Mukesh Doble

Nano-hydroxyapatite (nHA) reinforced magnesium composite (Mg-nHA) was fabricated by friction stir processing (FSP). The effect of smaller grain size and the presence of nHA particles on controlling the degradation of magnesium were investigated. Grain refinement from 1500 μm to 3.5 μm was observed after FSP. In vitro bioactivity studies by immersing the samples in supersaturated simulated body fluid (SBF 5 ×) indicate that the increased hydrophilicity and pronounced biomineralization are due to grain refinement and the presence of nHA in the composite respectively. Electrochemical test to assess the corrosion behavior also clearly showed the improved corrosion resistance due to grain refinement and enhanced biomineralization. Using MTT colorimetric assay, cytotoxicity study of the samples with rat skeletal muscle (L6) cells indicate marginal increase in cell viability of the FSP-Mg-nHA sample. The composite also showed good cell adhesion. © 2014 Elsevier B.V.

Nanotechnology has tremendous potential for the management of infectious diseases caused by multi-drug resistant bacteria, through the development of newer antibacterial materials and efficient modes of antibiotic delivery. Calcium phosphate (CaP) bioceramics are commonly used as bone substitutes due to their similarity to bone mineral and are widely researched upon for the treatment of bone infections associated with bone loss. CaPs can be used as local antibiotic delivery agents for bone infections and can be substituted with antibacterial ions in their crystal structure to have a wide spectrum, sustained antibacterial activity even against drug resistant bacteria. In the present work, a dual mode antibiotic delivery system with antibacterial ion substituted calcium deficient hydroxyapatite (CDHA) nanoparticles has been developed. Antibacterial ions such as zinc, silver, and strontium have been incorporated into CDHA at concentrations of 6, 0.25-0.75, and 2.5-7.5 at. %, respectively. The samples were found to be phase pure, acicular nanoparticles of length 40-50 nm and width 5-6 nm approximately. The loading and release profile of doxycycline, a commonly used antibiotic, was studied from the nanocarriers. The drug release was studied for 5 days and the release profile was influenced by the ion concentrations. The release of antibacterial ions was studied over a period of 21 days. The ion substituted CDHA samples were tested for antibacterial efficacy on Staphylococcus aureus and Escherichia coli by MIC/MBC studies and time-kill assay. AgCDHA and ZnCDHA showed high antibacterial activity against both bacteria, while SrCDHA was weakly active against S. aureus. Present study shows that the antibiotic release can provide the initial high antibacterial activity, and the sustained ion release can provide a long-term antibacterial activity. Such dual mode antibiotic and antibacterial ion release offers an efficient and potent way to treat an incumbent drug resistant infection.

A fibrous scaffold of curdlan/poly(vinyl alcohol) (PVA) blend is prepared by electrospinning technique and antimicrobial property is imparted to it by the addition of silver nitrate (1, 3, and 5 wt%). All the scaffolds except the PVA/curdlan with 5 wt% AgNO3 show good viability of Swiss 3T3 fibroblast cells. Significant reductions in the growth of Staphylococcus aureus and Escherichia coli are also observed in all the scaffolds. In vitro scratch assay and cell adhesion studies indicate that the scaffold containing 1% AgNO3 shows significant wound healing and better cell spreading. The in vivo results also show faster healing of excision wounds in diabetic rats treated with the same material when compared to the control and the commercial sample. Furthermore, downregulation of proinflammatory cytokines and upregulation of anti-inflammatory cytokines on the skin of the treated animals confirm that PVA/curdlan/1% AgNO3 electrospun mat could be a promising material for diabetic wound healing.

A dual local drug delivery system (DDS) composed of calcium phosphate bioceramic nanocarriers aimed at treating the antibacterial, anti-inflammatory, and bone-regenerative aspects of periodontitis has been developed. Calcium-deficient hydroxyapatite (CDHA, Ca/P = 1.61) and tricalcium phosphate (β-TCP) were prepared by microwave-accelerated wet chemical synthesis method. The phase purity of the nanocarriers was confirmed by x-ray diffraction (XRD) and Fourier transform infrared spectroscopy (FT-IR), while the transmission electron microscopy (TEM) confirmed their nanosized morphology. CDHA was selected as carrier for the antibiotic (tetracycline) while TCP was chosen as the anti-inflammatory drug (ibuprofen) carrier. Combined drug release profile was studied in vitro from CDHA/TCP (CTP) system and compared with a HA/TCP (BCP) biphasic system. The tetracycline and ibuprofen release rate was 71 and 23% from CTP system as compared to 63 and 20% from BCP system. CTP system also showed a more controlled drug release profile compared to BCP system. Modeling of drug release kinetics from CTP system indicated that the release follows Higuchi model with a non-typical Fickian diffusion profile. In vitro biological studies showed the CTP system to be biocompatible with significant antibacterial and anti-inflammatory activity. In vivo implantation studies on rat cranial defects showed greater bone healing and new bone formation in the drug-loaded CTP system compared to control (no carrier) at the end of 12 weeks. The in vitro and in vivo results suggest that the combined drug delivery platform can provide a comprehensive management for all bone infections requiring multi-drug therapy.

Curdlan (β-1,3 glucan) (7 wt%) with polyvinyl alcohol (PVA) (10 wt%) is blended at 1:2 weight ratio and electrospun to get nanofibers and is crosslinked with glutaraldehyde vapor to make it insoluble in water. It has a fiber diameter of less than 100 nm and is hydrophilic (contact angle = 35°). It is biodegradable (10% in 14 d) and also has a good swelling behavior (≈170%). More than 100% of L6 cells are viable on this scaffold after 3 d. The scanning electron microscope images also reveal that cells are able to attach and spread in the nanofibrous scaffolds. In vitro scratch assay indicates that the wound closure rate of curdlan/PVA scaffold is better than PVA scaffold probably due to the immunomodulatory properties of the biopolymer. Thus our results indicate that curdlan/PVA scaffold can be an ideal material for wound healing applications. (Figure presented.).

Hydroxyapatite (HA) nanoparticles were dispersed in AZ31 alloy by friction stir processing (FSP) to produce AZ31/HA metal matrix composites. The composite surface was acid treated using HNO3 and coated with polycaprolactone/HA (PCL/HA) mat by electrospinning. Coating parameters were optimized to obtain PCL/HA nanocomposite fibrous mat with an adhesion strength of 4B Grade (ASTM D3359-09 tape test) on AZ31/HA composite surface. Presence of HA on the substrate and in the coating helps in enhancing biomineralization and develop thick CaP layer on the surface which also facilitates controlled degradation in simulated body fluid at normal physiological conditions of pH 7.2 and 37 °C. Rat skeletal muscle cells culture study showed better adhesion and proliferation on coated samples compared to the uncoated samples. The combination of electrospun nanofibrous PCL/HA composite coating on acid pre-treated FSPed AZ31/HA composite surface seems to have the potential for biodegradable magnesium implant applications with enhanced bioactivity for tissue regeneration.

A novel and easy approach was attempted to developed a ready to use injectable macroporous apatite bone cement derived from eggshell under physiological conditions where, the solid phase contains hydroxyapatite and eggshell derived β-tricalcium phosphate and the liquid phase is the biopolymeric solution (gelatin and chitosan for improving injectability) with disodium hydrogen phosphate (as binding accelerator) in diluted acetic acid. Also, polysorbate as liquid porogen is incorporated in liquid phase (to enhance cement porosity) and it was compared with the cements containing mannitol as solid porogen. All are mixed in an optimized composition to get desired bone cement. The so-formed cements set within clinically acceptable setting time (≤20 min) and are good injectable (>75%), along with stability at physiological pH (∼7.3–7.4). The apatite phased bone cement formed when the after-set cement immersed in phosphate buffer solution (PBS) and incubated for 7 days at physiological conditions, confirmed by X-ray diffraction and Fourier transform Infrared spectroscopy analysis. The cements hold acceptable compressive strength (2.5–4 MPa), within the range of trabecular bone and are also degradable (19–25%) in PBS and simulated body fluid within 70 days. The average pore size of the eggshell derived apatite bone cements (ESDAPCs) falls in between 50 and 250 μm with interconnectivity, confirmed by scanning electron microscopy and micro-CT analysis verified its macroporous nature. The viability and alkaline phosphatase activity of MG63 cells incubated with the ESDAPCs was found to be significantly higher after 3rd and 14th day when compared to their respective controls. Also, the MG63 cells were fully grown over the surface of the ESDAPCs with increased proliferation and extended filopodia. In conclusion, the developed ESDAPC has the ability to become a potential material for repairing low or non-load bearing defects in orthopaedic applications.

Calcium phosphate cements (CPCs) are ideally suited for the local delivery of antibiotics in infected bone defects as they have multiple binding sites for loading various drugs. CPCs can also be substituted with ions such as Ag+, Zn2+, Mg2+, Sr2+, etc., to exhibit extended broad-spectrum antimicrobial activity. Strontium (Sr) in particular is known to enhance the new bone formation and decrease bone resorption. The current work aims to develop a dual action tetracalcium phosphate (TTCP) based cement which releases both the Sr2+ ion and ornidazole antibiotic drug for the treatment of bone infections. The TTCP with Sr2+ ion substitution was prepared by the solid state reaction method and it was used to form ornidazole loaded CPC. The ornidazole loaded cement prepared using 8 at% Sr substituted TTCP (8SCPC-O) showed complete hydroxyapatite (HA) formation in phosphate buffered solution at the end of 1 week. Fine needle-shaped HA crystals were observed in 8SCPC-O cement. In vitro drug release studies showed an accelerated ornidazole release from the 8SCPC-O sample when compared to samples without Sr substitution. Ornidazole releasing cements were found to be biocompatible with skeletal myoblast (L6) cells. Antibacterial activity of ornidazole releasing cement was evident from day 1 onwards against E. coli. The above results suggest 8SCPC-O as a good candidate for treating local bone infections.

Biodegradable metals are important for temporary implant applications. However, poor mechanical properties and higher degradation rate of Mg and its commercial alloys is a major disadvantage. In this study, the above properties were tailored using the groove pressing (GP) technique. Uniform fine-grained (UFG) pure Mg and ZE41 alloy were obtained using GP technique. The impact of GP-induced microstructural changes and surface characteristics on mechanical performance, degradation behavior, biomineralization, surface wettability, and cytocompatibility were investigated. The groove-pressed pure Mg and ZE41 alloy had grain sizes of approximately 30 ± 7.8 and 25 ± 7.5 μm, respectively. The hardness of groove-pressed pure Mg and ZE41 alloy increased by 40% and 26%, respectively, when compared to the annealed condition. Enhanced strength and elongation were observed with these samples. The potentiodynamic polarization test showed a higher Ecorr and lower Icorr values for groove-pressed sample when compared to the annealed condition indicating decreased degradation rate. The weight loss data from immersion tests also show that the degradation rate decreases with time confirming the surface passivity of the sample. The degradation rate decreased by 37% and 44% in simulated body fluid for pure Mg and ZE41 alloy, respectively, after GP. The higher hydroxyapatite deposition on the groove-pressed sample than the annealed sample, as observed from surface morphology studies after immersion in simulated body fluid, confirms the increased biomineralization tendency after GP. The higher wettability of the groove-pressed sample leads to higher cell adhesion and cell viability when compared to the annealed sample indicating higher biocompatibility.

Tetracalcium phosphate (TTCP) is the most basic phase among the calcium phosphate bioceramics. TTCP has been used as self setting bone cement and its relatively high surface pH seems to be very beneficial for cell adhesion and bone formation. Various attempts have been made to improve the cement properties for the ease of use during the surgical procedures. The effect of citric acid (CA) on the setting properties, apatite forming ability, strength and biological behavior of TTCP based bone cement has been studied in detail in the present study. The cement formulation containing 15 % CA has been found to have setting time between 9 and 16 min suitable for bone cement. It also showed complete conversion to apatite phase and highest compressive strength after 28 days of immersion in phosphate buffer at physiological conditions. Cell culture studies using rat skeletal muscle cells confirm higher cell viability in the CA containing cements compared to the pure cement without CA content. The results suggest that self-hardening tetracalcium phosphate based bone cement modified with CA holds a promise for its use in orthopaedic and trauma surgery.