versus hydropriming and bavistin seed treatment on chickpea seeds. A detailed physiological (germination percentage, root and shoot length, vigour index) and biochemical (amylase, protease, dehydrogenase, phytase, and lipid peroxidation) analysis was carried out in order to assess the effect of priming treatments. Turmeric-primed seeds showed better germination rate (94.5%), seedling length, enzyme activity, and lower malondialdehyde (MDA) content. Sodium dodecyl-sulfate polyacrylamide gel electrophoresis (SDS-PAGE) analysis revealed the expression of minor polypeptides of albumin and globulin in the primed seeds. Moreover, field experiments indicated increased crop growth, vigour, days to 50% flowering, yield and its attributing traits in turmeric-primed seeds. Botanical priming can increase chickpea yield by up to 16% over the control group. This low-cost and eco-friendly technique enhances seed and crop performance, making it a powerful tool for augmenting chickpea growth. Therefore, chickpea growers must adopt botanical priming techniques to enhance the quality of seed and crop performance. Moreover, this approach is environmentally sustainable and can help conserve natural resources in the long term. Therefore, this new approach must be widely adopted across the agricultural industry to ensure sustainable and profitable farming practices.

Residual protein analysis by SDS-PAGE in clinically manufactured BM-MSC products.

Residual substances that are considered hazardous to the recipient must be removed from final cellular therapeutic products manufactured for clinical purposes. In doing so, quality rules determined by competent authorities (CAs) for the clinical use of tissue- and cell-based products can be met. In our study, we carried out residual substance analyses, and purity determination studies of trypsin and trypsin inhibitor in clinically manufactured bone marrow-derived mesenchymal stromal/stem cell products, using the sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) method. Despite being a semiquantitative method, SDS-PAGE has several benefits over other methods for protein analysis, such as simplicity, convenience of use, and affordability. Due to its convenience and adaptability, SDS-PAGE is still a commonly used method in many laboratories, despite its limits in dynamic range and quantitative precision. Our goal in this work was to show that SDS-PAGE may be used effectively for protein measurement, especially where practicality and affordability are the major factors. The results of our study suggest a validated method to guide tissue and cell manufacturing sites for making use of an agreeable, accessible, and cost-effective method for residual substance analyses in clinically manufactured cellular therapies.

Determination of ROS Generated by Arabidopsis Xanthine Dehydrogenase1 (AtXDH1) Using Nitroblue Tetrazolium (NBT) and 3,3'-Diaminobenzidine (DAP).

Plants generate reactive oxygen species (ROS) during different metabolic processes, which play an essential role in coordinating growth and response. ROS levels are sensitive to environmental stresses and are often used as a marker for stress in plants. While various methods can detect ROS changes, histochemical staining with nitroblue tetrazolium (NBT) and 3,3'-diaminobenzidine (DAB) is a popular method, though it has faced criticism. This staining method is advantageous as it enables both the quantification and localization of ROS and the identification of the enzymatic origin of ROS in plants, cellular compartments, or gels. In this protocol, we describe the use of NBT and DAP staining to detect ROS generation under different stresses such as nitrogen starvation, wounding, or UV-C. Additionally, we describe the use of NBT staining for detecting enzymatic generation of ROS in native and native SDS PAGE gels. Our protocol also outlines the separation and comparison of the origin of ROS generated by xanthine dehydrogenase1 (XDH1) using different substrates.

An Evaluation of the Physicochemical Properties of Sesame Paste Produced by Ball Milling Compared against Conventional Colloid Milling.

The physicochemical characteristics and general food quality were greatly impacted by milling. In order to investigate the effect of milling technique for physicochemical properties of sesame paste of sesame paste, samples were prepared using ball mill and colloid mill by varying grinding times. The samples prepared by ball milling had the higher moisture contents (0.07% - 0.14%) than colloid milling (p < 0.05), except for colloid milling for one cycle (0.11%). The particle size curves showed the multimodal distributions. Compared to colloid milled samples, ball milled samples have smaller particle sizes and more uniform particle distribution. The L* values of samples prepared by ball milling were higher than colloid milling. The ball mill produced sesame paste with a wider range of hardness and silkier texture, and the samples made by ball milling for 30 min had the highest hardness. And the hardness of both CMS and BMS showed a decreasing trend with increasing grinding time. During ball milling, high-speed cutting and collision caused breakage of disulfide bonds, and the sesame proteins were decomposed to their subunits. In conclusions, ball milling may be an alternative and promising process for the preparation of sesame paste.

Computational Fluid Dynamics Analysis and Empirical Evaluation of Carboxymethylcellulose/Alginate 3D Bioprinting Inks for Screw-Based Microextrusion.

Three-dimensional microextrusion bioprinting technology uses pneumatics, pistons, or screws to transfer and extrude bioinks containing biomaterials and cells to print biological tissues and organs. Computational fluid dynamics (CFD) analysis can simulate the flow characteristics of bioinks in a control volume, and the effect on cell viability can be predicted by calculating the physical quantities. In this study, we developed an analysis system to predict the effect of a screw-based dispenser system (SDS) on cell viability in bioinks through rheological and CFD analyses. Furthermore, carboxymethylcellulose/alginate-based bioinks were used for the empirical evaluation of high-viscous bioinks. The viscosity of bioinks was determined by rheological measurement, and the viscosity coefficient for the CFD analysis was derived from a correlation equation by non-linear regression analysis. The mass flow rate derived from the analysis was successfully validated by comparison with that from the empirical evaluation. Finally, the cell viability was confirmed after bioprinting with bioinks containing C2C12 cells, suggesting that the developed SDS may be suitable for application in the field of bioengineering. Consequently, the developed bioink analysis system is applicable to a wide range of systems and materials, contributing to time and cost savings in the bioengineering industry.

Creating a novel genetic diversity of Trichoderma afroharzianum by γ-radiation for xylanase-cellulase production.

Creating novel sources of a microbial strain using induced mutation can increase enzyme production for industrial use. According to this, we have developed a mutant strain of Trichoderma afroharzianum by Co60 gamma irradiation. Trichoderma mutants were isolated from an optimum dose of 250 Gy. The qualitative and quantitative screening were used for evaluating their enzyme production and the DNA barcoding method was used to identify the best Trichoderma mutant isolates. The highest cellulase (exo-glucanase, endoglucanase, ß-glucosidase, and total cellulase) and xylanase activities were observed in superior mutant isolates of Trichoderma afroharzianum NAS107-M44 and Trichoderma afroharzianum NAS107-M82, which is approximately 1.6-2.5 times higher than its parent strain, respectively. The electrophoretic pattern of proteins showed that the exo-glucanase I, endo-glucanase III, and the xylanase I enzymes hydrolyzed the corn bran, synergistically. Overall, gamma irradiation-induced mutation could be an expedient technique to access such superior mutants for the bioconversion of corn bran wastes.