Chinmaya Mahapatra

Strategies for osteochondral repair: Focus on scaffolds.

Interest in osteochondral repair has been increasing with the growing number of sports-related injuries, accident traumas, and congenital diseases and disorders. Although therapeutic interventions are entering an advanced stage, current surgical procedures are still in their infancy. Unlike other tissues, the osteochondral zone shows a high level of gradient and interfacial tissue organization between bone and cartilage, and thus has unique characteristics related to the ability to resist mechanical compression and restoration. Among the possible therapies, tissue engineering of osteochondral tissues has shown considerable promise where multiple approaches of utilizing cells, scaffolds, and signaling molecules have been pursued. This review focuses particularly on the importance of scaffold design and its role in the success of osteochondral tissue engineering. Biphasic and gradient composition with proper pore configurations are the basic design consideration for scaffolds. Surface modification is an essential technique to improve the scaffold function associated with cell regulation or delivery of signaling molecules. The use of functional scaffolds with a controllable delivery strategy of multiple signaling molecules is also considered a promising therapeutic approach. In this review, we updated the recent advances in scaffolding approaches for osteochondral tissue engineering.

Mesoporous Silica-Layered Biopolymer Hybrid Nanofibrous Scaffold: A Novel Nanobiomatrix Platform for Therapeutics Delivery and Bone Regeneration

Nanoscale scaffolds that characterize high bioactivity and the ability to deliver biomolecules provide a 3D microenvironment that controls and stimulates desired cellular responses and subsequent tissue reaction. Herein novel nanofibrous hybrid scaffolds of polycaprolactone shelled with mesoporous silica (PCL@MS) were developed. In this hybrid system, the silica shell provides an active biointerface, while the 3D nanoscale fibrous structure provides cell-stimulating matrix cues suitable for bone regeneration. The electrospun PCL nanofibers were coated with MS at controlled thicknesses via a sol-gel approach. The MS shell improved surface wettability and ionic reactions, involving substantial formation of bone-like mineral apatite in body-simulated medium. The MS-layered hybrid nanofibers showed a significant improvement in mechanical properties, in terms of both tensile strength and elastic modulus, as well as in nanomechanical surface behavior, which is favorable for hard tissue repair. Attachment, growth, and proliferation of rat mesenchymal stem cells were significantly improved on the hybrid scaffolds, and their osteogenic differentiation and subsequent mineralization were highly up-regulated by the hybrid scaffolds. Furthermore, the mesoporous surface of the hybrid scaffolds enabled the loading of a series of bioactive molecules, including small drugs and proteins at high levels. The release of these molecules was sustainable over a long-term period, indicating the capability of the hybrid scaffolds to deliver therapeutic molecules. Taken together, the multifunctional hybrid nanofibrous scaffolds are considered to be promising therapeutic platforms for stimulating stem cells and for the repair and regeneration of bone.

Effect of Aminated Mesoporous Bioactive Glass Nanoparticles on the Differentiation of Dental Pulp Stem Cells

Mesoporous bioactive nanoparticles (MBNs) have been developed as promising additives to various types of bone or dentin regenerative material. However, biofunctionality of MBNs as dentin regenerative additive to dental materials have rarely been studied. We investigated the uptake efficiency of MBNs-NH2 with their endocytosis pathway and the role of MBNs-NH2 in odontogenic differentiation to clarify inherent biofunctionality. MBNs were fabricated by sol-gel synthesis, and 3% APTES was used to aminate these nanoparticles (MBNs-NH2) to reverse their charge from negative to positive. To characterize the MBNs-NH2, TEM, XRD, FTIR, zeta(ξ)-potential measurements, and Brunauer–Emmett–Teller analysis were performed. After primary cultured rat dental pulp stem cells (rDPSCs) were incubated with various concentrations of MBNs-NH2, stem cell viability (24 hours) with or without differentiated media, internalization of MBNs-NH2 in rDPSCs (~4 hours) via specific endocytosis pathway, intra or extracellular ion concentration and odontoblastic differentiation (~28 days) were investigated. Incubation with up to 50 μg/mL of MBNs-NH2 had no effect on rDPSCs viability with differentiated media (p>0.05). The internalization of MBNs-NH2 in rDPSCs was determined about 92% after 4 hours of incubation. Uptake was significantly decreased with ATP depletion and after 1 hour of pre-treatment with the inhibitor of macropinocytosis (p<0.05). There was significant increase of intracellular Ca and Si ion concentration in MBNs-NH2 treated cells compared to no-treated counterpart (p<0.05). The expression of odontogenic-related genes (BSP, COL1A, DMP-1, DSPP, and OCN) and the capacity for biomineralization (based on alkaline phosphatase activity and alizarin red staining) were significantly upregulated with MBNs-NH2. These results indicate that MBNs-NH2 induce odontogenic differentiation of rDPSCs and may serve as a potential dentin regenerative additive to dental material for promoting odontoblast differentiation.

Alginate-hyaluronic acid-collagen composite hydrogel favorable for the culture of chondrocytes and their phenotype maintenance

Articular cartilage has limited regeneration capacity, thus significant challenge has been made to restore the functions. The development of hydrogels that can encapsulate and multiply cells, and then effectively maintain the chondrocyte phenotype is a meaningful strategy to this cartilage repair. In this study, we prepared alginate-hyaluronic acid based hydrogel with type I collagen being incorporated, namely Alg-HA-Col composite hydrogel. The incorporation of Col enhanced the chemical interaction of molecules, and the thermal stability and dynamic mechanical properties of the resultant hydrogels. The primary chondrocytes isolated from rat cartilage were cultured within the composite hydrogel and the cell viability recorded revealed active proliferation over a period of 21 days. The mRNA levels of chondrocyte phenotypes, including SOX9, collagen type II, and aggrecan, were significantly up-regulated when the cells were cultured within the Alg-HA-Col gel than those cultured within the Alg-HA. Furthermore, the secretion of sulphated glycosaminoglycan, a cartilage-specific matrix molecule, was recorded higher in the collagen-added composite hydrogel. Although more in-depth studies are required such as the in vivo functions, the currently-prepared Alg-HA-Col composite hydrogel is considered to provide favorable 3-dimensional matrix conditions for the cultivation of chondrocytes. Moreover, the cell-cultured constructs may be useful for the cartilage repair and tissue engineering.

Preparation of Self-Activated Fluorescence Mesoporous Silica Hollow Nanoellipsoids for Theranostics

The newly developed multifunctional (self-activated fluorescent, mesoporous, and biocompatible) hollow mesoporous silica nanoellipsoids (f-hMS) are potentially useful as a delivery system of drugs for therapeutics and imaging purposes. For the synthesis of f-hMS, self-activated fluorescence hydroxyapatite (fHA) was used as a core template. A mesoporous silica shell was obtained by silica formation and subsequent removal of the fHA core, which resulted in a hollow-cored f-hMS. Although the silica shell provided a highly mesoporous structure, enabling an effective loading of drug molecules, the fluorescent property of fHA was also well-preserved in the f-hMS. Cytochrome c and doxorubicin, used as a model protein and anticancer drug, respectively, were shown to be effectively loaded onto f-hMS and were then released in a sustainable and controllable manner. The f-hMS was effectively taken up by the cells and exhibited fluorescent labeling while preserving excellent cell viability. Overall, the f-hMS nanoreservoir may be useful as a multifunctional carrier system for drug delivery and cell imaging.

Osteopromoting Reservoir of Stem Cells: Bioactive Mesoporous Nanocarrier/Collagen Gel through Slow-Releasing FGF18 and the Activated BMP Signaling

Providing an osteogenic stimulatory environment is a key strategy to construct stem cell-based bone-equivalent tissues. Here we design a stem cell delivering gel matrix made of collagen (Col) with bioactive glass nanocarriers (BGn) that incorporate osteogenic signaling molecule, fibroblast growth factor 18 (FGF18), a reservoir considered to cultivate and promote osteogenesis of mesenchymal stem cells (MSCs). The presence of BGn in the gel was shown to enhance the osteogenic differentiation of MSCs, possibly due to the therapeutic role of ions released. The mesoporous nature of BGn was effective in loading FGF18 at large quantity, and the FGF18 release from the BGn-Col gel matrix was highly sustainable with almost a zero-order kinetics, over 4 weeks as confirmed by the green fluorescence protein signal change. The released FGF18 was effective in accelerating osteogenesis (alkaline phosphatase activity and bone related gene expressions) and bone matrix formation (osteopontin, bone sialoprotein, and osteocalcin production) of MSCs. This was attributed to the bone morphogenetic protein (BMP) signaling pathway, where the FGF18 release stimulated the endogenous secretion of BMP2 and the downstream signal Smad1/5/8. Taken together, the FGF18-BGn/Col gel is considered an excellent osteopromoting depot to support and signal MSCs for bone tissue engineering.

Biocompatible Mesoporous Nanotubular Structured Surface to Control Cell Behaviors and Deliver Bioactive Molecules.

Biocompatible nanostructured surfaces control the cell behaviors and tissue integration process of medical devices and implants. Here we develop a novel biocompatible nanostructured surface based on mesoporous silica nanotube (MSNT) by means of an electrodeposition. MSNTs, replicated from carbon nanotubes of 25 nm × 1200 nm size, were interfaced in combination with fugitive biopolymers (chitosan or collagen) onto a Ti metallic substrate. The MSNT-biopolymer deposits uniformly covered the substrate with weight gains controllable by the electrodeposition conditions. Random nanotubular networks were generated successfully, which alongside the high mesoporosity provided unique nanotopological properties for the cell responses and the loading/delivery of biomolecules. Of note, the adhesion and spreading behaviors of mesenchymal stem cells (MSCs) were significantly altered, revealing more rapid cell anchorage and extensive nanofilopodia development along the nanotubular networks. Furthermore, the nanotubular surface improved the loading capacity of biomolecules (dexamethasone and bovine serum albumin) up to 5-7 times. The release of the biomolecules was highly sustained, exhibiting a diffusion-controlled pattern over 15 days. The therapeutic efficacy of the delivered biomolecules was also confirmed in the osteogenic differentiation of MSCs. While in vivo performance and applicability studies are needed further, the current biocompatible nanostructured surface may be considered as a novel biointerfacing platform to control cellular behaviors and biomolecular delivery.

Nano-shape varied cerium oxide nanomaterials rescue human dental stem cells from oxidative insult through intracellular or extracellular actions

Cerium oxide nanomaterials (CeNMs), due to their excellent scavenging properties of reactive oxygen species (ROS), have gained great promise for therapeutic applications. A high level of ROS often degrades the potential of stem cells in terms of survivability, maintenance and lineage differentiation. Here we hypothesize the CeNMs may play an important role in protecting the capacity of stem cells against the oxidative insult, and the suppression mechanism of ROS level may depend on the internalization of CeNMs. We synthesized CeNMs with different directional shapes (aspect ratios) by a pH-controlled hydrothermal method, and treated them to stem cells derived from human dental pulp at various doses. The short CeNMs (nanoparticles and nanorods) were internalized rapidly to cells whereas long CeNMs (nanowires) were slowly internalized, which led to different distributions of CeNMs and suppressed the ROS levels either intracellularly or extracellularly under the H2O2-exposed conditions. Resultantly, the stem cells, when dosed with the CeNMs, were rescued to have excellent cell survivability; the damages in intracellular components including DNA fragmentation, lipid rupture and protein degradation were significantly alleviated. The findings imply that the ROS-scavenging events of CeNMs need special consideration of aspect ratio-dependent cellular internalization, and also suggest the promising use of CeNMs to protect stem cells from the ROS-insult environments, which can ultimately improve the stem cell potential for tissue engineering and regenerative medicine uses. STATEMENT OF SIGNIFICANCE Oxidative stress governs many stem cell functions like self-renewal and lineage differentiation, and the biological conditions involving tissue repair and disease cure where stem cell therapy is often needed. Here we demonstrate the unique role of cerium oxide nanomaterials (CeNMs) in rescuing stem cell survivability, migration ability, and intracellular components from oxidative stress. In particular, we deliver a novel finding that nano-morphologically varied CeNMs show different mechanisms in their scavenging reactive oxygen species either intracellularly or extracellularly, and this is related with their different cellular internalizations. We used human dental pulp stem cells for the model study and proved the CeNMs were effective in controlling ROS level by means of scavenging intracellularly or extracellularly, which ultimately led to improving the intact therapeutic potential of stem cells. This work touches an important biological issue of nanomaterial interactions with stem cells under the conditions related with oxidative stress and the resultant damage. The correlation of shape factor in therapeutic nanomaterials with stem cell interaction and the oxidative stress-related functions will provide informative ideas in the design of CeNMs for cellular therapy.

C-Dot Generated Bioactive Organosilica Nanospheres in Theranostics: Multicolor Luminescent and Photothermal Properties Combined with Drug Delivery Capacity

Biocompatible nanomaterials that allow for labeling cells and tissues with the capacity to load and deliver drug molecules hold great promise for the therapeutic-diagnostic purposes in tissue repair and disease cure. Here a novel nanoplatform, called C-dot bioactive organosilica nanosphere (C-BON), is introduced to have excellent theranostic potential, such as controlled drug delivery, visible-light imaging, and NIR photothermal activity. C-dots with a few nanometers were in situ generated in the Ca-containing organosilica mesoporous nanospheres through the sol-gel and thermal-treatment processes. The C-BON exhibited multicolor luminescence over a wide visible-light range with strong emissions and high photostability over time and against acidity and the possible in vivo optical imaging capacity when injected in rat subcutaneous tissues. Moreover, the C-BON showed a photothermal heating effect upon the irradiation of near-infrared. The C-BON, thanks to the high mesoporosity and existence of Ca(2+) ions, demonstrated excellent loading capacity of anticancer drug doxorubicin (as high as 90% of carrier weight) and long-term (over a couple of weeks) and pH/NIR-dependent release ability. The C-BON preserved the compositional merit of Ca-Si glass, having excellent bioactivity and cell compatibility in vitro. Taken all, the multifunctional properties of C-BON-multicolor luminescence, photothermal activity, and high drug loading and controlled release-together with its excellent bioactivity and cell compatibility potentiate the future applications in theranostics (chemotherapy and photothermal therapy with optical imaging).

Functional Recovery of Contused Spinal Cord in Rat with the Injection of Optimal-Dosed Cerium Oxide Nanoparticles

Abstract Spinal cord injury (SCI) produces excess reactive oxygen species (ROS) that can exacerbate secondary injury and lead to permanent functional impairment. Hypothesizing that cerium oxide nanoparticles (CONPs) as an effective ROS scavenger may offset this damaging effect, it is first demonstrated in vitro that CONPs suppressed inducible nitric oxide synthase (iNOS) generation and enhanced cell viability of hydrogen peroxide (H2O2)‐insulted cortical neurons. Next, CONPs are administered at various does (50–4000 µg mL−1) to a contused spinal cord rat model and monitored the disease progression for up to eight weeks. At one day postinjury, the number of iNOS+ cells decreases in the treated groups compared with the control. At one week, the cavity size and inflammatory cells are substantially reduced, and the expression of proinflammatory and apoptotic molecules is downregulated with a concurrent upregulation of anti‐inflammatory cytokine. By eight weeks, the treated groups show significantly improved locomotor functions compared with the control. This study shows for the first time that injection of optimal‐dosed CONPs alone into contusion‐injured spinal cord of rats can reduce ROS level, attenuate inflammation and apoptosis, and consequently help locomotor functional recovery, adding a promising and complementary strategy to the other treatments of acute SCI.