Biomaterials / bioinks and extrusion bioprinting.

Bioinks are formulations of biomaterials and living cells, sometimes with growth factors or other biomolecules, while extrusion bioprinting is an emerging technique to apply or deposit these bioinks or biomaterial solutions to create three-dimensional (3D) constructs with architectures and mechanical/biological properties that mimic those of native human tissue or organs. Printed constructs have found wide applications in tissue engineering for repairing or treating tissue/organ injuries, as well as in vitro tissue modelling for testing or validating newly developed therapeutics and vaccines prior to their use in humans. Successful printing of constructs and their subsequent applications rely on the properties of the formulated bioinks, including the rheological, mechanical, and biological properties, as well as the printing process. This article critically reviews the latest developments in bioinks and biomaterial solutions for extrusion bioprinting, focusing on bioink synthesis and characterization, as well as the influence of bioink properties on the printing process. Key issues and challenges are also discussed along with recommendations for future research.

Spray-dried porcine plasma enhances feed efficiency, intestinal integrity, and immune response of broilers challenged with necrotic enteritis.

Re-emergence of enteric diseases in the postantibiotic era has imposed severe loss to the poultry industry leading to the urgent need for appropriate additives to maintain gut health. Recently, more attention has been paid to animal plasma due to its high concentrations of active components such as albumins and globulins. The objective of this study was to evaluate the effects of spray-dried porcine plasma (SDP) supplementation during the starter phase (d 0-10) on growth performance, intestine health, and immune response of broilers under necrotic enteritis (NE) challenge. A total of 720 day-old male broiler parental line chicks (Ross 308) were randomly assigned to a 2 (NE challenge: no, yes) × 2 (SDP: 0, 2%) factorial arrangement with 12 replications of 15 chicks each. To induce NE, birds were inoculated with live Eimeria vaccine on d 9 and Clostridium perfringens on d 14. The body weight of birds and feed consumption were measured per pen on d 8, 10, 24, and 29 to calculate performance parameters. On d 16, three birds per pen were sampled to analyse the intestinal lesion score, gut permeability, villi morphology, relative weight of organs, and immune response. Results showed that SDP improved (P < 0.001) FCR in the pre-challenge phase (d 0-8). The results indicated that supplementing SDP lowered (P < 0.01) FCR at the end of the experiment (d 29). Dietary SDP decreased (P < 0.05) the concentration of FITC-d in serum samples of challenged broilers, although it did not affect the intestinal morphology and lesion score. Birds fed with SDP had a higher (P < 0.05) relative weight of bursa (g/kg live body weight) compared to non-supplemented birds. Supplementing SDP reduced the concentration of interleukin-6 (P < 0.05) and α-1 acid glycoprotein (P = 0.051) in serum samples of broilers. In conclusion, supplementation of SDP in the starter phase enhanced feed efficiency and gut integrity in NE challenged broilers, possibly through manipulating the immune response, while further studies targeting intestinal microflora and key genes are required to explore the mode of action.

Protocol for single case experimental design for yoga and sleep quality and inflammation: A two-hit model of sleep intervention.

Background: Poor sleep quality is a neglected clinical condition in the elderly that could predispose them to morbidities and even mortality. Several lines of clinical evidence support the potential of Meditative Movement Interventions (MMIs) in the alleviation of sleep-related disturbances in the elderly population. However, further studies are needed to provide more definitive evidence regarding the effectiveness of yoga-based MMI. Objective: The primary objective of this study is to evaluate the effectiveness of a repeated course of a yoga-based meditative intervention in a home environment to improve the sleep quality of elderly subjects. Method: A single-case experimental design with multiple baselines will be used to assess the effectiveness of Yoga-based meditative movement as an intervention for the alleviation of poor sleep quality in the elderly. A concomitant study will also be conducted to test the effectiveness of walking as an intervention using the identical design. We will recruit 6 participants with self-rated poor sleep quality (PSQI>) with external validation using actigraphy. Each participant will be randomly allocated to a different baseline phase (i.e., 7, 10, or 14 days), which will then be followed by a daily 45-min intervention over 12 weeks. The walking group will undergo daily walking. Conclusion: This single-case, multiple-baseline, between-case intervention randomization design will be the first report, wherein yoga-based intervention would be longitudinally monitored for changes in the objective measure of sleep quality. Trial registration: The registration number for this trial is CTRI/2021/02/031466.

Fabrication of chitosan/alginate/hydroxyapatite hybrid scaffolds using 3D printing and impregnating techniques for potential cartilage regeneration.

Three-dimensional (3D) printed hydrogel scaffolds enhanced with ceramics have shown potential applications for cartilage regeneration, but leaving biological and mechanical properties to be desired. This paper presents our study on the development of chitosan /alginate scaffolds with nano hydroxyapatite (nHA) by combining 3D printing and impregnating techniques, forming a hybrid, yet novel, structure of scaffolds for potential cartilage regeneration. First, we incorporated nHA into chitosan scaffold printing and studied the printability by examining the difference between the printed scaffolds and their designs. Then, we impregnated alginate with nHA into the printed chitosan scaffolds to forming a hybrid structure of scaffolds; and then characterized the scaffolds mechanically and biologically, with a focus on identifying the influence of nHA and alginate for potential cartilage regeneration. The results of compression tests on the scaffolds showed that the inclusion of nHA increased the elastic moduli of scaffolds; while the live/dead assay illustrated that nHA had a great effect on improving attachment and viability of ATCD5 cells on the scaffolds. Also, our results illustrated scaffolds with nHA impregnated in alginate hydrogel enhanced the cell viability and attachment. Furthermore, antibacterial activity of hybrid scaffolds was characterized with results indicating that the chitosan scaffolds had favourable antibacterial ability, which was further enhanced with the impregnated nHA. Taken together, our study has illustrated that chitosan/HA/alginate hybrid scaffolds are promising for cartilage regeneration and the methods developed to create hybrid scaffolds based on 3D printing and impregnating techniques, which can also be extended to fabricating scaffolds for other tissue engineering applications.

Sexing chickens (Gallus gallus domesticus) with high-resolution melting analysis using feather crude DNA.

Identification of sex in broiler chickens allows researchers to reduce the level of variation in an experiment caused by the sex effect. Broiler breeds commonly used in research are no longer feather sexable because of the change in their genetics. Other alternate sexing methods are costly and difficult to apply on a large scale. Therefore, a sexing method is required that is both cost effective and highly sensitive as well as having the ability to offer high throughput genotyping. In this study, high-resolution melting (HRM) analysis was used to detect DNA variations present in the gene chromodomain helicase DNA binding 1 protein (CHD1) on the Z and W chromosomes (CHD1Z and CHD1W, respectively) of chickens. In addition, a simplified DNA extraction protocol, which made use of the basal part of chicken feathers, was developed to speed up the sexing procedure. Three pairs of primers, that is, CHD1UNEHRM1F/R, CHD1UNEHRM2F/R, and CHD1UNEHRM3F/R, flanking the polymorphic regions between CHD1Z and CHD1W were used to differentiate male and female chickens via distinct melting curves, typical of homozygous or heterozygous genotypes. The assay was validated by the HRM-sexing of 1,318 broiler chicks and verified by examining the sex of the birds after dissection. This method allows for the sexing of birds within a couple of days, which makes it applicable for use on a large scale such as in nutritional experiments.

A novel leaf rust resistance gene introgressed from Aegilops markgrafii maps on chromosome arm 2AS of wheat.

KEY MESSAGE: A novel leaf rust resistance gene, LrM, introgressed from Aegilops markgrafii and mapped on chromosome 2AS using SSR- and SNP-based PCR markers will aid in broadening the genetic base of rust resistance in wheat. A new leaf rust resistance gene tentatively named LrM was introgressed from the diploid non-progenitor species Ae. markgrafii (2n = 2x = 14, genome CC) into common wheat using the nulli-5B mechanism. The introgression line ER9-700 showed a high degree of resistance against a wide spectrum of Puccinia triticina pathotypes. Genetic analysis was performed using the F1, F2, F2:3 and BC1F1 generations derived from the cross ER9-700/Agra Local. The results showed a single dominant gene for leaf rust resistance. The resistance gene LrM was mapped on chromosome arm 2AS using SSR- and SNP-based PCR markers. Preliminary mapping with SSR markers in the F2:3 population from the cross ER9-700/Agra Local identified two SSR markers flanking the LrM. SNPs were identified in the genomic region flanked by SSR markers, and SNP-based PCR markers were developed to construct the final map. Three SNP-based PCR markers co-segregated and mapped closest to the resistance gene at a distance of 2 cM. The gene LrM was distinguished from all the other genes designated and mapped on chromosome arm 2AS by molecular markers and rust reaction. All five markers used in the mapping amplified identical alleles in the donor Ae. markgrafii accession and introgression line ER9-700. The chromosomal location and rust reaction suggest that LrM is a novel leaf rust resistance gene that may be useful in broadening the genetic base of leaf rust resistance in wheat.

Micromechanisms of Cortical Bone Failure Under Different Loading Conditions.

Bone being a hierarchical composite material has a structure varying from macro- to nanoscale. The arrangement of the components of bone material and the bonding between fibers and matrix gives rise to its unique material properties. In this study, the micromechanisms of cortical bone failure were examined under different loading conditions using scanning electron microscopy. The experimental tests were conducted in longitudinal and transverse directions of bone diaphysis under tensile as well as compressive loading. The results show that bone material has maximum stiffness under longitudinal tensile loading, while the strength is higher under transverse compressive loading. A reverse trend of compressive mechanical properties of bone is observed for longitudinal and transverse loading as compared to trends reported in the previous studies. Therefore, micromechanisms of cortical bone failure were analyzed for different loading conditions to reveal such type of behavior of cortical bone and to correlate bone microstructure with mechanical response of bone.

Assessment of Elastic-Plastic Fracture Behavior of Cortical Bone Using a Small Punch Testing Technique.

The fracture properties of cortical bone are directly coupled to its complex hierarchical structure. The limited availability of bone material from many anatomic locations creates challenges for assessing the effect of bone heterogeneity and anisotropy on fracture properties. The small punch technique was employed to examine the fracture behavior of cortical bone in terms of area under the curve values obtained from load-load point displacement behavior. Fracture toughness of cortical bone was also determined in terms of J-toughness values obtained using a compact tension (CT) test. Area under the curve values obtained from the small punch test were correlated with the J-toughness values of cortical bone. The effects of bone density and compositional parameters on area under the curve and Jtoughness values were also analyzed using linear and multiple regression analysis. Area under the curve and J-toughness values are strongly and positively correlated. Bone density and %mineral content are positively correlated with both area under the curve and J-toughness values. The multiple regression analysis outcomes support these results. Overall, the findings support the hypothesis that area under the curve values obtained from small punch tests can be used to assess the fracture behavior of cortical bone.

Bioprinting of Vascularized Tissue Scaffolds: Influence of Biopolymer, Cells, Growth Factors, and Gene Delivery.

Over the past decades, tissue regeneration with scaffolds has achieved significant progress that would eventually be able to solve the worldwide crisis of tissue and organ regeneration. While the recent advancement in additive manufacturing technique has facilitated the biofabrication of scaffolds mimicking the host tissue, thick tissue regeneration remains challenging to date due to the growing complexity of interconnected, stable, and functional vascular network within the scaffold. Since the biological performance of scaffolds affects the blood vessel regeneration process, perfect selection and manipulation of biological factors (i.e., biopolymers, cells, growth factors, and gene delivery) are required to grow capillary and macro blood vessels. Therefore, in this study, a brief review has been presented regarding the recent progress in vasculature formation using single, dual, or multiple biological factors. Besides, a number of ways have been presented to incorporate these factors into scaffolds. The merits and shortcomings associated with the application of each factor have been highlighted, and future research direction has been suggested.

Studies on the Stress-Strain Relationship Bovine Cortical Bone Based on Ramberg-Osgood Equation.

Bone is a complex material that exhibits an amount of plasticity before bone fracture takes place, where the nonlinear relationship between stress and strain is of importance to understand the mechanism behind the fracture. This brief presents our study on the examination of the stress-strain relationship of bovine femoral cortical bone and the relationship representation by employing the Ramberg-Osgood (R-O) equation. Samples were taken and prepared from different locations (upper, middle, and lower) of bone diaphysis and were then subjected to the uniaxial tensile tests under longitudinal and transverse loading conditions, respectively. The stress-strain curves obtained from tests were analyzed via linear regression analysis based on the R-O equation. Our results illustrated that the R-O equation is appropriate to describe the nonlinear stress-strain behavior of cortical bone, while the values of equation parameters vary with the sample locations (upper, middle, and lower) and loading conditions (longitudinal and transverse).

3D biofabrication of vascular networks for tissue regeneration: A report on recent advances.

Rapid progress in tissue engineering research in past decades has opened up vast possibilities to tackle the challenges of generating tissues or organs that mimic native structures. The success of tissue engineered constructs largely depends on the incorporation of a stable vascular network that eventually anastomoses with the host vasculature to support the various biological functions of embedded cells. In recent years, significant progress has been achieved with respect to extrusion, laser, micro-molding, and electrospinning-based techniques that allow the fabrication of any geometry in a layer-by-layer fashion. Moreover, decellularized matrix, self-assembled structures, and cell sheets have been explored to replace the biopolymers needed for scaffold fabrication. While the techniques have evolved to create specific tissues or organs with outstanding geometric precision, formation of interconnected, functional, and perfused vascular networks remains a challenge. This article briefly reviews recent progress in 3D fabrication approaches used to fabricate vascular networks with incorporated cells, angiogenic factors, proteins, and/or peptides. The influence of the fabricated network on blood vessel formation, and the various features, merits, and shortcomings of the various fabrication techniques are discussed and summarized.

Performance, egg quality, and liver lipid reserves of free-range laying hens naturally infected with Ascaridia galli.

A study was conducted to determine the performance, egg quality, and liver lipid reserves of laying hens exposed to ranges contaminated with Ascaridia galli. Sixteen-week-old Lohmann Brown laying hens (n = 200) were divided into 4 treatments with 5 replicates containing 10 hens per pen. Hens of treatment 1 [negative control (NC)] ranged on a decontaminated area, and hens of treatments 2 (low infection) and 3 (medium infection) ranged on areas previously contaminated by hens artificially infected with 250 and 1,000 embryonated A. galli eggs, respectively. The hens of treatment 4 [positive control (PC)] ranged on areas previously contaminated by hens artificially infected with 2,500 embryonated A. galli eggs, and in addition these hens were orally inoculated with 1,000 embryonated eggs. Results indicated that hens of the medium infection group had a higher number of intestinal A. galli worms and A. galli eggs in the coprodeum excreta (43.9 ± 4.0 and 3,437 ± 459 eggs/g) compared to hens of the low infection group (23.8 ± 4.0 and 1,820 ± 450 eggs/g) (P < 0.01) and similar worm counts to PC hens (34.4 ± 4.0 and 2,918 ± 474) (P > 0.05). Egg production, egg mass, feed intake, and feed conversion ratio (FCR) were not affected by A. galli infection (P > 0.05). Egg quality parameters (egg weight, shell reflectivity, shell weight, shell thickness, shell percentage, shell breaking strength, deformation, albumen height, Haugh unit, and yolk score) were not affected by A. galli infection (P > 0.05). Highly infected hens had lower liver lipid content (2.72 ± 0.51 g) compared to uninfected hens (4.46 ± 0.58 g, P < 0.01). The results indicate that exposure to ranges contaminated with A. galli resulted in infection of the ranging hens, but this did not affect egg production or egg quality. Infection with A. galli lowered the liver lipid reserves of the host significantly, suggesting infected hens use more energy reserves for maintenance and production.

Effect of an artificial Ascaridia galli infection on egg production, immune response, and liver lipid reserve of free-range laying hens.

This study was conducted to determine the effect of Ascaridia galli infection on free-range laying hens. Lohmann Brown laying hens (n = 200) at 17 wk of age were allocated to 4 treatment groups (n = 50 per group), each with 5 replicate pens of 10 hens. Hens in 3 treatment groups were orally inoculated with different doses of embryonated A. galli eggs: low (250 eggs), medium (1,000 eggs), and high (2,500 eggs) levels, whereas hens of the control group were not infected. Infection levels were monitored using excreta egg counts and mature A. galli worm counts in the intestine. Anti A. galli antibody titers (IgY) in the serum were measured prior to infection, and at 6, 11, 15, and 20 wk post infection (PI) and in egg yolk at 11 and 20 wk PI. Parameters evaluated included feed intake, egg production, egg weight, egg mass, FCR, liver weight, liver fat, and intra epithelial immune cell infiltration. The results showed no difference in feed intake, body weight, or FCR among any treatment groups (P > 0.05). Egg production was lower in the low infection group compared to other groups at 20 wk of age (P < 0.01). Serum IgY was higher in the infected groups' hens at 20 wk PI compared to control group hens (P < 0.01). Yolk IgY increased significantly over time and was higher in infected hens compared to hens of the control group at 11 and 20 wk PI (P < 0.001). No differences were observed in liver lipid content or intraepithelial lymphocytes infiltration among treatment groups. Ascaridia galli eggs in the coprodeum content and adult A. galli worm count were higher in infected hens compared to hens of the control group (P < 0.01). In conclusion, the effects of artificial infection with A. galli on the parameters investigated were minor, and egg yolk antibody may be a more reliable indicator of A. galli infection than serum antibody or excreta egg count.

Inflammasome gene profile is modulated in septic patients, with a greater magnitude in non-survivors.

Inflammasome signalling induces the processing and secretion of interleukin (IL)-1ß and IL-18 which, coupled with pyroptosis, activate further the inflammatory response. In the present study we evaluated the expression of genes involved in inflammasome signalling pathways in septic patients, their interaction networks and the predicted functions modulated in survivors and non-survivors. Twenty-seven patients with sepsis secondary to community-acquired pneumonia admitted to intensive care units from three general hospitals in São Paulo were included into the study. We performed a polymerase chain reaction (PCR) array encompassing 35 genes related to the nucleotide-binding oligomerization domain and leucine-rich repeat-containing (NLR)-inflammasome in peripheral blood mononuclear cells obtained at admission and after 7 days of follow-up. Eleven healthy volunteers were used as the reference group. Increased NLRC4 and NLRP3 and decreased nucleotide-binding oligomerization domain (NOD1), and NLRP1 expression was observed in septic patients compared to healthy individuals; the IL-1ß and IL-18 expression levels were also high in the patients. The gene expression changes followed the same patterns in surviving and non-surviving patients, with higher magnitudes observed in non-survivors. Functional analyses revealed, however, that activation and inhibition intensity for representing functions were different in survivors and non-survivors, as for production of reactive oxygen species, synthesis of nitric oxide and for the control of bacterial infections. Our results showed that the genes involved in the activation of the NLR-inflammasome cascades were altered substantially in septic patients, with a higher number of altered genes and a higher intensity in the disturbance of gene expression found among patients dying of sepsis.

User-independent EBSD parameters to study the progress of recovery and recrystallization in Cu-Zn alloy during in situ heating.

Microstructural evolution of cold-rolled Cu-5%Zn alloy during in situ heating inside field-emission scanning electron microscope was utilized to obtain user-independent parameters in order to trace the progress of static recovery and recrystallization. Electron back-scattered diffraction (EBSD)-based orientation imaging microscopy was used to obtain micrographs at various stages of in situ heating. It is shown that unlike the pre-existing methods, additional EBSD-based parameter can be used to trace the progress of recovery and recrystallization, which is not dependent on user input and hence less prone to error. True strain of 0.3 was imposed during cold rolling of alloy sample. Rolled sample was subjected to in situ heating from room temperature to 500°C (∼0.58 Tm) with soaking time of 10 min, at each of the intermediate temperatures viz. 100, 200, 300, 400 and 450°C. After reaching 500°C, the sample was kept at this temperature for a maximum duration of around 15 h. The sample showed clear signs of recovery for temperature up to 450°C, and at 500°C, recrystallization started to take place. Recrystallization kinetics was moderate, and full recrystallization was achieved in approximately 120 min. We found that EBSD parameter, namely, band contrast intensity can be used as an extra handle to map out the progress of recrystallization occurring in the sample. By contrast, mean angular deviation can be used to understand the evolution of recovery in samples. The parameters mentioned in the current study, unlike other pre-existing methods, can also be used for mapping local microstructural transformations due to recovery and recrystallization. We discuss the benefits and limitations in using these additional handles in understanding the changes taking place in the material during in situ heating.

Morpho-histological and ultra architectural changes during early development of endangered golden mahseer Tor putitora.

Ultrastructural and histological changes in the embryonic and larval surface during ontogenesis of the endangered golden mahseer Tor putitora is studied here for the first time. Embryonic development was completed 91-92 h after fertilization at an ambient temperature of 23° ± 1° C (mean ± s.d.). The gastrula stage was characterized by presence of the Kupffer's vesicle, notochord, ectoderm and endoderm cells. Primordial germ cells were clearly identifiable from c. 55 h post-fertilization at the organogenesis stage. Mean total length of newly hatched larvae was 7·0 ± 0·5 mm. Scanning electron microscopy of newly hatched larvae demonstrated vitelline arteries, microridged epithelial cells and mucous gland openings over much of the body surface. Eye, oral cavity, pharyngeal arches, heart, intestinal loop, prosencephalon, cephalic vesicle and nasal epithelium were clearly distinguished in 3 day old hatched individuals. In 6 day old individuals, caudal-fin rays and internal organs were evident. The dorsal fin became prominent at this stage and larvae began swimming at the surface. The reserved yolk material was totally absorbed 8-11 days after hatching and larvae began feeding exogenously. Tor putitora exhibited a longer early developmental period than other cyprinids reared at similar temperatures.

Economics, energy, and environmental assessment of diversified crop rotations in sub-Himalayas of India.

Reducing the carbon footprint and increasing energy use efficiency of crop rotations are the two most important sustainability issues of the modern agriculture. Present study was undertaken to assess economics, energy, and environmental parameters of common diversified crop rotations (maize-tomato, and maize-toria-wheat) vis-a-vis traditional crop rotations like maize-wheat, maize + ginger and rice-wheat of the north-western Himalayan region of India. Results revealed that maize-tomato and maize + ginger crop rotations being on par with each other produced significantly higher system productivity in terms of maize equivalent yield (30.2-36.2 t/ha) than other crop rotations (5.04-7.68 t/ha). But interestingly in terms of energy efficiencies, traditional maize-wheat system (energy efficiency 7.9, human energy profitability of 177.8 and energy profitability of 6.9 MJ/ha) was significantly superior over other systems. Maize + ginger rotation showed greater competitive advantage over other rotations because of less consumption of non-renewable energy resources. Similarly, maize-tomato rotation had ability of the production process to exploit natural resources due to 14-38% less use of commercial or purchased energy sources over other crop rotations. Vegetable-based crop rotations (maize + ginger and maize-tomato) maintained significantly the least carbon footprint (0.008 and 0.019 kg CO2 eq./kg grain, respectively) and the highest profitability (154,322 and 274,161 Rs./ha net return, respectively) over other crop rotations. As the greatest inputs of energy and carbon across the five crop rotations were nitrogen fertilizer (15-29% and 17-28%, respectively), diesel (14-24% and 8-19%, respectively) and irrigation (10-27% and 11-44%, respectively), therefore, alternative sources like organic farming, conservation agriculture practices, soil and water conservation measures, rain water harvesting etc. should be encouraged to reduce dependency of direct energy and external carbon inputs particularly in sub-Himalayas of India.

Novice lifters exhibit a more kyphotic lifting posture than experienced lifters in straight-leg lifting.

As torso flexion and repetitive lifting are known risk factors for low back pain and injury, it is important to investigate lifting techniques that might reduce injury during repetitive lifting. By normalizing lumbar posture to a subject's range of motion (ROM), as a function of torso flexion, this research examined when subjects approached their range of motion limits during dynamic lifting tasks. For this study, it was hypothesized that experienced lifters would maintain a more neutral lumbar angle relative to their range of motion, while novice lifters would approach the limits of their lumbar ROM during the extension phase of a straight-leg lift. The results show a statistically significant difference in lifting patterns for these two groups supporting this hypothesis. The novice group maintained a much more kyphotic lumbar angle for both the flexion (74% of the lumbar angle ROM) and extension phases (86% of the lumbar angle ROM) of the lifting cycle, while the experienced group retained a more neutral curvature throughout the entire lifting cycle (37% of lumbar angle ROM in flexion and 48% of lumbar angle ROM in extension). By approaching the limits of their range of motion, the novice lifters could be at greater risk of injury by placing greater loads on the supporting soft tissues of the spine. Future research should examine whether training subjects to assume more neutral postures during lifting could indeed lower injury risks.