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Cytotoxicity of Dental Materials

New this year · #32 of 33 · on the outline, never on a past final · topic 4.11 — every high-yield fact from the book chapter, exam-ready.

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  1. Front
    What single axiom does all of toxicology rest on?
    Back
    The dose makes the poison
  2. Front
    IC50 stands for the half-maximal _____ concentration.
    Back
    inhibitory
  3. Front
    What does IC50 specifically measure?
    Back
    The concentration that reduces a measured cellular process to 50% of the uninhibited control
  4. Front
    A material with a LOW IC50 is more or less potent as a toxin?
    Back
    More potent (it takes very little to half-shut down cell function)
  5. Front
    Necrosis is the passive form of cell death; _____ is the active, regulated form.
    Back
    apoptosis
  6. Front
    Which mode of cell death ( necrosis vs apoptosis ) requires ATP / is energy-dependent?
    Back
    Apoptosis ( necrosis is passive, requires no energy)
  7. Front
    What is EC50 (half-maximal effective concentration)?
    Back
    The concentration at which 50% of the maximum POSITIVE (stimulatory/beneficial) effect is achieved
  8. Front
    In necrosis , why does the cell swell?
    Back
    Ion pumps fail , the cell loses osmotic control, and water rushes in along its gradient ( oncosis )
  9. Front
    LD50 is expressed in what units, and what does it measure?
    Back
    mg of substance per kg body weight ; the dose lethal to 50% of a test (whole-animal) population
  10. Front
    What term describes the cell-and-organelle swelling that occurs in necrosis ?
    Back
    Oncosis
  11. Front
    Which death mode releases DAMPs (e.g. HMGB1, ATP, heat shock proteins) and triggers inflammation ?
    Back
    Necrosis
  12. Front
    Therapeutic index (TI) is calculated as TI = _____ / ED50.
    Back
    LD50
  13. Front
    In apoptosis , does the cell shrink or swell?
    Back
    Shrinks ( necrosis swells)
  14. Front
    A WIDE therapeutic index means what about a material's safety?
    Back
    You can achieve the therapeutic/effective benefit at concentrations far below those that kill cells ( safer margin )
  15. Front
    Which death mode keeps its plasma membrane intact until the cell is engulfed?
    Back
    Apoptosis
  16. Front
    Why is necrosis described as self-amplifying in the confined pulp chamber ?
    Back
    DAMP -driven inflammatory edema raises intrapulpal pressure , compromising blood flow and propagating injury beyond the original site
  17. Front
    What are the membrane-bound packets formed when an apoptotic cell blebs called?
    Back
    Apoptotic bodies
  18. Front
    Which phospholipid flips from the inner to the outer membrane leaflet to flag an apoptotic cell for phagocytosis?
    Back
    Phosphatidylserine
  19. Front
    Caspases are what class of enzyme, and in what inactive form do they exist?
    Back
    Cysteine proteases ; they exist as inactive zymogens ( procaspases )
  20. Front
    The intrinsic apoptotic pathway is initiated at which organelle?
    Back
    The mitochondrion
  21. Front
    In intrinsic apoptosis , what mitochondrial protein leaks into the cytoplasm to start the cascade?
    Back
    Cytochrome c
  22. Front
    Cytochrome c joins with Apaf-1 and procaspase-9 to form what complex?
    Back
    The apoptosome
  23. Front
    The apoptosome activates which initiator caspase?
    Back
    Caspase-9
  24. Front
    Caspase-9 cleaves and activates which executioner caspases?
    Back
    Caspases 3, 6, and 7
  25. Front
    Caspase-3 activates _____, the nuclease that cuts genomic DNA between nucleosomes.
    Back
    CAD (caspase-activated DNase)
  26. Front
    What characteristic gel-electrophoresis pattern is a definitive biochemical marker of apoptosis ?
    Back
    A DNA ladder (evenly spaced bands of 180–200 bp and multiples)
  27. Front
    DNA cut by CAD produces fragments of what size (and multiples)?
    Back
    180–200 base pairs
  28. Front
    On a gel, what DNA pattern does NECROSIS produce (vs the apoptotic ladder)?
    Back
    A random smear
  29. Front
    Within a single restoration, which death mode predominates closest to the margin where monomer concentration is highest?
    Back
    Necrosis ( apoptosis occurs deeper where concentration is sublethal)
  30. Front
    What biochemical principle is shared by the MTT, XTT, MTS, and WST-1 assays?
    Back
    Mitochondrial dehydrogenases in viable cells reduce a tetrazolium salt to a colored formazan product
  31. Front
    The MTT assay measures _____ activity as a proxy for cell viability.
    Back
    mitochondrial dehydrogenase (metabolic)
  32. Front
    What colored product does the MTT assay generate, and at what wavelength is it read?
    Back
    Insoluble purple formazan ; absorbance read at 570 nm
  33. Front
    Below what % of control viability is a material classified as cytotoxic by ISO 10993-5 (MTT)?
    Back
    Below 70% of control
  34. Front
    What operational advantage do XTT and WST-1 have over the original MTT assay?
    Back
    They form water-soluble formazan , so no lysis/solubilization (DMSO) step is needed
  35. Front
    The LDH release assay specifically detects which type of cell damage?
    Back
    Membrane rupture (the hallmark of necrosis )
  36. Front
    Why does lactate dehydrogenase appear in the culture medium only when cells are damaged?
    Back
    LDH is a cytoplasmic enzyme confined inside intact cells; it leaks out only when the plasma membrane ruptures
  37. Front
    A material shows preserved MTT activity but ELEVATED LDH release. What does this indicate?
    Back
    Early-phase necrosis (or pyroptosis ) — mitochondria intact but membrane permeabilized
  38. Front
    What does the neutral red uptake assay exploit?
    Back
    Lysosomal integrity — viable cells trap the protonated dye in their acidic lysosomes; damaged cells cannot
  39. Front
    In live/dead staining, calcein-AM yields a green signal in which cells, and why?
    Back
    Live cells — esterases convert it to fluorescent calcein , which their intact membranes retain
  40. Front
    In live/dead staining, ethidium homodimer or propidium iodide marks which cells (and what color)?
    Back
    Dead cells with compromised membranes (red/orange) — these dyes are membrane-impermeant DNA intercalators excluded from live cells
  41. Front
    Which flow-cytometry probe binds externalized phosphatidylserine to mark EARLY apoptosis ?
    Back
    Annexin V
  42. Front
    In Annexin V / PI flow cytometry, what staining pattern indicates early apoptosis ?
    Back
    Annexin V positive / PI negative (AnnV+/PI–)
  43. Front
    In Annexin V / PI flow cytometry, AnnV+/PI+ indicates what?
    Back
    Late apoptosis or necrosis
  44. Front
    In the ISO agar overlay method, what does the agar layer simulate?
    Back
    The dentin barrier — a diffusion barrier eluates must cross to reach the cells
  45. Front
    Which ISO contact configuration (agar overlay vs direct contact) is the harshest screen?
    Back
    Direct contact — the material sits directly on the cell monolayer (maximum exposure)
  46. Front
    What is the typical degree of conversion of even a well-cured composite?
    Back
    Roughly 55–75% (so 25–45% of monomer remains unreacted )
  47. Front
    Why do residual monomers in a composite continuously elute rather than staying locked in place?
    Back
    They are small molecules in a polymer network that swells in aqueous environments , releasing them into any contacting fluid
  48. Front
    What do HEMA , TEGDMA , and BisGMA stand for? (full names)
    Back
    HEMA = 2-hydroxyethyl methacrylate; TEGDMA = triethylene glycol dimethacrylate; BisGMA = bisphenol A glycidyl methacrylate
  49. Front
    Which resin monomer is the smallest and most hydrophilic, diffusing most readily to the pulp?
    Back
    HEMA
  50. Front
    Rank HEMA , TEGDMA , BisGMA by ability to DIFFUSE through dentinal tubules to the pulp.
    Back
    HEMA > TEGDMA > BisGMA (smaller, more hydrophilic = better penetration)
  51. Front
    Rank HEMA , TEGDMA , BisGMA by intrinsic cytotoxicity PER MOLECULE.
    Back
    BisGMA > TEGDMA > HEMA (BisGMA has the lowest IC50)
  52. Front
    By depleting which tripeptide antioxidant does HEMA tip the cell toward oxidative stress?
    Back
    Glutathione (GSH)
  53. Front
    What is the amino acid sequence of glutathione , and which residue carries the active reducing thiol?
    Back
    Glu–Cys–Gly ; the thiol (–SH) on cysteine is the active reducing species
  54. Front
    When glutathione is oxidized it forms _____.
    Back
    GSSG (glutathione disulfide)
  55. Front
    Which common resin monomer is reported as the MOST cytotoxic on a molar basis?
    Back
    TEGDMA
  56. Front
    Besides generating ROS , how does TEGDMA damage intracellular proteins?
    Back
    It alkylates them through its epoxide-like reactivity
  57. Front
    Which resin monomer has estrogenic activity, and why?
    Back
    BisGMA — due to its bisphenol A backbone
  58. Front
    Beyond ROS generation, BisGMA activates which ER-stress pathway?
    Back
    The unfolded protein response (UPR) / endoplasmic reticulum stress
  59. Front
    How does remaining dentin thickness (RDT) relate to monomer concentration reaching the pulp — linearly or exponentially?
    Back
    Exponentially (not linear)
  60. Front
    Roughly how much lower is pulpal monomer exposure under 2 mm of dentin vs 0.5 mm?
    Back
    About 100-fold lower
  61. Front
    Name the two liner materials used in deep preparations to add a diffusion barrier and stimulate tertiary dentin.
    Back
    Calcium hydroxide and MTA
  62. Front
    What is the primary mercury form released from dental amalgam during service?
    Back
    Elemental mercury vapor (Hg⁰) , with lesser inorganic Hg²⁺
  63. Front
    Mechanistically, how does Hg²⁺ disable a cell's entire redox defense system?
    Back
    It binds cysteine thiols on glutathione and antioxidant enzymes ( glutathione peroxidase , thioredoxin , thioredoxin reductase ), inhibiting them all simultaneously
  64. Front
    Why is clinical amalgam considered acceptable despite mercury's real cytotoxic mechanism?
    Back
    Pulpal mercury concentrations stay well below the IC50 for pulp cells (wide therapeutic index)
  65. Front
    Eugenol is the active component of which cement/material?
    Back
    Zinc oxide–eugenol (ZOE)
  66. Front
    Name two targets eugenol inhibits that explain its anti-inflammatory and analgesic effects.
    Back
    Cyclooxygenase (COX) and NF-κB signaling (plus voltage-gated Na⁺ channels for analgesia)
  67. Front
    Why is eugenol (ZOE) contraindicated for DIRECT pulp capping?
    Back
    It is directly cytotoxic to odontoblasts AND blocks their adhesion to ECM proteins , preventing reparative dentin bridge formation
  68. Front
    What is the pH of freshly mixed glass-ionomer cement, and what causes it?
    Back
    pH 3–4 (acidic) , due to the polyacrylic acid component
  69. Front
    Why is freshly mixed GIC only TRANSIENTLY cytotoxic?
    Back
    As it sets, the acid is consumed forming ionic crosslinks with the glass; pH rises to neutral within minutes-to-hours, leaving the set material essentially inert
  70. Front
    Set GIC releases fluoride , which inhibits which glycolytic enzyme in oral bacteria?
    Back
    Enolase
  71. Front
    Why does GIC have no eluted-monomer problem like composite?
    Back
    It bonds chemically to tooth structure and needs no adhesive resin system
  72. Front
    Which ISO 10993 part governs cytotoxicity testing specifically?
    Back
    ISO 10993-5
  73. Front
    What is the ISO reference cell line for Tier 1 in vitro cytotoxicity screening?
    Back
    L929 mouse fibroblasts
  74. Front
    List the three tiers of ISO 10993 biocompatibility testing in order.
    Back
    Tier 1 in vitro cytotoxicity → Tier 2 in vivo animal implantation → Tier 3 clinical/usage studies
  75. Front
    For pulp-contacting materials, Tier 2 animal studies assess pulpal response at which time points?
    Back
    7, 30, and 90 days
  76. Front
    What standardized surface-area-to-volume ratio is typically used for ISO elution testing?
    Back
    About 1.25 cm²/mL (eluted for 24 or 72 hours)
  77. Front
    Why must even a material with a favorable in vitro profile still undergo animal validation?
    Back
    A cell monolayer cannot recapitulate the dentin diffusion barrier , pulpal blood flow, immune competence, or the 3-D pulp-dentin organization
  78. Front
    A resin cement shows MTT viability of 48% but no significant LDH elevation. Which death pathway is most likely active?
    Back
    Early-to-mid apoptosis (mitochondria impaired, membrane still intact); had LDH been elevated, necrosis would dominate
  79. Front
    What does opening the mitochondrial permeability transition pore release to start the apoptosome cascade?
    Back
    Cytochrome c
  80. Front
    Which specific ECM proteins does eugenol prevent odontoblasts from adhering to?
    Back
    Fibronectin and laminin
  81. Front
    What overall SHAPE does the dose-response curve take?
    Back
    Sigmoidal (S-shaped)
  82. Front
    Name the three regions of the sigmoidal dose-response curve in order of increasing dose.
    Back
    Low-dose tolerance zone (no measurable effect) -> steep inflection point -> high-dose plateau (maximum effect)
  83. Front
    On a dose-response curve, what is plotted on the x-axis (and on what scale) vs the y-axis?
    Back
    x-axis = concentration/dose on a logarithmic scale ; y-axis = biological effect as % of maximum
  84. Front
    On the sigmoidal curve, what is happening at the inflection point?
    Back
    A small increase in dose produces a steep increase in effect
  85. Front
    Two materials with IDENTICAL IC50 values can differ greatly in safety because of which curve property?
    Back
    The slope of the dose-response curve (its Hill coefficient)
  86. Front
    What does a STEEP dose-response slope imply for clinical safety vs a shallow slope?
    Back
    Steep slope = cells shift from ~10% to ~90% effect over a tiny dose window ( little margin for error ); shallow slope = wider safe dose window
  87. Front
    What parameter describes the steepness of a dose-response curve and should be noted alongside any reported IC50 ?
    Back
    The Hill coefficient
  88. Front
    Which liner material is the chapter's example of a WIDE (high) therapeutic index ?
    Back
    Calcium hydroxide (the tertiary-dentin-stimulating concentration is well below the necrosis-causing concentration)
  89. Front
    Which resin monomer is the chapter's example of a NARROW therapeutic margin ?
    Back
    TEGDMA (eluted concentrations from a poorly cured composite can approach its IC50 for pulp cells)
  90. Front
    Why is dental amalgam clinically acceptable despite a real cytotoxic mechanism (in therapeutic-index terms)?
    Back
    It has a WIDE therapeutic index - clinically achieved pulpal mercury stays well below the cytotoxic (IC50) concentration
  91. Front
    In NECROSIS , what two terms describe the random nuclear changes?
    Back
    Karyolysis and karyorrhexis
  92. Front
    Contrast the nuclear changes of necrosis vs apoptosis .
    Back
    Necrosis = random karyolysis/karyorrhexis ; apoptosis = orderly chromatin condensation and fragmentation
  93. Front
    At roughly what concentration does TEGDMA begin triggering cytochrome c release and caspase-3 activation in pulp fibroblasts?
    Back
    As low as ~0.1 mM
  94. Front
    At roughly what TEGDMA concentration does the death mode switch to rapid NECROSIS ?
    Back
    ~1 mM
  95. Front
    What is the typical IC50 range for TEGDMA in pulp-derived cell lines?
    Back
    ~0.5-1.5 mM (depending on exposure time)
  96. Front
    BisGMA 's IC50 in cell culture is often below what value?
    Back
    Below 0.1 mM
  97. Front
    In what concentration range does HEMA reach pulpal cells in deep preparations?
    Back
    ~0.5-2 mM
  98. Front
    Which monomer has the LOWEST IC50 (most potent per molecule): HEMA , TEGDMA , or BisGMA ?
    Back
    BisGMA (IC50 often <0.1 mM)
  99. Front
    On what numeric scale is the zone of cytotoxicity graded in the ISO agar overlay method?
    Back
    A 0-4 scale (analogous to an antibiotic disk assay)
  100. Front
    A material is cytotoxic only in DIRECT contact but not in its 24/72-hour eluate. Is it generally acceptable for clinical use?
    Back
    Yes - generally acceptable provided there is an adequate dentin barrier (the eluate, not direct contact, models clinical exposure)
  101. Front
    Why does ISO testing use a material's ELUATE rather than extracting all residual monomer at once?
    Back
    The eluate (standardized surface-area/volume for 24-72 h) simulates long-term, clinically realistic elution rather than acute total extraction
  102. Front
    Name three specific DAMPs released by a necrotic cell that activate innate immunity.
    Back
    HMGB1, ATP, and heat shock proteins
  103. Front
    In apoptosis , where does the condensing chromatin migrate within the nucleus?
    Back
    To the nuclear periphery (peripheral chromatin condensation)
  104. Front
    Which phagocyte-displayed signal on the apoptotic cell is recognized so apoptotic bodies are engulfed before they leak?
    Back
    Externalized phosphatidylserine (flipped to the outer membrane leaflet)
  105. Front
    By what mechanism do resin monomers initially damage the mitochondrial outer membrane to start intrinsic apoptosis ?
    Back
    They raise intracellular reactive oxygen species (ROS) , which damage the mitochondrial outer membrane
  106. Front
    Which original- MTT -assay mitochondrial enzymes reduce the tetrazolium salt to formazan ?
    Back
    Mitochondrial succinic dehydrogenase and NADH-dependent reductases
  107. Front
    In the LDH release assay, the leaked LDH oxidizes lactate to pyruvate , reducing NAD+ to _____, which then reduces a tetrazolium salt .
    Back
    NADH
  108. Front
    Why does neutral red accumulate and become trapped inside lysosomes ?
    Back
    It is a weak base; the acidic lysosomal environment protonates it , and the protonated form is trapped inside
  109. Front
    Which organelle is among the first to fail in BOTH necrosis and late apoptosis (the basis of the neutral red assay)?
    Back
    The lysosome
  110. Front
    In live/dead staining, calcein-AM is a substrate for which enzyme that converts it to fluorescent calcein ?
    Back
    Esterases (present in live cells)
  111. Front
    How does HEMA 's oxidation of glutathione lead to mitochondrial cytochrome c release?
    Back
    GSH is oxidized to GSSG , ROS accumulate, lipid peroxidation opens the mitochondrial permeability transition pore , releasing cytochrome c
  112. Front
    Besides suppressing lymphocyte proliferation, what immune-relevant effect does BisGMA have via its bisphenol A backbone?
    Back
    Estrogenic activity
  113. Front
    Eugenol belongs to what class of compound?
    Back
    A phenylpropanoid
  114. Front
    Name three mechanisms of eugenol 's direct cytotoxicity at higher concentrations.
    Back
    Membrane disruption , mitochondrial uncoupling , and protein denaturation
  115. Front
    Besides reaching the pulp in lower concentration, why does BisGMA elute and diffuse slowly?
    Back
    Its high molecular weight and relatively hydrophobic character limit its elution rate and tubular diffusion
  116. Front
    Beyond being a diffusion barrier and stimulating tertiary dentin, how does a calcium hydroxide/MTA liner aid pulp healing?
    Back
    It provides an alkaline microenvironment that accelerates pulp healing
  117. Front
    For amalgam at LOW, chronic in-service mercury concentrations, what is the cellular picture (vs frank necrosis at high dose)?
    Back
    Slow oxidative stress , mitochondrial dysfunction, and apoptotic cell loss
  118. Front
    Which ISO 10993 tier (and animal model) uses subcutaneous and intrapulpal implantation in rats/monkeys?
    Back
    Tier 2 (in vivo animal studies)
  119. Front
    What three things does histological scoring evaluate in Tier 2 intrapulpal implantation studies?
    Back
    Inflammation grade , reparative dentin bridge formation, and necrosis extent
  120. Front
    Which ISO tier is required specifically for materials intended for direct pulp contact, root canal filling, or osseointegration?
    Back
    Tier 3 (usage / clinical studies)
  121. Front
    Why does fluoride from set GIC reach the pulp without being cytotoxic, while still being antibacterial?
    Back
    Its concentration reaching pulpal cells stays below cytotoxic levels , yet is enough to inhibit bacterial enolase and promote remineralization
  122. Front
    According to the chapter, what is the single MOST important variable in dental material biocompatibility?
    Back
    The dentin thickness above the pulp (remaining dentin thickness) — it determines the monomer concentration reaching pulpal cells
  123. Front
    Within one restoration, what happens to pulp cells in the DEEPEST zone, where monomer concentration is below the IC50 (deeper than the apoptosis zone)?
    Back
    Cells survive but may upregulate stress-response genes (necrosis at the margin → apoptosis deeper → survival + stress-gene upregulation deepest)

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