Protocell Flow Reactors for Enzyme and Whole-cell Mediated Biocatalysis.

The design and construction of continuous flow biochemical reactors comprising immobilized biocatalysts have generated great interest in the efficient synthesis of value-added chemicals. Living cells use compartmentalization and reaction-diffusion processes for spatiotemporal regulation of biocatalytic reactions and implementing these strategies into continuous flow reactors could offer new opportunities in reactor design and application. Herein, we demonstrate the fabrication of protocell-based continuous flow reactors for enzyme and whole-cell mediated biocatalysis. We employ semipermeable membranized coacervate vesicles as model protocells that spontaneously sequester enzymes or accumulate living bacteria to produce embodied microreactors capable of single- or multiple-step catalytic reactions. By packing millions of the enzyme/bacteria-containing coacervate vesicles in a glass column, we demonstrate a facile, cost-effective, and modular methodology capable of performing oxidoreductase, peroxidase and lipolytic reactions, enzyme-mediated L-DOPA synthesis and whole-cell glycolysis under continuous flow conditions. We show that the protocell-nested enzymes and bacterial cells exhibit enhanced activities and stability under deleterious operating conditions compared with their non-encapsulated counterparts. Our results provide a step towards the engineering of continuous flow reactors based on cell-like microscale agents and offer opportunities in the development of green and sustainable industrial bioprocessing. This article is protected by copyright. All rights reserved.

Bacterial Surface Appendages Modulate the Antimicrobial Activity Induced by Nanoflake Surfaces on Titanium.

Bioinspired nanotopography is a promising approach to generate antimicrobial surfaces to combat implant-associated infection. Despite efforts to develop bactericidal 1D structures, the antibacterial capacity of 2D structures and their mechanism of action remains uncertain. Here, hydrothermal synthesis is utilized to generate two 2D nanoflake surfaces on titanium (Ti) substrates and investigate the physiological effects of nanoflakes on bacteria. The nanoflakes impair the attachment and growth of Escherichia coli and trigger the accumulation of intracellular reactive oxygen species (ROS), potentially contributing to the killing of adherent bacteria. E. coli surface appendages type-1 fimbriae and flagella are not implicated in the nanoflake-mediated modulation of bacterial attachment but do influence the bactericidal effects of nanoflakes. An E. coli ΔfimA mutant lacking type-1 fimbriae is more susceptible to the bactericidal effects of nanoflakes than the parent strain, while E. coli cells lacking flagella (ΔfliC) are more resistant. The results suggest that type-1 fimbriae confer a cushioning effect that protects bacteria upon initial contact with the nanoflake surface, while flagella-mediated motility can lead to elevated membrane abrasion. This finding offers a better understanding of the antibacterial properties of nanoflake structures that can be applied to the design of antimicrobial surfaces for future medical applications.

Nanotextured titanium inhibits bacterial activity and supports cell growth on 2D and 3D substrate: A co-culture study.

Medical implant-associated infections pose a significant challenge to modern medicine, with aseptic loosening and bacterial infiltration being the primary causes of implant failure. While nanostructured surfaces have demonstrated promising antibacterial properties, the translation of their efficacy from 2D to 3D substrates remains a challenge. Here, we used scalable alkaline etching to fabricate nanospike and nanonetwork topologies on 2D and laser powder-bed fusion printed 3D titanium. The fabricated surfaces were compared with regard to their antibacterial properties against Staphylococcus aureus, Escherichia coli, and Pseudomonas aeruginosa, and mesenchymal stromal cell responses with and without the presence of bacteria. Finite elemental analysis assessed the mechanical properties and permeability of the 3D substrate. Our findings suggest that 3D nanostructured surfaces have potential to both prevent implant infections and allow host cell integration. This work represents a significant step towards developing effective and scalable fabrication methods on 3D substrates with consistent and reproducible antibacterial activity, with important implications for the future of medical implant technology.

Enhanced and Stem-Cell-Compatible Effects of Nature-Inspired Antimicrobial Nanotopography and Antimicrobial Peptides to Combat Implant-Associated Infection.

Nature-inspired antimicrobial surfaces and antimicrobial peptides (AMPs) have emerged as promising strategies to combat implant-associated infections. In this study, a bioinspired antimicrobial peptide was functionalized onto a nanospike (NS) surface by physical adsorption with the aim that its gradual release into the local environment would enhance inhibition of bacterial growth. Peptide adsorbed on a control flat surface exhibited different release kinetics compared to the nanotopography, but both surfaces showed excellent antibacterial properties. Functionalization with peptide at micromolar concentrations inhibited Escherichia coli growth on the flat surface, Staphylococcus aureus growth on the NS surface, and Staphylococcus epidermidis growth on both the flat and NS surfaces. Based on these data, we propose an enhanced antibacterial mechanism whereby AMPs can render bacterial cell membranes more susceptible to nanospikes, and the membrane deformation induced by nanospikes can increase the surface area for AMPs membrane insertion. Combined, these effects enhance bactericidal activity. Since functionalized nanostructures are highly biocompatible with stem cells, they make promising candidates for next generation antibacterial implant surfaces.

Does Self-Sacrifice Make Me Great? Research on the Relationship Between Employee Conscientiousness and Pro-Social Rule Breaking.

Based on the theory of purposeful work behavior, this study proposed that the two facets of employee conscientiousness, namely duty orientation and achievement orientation, have opposite effects on pro-social rule breaking (PSRB). We also explored the moderating effect of employees' task characteristic (job autonomy) and social characteristic (leader reward omission) on the above relationships. Using two-wave data collected from 216 employee-supervisor dyads, we found that duty orientation was positively related to PSRB, while achievement orientation was negatively related to PSRB. Further, job autonomy, by amplifying employees' perceived meaningfulness of their higher-order implicit goals, can strengthen the positive effect of duty orientation and the negative effect of achievement orientation on PSRB. Similarly, leader reward omission could also activate the negative effect of achievement orientation and PSRB, but not significantly moderate the positive relationship between duty orientation and PSRB. By separating the distinct role of facet-specific personality, our study sheds light on the relationship between employee conscientiousness and PSRB.

DDIT3/CHOP promotes autophagy in chondrocytes via SIRT1-AKT pathway.

Endoplasmic reticulum (ER) stress can initiate autophagy via unfolded protein response (UPR). As a key downstream gene of UPR, DDIT3/CHOP is expressed in chondrocytes. However, the regulation mechanism of DDIT3/CHOP on autophagy in chondrocytes remains unclear. In this study, the expression levels of autophagic markers Beclin1 and LC3B were found to decrease while p62 increase in the tibial growth plate and costal primary chondrocytes from DDIT3/CHOP KO mice. In vitro, overexpressing DDIT3/CHOP induced autophagy in ATDC5 chondrocytes, displaying an elevated immunofluorescence signal of LC3B and elevated numbers of autophagosomes and autolysosomes. Analysis of the gain- and loss-of-function indicated that the protein level of Beclin1 and the ratio of LC3BII/I increased in DDIT3/CHOP overexpression cells, whereas decreased in DDIT3/CHOP knockdown cells. The decreased level of p62 and additional accumulation of LC3BII caused by chloroquine (CQ) further indicated that DDIT3/CHOP enhanced autophagic flux. Mechanistically, we found that DDIT3/CHOP binds directly to the promoter of SIRT1 to promote its expression by CHIP, qRT-PCR, and Western blot analysis. In addition, SIRT1 enhanced autophagic activity in ATDC5 cells, and inhibition or activation of SIRT1 partially reversed the effect of overexpressing or downregulating DDIT3/CHOP on autophagy. Furthermore, AKT signaling was found to be responsible for DDIT3/CHOP-regulated autophagy in ATDC5 cells. SIRT1 knockdown reversed the effect of DDIT3/CHOP overexpression on AKT signaling. In conclusion, our data clarifies that DDIT3/CHOP promotes autophagy in ATDC5 chondrocytes through the SIRT1-AKT pathway. These results were also confirmed in the primary chondrocytes.

When and How Workplace Helping Promotes Deviance? An Actor-Centric Perspective.

Despite the vast academic interest in workplace helping, little is known about the impact of different types of helping behaviors on physiological and behavioral ramifications of helpers. By taking the actor-centric perspective, this study attempts to investigate the differential impacts of three kinds of helping behaviors (caring, coaching, and substituting helping) on helpers themselves from the theory of resource conservation. To test our model, 512 Chinese employees were surveyed, utilizing a three-wave time-lagged design, and we found that caring and coaching helping were negatively associated with workplace deviance, whereas substituting helping was positively associated with subsequent workplace deviance. Emotional exhaustion mediated the effects of three helping behaviors on subsequent workplace deviance. Moreover, employees' extrinsic career goals influenced the strength of the relationship between three helping behaviors and emotional exhaustion and the indirect effects of three helping behaviors on subsequent workplace deviance via emotional exhaustion. We discuss the implications of our findings for both theories and practices.

Identifying Candidate Genes for Hypoxia Adaptation of Tibet Chicken Embryos by Selection Signature Analyses and RNA Sequencing.

The Tibet chicken (Gallus gallus) lives on the Qinghai-Tibet Plateau and adapts to the hypoxic environment very well. The objectives of this study was to obtain candidate genes associated with hypoxia adaptation in the Tibet chicken embryos. In the present study, we used the fixation index (Fst) and cross population extended haplotype homozygosity (XPEHH) statistical methods to detect signatures of positive selection of the Tibet chicken, and analyzed the RNA sequencing data from the embryonic liver and heart with HISAT, StringTie and Ballgown for differentially expressed genes between the Tibet chicken and White leghorn (Gallus gallus, a kind of lowland chicken) embryos hatched under hypoxia condition. Genes which were screened out by both selection signature analysis and RNA sequencing analysis could be regarded as candidate genes for hypoxia adaptation of chicken embryos. We screened out 1772 genes by XPEHH and 601 genes by Fst, and obtained 384 and 353 differentially expressed genes in embryonic liver and heart, respectively. Among these genes, 89 genes were considered as candidate genes for hypoxia adaptation in chicken embryos. ARNT, AHR, GSTK1 and FGFR1 could be considered the most important candidate genes. Our findings provide references to elucidate the molecular mechanism of hypoxia adaptation in Tibet chicken embryos.

Stick to Convention or Bring Forth the New? Research on the Relationship Between Employee Conscientiousness and Job Crafting.

Integrating regulatory focus theory and personality literature, we develop and test a moderated mediation model to specify the mediation mechanisms and boundary conditions of the association between employee conscientiousness and job crafting. Two-wave data collected from 389 employees and 95 supervisors showed that: Employee conscientiousness was positively associated with work promotion focus and work prevention focus. Employee conscientiousness was positively related to job crafting via work promotion focus, negatively related to job crafting via work prevention focus. Error management climate positively moderated the relationship between employee conscientiousness and work promotion focus, negatively moderated the relationship between employee conscientiousness and work prevention focus. The indirect relationship between employee conscientiousness and job crafting through work promotion focus was more pronounced under positive error management climate rather than negative, whereas the indirect relationship through work prevention focus was more pronounced under negative error management climate rather than positive.

Genome-Wide Association Study and Transcriptome Differential Expression Analysis of the Feather Rate in Shouguang Chickens.

The feather rate phenotype in chicks, including early-feathering and late-feathering phenotypes, are widely used as a sexing system in the poultry industry. The objective of this study was to obtain candidate genes associated with the feather rate in Shouguang chickens. In the present study, we collected 56 blood samples and 12 hair follicle samples of flight feathers from female Shouguang chickens. Then we identified the chromosome region associated with the feather rate by genome-wide association analysis (GWAS). We also performed RNA sequencing and analyzed differentially expressed genes between the early-feathering and late-feathering phenotypes using HISAT2, StringTie, and DESeq2. We identified a genomic region of 10.0-13.0 Mb of chromosome Z, which is statistically associated with the feather rate of Shouguang chickens at one-day old. After RNA sequencing analysis, 342 differentially expressed known genes between the early-feathering (EF) and late-feathering (LF) phenotypes were screened out, which were involved in epithelial cell differentiation, intermediate filament organization, protein serine kinase activity, peptidyl-serine phosphorylation, retinoic acid binding, and so on. The sperm flagellar 2 gene (SPEF2) and prolactin receptor (PRLR) gene were the only two overlapping genes between the results of GWAS and differential expression analysis, which implies that SPEF2 and PRLR are possible candidate genes for the formation of the chicken feathering phenotype in the present study. Our findings help to elucidate the molecular mechanism of the feather rate in chicks.

YAP1 influences differentiation of osteoblastic MC3T3-E1 cells through the regulation of ID1.

Yes-associated protein 1 (YAP1) transcriptional coactivator has recently been identified to regulate skeletal lineage cell differentiation and bone development. However, the role and molecular mechanisms of YAP1 in the regulation of osteoblastic differentiation remains to be elucidated. In this study, we demonstrated that YAP1 expression was increased during osteogenic differentiation of rat bone mesenchymal stem cells and MC3T3-E1. YAP1 overexpression MC3T3-E1 showed increased expression of osteogenesis markers, such as runt-related transcription factor 2, osteocalcin, and osteopontin, as well as alkaline phosphatase and alizarin red staining. Conversely, YAP1 knockdown significantly suppressed MC3T3-E1 osteoblastic differentiation. Mechanistically, we found that YAP1 overexpression upregulated the mRNA and protein expression of the inhibitor of differentiation/DNA binding 1 (ID1), which was contrary to the results of YAP1-knockdown group. Moreover, the early osteogenic differentiation of MC3T3-E1 cells was enhanced by ID1 overexpression. Furthermore, transient transfection with exogenous ID1 overexpression plasmid completely recaptured the decreased effects of YAP1 knockdown on MC3T3-E1 cell differentiation. In addition, ß-catenin and AMP-activated protein kinase signaling pathways participated in YAP1 regulation processes. Taken together, our study suggests that YAP1 is a crucial modulator of osteoblast differentiation in vitro, and provides insight into the mechanism by which YAP1 regulates osteoblast differentiation.

DDIT3 regulates cementoblast mineralization by isocitrate dehydrogenase 1 through nuclear factor-κB pathway.

DDIT3 is of great importance in endoplasmic reticulum stress and is involved in many inflammatory diseases and mineralization processes. The cementum protects teeth from periodontitis and provides attachment for Sharpey's fibers of the periodontal ligament. However, the effect of DDIT3 on cementoblast differentiation remains largely unknown. In this study, we found that DDIT3 was suppressed during cementoblast differentiation. Knockdown of DDIT3 increased the messenger RNA (mRNA) and protein levels of several key osteogenic markers in vitro, including alkaline phosphatase, runt-related transcription factor 2, and osteocalcin (OCN). In addition, isocitrate dehydrogenase 1 (IDH1) was increased during cementoblast differentiation, and knockdown of DDIT3 increased the protein and mRNA levels of IDH1. Furthermore, inhibition of IDH1 could partially reduce the effect of DDIT3 on cementoblast differentiation. The DDIT3 knockdown activated nuclear factor-κB (NF-κB) transcriptional activity and upregulated the expression of p-p65 and p-IκBα. The increased osteogenic differentiation ability and IDH1 expression, as induced by the DDIT3 knockdown, could be partially turned over by the addition of NF-κB inhibitor BAY 11-7082. Overall, our data clarified that DDIT3 suppresses cementoblast differentiation through IDH1, via the NF-κB pathway.

Image Reconstruction Under Contact Impedance Effect in Micro Electrical Impedance Tomography Sensors.

Contact impedance has an important effect on micro electrical impedance tomography (EIT) sensors compared to conventional macro sensors. In the present work, a complex contact impedance effect ratio ξ is defined to quantitatively evaluate the effect of the contact impedance on the accuracy of the reconstructed images by micro EIT. Quality of the reconstructed image under various ξ is estimated by the phantom simulation to find the optimum algorithm. The generalized vector sampled pattern matching (GVSPM) method reveals the best image quality and the best tolerance to ξ. Moreover, the images of yeast cells sedimentary distribution in a multilayered microchannel are reconstructed by the GVSPM method under various mean magnitudes of contact impedance effect ratio |ξ|. The result shows that the best image quality that has the smallest voltage error UE = 0.581 is achieved with measurement frequency f = 1 MHz and mean magnitude |ξ| = 26. In addition, the reconstructed images of cells distribution become improper while f < 10 kHz and mean value of |ξ| > 2400.