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Open Access Publications from the University of California

Scholarly Works (36 results)

  • Thesis
  • Peer Reviewed
UCLA Electronic Theses and Dissertations (2024)

Titanium implants are the standard therapeutic option for restoring missing teeth and reconstructing fractured or diseased bone. Despite the advancements in implant surface technology over the past 30 years, the osseointegration capabilities of titanium have not significantly improved. This study aimed to develop a novel microrough titanium surface with smooth, rounded edges to enhance osteoblast proliferation and differentiation while improving osseointegration [1,2,5].

In this study, the surface characterization involved flame-heating titanium surfaces thus transforming sharp, knife-like edges into smooth, rounded forms. Scanning Electron Microscopy (SEM) and Atomic Force Microscopy (AFM) were employed to visualize and quantify surface morphology changes. X-ray Photoelectron Spectroscopy (XPS) confirmed that the surface chemistry remained consistent post-treatment. Wettability was assessed via water contact angle measurements, and cell attachment and proliferation were evaluated using WST-1 assays, fluorescent microscopy, BrdU incorporation, and flow cytometry.

Our findings indicated a substantial enhancement in osteoblast attachment and proliferation on the soft edge microrough surface compared to the knife-edge surface. The WST-1 assay, and fluorescent microscopy confirmed higher cell attachment on the newly created surface. Enhanced cell spreading behavior was observed, with larger cell area, perimeter, and Feret's diameter. Proliferation assays showed sustained increases in cell numbers, and BrdU incorporation confirmed higher DNA synthesis rates. Flow cytometric analysis indicated improved cell cycle progression.

Osteoblast differentiation and function were assessed through ALP activity, matrix mineralization assays, and gene expression analysis. Alizarin red staining demonstrated robust extracellular matrix formation and mineralization on the new surface. Gene expression analysis was also performed to compare the osteogenic potential.

Thus, we demonstrated that our novel micro- rough titanium surface generates substantially increased Osseo integrative ability over the previously available micro-rough titanium surface. This novel surface can have vivid applications in dental and orthopedic fields as well as titanium-based bone engineering scaffolds.

    Cover page: A novel, soft edge microrough titanium surface to overcome osteoblasts kinetics dichotomy
    • Thesis
    • Peer Reviewed
    UCLA Electronic Theses and Dissertations (2012)

    Although dental implants have become a standard treatment procedure, there is still a growing demand to enhance their osseointegration capability to address complex cases, such as host sites with bone grafts and type IV bone. This study explored the viability and effectiveness of pre-osseointegrated implants to achieve faster establishment of bone-implant integration using cell sheet technology. Cells harvested from the rat bone marrow were cultured on a poly (N-isopropylacrylamide) culture dish. Taking advantage of the ability of temperature responsive conversion of the dish from hydrophobic to hydrophilic, the layer of the grown cells was detached as a sheet. For in-vitro experiments, the cell sheet (single or double layered) was transferred to acid-etched titanium disks and examined for its capability to show cell metabolic

    activity, osteoblastic phenotypes and responsiveness to an osteogenic enhancer molecule, N-acetyl cysteine (NAC). For in vivo study, the experimental titanium implants (1 mm in diameter and 2 mm in length) were coated with the cell sheet and placed into rat femurs. After 2 weeks of healing, the implants were subjected to the biomechanical push-in test to evaluate the strength of bone-implant integration. Cell sheet grown on temperature sensitive dishes showed the expected cell metabolic activity levels and osteoblastic phenotypes of cells grown on regular culture dishes. Cell sheets, created from bone marrow cells, were successfully transferred as an intact sheet, to titanium disks and showed the alkaline phosphatase (ALP) activity. Double layers of the sheets showed 2-fold greater cell metabolic activity than single layered ones. The cell sheets treated with NAC showed significantly greater ALP activity in a NAC dose-dependent manner. The cell sheet-coated implants showed a 2.2-fold greater push-in value than uncoated implants. This is the first study that has established an osteogenic cell sheet applicable to titanium implants. Transferred cell sheets to titanium surfaces are capable to show reliable osteogenic phenotypes and to accelerate the bone-titanium integration, can be enhanced by foreign molecules and show increased cell metabolic activity levels depending on the number of layers of cell sheets transferred.

      Cover page: Cell-Sheet Technology: A Novel Method To Enhance Bone-Implant Integration
      • Article
      • Peer Reviewed
      UCLA Previously Published Works (2022)
      The fibroblast-rich gingival tissue is usually in contact with or adjacent to cytotoxic polymer-based dental restoration materials. The objective of this study was to determine whether the antioxidant amino acid, N-acetyl cysteine (NAC), reduces the toxicity of dental restorative materials. Human oral fibroblasts were cultured with bis-acrylic, flowable composite, bulk-fill composite, self-curing acrylic, and titanium alloy test specimens. Cellular behavior and function were analyzed on and around the materials. Impregnation of the bulk-fill composite and self-curing acrylic with NAC reduced their toxicity, improving the attachment, growth, and function of human oral fibroblasts on and around the materials. These mitigating effects were NAC dose dependent. However, NAC impregnation of the bis-acrylic and flowable composite was ineffective, with no cells attaching to nor around the materials. Although supplementing the culture medium with NAC also effectively improved fibroblast behaviors, direct impregnation of materials with NAC was more effective than supplementing the cultures. NAC-mediated improvements in fibroblast behavior were associated with reduced production of reactive oxygen species and oxidized glutathione together with increased glutathione reserves, indicating that NAC effectively directly scavenged ROS from materials and reinforced the cellular antioxidant defense system. These results establish a proof of concept of NAC-mediated improvements in biocompatibility in the selected dental restorative materials.
        Cover page: N-Acetyl Cysteine-Mediated Improvements in Dental Restorative Material Biocompatibility
        • Article
        • Peer Reviewed
        UCLA Previously Published Works (2019)
        Oxidative stress hinders tissue regeneration in cell therapy by inducing apoptosis and dysfunction in transplanted cells. N-acetyl-L-cysteine (NAC) reinforces cellular antioxidant capabilities by increasing a major cellular endogenous antioxidant molecule, glutathione, and promotes osteogenic differentiation. This study investigates the effects of pretreatment of osteoblast-like cells with NAC on oxidative stress-induced apoptosis and dysfunction and bone regeneration in local transplants. Rat femur bone marrow-derived osteoblast-like cells preincubated for 3 h with and without 5 mM NAC were cultured in a NAC-free osteogenic differentiation medium with continuous exposure to 50 μM hydrogen peroxide to induce oxidative stress. NAC preincubation prevented disruption of intracellular redox balance and alleviated apoptosis and negative impact on osteogenic differentiation, even under oxidative stress. Autologous osteoblast-like cells with and without NAC pretreatment in a collagen sponge vehicle were implanted in critical-size defects in rat femurs. In the third week, NAC-pretreated cells yielded complete defect closure with significantly matured lamellar bone tissue in contrast with poor bone healing by cells without pretreatment. Cell-tracking analysis demonstrated direct bone deposition by transplanted cells pretreated with NAC. Pretreatment of osteoblast-like cells with NAC enhances bone regeneration in local transplantation by preventing oxidative stress-induced apoptosis and dysfunction at the transplanted site.
          Cover page: Cytoprotective Preconditioning of Osteoblast-Like Cells with N-Acetyl-L-Cysteine for Bone Regeneration in Cell Therapy.
          • Article
          • Peer Reviewed
          UCLA Previously Published Works (2023)
          Ultraviolet (UV) photofunctionalization counteracts the biological aging of titanium to increase the bioactivity and osseointegration of titanium implants. However, UV photofunctionalization currently requires long treatment times of between 12 min and 48 h, precluding routine clinical use. Here, we tested the ability of a novel, xenon excimer lamp emitting 172 nm vacuum UV (VUV) to decompose organic molecules coated on titanium as a surrogate of photofunctionalization. Methylene blue as a model organic molecule was coated on grade 4 commercially pure titanium and treated with four UV light sources: (i) ultraviolet C (UVC), (ii) high-energy UVC (HUVC), (iii) proprietary UV (PUV), and (iv) VUV. After one minute of treatment, VUV decomposed 57% of methylene blue compared with 2%, 36%, and 42% for UVC, HUVC, and PUV, respectively. UV dose-dependency testing revealed maximal methylene blue decomposition with VUV within one minute. Equivalent decomposition was observed on grade 5 titanium alloy specimens, and placing titanium specimens in quartz ampoules did not compromise efficacy. Methylene blue was decomposed even on polymethyl methacrylate acrylic specimens at 20-25% lower efficiency than on titanium specimens, indicating a relatively small contribution of titanium dioxide-mediated photocatalytic decomposition to the total decomposition. Load-testing revealed that VUV maintained high efficacy of methylene blue decomposition regardless of the coating density, whereas other UV light sources showed low efficacy with thin coatings and plateauing efficacy with thicker coatings. This study provides foundational data on rapid and efficient VUV-mediated organic decomposition on titanium. In synergy with quartz ampoules used as containers, VUV has the potential to overcome current technical challenges hampering the clinical application of UV photofunctionalization.
            Cover page: Decomposing Organic Molecules on Titanium with Vacuum Ultraviolet Light for Effective and Rapid Photofunctionalization
            • Article
            • Peer Reviewed
            UCLA Previously Published Works (2016)
            Titanium undergoes time-dependent degradation in biological capability, or "biological aging". It is unknown whether the biological aging of titanium occurs beyond four weeks and whether age-related changes are definitely associated with surface hydrophilicity. We therefore measured multiple biological parameters of bone marrow-derived osteoblasts cultured on newly prepared, one-month-old, three-month-old, and six-month-old acid-etched titanium surfaces, as well as the hydrophilicity of these surfaces. New surfaces were superhydrophilic with a contact angle of ddH₂O of 0°, whereas old surfaces were all hydrophobic with the contact angle of around 90°. Cell attachment, cell spread, cell density, and alkaline phosphatase activity were highest on new surfaces and decreased in a time-dependent manner. These decreases persisted and remained significant for most of the biological parameters up to six-months. While the number of attached cells was negatively correlated with hydrophilicity, the other measured parameters were not. The biological capability of titanium continues to degrade up to six months of aging, but these effects are not directly associated with time-dependent reductions in hydrophilicity. A full understanding of the biological aging will help guide regulatory improvements in implant device manufacturing and develop countermeasures against this phenomenon in order to improve clinical outcomes.
              Cover page: Long-Term Progressive Degradation of the Biological Capability of Titanium
              • Article
              • Peer Reviewed
              UCLA Previously Published Works (2023)
              This is the first genome-wide transcriptional profiling study using RNA-sequencing to investigate osteoblast responses to different titanium surface topographies, specifically between machined, smooth and acid-etched, microrough surfaces. Rat femoral osteoblasts were cultured on machine-smooth and acid-etched microrough titanium disks. The culture system was validated through a series of assays confirming reduced osteoblast attachment, slower proliferation, and faster differentiation on microrough surfaces. RNA-sequencing analysis of osteoblasts at an early stage of culture revealed that gene expression was highly correlated (r = 0.975) between the two topographies, but 1.38 % genes were upregulated and 0.37 % were downregulated on microrough surfaces. Upregulated transcripts were enriched for immune system, plasma membrane, response to external stimulus, and positive regulation to stimulus processes. Structural mapping confirmed microrough surface-promoted gene sharing and networking in signaling pathways and immune system/responses. Target-specific pathway analysis revealed that Rho family G-protein signaling pathways and actin genes, responsible for the formation of stress fibers, cytoplasmic projections, and focal adhesion, were upregulated on microrough surfaces without upregulation of core genes triggered by cell-to-cell interactions. Furthermore, disulfide-linked or -targeted extracellular matrix (ECM) or membranous glycoproteins such as laminin, fibronectin, CD36, and thrombospondin were highly expressed on microrough surfaces. Finally, proliferating cell nuclear antigen (PCNA) and cyclin D1, whose co-expression reduces cell proliferation, were upregulated on microrough surfaces. Thus, osteoblasts on microrough surfaces were characterized by upregulation of genes related to a wide range of functions associated with the immune system, stress/stimulus responses, proliferation control, skeletal and cytoplasmic signaling, ECM-integrin receptor interactions, and ECM-membranous glycoprotein interactions, furthering our knowledge of the surface-dependent expression of osteoblastic biomarkers on titanium.
                Cover page: Genome-wide transcriptional responses of osteoblasts to different titanium surface topographies.
                • Article
                • Peer Reviewed
                UCLA Previously Published Works (2023)
                Titanium undergoes biological aging, represented by increased hydrophobicity and surface accumulation of organic molecules over time, which compromises the osseointegration of dental and orthopedic implants. Here, we evaluated the efficacy of a novel UV light source, 172 nm wavelength vacuum UV (VUV), in decomposing organic molecules around titanium. Methylene blue solution used as a model organic molecule placed in a quartz ampoule with and without titanium specimens was treated with four different UV light sources: (i) ultraviolet C (UVC), (ii) high-energy UVC (HUVC), (iii) proprietary UV (PUV), and (iv) VUV. After one minute of treatment, VUV decomposed over 90% of methylene blue, while there was 3-, 3-, and 8-fold more methylene blue after the HUVC, PUV, and UVC treatments, respectively. In dose-dependency experiments, maximal methylene blue decomposition occurred after one minute of VUV treatment and after 20-30 min of UVC treatment. Rapid and effective VUV-mediated organic decomposition was not influenced by the surface topography of titanium or its alloy and even occurred in the absence of titanium, indicating only a minimal photocatalytic contribution of titanium dioxide to organic decomposition. VUV-mediated but not other light source-mediated methylene blue decomposition was proportional to its concentration. Plastic tubes significantly reduced methylene blue decomposition for all light sources. These results suggest that VUV, in synergy with quartz ampoules, mediates rapid and effective organic decomposition compared with other UV sources. This proof-of-concept study paves the way for rapid and effective VUV-powered photofunctionalization of titanium to overcome biological aging.
                  Cover page: A Novel High-Energy Vacuum Ultraviolet Light Photofunctionalization Approach for Decomposing Organic Molecules around Titanium
                  • Article
                  • Peer Reviewed
                  UCLA Previously Published Works (2024)
                  Considering the biological activity of osteoblasts is crucial when devising new approaches to enhance the osseointegration of implant surfaces, as their behavior profoundly influences clinical outcomes. An established inverse correlation exists between osteoblast proliferation and their functional differentiation, which constrains the rapid generation of a significant amount of bone. Examining the surface morphology of implants reveals that roughened titanium surfaces facilitate rapid but thin bone formation, whereas smooth, machined surfaces promote greater volumes of bone formation albeit at a slower pace. Consequently, osteoblasts differentiate faster on roughened surfaces but at the expense of proliferation speed. Moreover, the attachment and initial spreading behavior of osteoblasts are notably compromised on microrough surfaces. This review delves into our current understanding and recent advances in nanonodular texturing, meso-scale texturing, and UV photofunctionalization as potential strategies to address the biological dilemma of osteoblast kinetics, aiming to improve the quality and quantity of osseointegration. We discuss how these topographical and physicochemical strategies effectively mitigate and even overcome the dichotomy of osteoblast behavior and the biological challenges posed by microrough surfaces. Indeed, surfaces modified with these strategies exhibit enhanced recruitment, attachment, spread, and proliferation of osteoblasts compared to smooth surfaces, while maintaining or amplifying the inherent advantage of cell differentiation. These technology platforms suggest promising avenues for the development of future implants.
                    Cover page: Beyond microroughness: novel approaches to navigate osteoblast activity on implant surfaces.
                    • Article
                    • Peer Reviewed
                    UCLA Previously Published Works (2022)
                    Bone cement containing benzoyl peroxide (BPO) as a polymerization initiator are commonly used to fix orthopedic metal implants. However, toxic complications caused by bone cement are a clinically significant problem. Poly (methyl methacrylate) tri-n-butylborane (PMMA-TBB), a newly developed material containing TBB as a polymerization initiator, was found to be more biocompatible than conventional PMMA-BPO bone cements due to reduced free radical generation during polymerization. However, free radicals might not be the only determinant of cytotoxicity. Here, we evaluated the response and functional phenotypes of cells exposed to extracts derived from different bone cements. Bone cement extracts were prepared from two commercial PMMA-BPO cements and an experimental PMMA-TBB. Rat bone marrow-derived osteoblasts and osteoclasts were cultured in a medium supplemented with bone cement extracts. More osteoblasts survived and attached to the culture dish with PMMA-TBB extract than in the culture with PMMA-BPO extracts. Osteoblast proliferation and differentiation were higher in the culture with PMMA-TBB extract. The number of TRAP-positive multinucleated cells was significantly lower in the culture with PMMA-TBB extract. There was no difference in osteoclast-related gene expression in response to different bone cement extracts. In conclusion, PMMA-TBB extract was less toxic to osteoblasts than PMMA-BPO extracts. Although extracts from the different cement types did not affect osteoclast function, PMMA-TBB extract seemed to reduce osteoclastogenesis, a possible further advantage of PMMA-TBB cement. These implied that the reduced radical generation during polymerization is not the only determinant for the improved biocompatibility of PMMA-TBB and that the post-polymerization chemical elution may also be important.
                      Cover page: Novel Tuning of PMMA Orthopedic Bone Cement Using TBB Initiator: Effect of Bone Cement Extracts on Bioactivity of Osteoblasts and Osteoclasts
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