Alfredo Rondinella

Bioactive silicon nitride: A new therapeutic material for osteoarthropathy

While the reciprocity between bioceramics and living cells is complex, it is principally governed by the implant’s surface chemistry. Consequently, a deeper understanding of the chemical interactions of bioceramics with living tissue could ultimately lead to new therapeutic strategies. However, the physical and chemical principles that govern these interactions remain unclear. The intricacies of this biological synergy are explored within this paper by examining the peculiar surface chemistry of a relatively new bioceramic, silicon nitride (Si3N4). Building upon prior research, this paper aims at obtaining new insights into the biological interactions between Si3N4 and living cells, as a consequence of the off-stoichiometric chemical nature of its surface at the nanometer scale. We show here yet unveiled details of surface chemistry and, based on these new data, formulate a model on how, ultimately, Si3N4 influences cellular signal transduction functions and differentiation mechanisms. In other words, we interpret its reciprocity with living cells in chemical terms. These new findings suggest that Si3N4 might provide unique new medicinal therapies and effective remedies for various bone or joint maladies and diseases.

Human osteoblasts grow transitional Si/N apatite in quickly osteointegrated Si3N4 cervical insert

Silicon nitride (Si3N4) ceramics possesses surface chemistry that accelerates bone repair, as previously established by in vitro experiments using both osteosarcoma and mesenchymal cells. The release of silicic acid and nitrogen compounds from the surface Si3N4 enhanced in vitro cellular activity. The results of this study demonstrate for the first time that the osseointegration behavior previously observed is operative with a peculiar chemistry within the human milieu. Si and N elements stimulated progenitor cell differentiation and osteoblastic activity, which ultimately resulted in accelerated bone ingrowth. At the molecular scale, insight into the effect of silicon and nitrogen ions released from the Si3N4 surface was obtained through combined histomorphometric analyses, Raman, Fourier-transform-infrared, and X-ray photoelectron spectroscopies. Identical analyses conducted on a polyetheretherketone (PEEK) spinal explant showed no chemical changes and a lower propensity for osteogenic activity. Silicon and nitrogen are key elements in stimulating cells to generate bony apatite with crystallographic imperfections, leading to enhanced bioactivity of Si3N4 biomedical devices. STATEMENT OF SIGNIFICANCE This research studies osseointegration processes comparing results from explanted PEEK and Si3N4 spinal spacers. Data show that the formation of hydroxyapatite on silicon nitride bio-ceramic surfaces happens with a peculiar mechanism inside the human body. Silicon and nitrogen were incorporated inside the bony tissue structure allowing the developing of off-stoichiometric bony apatite and stimulating progenitor cell differentiation/osteoblastic activity. Silicon and nitrogen ions released from the Si3N4 surface were detected through combined histologic analyses, Raman microspectroscopy, Fourier-transform-infrared, and X-ray photoelectron spectroscopies.

Raman spectroscopic investigation on the molecular structure of apatite and collagen in osteoporotic cortical bone

This study employed highly spectrally resolved Raman spectroscopy to examine the molecular composition of cortical bone tissue obtained from murine females in their healthy and ovariectomy- (OVX-) induced osteoporotic states. The aim of the study was to identify structural differences at the molecular scale both in apatite mineral and collagen fibrils between the two groups of samples. Raman spectroscopy was used to determine the chemical composition of cortical bone in regions including characteristic bands of both bone mineral and bone matrix. The results demonstrated that the mineral apatite of bone did not undergo significant amorphization in its diseased state, with the Raman microprobe also failing in recognizing a direct role of carbonate content in the embrittlement of OVX-diseased bone. On the other hand, complex off-stoichiometry variations could be detected in the columnar Ca-structure of the bony hydroxyapatite according to morphological variations of the Raman band belonging to the symmetric phosphate stretching (A1) band at ~959cm-1. A fundamental role was also recognized for collagen quality on the process of bone embrittlement. The so-called matrix maturity ratio, as systematically measured on Raman spectra in the Amide I region, increased with statistical significance in OVX-treated samples as compared to control samples. An 8% increase could be associated to a 115% increase in elastic stress intensification in the mineral phase of OVX-diseased tissue as compared to the control one, thus proving a degradation in the (elastic) energy-dissipative capacity of a diseased bone matrix.

Wear and surface degradation of commercial ZTA femoral heads under boundary lubrication conditions

The effect of frictional sliding on the surface degradation of commercially available zirconia-toughened alumina (ZTA) femoral heads was studied using a pin-on-ball wear tester under three different lubricating conditions: dry, water, and squalene. Water and squalene were employed under boundary lubrication regimes. Despite the unique (non-standard) character of this apparatus, standard loading conditions could be applied, which effectively determined dynamic friction coefficients as basic material properties. Two types of surface degradation were observed: (i) the polymorphic tetragonal-to-monoclinic (t→m) transformation of the zirconia (ZrO2) dispersoids, and (ii) the off-stoichiometry drift caused by oxygen vacancy formation within the alumina matrix. Scanning laser microscopy (SLM), Raman spectroscopy (RS), scanning electron microscopy (SEM), cathodoluminescence (CL), and X-ray photoelectron spectroscopy (XPS) were utilized to evaluate the fractions of transformed zirconia phase and the stoichiometric evolution of the oxygen sub-lattice at the surface of wear-tested ZTA components. Wear tracks on the surface of the femoral heads were only detected under dry conditions. Dry wear also showed the highest frictional forces and the most pronounced formation of oxygen vacancies among the tested conditions. Conversely, equivalent or greater amounts of the t→m transformation were observed with water and squalene lubrication when compared to the dry wear condition.

Development of a SiYAlON glaze for improved osteoconductivity of implantable medical devices

The application of bioactive coatings onto orthopaedic appliances is commonly performed to compensate for the otherwise bioinert nature of medical devices and to improve their osseointegration. Calcium phosphates, hydroxyapatite (HAp), and bioglasses are commercially available for this purpose. Until recently, few other inorganic compounds have been identified with similar biofunctionality. However, silicon nitride (Si3 N4 ) has emerged as a new orthopaedic material whose unique surface chemistry also enhances osteoconductivity. Recent research has confirmed that its minority intergranular phase, consisting of silicon yttrium aluminum oxynitride (SiYAlON), is principally responsible for this improvement. As a result, it was hypothesized that SiYAlON itself might serve as an effective osteoconductive coating or glaze for medical devices. To test this hypothesis, a process inspired by traditional ceramic whiteware glazing was developed. A slurry containing ingredients similar to the intergranular SiYAlON composition was applied to a Si3 N4 surface, which was then subjected to a heat treatment to form a glaze. Various analytical tools were employed to assess its chemistry and morphology. It was found that the glaze was comprised predominately of Y5 Si3 O12 N, a compound commonly referred to as N-apatite, which is isostructural to native HAp. Subsequent exposure of the glazed surface to acellular simulated body fluid led to increased deposition of biomimetic HAp-like crystals, while exposure to Saos-2 osteosarcoma cells in vitro resulted in greater HAp deposition relative to control samples. The observation that SiYAlON exhibits enhanced osteoconductivity portends its potential as a therapeutic aid in bone and tissue repair.

In Vitro versus In Vivo Phase Instability of Zirconia-Toughened Alumina Femoral Heads: A Critical Comparative Assessment

A clear discrepancy between predicted in vitro and actual in vivo surface phase stability of BIOLOX®delta zirconia-toughened alumina (ZTA) femoral heads has been demonstrated by several independent research groups. Data from retrievals challenge the validity of the standard method currently utilized in evaluating surface stability and raise a series of important questions: (1) Why do in vitro hydrothermal aging treatments conspicuously fail to model actual results from the in vivo environment? (2) What is the preponderant microscopic phenomenon triggering the accelerated transformation in vivo? (3) Ultimately, what revisions of the current in vitro standard are needed in order to obtain consistent predictions of ZTA transformation kinetics in vivo? Reported in this paper is a new in toto method for visualizing the surface stability of femoral heads. It is based on CAD-assisted Raman spectroscopy to quantitatively assess the phase transformation observed in ZTA retrievals. Using a series of independent analytical probes, an evaluation of the microscopic mechanisms responsible for the polymorphic transformation is also provided. An outline is given of the possible ways in which the current hydrothermal simulation standard for artificial joints can be improved in an attempt to reduce the gap between in vitro simulation and reality.

Incorporating Si3N4 into PEEK to Produce Antibacterial, Osteocondutive, and Radiolucent Spinal Implants

Polyetheretherketone (PEEK) is a popular polymeric biomaterial which is primarily used as an intervertebral spacer in spinal fusion surgery; but it is developed for trauma, prosthodontics, maxillofacial, and cranial implants. It has the purported advantages of an elastic modulus which is similar to native bone and it can be easily formed into custom 3D shapes. Nevertheless, PEEKs disadvantages include its poor antibacterial resistance, lack of bioactivity, and radiographic transparency. This study presents a simple approach to correcting these three shortcomings while preserving the base polymers biocompatibility, chemical stability, and elastic modulus. The proposed strategy consists of preparing a PEEK composite by dispersing a minor fraction (i.e., 15 vol%) of a silicon nitride (Si3 N4 ) powder within its matrix. In vitro tests of PEEK composites with three Si3 N4 variants-β-Si3 N4 , α-Si3 N4 , and β-SiYAlON-demonstrate significant improvements in the polymers osteoconductive versus SaOS-2 cells and bacteriostatic properties versus gram-positive Staphylococcus epidermidis bacteria. These properties are clearly a consequence of adding the bioceramic dispersoids, according to chemistry similar to that previously demonstrated for bulk Si3 N4 ceramics in terms of osteogenic behavior (vs both osteosarcoma and mesenchymal progenitor cells) and antibacterial properties (vs both gram-positive and gram-negative bacteria).

Oxide ceramic femoral heads contribute to the oxidation of polyethylene liners in artificial hip joints

Experimental evidence demonstrates that a loss of stoichiometry at the surface of oxide bioceramic femoral heads enhances the oxidation rate of polyethylene acetabular liners in artificial hip joints. Contradicting the common notion that ceramics are bioinert, three independent experiments confirmed substantial chemical interactions between the ceramic femoral heads and their polyethylene counterparts. The experiments reported herein included hydrothermal tests, frictional tests, and hip-simulator experiments. It was discovered that oxide and non-oxide femoral heads differently affected the oxidation processes at the surface of the polyethylene liners, all other testing parameters being equal. Analytical data from X-ray photoelectron (XPS), cathodoluminescence (CL), Fourier-transform infrared (FTIR), and Raman spectroscopies unequivocally and consistently showed that the oxidation rate of polyethylene liners was greater when coupled with oxide as opposed to non-oxide ceramic heads. XPS analyses of O-Al-O bond fractions at the surface of a zirconia-toughened alumina (ZTA) short-term (20 months in vivo) femoral heads retrieval showed a ~50% reduction in favor of oxygen vacancy O-Al-VO and hydroxylated Al-O-H bonds. Off-stoichiometry drifts were confirmed in vitro under both static and dynamic conditions. They triggered oxidation and tangibly affected an advanced highly cross-linked sequentially irradiated and annealed ultra-high molecular weight polyethylene (UHMWPE) liner (increase in oxidation index up to ΔOI~1.2 after 5 × 105 cycles under dynamic swing conditions). Second-generation UHMWPE liners infused with vitamin E were also affected by the free flow of oxygen from the oxide femoral heads, although to a lesser extent. The fundamental findings of this study, which were also confirmed on retrievals, call for revised standards in material design and testing. Adopting these new criteria will provide an improved understanding of the importance of off-stoichiometry at the head/liner interface and may lead to significant extensions in artificial joint lifetimes.

Green Tea Polyphenols Coupled with a Bioactive Titanium Alloy Surface: In Vitro Characterization of Osteoinductive Behavior through a KUSA A1 Cell Study

A chemically-treated titanium alloy (Ti6Al4V) surface, able to induce hydroxyapatite precipitation from body fluids (inorganic mineralization activity), was functionalized with a polyphenolic extract from green tea (tea polyphenols, TPH). Considering that green tea polyphenols have stimulating effects on bone forming cells (biological mineralization), the aim was to test their osteoinductive behavior due to co-operation of inorganic and biological mineralization on mesenchymal stem cells KUSA A1. The functionalized surfaces were characterized by using the Folin–Ciocalteu method and X-ray photoelectron spectroscopy to confirm the successful outcome of the functionalization process. Two cell cultures of mesenchymal stem cells, KUSA A1 were performed, with or without osteoinductive factors. The cells and surfaces were characterized for monitoring cell viability and hydroxyapatite production: Fourier Transform Infrared Spectroscopy and Raman spectroscopy analyses showed deposition of hydroxyapatite and collagen due to the cell activity, highlighting differentiation of KUSA A1 into osteoblasts. A higher production of extracellular matrix was highlighted on the functionalized samples by laser microscope and the fluorescence images showed higher viability of cells and greater presence of osteocalcin in these samples. These results highlight the ability of polyphenols to improve cell differentiation and to stimulate biological mineralization, showing that surface functionalization of metal implants could be a promising way to improve osteointegrability.

Bioceramics are Not Bioinert: The Role of Oxide and Non-Oxide Bioceramics on the Oxidation of UHMWPE Components in Artificial Joints

The following research is aimed at understanding the influence of Zirconia-Toughened Alumina (ZTA) and Silicon Nitride (Si3N4) on Ultra-High Molecular Weight Polyethylene (UHMWPE) acetabular liners. Bioceramic femoral heads were systematically tested against UHMWPE in controlled environment according to static/load-free coupling in hydrothermal environment, pin-on-ball wear testing, and hip-simulator wear testing. In addition, a retrieved ZTA femoral head has been analyzed and results have been compared to the simulations. Experimental results from X-ray photoelectron (XPS), cathodoluminescence (CL), Raman and Fourier-Transformed Infrared spectroscopy suggest that, despite conventional notions imply that bioceramics are inert, the surface chemistry of bioceramics was relevant to the oxidation rate of polyethylene liners. Non-biointertness could either be advantageous or disadvantageous toward polyethylene oxidation. The main reason resides in the peculiar chemical interactions between polyethylene and different ceramics, and, more specifically, depends on the direction of oxygen flow at the interface between the ceramic and the polymer. ZTA femoral heads were found to release a non-negligible amount of oxygen moieties from their surfaces, thus accelerating oxidative degradation of polyethylene. Conversely, Si3N4 ceramics exerted a protective role towards the polyethylene liner by scavenging oxygen from the tribolayer. The results of this work provide new insights into the interaction between bioceramics and polymers, which should also be considered when designing the next generation artificial hip joints with significantly elongated lifetimes.