Increased recruitment of endogenous stem cells and chondrogenic differentiation by a composite scaffold containing bone marrow homing peptide for cartilage regeneration

Even small cartilage defects could finally degenerate to osteoarthritis if left untreated, owing to the poor self-healing ability of articular cartilage. Stem cell transplantation has been well implemented as a common approach in cartilage tissue engineering but has technical complexity and safety concerns. The stem cell homing-based technique emerged as an alternative promising therapy for cartilage repair to overcome traditional limitations. In this study, we constructed a composite hydrogel scaffold by combining an oriented acellular cartilage matrix (ACM) with a bone marrow homing peptide (BMHP)-functionalized self-assembling peptide (SAP). We hypothesized that increased recruitment of endogenous stem cells by the composite scaffold could enhance cartilage regeneration. Methods: To test our hypothesis, in vitro proliferation, attachment and chondrogenic differentiation of rabbit mesenchymal stem cells (MSCs) were tested to confirm the bioactivities of the functionalized peptide hydrogel. The composite scaffold was then implanted into full-thickness cartilage defects on rabbit knee joints for cartilage repair, in comparison with microfracture or other sample groups. Stem cell recruitment was monitored by dual labeling with CD29 and CD90 under confocal microcopy at 1 week after implantation, followed by chondrogenic differentiation examined by qRT-PCR. Repaired tissue of the cartilage defects was evaluated by histological and immunohistochemistry staining, microcomputed tomography (micro-CT) and magnetic resonance imaging (MRI) at 3 and 6 months post-surgery. Macroscopic and histological scoring was done to evaluate the optimal in vivo repair outcomes of this composite scaffold. Results: The functionalized SAP hydrogels could stimulate rabbit MSC proliferation, attachment and chondrogenic differentiation during in vitro culture. At 7 days after implantation, increased recruitment of MSCs based on CD29(+)/CD90(+) double-positive cells was found in vivo in the composite hydrogel scaffold, as well as upregulation of cartilage-associated genes (aggrecan, Sox9 and type II collagen). After 3 and 6 months post-surgery, the articular cartilage defect in the composite scaffold-treated group was fully covered with cartilage-like tissue with a smooth surface, which was similar to the surrounding native cartilage, according to the results of histological and immunohistochemistry staining, micro-CT and MRI analysis. Macroscopic and histological scoring confirmed that the quality of cartilage repair was significantly improved with implantation of the composite scaffold at each timepoint, in comparison with microfracture or other sample groups. Conclusion: Our findings demonstrated that the composite scaffold could enhance endogenous stem cell homing and chondrogenic differentiation and significantly improve the therapeutic outcome of chondral defects. The present study provides a promising approach for in vivo cartilage repair without cell transplantation. Optimization of this strategy may offer great potential and benefits for clinical application in the future

Robust estimation of bacterial cell count from optical density

Optical density (OD) is widely used to estimate the density of cells in liquid culture, but cannot be compared between instruments without a standardized calibration protocol and is challenging to relate to actual cell count. We address this with an interlaboratory study comparing three simple, low-cost, and highly accessible OD calibration protocols across 244 laboratories, applied to eight strains of constitutive GFP-expressing E. coli. Based on our results, we recommend calibrating OD to estimated cell count using serial dilution of silica microspheres, which produces highly precise calibration (95.5% of residuals <1.2-fold), is easily assessed for quality control, also assesses instrument effective linear range, and can be combined with fluorescence calibration to obtain units of Molecules of Equivalent Fluorescein (MEFL) per cell, allowing direct comparison and data fusion with flow cytometry measurements: in our study, fluorescence per cell measurements showed only a 1.07-fold mean difference between plate reader and flow cytometry data

Effect of Different Mineralization Modes on Strengthening Calcareous Sand under Simulated Seawater Conditions

Calcareous sand, as a blow-fill or construction material, is widely used in island and reef construction projects in marine environments after treatment. When microorganism-induced mineralization is used to strengthen calcareous sand, salinity and other conditions in the marine environment will adversely affect microorganisms or their mineralization process. For this reason, the two environmental conditions created by deionized water and simulated seawater were introduced to explore their effects on the growth and urease activity of Sporosarcina pasteurii. Then, the changes in the permeability and mechanical strength of calcareous sand under different mineralization methods were compared by one-dimensional sand column tests. Finally, the reinforcement mechanism was compared and analyzed based on the results of scanning electron microscopy and X-ray diffraction tests. The results show that Sporosarcina pasteurii can induce carbonate and phosphate precipitation and mineralization to strengthen calcareous sand in simulated seawater. The mineralized products greatly reduce the permeability of calcareous sand and significantly improve the mechanical strength by wrapping calcareous sand particles, filling water seepage channels and cementing adjacent particles. The reinforcement effect of carbonate mineralization is better than that of phosphate mineralization, but phosphate mineralization has less impact on the environment during the treatment process

Variable-temperature 17O NMR studies allow quantitative evaluation of molecular dynamics in organic solids

We report a comprehensive variable-temperature solid-state 17O NMR study of three 17O-labeled crystalline sulfonic acids: 2-aminoethane-1-sulfonic acid (taurine, T), 3- aminopropane-1-sulfonic acid (homotaurine, HT), and 4-aminobutane- 1-sulfonic acid (ABSA). In the solid state, all three compounds exist as zwitterionic structures, NH3+ 12R 12SO3 12, in which the SO3 12 group is involved in various degrees of O\ub7\ub7\ub7H 12N hydrogen bonding. High-quality 17O NMR spectra have been obtained for all three compounds under both static and magic angle spinning (MAS) conditions at 21.1 T, allowing the complete set of 17O NMR tensor parameters to be measured. Assignment of the observed 17O NMR parameters to the correct oxygen sites in the crystal lattice was achieved with the aid of DFT calculations. By modeling the temperature dependence of 17O NMR powder line shapes, we have not only confirmed that the SO3 12 groups in these compounds undergo a 3-fold rotational jump mechanism but also extracted the corresponding jump rates (102 12105 s 121) and the associated activation energies (Ea) for this process (Ea = 48 \ub1 7, 42 \ub1 3, and 45 \ub1 1 kJ mol 121 for T, HT, and ABSA, respectively). This is the first time that SO3 12 rotational dynamics have been directly probed by solid-state 17O NMR. Using the experimental activation energies for SO3 12 rotation, we were able to evaluate quantitatively the total hydrogen bond energy that each SO3 12 group is involved in within the crystal lattice. The activation energies also correlate with calculated rotational energy barriers. This work provides a clear illustration of the utility of solid-state 17O NMR in quantifying dynamic processes occurring in organic solids. Similar studies applied to selectively 17O-labeled biomolecules would appear to be very feasible.Peer reviewed: YesNRC publication: Ye