Fabrication of a 3D bioprinting model for posterior capsule opacification using GelMA and PLMA hydrogel-coated resin.

Posterior capsule opacification (PCO) remains the predominant complication following cataract surgery, significantly impairing visual function restoration. In this study, we developed a PCO model that closely mimics the anatomical structure of the crystalline lens capsule post-surgery. The model incorporated a threaded structure for accurate positioning and observation, allowing for opening and closing. Utilizing 3D printing technology, a stable external support system was created using resin material consisting of a rigid, hollow base and cover. To replicate the lens capsule structure, a thin hydrogel coating was applied to the resin scaffold. The biocompatibility and impact on cellular functionality of various hydrogel compositions were assessed through an array of staining techniques, including calcein-AM/PI staining, rhodamine staining, BODIPY-C11 staining and EdU staining in conjunction with transwell assays. Additionally, the PCO model was utilized to investigate the effects of eight drugs with anti-inflammatory and anti-proliferative properties, including 5-aminoimidazole-4-carboxamide ribonucleotide (AICAR), THZ1, sorbinil, 4-octyl itaconate (4-OI), xanthohumol, zebularine, rapamycin and caffeic acid phenethyl ester, on human lens epithelial cells (HLECs). Confocal microscopy facilitated comprehensive imaging of the PCO model. The results demonstrated that the GelMA 60 5% + PLMA 2% composite hydrogel exhibited superior biocompatibility and minimal lipid peroxidation levels among the tested hydrogels. Moreover, compared to using hydrogel as the material for 3D printing the entire model, applying surface hydrogel spin coating with parameters of 2000 rpm × 2 on the resin-based 3D printed base yielded a more uniform cell distribution and reduced apoptosis. Furthermore, rapamycin, 4-OI and AICAR demonstrated potent antiproliferative effects in the drug intervention study. Confocal microscopy imaging revealed a uniform distribution of HLECs along the anatomical structure of the crystalline lens capsule within the PCO model, showcasing robust cell viability and regular morphology. In conclusion, the PCO model provides a valuable experimental platform for studying PCO pathogenesis and exploring potential therapeutic interventions.

Rational Design of a Circularly Permuted Flavin-Based Fluorescent Protein.

Flavin-based fluorescent proteins are oxygen-independent reporters that hold great promise for imaging anaerobic and hypoxic biological systems. In this study, we explored the feasibility of applying circular permutation, a valuable method for the creation of fluorescent sensors, to flavin-based fluorescent proteins. We used rational design and structural data to identify a suitable location for circular permutation in iLOV, a flavin-based reporter derived from A. thaliana. However, relocating the N- and C-termini to this position resulted in a significant reduction in fluorescence. This loss of fluorescence was reversible, however, by fusing dimerizing coiled coils at the new N- and C-termini to compensate for the increase in local chain entropy. Additionally, by inserting protease cleavage sites in circularly permuted iLOV, we developed two protease sensors and demonstrated their application in mammalian cells. In summary, our work establishes the first approach to engineer circularly permuted FbFPs optimized for high fluorescence and further showcases the utility of circularly permuted FbFPs to serve as a scaffold for sensor engineering.

Preferred penetration of active nano-rods into narrow channels and their clustering.

In a channel connected to a reservoir, passive particles prefer staying in the reservoir than the channel due to the entropic effect, as the size of the particles is comparable to that of the channel. Self-propelled rods can exhibit out-of-equilibrium phenomena, and their partition behavior may differ from that of passive rods due to their persistent swimming ability. In this work, the distribution of active nano-rods between the nanoscale channel and reservoir is explored using dissipative particle dynamics. The ratio of the nano-rod concentration in the slit to that in the reservoir, defined as the partition ratio Ψ, is a function of active force, channel width, and rod length. Although passive nano-rods prefer staying in bulk (Ψ < 1), active rods can overcome the entropic barrier and show favorable partition toward narrow channels (Ψ > 1). As the slit width decreases to about the rod's width, active rods entering the slit behave like a quasi-two-dimensional system dynamically. At sufficiently high concentrations and Peclet numbers, nano-rods tend to align and move together in the same direction for a certain time. The distribution (PM) of the cluster size (M) follows a power law, PM ∝ M-2, for small clusters.

Systematic Investigation of mRNA N 6-Methyladenosine Machinery in Primary Prostate Cancer.

BACKGROUND: Appreciable findings have pointed out pivotal roles of N 6-methyladenosine (m6A) machinery in cancer onset and progression. However, limited efforts have been directed towards relevant research in the prostate cancer area. METHODS: A PubMed search was conducted to acquire components of the mRNA m6A machinery. Multiomics integration was performed to systematically investigate the mRNA m6A machinery in primary prostate cancer. Furthermore, RNA interference assays of two prognostic m6A readers EIF3D and HNRNPA2B1 were conducted to explore m6A dependence of their functions in prostate cancer cell proliferation and migration. RESULTS: A total of 41 mRNA m6A regulators have been identified to date. A small degree of copy number aberrations and an extremely low frequency of somatic mutations were observed in the regulators across prostate tumors. Enrichment of CpG sites and extensive changes of DNA methylation in the m6A machinery were also found. Impact of copy number variation on m6A regulator expression was stronger than that of DNA methylation disturbance. Furthermore, our study identified a set of m6A regulators related to clinical features and/or survival which were largely m6A-binding proteins. The translation initiation factor subunit EIF3D and the splicing factor HNRNPA2B1 can be independent prognostic factors which may contribute to retardation and promotion of cancer progression, respectively, through affecting cancer-related processes such as cell cycle. Moreover, in vitro assays demonstrated that m6A impacted the EIF3D and HNRNPA2B1 roles in proliferation and migration of prostate cancer cells. CONCLUSIONS: Our report systematically described molecular features of the mRNA m6A machinery and their potential roles in primary prostate cancer. Knowledge gained from this work may pave the way for further studies on the m6A system in prostate cancer.

Size-dependence and interfacial segregation in nanofilms and nanodroplets of homologous polymer blends.

The size-dependent behavior of nanofilms and nanodroplets of homologous polymer blends was explored by many-body dissipative particle dynamics. Although a homologous blend can be regarded as a completely miscible and athermal system, enrichment of the surface in short polymers always takes place. First, liquid-gas and solid-liquid interfacial tensions of polymer melts were acquired. It is found that they increase and approach asymptotes with increasing chain lengths. The molecular weight dependence can be depicted using two semi-empirical expressions. Second, the variation of surface tension and surface excess of polymer blend nanofilms with the thickness was observed. Surface tension of the blend is observed to increase but the extent of surface segregation decreases upon increasing the film thickness. Finally, the wetting phenomenon of nanodroplets of homologous blends was examined. The contact angle is found to increase as the droplet size is reduced. Our simulation results indicate that the size-dependence of nanofilms and nanodroplets is closely related to surface segregation in homologous blends.

Size-dependent behavior and failure of young's equation for wetting of two-component nanodroplets.

HYPOTHESIS: For macroscopic systems, the interfacial properties are size-independent and Young's equation is generally valid for smooth substrates. For nanoscale systems, however, size-dependence and failure of Young's equation may emerge. EXPERIMENTS: The wetting behavior of a nanodroplet containing two miscible liquids on a smooth substrate is explored by many-body dissipative particle dynamics simulations. The size-dependent surface tension of nanofilms is investigated as well. FINDINGS: It is found that Young's equation is valid for nanodroplets of pure fluids but fails for two-component nanodroplets. The actual contact angle is always larger than the Young's contact angle, and their difference is getting smaller as the composition approaches pure fluids or the compatibility of the mixture is increased. The failure of Young's equation is closely associated with the size-dependent behavior in two-component nanodroplets and nanofilms. As the nanodroplet size is increased, the actual contact angle is found to decline but approaches a constant expected in macroscopic systems. Similarly, as the nanofilm thickness is increased, surface tension decreases and reaches its macroscopic value. The change of surface tension is attributed to the size-dependent surface composition, which is responsible for the failure of Young's equation.

Pressure-gated capillary nanovalves based on liquid nanofilms.

HYPOTHESIS: Nanoscale valving is essential to expand the potentiality of the nanodevices. However, it is difficult to fabricate valves with movable control elements in nanoscale systems and thus it is desirable to design a nanovalve which can manipulate opening and blocking a gate without moving parts. EXPERIMENTS: A pressure-gated capillary valve which contains a nanoscale liquid layer sandwiched between two plates with two aligned orifices was designed and the proof-of-concept demonstration was achieved by Many-body Dissipative Particle Dynamics. FINDINGS: The concave or convex meniscus appears naturally within the orifice of the capillary valve and can be controlled by the pressure difference between liquid and gas phases based on Young-Laplace equation and the edge effect. The closed state can be transformed into the open state (hole) when the two concave menisci are allowed to touch each other. The on/off switch is reversible by Laplace pressure manipulation and the passing and blocking of the fluid particles through the valve is verified.

Beyond the Green Fluorescent Protein: Biomolecular Reporters for Anaerobic and Deep-Tissue Imaging.

Fluorescence imaging represents cornerstone technology for studying biological function at the cellular and molecular levels. The technology's centerpiece is a prolific collection of genetic reporters based on the green fluorescent protein (GFP) and related analogs. More than two decades of protein engineering have endowed the GFP repertoire with an incredible assortment of fluorescent proteins, allowing scientists immense latitude in choosing reporters tailored to various cellular and environmental contexts. Nevertheless, GFP and derivative reporters have specific limitations that hinder their unrestricted use for molecular imaging. These challenges have inspired the development of new reporter proteins and imaging mechanisms. Here, we review how these developments are expanding the frontiers of reporter gene techniques to enable nondestructive studies of cell function in anaerobic environments and deep inside intact animals-two important biological contexts that are fundamentally incompatible with the use of GFP-based reporters.

Favorable partition of nanoswimmers toward a confined slit.

The partition of nanoswimmers between a narrow channel and a reservoir is explored by dissipative particle dynamics. In contrast to passive colloids, nanoswimmers prefer to stay in the slit rather than in the reservoir for sufficiently large active force (F_{a}) or run time (τ). The partition ratio (φ) increases with F_{a} and τ. Interestingly, as the slit height decreases, φ grows accordingly until the confinement effect dominates. Three types of the concentration profile in the slit are identified as a consequence of the competition between surface accumulation and entropic barrier.

Strong competition between adsorption and aggregation of surfactant in nanoscale systems.

HYPOTHESIS: The size-dependent behavior including surface tension, surface density (Γ), and critical micelle concentration (CMC) of a nanoscale liquid film containing surfactant has not been investigated until now. EXPERIMENTS: Strong competition between surface adsorption and bulk aggregation of surfactant in nanoscale systems was explored by Many-body Dissipative Particle Dynamics simulations. FINDINGS: In nanoscale systems, as the surfactant concentration increases, Γ continues rising even after CMC is exceeded. The saturation level of Γ is achieved only when the surfactant bulk concentration is over ten times of CMC. Moreover, both surface micelles formed by adsorbed surfactant and the sublayer below the adsorbed layer are clearly identified. The former can reduce the contacts of adsorbed surfactant with water, while the latter has the surfactant concentration significantly higher than that in bulk. The strong coupling between adsorption and micellization is attributed to large surface-to-volume ratio compared to macroscopic systems, and can be simply realized by the fact that the ratio of the numbers of surfactant distributed in bulk (nbulk) and at interface (nads) is always less than unity (nbulk/nads < 1) in nanoscale systems.

Penetration dynamics through nanometer-scale hydrophilic capillaries: Beyond Washburn's equation and extended menisci.

HYPOTHESIS: Recently, the naoncapillary devices with the channel width about 2-3 water molecules have been fabricated. Water transport through these nanoslits showed unexpectedly fast flow, revealing the failure of Washburn's equation. EXPERIMENTS: Liquid penetration into a nanocapillary made of two parallel walls is explored by many-body dissipative particle dynamics. Both partial wetting and total wetting walls are considered and the no-slip boundary condition is satisfied. FINDINGS: The wicking velocity generally obeys Washburn's equation, but the dynamic contact angle (CA) has to be employed. The dynamic CA (θD) relies on the penetration rate and is always larger than the equilibrium CA. The breakdown of Washburn's equation occurs under two conditions, (i) the channel width close to molecular size and (ii) the positive spreading coefficient is large enough. Both cases come about when the wicking velocity in a nanoslit exceeds the maximum value corresponding to cos(θD) = 1. The failure of Washburn's equation is attributed to the invalidity of Young-Laplace equation associated with undefined meniscus. The extended meniscus will be developed as a wall of the nanoslit continues to extend outside the exit mouth. The shapes of extended menisci are discussed for both partial wetting and total wetting surfaces.

Hydrodynamic interaction induced breakdown of the state properties of active fluids.

The mechanical pressure of active fluids in which swimmers are modeled by soft run-and-tumble spheres is investigated by dissipative particle dynamics simulations. The incremental pressure (Π) with respect to the system pressure with inactive swimmers comprises the direct contribution of the swimmers (π) and the indirect contribution of fluids associated with hydrodynamic interactions (HIs). The pressure can be determined from the bulk and confining wall and the former is always less than the latter. The π of dilute active dispersions is proportional to their active diffusivity while Π grows generally with propulsive force and run time. However, Π is always substantially less than π because of negative contributions to pressure by HIs. The wall pressure depends on the swimmer-wall interactions, verifying that pressure is not a state function for active spheres due to the HIs. Owing to the distinct flow patterns, Π varies with the swim-type (pusher and puller) subject to the same run-and-tumble parameters at high concentrations.

Derivation of porcine pluripotent stem cells for biomedical research.

Pluripotent stem cells including embryonic stem cells (ESCs), embryonic germ cells (EGCs), and induced pluripotent stem cells (iPSCs) are capable of self-renew and limitlessly proliferating in vitro with undifferentiated characteristics. They are able to differentiate in vitro, spontaneously or responding to suitable signals, into cells of all three primary germ layers. Consequently, these pluripotent stem cells will be valuable sources for cell replacement therapy in numerous disorders. However, the promise of human ESCs and EGCs is cramped by the ethical argument about destroying embryos and fetuses for cell line creation. Moreover, there are still carcinogenic risks existing toward the goal of clinical application for human ESCs, EGCs, and iPSCs. Therefore, a suitable animal model for stem cell research will benefit the further development of human stem cell technology. The pigs, on the basis of their similarity in anatomy, immunology, physiology, and biochemical properties, have been wide used as model animals in the study of various human diseases. The development of porcine pluripotent stem cell lines will hold the opportunity to provide an excellent material for human counterpart to the transplantation in biomedical research and further development of cell-based therapeutic strategy.

Metformin regulates hepatic lipid metabolism through activating AMP-activated protein kinase and inducing ATGL in laying hens.

Although many clinical trials have showed that metformin improves non-alcoholic fatty liver disease, which is a common liver disease associated with hepatic enzyme abnormalities, an animal model is required to investigate the effects of altered gene expression and post-translational processing (proteins) in mediating the observed responses. Laying hens appear to develop fatty livers, as in the case in human beings, when ingesting energy in excess of maintenance, and they can be used as an animal model for observing hepatic steatosis. The aim of this study was to investigate whether metformin could improve the non-alcoholic fatty liver of laying hens and to examine the possible mechanisms of lipid-lowering effects. Forty-eight Leghorn laying hens of Hy-Line variety W-36 - 44 weeks with 64.8% hen-day egg production - were randomly assigned into 4 treatments, each receiving 0, 10, 30, or 100mg of metformin with saline per kg body weight by daily wing vein injection. Results showed that, compared with the control, significant decreases existed in the laying rates; plasma triglyceride, cholesterol, and insulin levels; body weights; abdominal fat weights; hepatic lipid contents; and hepatic fatty acid synthase expression of layers receiving 30 or 100mg per kg body weight, whereas significant increases in their hepatic 5'adenosine monophosphate-activated protein kinase, acyl-CoA carboxylase phosphorylation, adipose triglyceride lipase, and carnitine palmitoyl transferase-1 expression were observed. These data suggest that metformin could reduce lipid deposits in the liver and that the laying hen is a valuable animal model for studying hepatic steatosis.