Topographic Guidance in Melt-Electrowritten Tubular Scaffolds Enhances Engineered Kidney Tubule Performance

Introduction: To date, tubular tissue engineering relies on large, non-porous tubular scaffolds (Ø > 2 mm) for mechanical self-support, or smaller (Ø 150–500 μm) tubes within bulk hydrogels for studying renal transport phenomena. To advance the engineering of kidney tubules for future implantation, constructs should be both self-supportive and yet small-sized and highly porous. Here, we hypothesize that the fabrication of small-sized porous tubular scaffolds with a highly organized fibrous microstructure by means of melt-electrowriting (MEW) allows the development of self-supported kidney proximal tubules with enhanced properties.Materials and Methods: A custom-built melt-electrowriting (MEW) device was used to fabricate tubular fibrous scaffolds with small diameter sizes (Ø = 0.5, 1, 3 mm) and well-defined, porous microarchitectures (rhombus, square, and random). Human umbilical vein endothelial cells (HUVEC) and human conditionally immortalized proximal tubular epithelial cells (ciPTEC) were seeded into the tubular scaffolds and tested for monolayer formation, integrity, and organization, as well as for extracellular matrix (ECM) production and renal transport functionality.Results: Tubular fibrous scaffolds were successfully manufactured by fine control of MEW instrument parameters. A minimum inner diameter of 1 mm and pore sizes of 0.2 mm were achieved and used for subsequent cell experiments. While HUVEC were unable to bridge the pores, ciPTEC formed tight monolayers in all scaffold microarchitectures tested. Well-defined rhombus-shaped pores outperformed and facilitated unidirectional cell orientation, increased collagen type IV deposition, and expression of the renal transporters and differentiation markers organic cation transporter 2 (OCT2) and P-glycoprotein (P-gp).Discussion and Conclusion: Here, we present smaller diameter engineered kidney tubules with microgeometry-directed cell functionality. Due to the well-organized tubular fiber scaffold microstructure, the tubes are mechanically self-supported, and the self-produced ECM constitutes the only barrier between the inner and outer compartment, facilitating rapid and active solute transport.

Topographic Guidance in Melt-Electrowritten Tubular Scaffolds Enhances Engineered Kidney Tubule Performance

To advance the engineering of kidney tubules for future implantation, constructs should be both self-supportive and yet small-sized and highly porous. Here, we hypothesize that the fabrication of small-sized porous tubular scaffolds with a highly organized fibrous microstructure by means of melt-electrowriting (MEW) allows the development of self-supported kidney proximal tubules with enhanced properties. A custom-built MEW device was used to fabricate tubular fibrous scaffolds with small diameter sizes (Ø = 0.5, 1, 3 mm) and well-defined, porous microarchitectures (rhombus, square, and random). Human umbilical vein endothelial cells (HUVEC) and human conditionally immortalized proximal tubular epithelial cells (ciPTEC) were seeded into the scaffolds and tested for monolayer formation, integrity, and organization, as well as for extracellular matrix (ECM) production and renal transport functionality. Tubular scaffolds were successfully manufactured by fine control of MEW instrument parameters. A minimum inner diameter of 0.5 mm and pore sizes of 0.2 mm were achieved. CiPTEC formed tight monolayers in all scaffold microarchitectures tested, but well-defined rhombus-shaped pores outperformed and facilitated unidirectional cell orientation, increased collagen type IV deposition, and expression of the renal transporters and differentiation markers organic cation transporter 2 (OCT2) and P-glycoprotein (P-gp). To conclude, we present smaller diameter engineered kidney tubules with microgeometry-directed cell functionality. Due to the well-organized tubular fiber scaffold microstructure, the tubes are mechanically self-supported, and the self-produced ECM constitutes the only barrier between the inner and outer compartment, facilitating rapid and active solute transport.

Sox2 and FGF20 interact to regulate organ of Corti hair cell and supporting cell development in a spatially-graded manner

The mouse organ of Corti develops in two steps: progenitor specification and differentiation. Fibroblast Growth Factor (FGF) signaling is important in this developmental pathway, as deletion of FGF receptor 1 (Fgfr1) or its ligand, Fgf20, leads to the loss of hair cells and supporting cells from the organ of Corti. However, whether FGF20-FGFR1 signaling is required during specification or differentiation, and how it interacts with the transcription factor Sox2, also important for hair cell and supporting cell development, has been a topic of debate. Here, we show that while FGF20-FGFR1 signaling functions during progenitor differentiation, FGFR1 has an FGF20-independent, Sox2-dependent role in specification. We also show that a combination of reduction in Sox2 expression and Fgf20 deletion recapitulates the Fgfr1-deletion phenotype. Furthermore, we uncovered a strong genetic interaction between Sox2 and Fgf20, especially in regulating the development of hair cells and supporting cells towards the basal end and the outer compartment of the organ of Corti. To explain this genetic interaction and its effects on the basal end of the organ of Corti, we provide evidence that decreased Sox2 expression delays specification, which begins at the organ of Corti apex, while Fgf20-deletion results in premature onset of differentiation, which begins near the organ of Corti base. Thereby, Sox2 and Fgf20 interact to ensure that specification occurs before differentiation towards the cochlear base. These findings reveal an intricate developmental program regulating organ of Corti development along the basal-apical axis of the cochlea.Author summaryThe mammalian cochlea contains the organ of Corti, a specialized sensory epithelium populated by hair cells and supporting cells that detect sound. Hair cells are susceptible to injury by noise, toxins, and other insults. In mammals, hair cells cannot be regenerated after injury, resulting in permanent hearing loss. Understanding genetic pathways that regulate hair cell development in the mammalian organ of Corti will help in developing methods to regenerate hair cells to treat hearing loss. Many genes are essential for hair cell and supporting cell development in the mouse organ of Corti. Among these are Sox2, Fgfr1, and Fgf20. Here, we investigate the relationship between these three genes to further define their roles in development.Interestingly, we found that Sox2 and Fgf20 interact to affect hair cell and supporting cell development in a spatially-graded manner. We found that cells toward the outer compartment and the base of the organ of Corti are more strongly affected by the loss of Sox2 and Fgf20. We provide evidence that this spatially-graded effect can be partially explained by the roles of the two genes in the precise timing of two sequential stages of organ of Corti development, specification and differentation.

Structural and histochemical profile of Lopesia sp. Rübsaamen 1908 pinnula galls on Mimosa tenuiflora (Willd.) Poir. in a Caatinga environment

ABSTRACT Gall-inducing insects can change the anatomical pattern of host plant tissues by inducing peculiar gall morphotypes. In this study, the structural changes observed in Lopesia galls on Mimosa tenuiflora resemble those found in other Cecidomyiidae, with two tissue compartments. Nevertheless, the parenchyma layers of the inner compartment, between the mechanical zone and the nutritive tissue, are peculiar. Gall development does not impair the synthesis of any compounds detected by histochemical tests on non-galled tissues of M. tenuiflora. Lignin, polyphenols, alkaloids and terpenoids were detected in the outer compartment, suggesting their involvement in chemical defence of galls. Proteins, reducing sugars and lipids were detected both in outer and inner compartments, whereas nutritive tissue is rich in reducing sugar. This profile is linked with the nutrition of the gall-inducing insect. The Caatinga environment does not seem to constrain the development of galls, but the thick periclinal cell wall and homogeneous parenchyma may contribute to the control of humidity and light radiation, thus favouring the survival of the gall-inducing insect.

Development and Validation of anIn VitroPharmacokinetic/Pharmacodynamic Model To Test the Antibacterial Efficacy of Antibiotic Polymer Conjugates

ABSTRACTThis study describes the use of a novel, two-compartment, static dialysis bag model to study the release, diffusion, and antibacterial activity of a novel, bioresponsive dextrin-colistin polymer conjugate against multidrug resistant (MDR) wild-typeAcinetobacter baumannii. In this model, colistin sulfate, at its MIC, produced a rapid and extensive drop in viable bacterial counts (<2 log10CFU/ml at 4 h); however, a marked recovery was observed thereafter, with regrowth equivalent to that of control by 48 h. In contrast, dextrin-colistin conjugate, at its MIC, suppressed bacterial growth for up to 48 h, with 3 log10CFU/ml lower bacterial counts after 48 h than those of controls. Doubling the concentration of dextrin-colistin conjugate (to 2× MIC) led to an initial bacterial killing of 3 log10CFU/ml at 8 h, with a similar regrowth profile to 1× MIC treatment thereafter. The addition of colistin sulfate (1× MIC) to dextrin-colistin conjugate (1× MIC) resulted in undetectable bacterial counts after 4 h, followed by suppressed bacterial growth (3.5 log10CFU/ml lower than that of control at 48 h). Incubation of dextrin-colistin conjugates with infected wound exudate from a series of burn patients (n= 6) revealed an increasing concentration of unmasked colistin in the outer compartment (OC) over time (up to 86.3% of the initial dose at 48 h), confirming that colistin would be liberated from the conjugate by endogenous α-amylase within the wound environment. These studies confirm the utility of this model system to simulate the pharmacokinetics of colistin formation in humans administered dextrin-colistin conjugates and further supports the development of antibiotic polymer conjugates in the treatment of MDR infections.

Contact Mechanics Model of Lateral Resurfacing Elbow With Inclusion of Rough Surfaces

The study of joint contact mechanics to better understanding of all processes leading to cartilage degradation is very necessary. Elbow replacement will be the only option if both inner and outer components of the elbow have severe arthritis, or usually rheumatoid arthritis. It may also be recommended if there is osteoarthritis which makes the elbow stiff and painful, or a severe fracture of the elbow. Recently the operation known as Lateral resurfacing elbow replacement has been developed and introduced in hospitals. This operation has been designed for those people whose disease in the elbow involves mainly the outer compartment of the joint. The components are made from the metal and polyethylene and are fixed to the bones. An improper design of implants can lead to toxicity for patients caused by excessive amount of metal debris. The purpose of this paper is to investigate the effect of roughness in Lateral resurfacing elbow implants. This paper develops a contact model to treat the interaction of the new surfaces fixed to the top of radius and humeral. The contact model describes the interaction of implant rough surfaces including both elastic and plastic deformations. In the model, surfaces are investigated as macroscopically conforming semi-spheres containing micron-scale roughness. The derived equations relate contact force on the implant and the minimum mean surface separation of the rough surfaces. Based on the distribution of asperity heights, the force is expressed using statistical integral function of asperity heights over the possible region of interaction of the roughness of the implant surfaces. Closed-form approximate equation relating contact force and minimum separation is used to obtain energy loss per cycle in a load-unload sequence applied to the implant.