Role of calcium phosphate and bioactive glass coating on in vivo bone healing of new Mg-Zn-Ca implant.

Present investigation focuses on development and detailed characterization of a new Mg alloy sample (BM) with and without coating of hydroxyapatite (BMH) and bioactive glass (BMG) by air plasma spray method. After detailed mechano-physico-chemical characterization of powders and coated samples, electrochemical corrosion and SBF immersion tests were carried out. Detailed in vitro characterizations for cell viability were undertaken using MG-63 cell line followed by in vivo tests in rabbit model for studying bone healing up to 60 days. Starting current density increases from BM to BMH to BMG indicating highest resistance towards corrosion in case of BMG samples, however BMH also showed highest icorr value suggesting slowest rate of corrosion than BM and BMG samples. Dissolution of calcium ion in case of BMH and BMG control formation of apatite phases on surface. Ca2+ ions of coatings and from SBF solution underwent reduction reaction simultaneously with conversion of Mg to MgCl2 releasing OH- in the solution, which increases pH. Viability and propagation of human osteoblast-like cells was verified using confocal microscopy observations and from expression of bone specific genes. Alkaline phosphatase assay and ARS staining indicate cell proliferation and production of neo-osseous tissue matrix. In vivo, based on histology of heart, kidney and liver, and immune response of IL-2, IL-6 and TNFα, all the materials show no adverse effects in body system. The bone creation was observed to be more for BMH. Although both BMH and BMG show rays of possibilities in early new bone formation and tough bone-implant bonding at interface as compared to bare Mg alloy, however, BMG showed better well-sprayed coating covering on substrate and resistance against corrosion prior implanting in vivo. Also, better apatite formation on this sample makes it more favourable implant.

Development of nano-porous hydroxyapatite coated e-glass for potential bone-tissue engineering application: An in vitro approach.

To reconstruct the defects caused by craniectomies autologous, bone grafting was usually used, but they failed most commonly due to bone resorption, infections and donor-site morbidity. In the present investigation, an effort has been made for the first time to check the feasibility and advantage of using hydroxyapatite (HAp) coated e-glass as component of bone implants. Sol-gel synthesized coatings were found to be purely hydroxyapatite from XRD with graded and interconnected pores all over the surface observable in TEM. The interconnected porous nature of ceramics are found to increase bioactivity by acting to up-regulate the process of osseointegration through enhanced nutrient transfer and induction of angiogenesis. From TEM studies and nano indentation studies, we have shown that pores were considered to be appropriate for nutrient supply without compromising the strength of sample while in contact with physiological fluid. After SBF immersion test, porous surface was found to be useful for nucleation of apatite crystals, hence increasing the feasibility and bioactivity of sample. However, our quasi-dynamic study showed less crystallization but had significant formation of apatite layer. Overall, the in vitro analyses show that HAp coated e-glass leads to significant improvement of implant properties in terms of biocompatibility, cell viability and proliferation, osteoinductivity and osteoconductivity. HAp coating of e-glass can potentially be utilized in fabricating durable and strong bioactive non-metallic implants and tissue engineering scaffolds.

Influence of single and binary doping of strontium and lithium on in vivo biological properties of bioactive glass scaffolds.

Effects of strontium and lithium ion doping on the biological properties of bioactive glass (BAG) porous scaffolds have been checked in vitro and in vivo. BAG scaffolds were prepared by conventional glass melting route and subsequently, scaffolds were produced by evaporation of fugitive pore formers. After thorough physico-chemical and in vitro cell characterization, scaffolds were used for pre-clinical study. Soft and hard tissue formation in a rabbit femoral defect model after 2 and 4 months, were assessed using different tools. Histological observations showed excellent osseous tissue formation in Sr and Li + Sr scaffolds and moderate bone regeneration in Li scaffolds. Fluorochrome labeling studies showed wide regions of new bone formation in Sr and Li + Sr doped samples as compared to Li doped samples. SEM revealed abundant collagenous network and minimal or no interfacial gap between bone and implant in Sr and Li + Sr doped samples compared to Li doped samples. Micro CT of Li + Sr samples showed highest degree of peripheral cancellous tissue formation on periphery and cortical tissues inside implanted samples and vascularity among four compositions. Our findings suggest that addition of Sr and/or Li alters physico-chemical properties of BAG and promotes early stage in vivo osseointegration and bone remodeling that may offer new insight in bone tissue engineering.

Converted marine coral hydroxyapatite implants with growth factors: in vivo bone regeneration.

Herein we report rabbit model in vivo bone regeneration of hydrothermally converted coralline hydroxyapatite (HCCHAp) scaffolds without (group I) and with growth factors namely insulin like growth factor-1 (IGF-1) (group II) and bone morphogenetic protein-2 (BMP-2) (group III). All HCCHAp scaffolds have been characterized for phase purity and morphology before implantation. Calcined marine coral was hydrothermally converted using a mineralizer/catalyst to phase pure HAp retaining original pore structure and geometry. After sintering at 1250°C, the HCCHAp found to have ~87% crystallinity, 70-75% porosity and 2±0.5MPa compressive strength. In vitro growth factor release study at day 28 revealed 77 and 98% release for IGF-1 and BMP-2, respectively. The IGF-1 release was more sustained than BMP-2. In vivo bone healing of different groups was compared using chronological radiology, histological evaluations, scanning electron microscopy and fluorochrome labeling up to 90days of implantation. In vivo studies showed substantial reduction in radiolucent zone and decreased radiodensity of implants in group II followed by group III and group I. These observations clearly suggest in-growth of osseous tissue, initiation of bone healing and complete union between implants and natural bone in group II implants. A statistical score sheet based on histological observations showed an excellent osseous tissue formation in group II and group III scaffolds and moderate bone regeneration in group I scaffolds.

In vitro and in vivo evaluation of the marine sponge skeleton as a bone mimicking biomaterial.

This investigation was carried out to identify and characterize marine sponges as potential bioscaffolds in bone tissue engineering. The marine sponge (Biemna fortis) samples were collected from the rocky intertidal region of Anjuna, Goa, India, freeze-dried and converted to pure cristobalite at low temperature. After thorough evaluation of sponge samples by DTA-TGA thermography, XRD, FTIR, SEM and cell cytotoxicity by MTT assay, bare sponge scaffolds were fabricated by firing at 1190 °C. These scaffolds were loaded with growth factors (IGF-1 and BMP-2), checked for quasi-dynamic in vitro release kinetics and finally implanted into femoral bone defects in rabbits for up to 90 days, by keeping an empty defect as a control. The in vivo bone healing process was evaluated and compared using chronological radiology, histology, SEM and fluorochrome labeling studies. SEM revealed that the sponge skeleton possesses a collagenous fibrous network consisting of highly internetworked porosity in the size range of 10-220 µm. XRD and FTIR analysis showed a cristobalite phase with acicular crystals of high aspect ratio, and crystallinity was found to increase from 725 to 1190 °C. MTT assay demonstrated the non-cytotoxicity of the samples. A combination of burst and sustained release profile was noticed for both the growth factors and about 74.3% and 83% total release at day 28. In the radiological, histological, scanning electron microscopy and fluorochrome labeling analysis, the IGF-1 impregnated converted sponge scaffold promoted excellent osseous tissue formation followed by the BMP-2 loaded and bare one. These observations suggest that the marine sponge alone and in combination with growth factors is a promising biomaterial for bone repair and bone augmentation.

Development and effect of different bioactive silicate glass scaffolds: in vitro evaluation for use as a bone drug delivery system.

Local drug delivery systems to bone have attracted appreciable attention due to their efficacy to improve drug delivery, healing and regeneration. In this paper, development and characterization of new formulations of bioactive glass into a porous scaffold has been reported for its suitability to act as a drug delivery system in the management of bone infections, in vitro. Two new glass compositions based on SiO2-Na2O-ZnO-CaO-MgO-P2O5 system (BGZ and MBG) have been developed which after thorough chemical and phase evaluation, studied for acellular static in vitro bioactivity in SBF. Porous scaffolds made of these glasses have been fabricated and characterized thoroughly for bioactivity study, SEM, XRD, in vitro cytotoxicity, MTT assay and wound healing assay using human osteocarcoma cells. Finally, gatifloxacin was loaded into the porous scaffold by vacuum infiltration method and in vitro drug release kinetics have been studied with varying parameters including dissolution medium (PBS and SBF) and with/without impregnation chitosan. Suitable model has also been proposed for the kinetics. 63-66% porous and 5-50µm almost unimodal porous MBG and BGZ bioactive glass scaffolds were capable of releasing drugs successfully for 43 days at concentrations to treat orthopedic infections. In addition, it was also observed that the release of drug followed Peppas-Korsmeyer release pattern based on Fickian diffusion, while 0.5-1% chitosan coating on the scaffolds decreased the burst release and overall release of drug. The results also indicated that MBG based scaffolds were bioactive, biocompatible, noncytotoxic and exhibited excellent wound healing potential while BGZ was mildly cytotoxic with moderate wound healing potential. These results strongly suggest that MBG scaffolds appear to be a suitable bone drug delivery system in orthopedic infections treatment and as bone void fillers, but BGZ should be handled with caution or studied elaborately in detail further to ascertain and confirm the cytotoxic nature and wound healing potential of this glass.