Growth of Large Single-Crystalline Monolayer Hexagonal Boron Nitride by Oxide-Assisted Chemical Vapor Deposition

We show how an oxide passivating layer on the Cu surface before the growth of h-BN by chemical vapor deposition (CVD) can lead to increased domain sizes from 1 to 20 μm by reducing the nucleation density from 10⁶ to 10³ mm^–2. The h-BN domains within each Cu grain are well-oriented, indicating an epitaxial relationship between the h-BN crystals and the Cu growth substrates that leads to larger crystal domains within the film of ∼100 μm. Continuous films are grown and show a high degree of monolayer uniformity. This CVD approach removes the need for low pressures, electrochemical polishing, and expensive substrates for large-area continuous films of h-BN monolayers, which is beneficial for industrial applications that require scalable synthesis

Characterization of Cross-Linked Porous Gelatin Carriers and Their Interaction with Corneal Endothelium: Biopolymer Concentration Effect

<div><p>Cell sheet-mediated tissue regeneration is a promising approach for corneal reconstruction. However, the fragility of bioengineered corneal endothelial cell (CEC) monolayers allows us to take advantage of cross-linked porous gelatin hydrogels as cell sheet carriers for intraocular delivery. The aim of this study was to further investigate the effects of biopolymer concentrations (5–15 wt%) on the characteristic and safety of hydrogel discs fabricated by a simple stirring process combined with freeze-drying method. Results of scanning electron microscopy, porosity measurements, and ninhydrin assays showed that, with increasing solid content, the pore size, porosity, and cross-linking index of carbodiimide treated samples significantly decreased from 508±30 to 292±42 µm, 59.8±1.1 to 33.2±1.9%, and 56.2±1.6 to 34.3±1.8%, respectively. The variation in biopolymer concentrations and degrees of cross-linking greatly affects the Young’s modulus and swelling ratio of the gelatin carriers. Differential scanning calorimetry measurements and glucose permeation studies indicated that for the samples with a highest solid content, the highest pore wall thickness and the lowest fraction of mobile water may inhibit solute transport. When the biopolymer concentration is in the range of 5–10 wt%, the hydrogels have high freezable water content (0.89–0.93) and concentration of permeated glucose (591.3–615.5 µg/ml). These features are beneficial to the in vitro cultivation of CECs without limiting proliferation and changing expression of ion channel and pump genes such as ATP1A1, VDAC2, and AQP1. In vivo studies by analyzing the rabbit CEC morphology and count also demonstrate that the implanted gelatin discs with the highest solid content may cause unfavorable tissue-material interactions. It is concluded that the characteristics of cross-linked porous gelatin hydrogel carriers and their triggered biological responses are in relation to biopolymer concentration effects.</p></div

Determination of freezable water content.

<p>(a) Typical DSC thermograms of swollen gelatin hydrogel discs. (b) Freezable water content (<i>W</i>_fH/<i>W</i>_s) of various gelatin samples. An asterisk indicates statistically significant differences (*<i>P</i><0.05; <i>n</i> = 6) as compared with the G5 groups.</p

Quantitative real-time reverse transcription polymerase chain reaction and Western blot analyses.

<p>(a) Gene expression level of ATP1A1, VDAC2, and AQP1 in rabbit corneal endothelial cells after 12 h of direct contact with various gelatin samples, measured by real-time RT-PCR. Normalization was done by using GAPDH. Data in the experimental groups are percentages relative to that of control groups (cells cultured in the absence of gelatin materials). An asterisk indicates statistically significant differences (*<i>P</i><0.05; <i>n</i> = 3) as compared with the control groups. (b) Western blot analysis of Na⁺,K⁺-ATPase expression in the rabbit corneal endothelial cells after 12 h of direct contact with gelatin samples. Lane 1: control (without gelatin materials), Lane 2: G5, Lane 3: G10, and Lane 4: G15 groups.</p

In vivo studies.

<p>Specular microscopy measurements of rabbit corneal endothelium 3 days after surgical insertion of various gelatin implants in the ocular anterior chamber. (a) Typical images; (b) graph of corneal endothelial cell count. An asterisk indicates statistically significant differences (*<i>P</i><0.05; <i>n</i> = 6) between the preoperative (Pre) and postoperative (Post) cell density for each type of gelatin disc. The rabbits received no implant (only corneal/limbal incision) and served as a control (sham-operated) group.</p

Morphological observations and cross-linking analyses.

<p>(a) Representative scanning electron microscopic images of various gelatin discs after cross-linking. Scale bars: 100 µm. (b) Cross-linking index of various gelatin discs. An asterisk indicates statistically significant differences (*<i>P</i><0.05; <i>n</i> = 5) as compared with the G5 groups.</p

Glucose permeation studies.

<p>Concentration of glucose permeated through various gelatin discs at 34°C. An asterisk indicates statistically significant differences (*<i>P</i><0.05; <i>n</i> = 6) as compared with the G5 groups.</p

Characterization of porous structure.

<p>(a) Pore size and (b) porosity of various gelatin discs. An asterisk indicates statistically significant differences (*<i>P</i><0.05; <i>n</i> = 4) between the non-cross-linked and cross-linked groups for each type of gelatin disc. ^#<i>P</i><0.05 versus all groups (compared only within non-cross-linked or cross-linked groups).</p

Cell viability and proliferation assays.

<p>(a) Cell viability of rabbit corneal endothelial cell cultures was determined by staining with Live/Dead Viability/Cytotoxicity Kit in which the live cells fluoresce green and dead cells fluoresce red. Fluorescence images of cells in controls (without gelatin materials) after 12 h of direct contact with different types of gelatin samples. Scale bars: 100 µm. (b) Cell proliferation assay of rabbit corneal endothelial cell cultures after 12 h of direct contact with various gelatin samples. Results are expressed as percentage of control groups (MTS activity of cells cultured in the absence of gelatin materials). An asterisk indicates statistically significant differences (*<i>P</i><0.05; <i>n</i> = 5) as compared with the control groups.</p

Mechanical tests.

<p>Young’s modulus of various gelatin samples. An asterisk indicates statistically significant differences (*<i>P</i><0.05; <i>n</i> = 10) between the non-cross-linked and cross-linked groups for each type of gelatin sample. ^#<i>P</i><0.05 versus all groups (compared only within non-cross-linked or cross-linked groups).</p