The elastic component of anisotropic strain dominates the observed shift in the F2g Raman mode of anelastic ceria thin films.

Raman spectroscopy is applied for non-destructive characterization of strain in crystalline thin films. The analysis makes use of the numerical value of the mode Grüneisen parameter γ, which relates the fractional change in the frequency of a Raman-active vibrational mode and the strain-induced fractional change in the unit cell volume. When in-plane, compressive biaxial strain in aliovalent doped CeO2-films is relieved by partial substrate removal, the films exhibit values of γ for the F2g vibrational mode which are ∼30% of the literature values for bulk ceramics under isostatic stress. This discrepancy has been attributed to a negative contribution from the anelastic (time-dependent) mechanical properties of aliovalent-doped ceria. Here we propose a way to "separate" anelastic and elastic contributions to the F2g mode Grüneisen parameter. Mechanically elastic yttria (Y2O3) films on Ti/SiO2/Si substrate serve as "control". The values of γ calculated from the change in frequency of the ∼375 cm-1 F2g Raman-active mode are close to the literature values for bulk yttria under isostatic stress. This work should serve to provide a protocol for characterization of selective sensitivity to different strain components of doped ceria thin films.

Conjugated detergent micelles as a platform for IgM purification.

Immunoglobulin M (IgM) antibodies hold promise as anticancer drugs and as agents for promoting immune homeostasis. This promise has not been realized due to low expression levels in mammalian cells producing IgM class antibodies, and the failure of protein A chromatography for IgM purification. Here, we describe a nonchromatographic platform for quantitatively capturing IgMs at neutral pH, which is then recovered with 86%-94% yield and >95% purity at pH 3. The platform contains micelles conjugated with the [(bathophenanthroline)3 :Fe2+ ] amphiphilic complex. Inclusion of amino acid monomers, for example, phenylalanine or tyrosine, during conjugation of detergent micelles, allows subsequent extraction of IgMs at close to neutral pH. With the successful implementation of this purification platform for both polyclonal humans and bovine IgMs, we anticipate similar results for monoclonal IgMs, most relevant for the pharmaceutical industry.

Reversible Conjugation of Non-ionic Detergent Micelles Promotes Partitioning of Membrane Proteins under Non-denaturing Conditions.

In the decades'-long quest for high-quality membrane protein (MP) crystals, non-ionic detergent micelles have primarily served as a passive shield against protein aggregation in aqueous solution and/or as a conformation stabilizing environment. We have focused on exploiting the physical chemistry of detergent micelles in order to direct intrinsic MP/detergent complexes to assemble via conjugation under ambient conditions, thereby permitting finely tuned control over the micelle cloud point. In the current work, three commercially available amphiphilic, bipyridine chelators in combination with Fe2+ or Ni2+ were tested for their ability to conjugate non-ionic detergent micelles both in the presence and absence of an encapsulated bacteriorhodopsin molecule. Water-soluble chelators were added, and results were monitored with light microscopy and dynamic light scattering (DLS). [Bipyridine:metal] complexes produced micellar conjugates, which appeared as oil-rich globules (10-200 µm) under a light microscope. DLS analysis demonstrated that micellar conjugation is complete 20 min after the introduction of the amphiphilic complex, and that the conjugation process can be fully or partially reversed with water-soluble chelators. This process of controlled conjugation/deconjugation under nondenaturing conditions provides broader flexibility in the choice of detergent for intrinsic MP purification and conformational flexibility during the crystallization procedure.

C-Axis Textured, 2-3 µm Thick Al0.75Sc0.25N Films Grown on Chemically Formed TiN/Ti Seeding Layers for MEMS Applications.

A protocol for successfully depositing [001] textured, 2−3 µm thick films of Al0.75Sc0.25N, is proposed. The procedure relies on the fact that sputtered Ti is [001]-textured α-phase (hcp). Diffusion of nitrogen ions into the α-Ti film during reactive sputtering of Al0.75,Sc0.25N likely forms a [111]-oriented TiN intermediate layer. The lattice mismatch of this very thin film with Al0.75Sc0.25N is ~3.7%, providing excellent conditions for epitaxial growth. In contrast to earlier reports, the Al0.75Sc0.25N films prepared in the current study are Al-terminated. Low growth stress (<100 MPa) allows films up to 3 µm thick to be deposited without loss of orientation or decrease in piezoelectric coefficient. An advantage of the proposed technique is that it is compatible with a variety of substrates commonly used for actuators or MEMS, as demonstrated here for both Si wafers and D263 borosilicate glass. Additionally, thicker films can potentially lead to increased piezoelectric stress/strain by supporting application of higher voltage, but without increase in the magnitude of the electric field.

Modeling Strain Distribution at the Atomic Level in Doped Ceria Films with Extended X-ray Absorption Fine Structure Spectroscopy.

Ceria doped with trivalent dopants exhibits nonclassical electrostriction, strong anelasticity, and room-temperature (RT) mechanical creep. These phenomena, unexpected for a ceramic material with a large Young's modulus, have been attributed to the generation of local strain in the vicinity of the host Ce cations due to symmetry-breaking point defects, including oxygen vacancies. However, understanding why strain is generated at the host rather than at the dopant site, as well as predicting these effects as a function of dopant size and concentration, remains a challenge. We have used the evolutionary-algorithm-based reverse Monte Carlo modeling to reconcile the experimental data of extended X-ray absorption fine structure and X-ray diffraction in a combined model structure. By extracting the details of the radial distribution function (RDF) around the host (Ce) and trivalent dopants (Sm or Y), we find that RDF of the first-nearest neighbor (1NN) of host and dopant cations as well as the second-nearest neighbor (2NN) of the dopant are each best modeled with two separate populations corresponding to short and long interatomic distances. This heterogeneity indicates that fluorite symmetry is not preserved locally, especially for the dopant first-and second-NN sites, appearing at surprisingly low doping fractions (5 mol % Sm and 10 mol % Y). Given that Ce rather than dopant sites act as the source of local strain for electrostriction and RT creep, we conclude that the environment around the dopant does not respond to electrical and mechanical excitations, likely because of its similarity to the double fluorite structure which has poor electrostrictive and anelastic properties. The trends we observe in the RDFs around the Ce sites as a function of dopant size and concentration suggest that the response of these sites can be controlled by the extent of doping: Increasing dopant size to increase strain magnitude at the 1NN shell of Ce and decreasing dopant fraction to decrease strain propagation to the 2NN shell of Ce should produce stronger electrostrictive response and RT creep.

Controlled micelle conjugation via charged peptide amphiphiles.

We report the first demonstration of nonionic detergent micelle conjugation and phase separation using purpose-synthesized, peptide amphiphiles, C10 -(Asp)5 and C10 -(Lys)5 . Clustering is achieved in two different ways. Micelles containing the negatively charged peptide amphiphile C10 -(Asp)5 are conjugated (a) via a water-soluble, penta-Lys mediator or (b) to micelles containing the C10 -(Lys)5 peptide amphiphile. Both routes lead to phase separation in the form of oil-rich globules visible in the light microscope. The hydrophobic nature of these regions leads to spontaneous partitioning of hydrophobic dyes into globules that were found to be stable for weeks to months. Extension of the conjugation mechanism to micelles containing a recently discovered, light-driven proton pump King Sejong 1-2 (KS1-2) demonstrates that a membrane protein may be concentrated using peptide amphiphiles while preserving its native conformation as determined by characteristic UV absorption. The potential utility of these peptide amphiphiles for biophysical and biomedical applications is discussed.

Van Vleck paramagnetism in undoped and Lu-doped bulk ceria.

The magnetic properties of undoped, bulk CeO2 are not fully understood. In contrast to nanocrystalline ceria that exhibits paramagnetism attributed to Ce3+ at grain surfaces, bulk ceria is weakly paramagnetic, despite the absence of magnetic ions. In the present work, the magnetic susceptibility of bulk ceria ceramics doped with Lu3+, which has neither spin nor orbital angular momentum, was measured in order to assess the relative contributions of the crystal lattice, residual Ce3+ and oxygen vacancies to the overall bulk magnetization. We observed a magnetic response consisting of two parts: temperature independent (5-300 K) magnetic susceptibility, and Curie-Weiss paramagnetism. The temperature independent susceptibility decreases linearly with Lu content, and becomes diamagnetic at 30 mol% Lu. The Curie-Weiss magnetism visible at low temperatures was identified as resulting from a few ppm of Fe contaminant. However, Fe contamination does not contribute to the temperature independent paramagnetism. No contribution from Ce3+ could be detected. The fact that the magnetization decreases with Lu content, even though the concentration of oxygen vacancies, and the lattice defects associated with them, increases, indicates that neither is coupled to the magnetic field. Weak, temperature-independent paramagnetism in non-metals is usually attributed to a second order, Van Vleck-type magnetization. However, Van Vleck paramagnetism requires that the population of the first excited state be constant within the range of temperatures investigated. We discuss possible modifications of the large band gap electronic structure of undoped ceria which could account for our observations.

Anomalous viscosity-time behavior of polysaccharide dispersions.

Using viscosity and dynamic light scattering (DLS) measurements, we monitored the changes in the properties of dispersions of chitosan (a cationic polysaccharide) in acidic solution over a period of up to 700 h. Different polymer concentrations, weight average molecular weights, and degrees of deacetylation were examined. We found that the solution rheology and chitosan aggregates continue to change even up to 700 h. It was observed, remarkably, using both capillary and cone and plate viscometry that the viscosity decreased significantly during the storage period of the chitosan dispersions, with a rapid initial decrease and a slow approach to the steady state value. DLS measurements over this period could be interpreted in terms of a gradual decrease in the size of the chitosan aggregates in the dispersion. This behavior is puzzling, insofar as one expects the dissolution of compact polymer aggregates with time into individual polymer chains to increase the viscosity rather than decrease it as observed: We attribute this apparently anomalous behavior to the fact that the chitosan aggregates are rigid crystalline rod-like entities, which dissolved with time from dispersion of overlapping rods (with high viscosity) into solution of individual random coils (with lower viscosity). A detailed model comparing the hydrodynamic behavior of the initial overlapping rod-like aggregates with the subsequent free coils in solution is in semi-quantitative agreement with our observation.

Membrane protein crystallization in micelles conjugated by nucleoside base-pairing: A different concept.

The dearth of high quality, three dimensional crystals of membrane proteins, suitable for X-ray diffraction analysis, constitutes a serious barrier to progress in structural biology. To address this challenge, we have developed a new crystallization medium that relies on the conjugation of surfactant micelles via base-pairing of complementary hydrophobic nucleosides. Base-pairs formed at the interface between micelles bring them into proximity with each other; and when the conjugated micelles contain a membrane protein, crystal nucleation centers can be stabilized, thereby promoting crystal growth. Accordingly, two hydrophobic nucleoside derivatives - deoxyguanosine (G) and deoxycytidine (C), each covalently bonded to a 10 carbon chain were synthesized and added to an aqueous solution containing octyl ß-d-thioglucopyranoside micelles. These hydrophobic nucleosides induced the formation of oil-rich globules after 2days incubation at 19°C or after a few hours in the presence of ammonium sulfate; however, phase separation was inhibited by 100mM GMP. The presence of the membrane protein bacteriorhodopsin in the conjugated - micellar dispersion resulted in the growth within the colorless globules of a variety of purple crystals, the color indicating a functional protein. On this basis, we suggest that conjugation of micelles via base-pair complementarity may provide significant assistance to the structural determination of integral membrane proteins.

Structure, function, aging and turnover of aggrecan in the intervertebral disc.

BACKGROUND: Aggrecan is the major non-collagenous component of the intervertebral disc. It is a large proteoglycan possessing numerous glycosaminoglycan chains and the ability to form aggregates in association with hyaluronan. Its abundance and unique molecular features provide the disc with its osmotic properties and ability to withstand compressive loads. Degradation and loss of aggrecan result in impairment of disc function and the onset of degeneration. SCOPE OF REVIEW: This review summarizes current knowledge concerning the structure and function of aggrecan in the normal intervertebral disc and how and why these change in aging and degenerative disc disease. It also outlines how supplementation with aggrecan or a biomimetic may be of therapeutic value in treating the degenerate disc. MAJOR CONCLUSIONS: Aggrecan abundance reaches a plateau in the early twenties, declining thereafter due to proteolysis, mainly by matrix metalloproteinases and aggrecanases, though degradation of hyaluronan and non-enzymic glycation may also participate. Aggrecan loss is an early event in disc degeneration, although it is a lengthy process as degradation products may accumulate in the disc for decades. The low turnover rate of the remaining aggrecan is an additional contributing factor, preventing protein renewal. It may be possible to retard the degenerative process by restoring the aggrecan content of the disc, or by supplementing with a bioimimetic possessing similar osmotic properties. GENERAL SIGNIFICANCE: This review provides a basis for scientists and clinicians to understand and appreciate the central role of aggrecan in the function, degeneration and repair of the intervertebral disc.

Comment on "Cholesterol solubility limit in lipid membranes probed by small angle neutron scattering and MD simulations" by S. Garg et al., Soft Matter, 2014, 10, 9313.

In a recent article, Garg et al. used neutron scattering techniques to determine the limiting amount of cholesterol which vesicles of either POPS or POPC can accommodate. This amount was called "the cholesterol solubility limit". In light of extensive literature on cholesterol phase separation in phospholipid bilayers, the way in which "solubility limit" is defined in this article and the conclusions derived are misleading and require some clarification.

Engineered-membranes and engineered-micelles as efficient tools for purification of halorhodopsin and bacteriorhodopsin.

We describe two alternative and complementary purification methods for halorhodopsin and bacteriorhodopsin. The first relies on a unique form of detergent micelles which we have called engineered-micelles. These are specifically conjugated in the presence of [hydrophobic chelator:Fe(2+)] complexes and form detergent aggregates into which membrane proteins partition, but hydrophilic water-soluble proteins do not. The approach was tested on the membrane protein, bacteriorhodopsin (bR), with five non-ionic detergents (OG, OTG, NG, DM, and DDM), commonly used in purification and crystallization of membrane proteins, in combination with the commercially available bathophenanthroline or with one of the three synthesized phenanthroline derivatives (Phen-C10, Phen-C8 and Phen-C6). Our results show that bR is extracted efficiently (60-86%) and directly from its native membrane into diverse detergent aggregates with preservation of its native conformation, while 90-95% of an artificial contaminating background is excluded. For implementation of the second method, based on engineered-membranes, the use of detergents, which in some cases may produce protein denaturation, is not required at all. Protein-containing membranes are conjugated via the same hydrophobic [chelator:metal ion] complexes but maintain the membrane protein in its native bilayer environment throughout the process. This method is demonstrated on the membrane protein halorhodopsin from Natronomonas pharaonis (phR) and leads to good recovery yields (74-89%) and removal of >85% of artificial background impurities while preserving the native state of phR. The detailed structure of the hydrophobic chelator used has been found to have a marked effect on the purity and yield of both methods.

Cryo-TEM structural analysis of conjugated nonionic engineered-micelles.

Conjugated engineered-micelles, i.e. micelles that are composed of nonionic detergents and hydrophobic chelators and subsequently conjugated in the presence of divalent metal ions, have been shown to be remarkably suited to the task of membrane protein purification, maintaining these proteins in their native state. They also efficiently solubilize highly hydrophobic antibiotics. To date, however, the morphological changes induced in the initially spherical or ellipsoidal micelles by conjugation have not been explored. In this study, the very rapid sample-vitrification protocol of cryogenic transmission electron microscopy (cryo-TEM) has been used to capture structural transformations that engineered-micelles undergo immediately following conjugation with the [(bathophenanthroline)3:Fe(2+)] hydrophobic complex. We found that condensed thread-like aggregates are formed when the detergents used are: octyl ß-D-glucopyranoside (OG), octyl ß-D-thioglucopyranoside (OTG) or pentaethylene glycol monododecyl ether (C12E5). However, with ß-D-maltoside (DM), n-dodecyl ß-D-maltoside (DDM) or ß-D-glucopyranoside (DDG), lamellar structures, some of which appear as stacked lamellae or multilamellar vesicles (MLV's), were observed. Such architectural changes occur under very mild conditions i.e. low detergent concentration, no temperature or pH alterations and without the presence of any precipitants such as PEG or ammonium sulfate.

Blue-violet photoluminescence of 4-isopropyl-pyridine hydroxide crystals.

There is continuing interest in determining essential structural features of polymer gels, which display photoelectric and/or thermoelectric behavior. One such gel is the blend, poly(4-vinylpyridine-co-butyl methacrylate)/poly(4-vinylpyridine), dissolved in liquid pyridine. Following extended aeration of a three-component mixture, which serves as a model for the gel side chain interactions, crystallization of a new molecule, 4-isopropylpyridine hydroxide (IPPOH), occurs. X-ray diffraction, DFT modeling, and spectroscopy were used to determine the structural, electronic, and luminescent properties of the crystal. The crystal structure reveals molecules forming head-to-tail, hydrogen-bonded chains without base stacking or marked interchain interaction. The molecular chains are characterized by moderately long-lived, blue-violet luminescence excited in the near-UV. Because these photoluminescent properties resemble those of the gel from which the crystals are derived, we may posit similar structural features in the gel for which direct structural analysis is not available.

Biochemical composition and turnover of the extracellular matrix of the normal and degenerate intervertebral disc.

BACKGROUND: The intervertebral disc (IVD) is a complex cartilaginous structure which functions to resist biomechanical loads during spinal movement. It consists of the highly viscous cartilaginous nucleus pulposus, which is surrounded laterally by a thick outer ring of fibrous cartilage-the annulus fibrosus-and sandwiched inferiorly and superiorly by the cartilage end-plates. The main extracellular matrix molecules of the disc are collagens, proteoglycans, glycoproteins and elastin. The disc also contains appreciable amounts of water, matrix-degrading protease enzymes and their inhibitors, soluble signalling molecules and various metabolic breakdown products. METHODS: This review provides a comprehensive description of the biochemical composition of the extracellular matrix of the IVD and, specifically, the proteases involved in its molecular turnover. Quantitation of the turnover rates using racemization of aspartic acid as a molecular clock is also discussed. CONCLUSIONS: Molecular turnover rates of the major constituent matrix macromolecules of the IVD are found to be particularly slow, especially in the case of collagen. Over a normal human life span, this slow turnover may compromise the structural integrity of the IVD extracellular matrix essential for normal physiological functioning.

Advances in the diagnosis of degenerated lumbar discs and their possible clinical application.

PURPOSE: One possible source of chronic low back pain is a degenerated intervertebral disc. In this review, various diagnostic methods for the assessment of the presence of degenerative changes are described. These include clinical MRI, a number of novel MRI techniques and nuclear magnetic resonance spectroscopy. METHODS: Non-systematic literature review. RESULTS: Clinical MRI is the most commonly employed technique to determine the general "health status" of the intervertebral disc. Novel MRI techniques, such as quantitative MRI, T1ρ MRI, sodium MRI and nuclear magnetic resonance spectroscopy, are more sensitive in quantifying the biochemical changes of disc degeneration, as measured by alteration in collagen structure, as well as water and proteoglycan loss. As potential future diagnostic alternatives, miniature sensors are currently being developed to measure parameters associated with the disc degeneration cascade, such as intradiscal pressure and PG concentration. However, none of the methods listed above show sufficient specificity to identify a degenerated disc as the actual source of the pain. Provocative discography is the only test aimed at a direct diagnosis of discogenic pain, but it has a high false positive rate and there is some evidence of long-term adverse effects. Imaging techniques have also been tested for this purpose, but their validity has not been confirmed and they do appear to be problematic. CONCLUSIONS: A reliable diagnostic tool that could help a clinician to determine if a disc is the source of the pain in patients with chronic LBP is still not available. New MRI techniques are under investigation that could result in a significant improvement over current methods, particularly as they can allow monitoring, not only of morphological but also of biochemical changes.

Conjugated Nonionic Detergent Micelles: An Efficient Purification Platform for Dimeric Human Immunoglobulin A.

The SARS-COV-2 virus is a deadly agent of inflammatory respiratory disease. Since 2020, studies have focused on developing new therapies based on galactose-rich IgA antibodies. Clinical surveys have also revealed that galactose-deficient IgA1 polymerizes in serum, producing IgA nephropathy, which is a common cause of kidney failure in young adults. Here we show that IgA1-IgA2 dimers are efficiently and economically purified in solution via conjugated nonionic surfactant micellar aggregates. Quantitative capture at pH 7 and extraction at pH 6.5 can avoid antibody exposure to acidic, potentially denaturing conditions. Brij-O20 aggregates lead to the highest process yields (88-91%) and purity (94%). Recovered IgA dimers preserve their native secondary structure and do not self-associate. Increasing the reaction volume has little impact on yield or purity. By introducing an efficient, inexpensive IgA purification protocol, we assist pharmaceutical firms and research laboratories in developing new IgA-based therapies as well as in increasing our understanding of IgA1 polymerization.

The [(bathophenanthroline)3:Fe2+] complex as an aromatic non-polymeric medium for purification of human lactoferrin.

We describe a non-chromatographic, ligand-free platform for the efficient purification of recombinant human lactoferrin (LF). The platform consists of a [metal:chelator] complex precipitate in the presence of osmotically active polyethylene glycol 6000 (PEG-6000). Purification is achieved in three stages. Following formation of the complex, LF is captured under neutral conditions by the aggregated complexes (Step I), a washing step follows (Step II) and then, (Step III) LF is extracted in pure form with 100 mM tribasic Na citrate buffer (pH 7). Of the four complexes investigated, [bathophenanthroline (batho)3:Fe2+] was determined to be the most efficient. LF is recovered with high yield (∼90%, by densitometry) and purity (≥97%, by SDS polyacrylamide gel electrophoresis (SDS-PAGE)) from an artificial contamination background comprising E. coli lysate proteins. Purified LF is demonstrated to be monomeric by dynamic light scattering (DLS); to preserve its native secondary structure by circular dichroism (CD) spectroscopy; and, as apo-LF, to efficiently inhibit bacterial growth. Process yield is not affected by a 45-fold increase in LF concentration from 0.2 to 9 mg/mL. We provide evidence that protein capture relies on [cation:π] interactions between the lysine and arginine residues of LF with the fully aromatic [(batho)3:Fe2+] complexes. The use of [metal:chelator] complex aggregates is demonstrated to provide an economical and efficient avenue for LF purification.

Local Environment of Sc and Y Dopant Ions in Aluminum Nitride Thin Films.

The local environments of Sc and Y in predominantly ⟨002⟩ textured, Al1-xDoxN (Do = Sc, x = 0.25, 0.30 or Y, x = 0.25) sputtered thin films with wurtzite symmetry were investigated using X-ray absorption (XAS) and photoelectron (XPS) spectroscopies. We present evidence from the X-ray absorption fine structure (XAFS) spectra that, when x = 0.25, both Sc3+ and Y3+ ions are able to substitute for Al3+, thereby acquiring four tetrahedrally coordinated nitrogen ligands, i.e., coordination number (CN) of 4. On this basis, the crystal radius of the dopant species in the wurtzite lattice, not available heretofore, could be calculated. By modeling the scandium local environment, extended XAFS (EXAFS) analysis suggests that when x increases from 0.25 to 0.30, CN for a fraction of the Sc ions increases from 4 to 6, signaling octahedral coordination. This change occurs at a dopant concentration significantly lower than the reported maximum concentration of Sc (42 mol % Sc) in wurtzite (Al, Sc)N. XPS spectra provide support for our observation that the local environment of Sc in (Al, Sc)N may include more than one type of coordination.

Longevity of elastin in human intervertebral disc as probed by the racemization of aspartic acid.

BACKGROUND: Aging and degeneration of human intervertebral disc (IVD) are associated with biochemical changes, including racemization and glycation. These changes can only be counteracted by protein turnover. Little is known about the longevity of IVD elastin in health or disease. Yet, such knowledge is important for a quantitative understanding of tissue synthesis and degradation. METHODS: We have measured the accumulation of d-Asp and pentosidine in IVD elastin. Samples representing a broad range of ages (28-82years) and degeneration grades (1-5) were analyzed. RESULTS: d/l-Asp for elastin increased linearly with age from 3.2% (early 30s) to 14.8% (early 80s) for normal tissue (grades 1-2) and from 1.7% (late 20s) to 6.0% (until the mid 50s) for degenerate tissue (grades 3-5) with accumulation rates of 16.2±3.1×10(-4) and 11.7±3.8×10(-4)year(-1), respectively; no significant difference was found between these values (p<0.05). Above the mid 50s, a decrease in d-Asp accumulation was recorded in the degenerate tissue. d-Asp accumulation correlated well with pentosidine content for elastin from healthy and degenerate tissues combined. We conclude that IVD elastin is metabolically-stable and long-lived in both healthy and degenerate human IVDs, with signs of new synthesis in the latter. The correlation of d-Asp with pentosidine content suggests that both these agents may be used as markers in the overall aging process of IVD. GENERAL SIGNIFICANCE: Accumulation of modified IVD elastin argues for its longevity and may have a negative impact on its role in disc function. Weak signs of newly synthesized molecules may act to counteract this effect in degenerate tissue.