Rapid, continuous additive manufacturing by volumetric polymerization inhibition patterning.

Contemporary, layer-wise additive manufacturing approaches afford sluggish object fabrication rates and often yield parts with ridged surfaces; in contrast, continuous stereolithographic printing overcomes the layer-wise operation of conventional devices, greatly increasing achievable print speeds and generating objects with smooth surfaces. We demonstrate a novel method for rapid and continuous stereolithographic additive manufacturing by using two-color irradiation of (meth)acrylate resin formulations containing complementary photoinitiator and photoinhibitor species. In this approach, photopatterned polymerization inhibition volumes generated by irradiation at one wavelength spatially confine the region photopolymerized by a second concurrent irradiation wavelength. Moreover, the inhibition volumes created using this method enable localized control of the polymerized region thickness to effect single-exposure, topographical patterning.

Extracellular matrix regulation of fibroblast function: redefining our perspective on skin aging.

The dermal extracellular matrix (ECM) comprises the bulk of skin and confers strength and resiliency. In young skin, fibroblasts produce and adhere to the dermal ECM, which is composed primarily of type I collagen fibrils. Adherence allows fibroblasts to spread and exert mechanical force on the surrounding ECM. In this state, fibroblasts display a "youthful" phenotype characterized by maintenance of the composition and structural organization of the dermal ECM. During aging, fibroblast-ECM interactions become disrupted due to fragmentation of collagen fibrils. This disruption causes loss of fibroblast spreading and mechanical force, which inextricably lead to an "aged" phenotype; fibroblasts synthesize less ECM proteins and more matrix-degrading metalloproteinases. This imbalance of ECM homeostasis further drives collagen fibril fragmentation in a self-perpetuating cycle. This article summarizes age-related changes in the dermal ECM and the mechanisms by which these changes alter the interplay between fibroblasts and their extracellular matrix microenvironment that drive the aging process in human skin.

Alopecia and platelet-derived therapies.

BACKGROUND: Platelet rich plasma (PRP) injections have emerged as a promising regenerative therapy for androgenetic alopecia (AGA). To date, injections of both autologous native and activated PRP have been administered to hair loss patients, and positive results have been observed. However, little to no work has yet to be seen wherein PRP treatments are combined with hair restoration surgeries. Furthermore, the PRP activation protocol in the hair restoration setting employs compounds with potentially deleterious side effects, namely thrombin or calcium gluconate. Therefore, the objectives of this work are to evaluate the effectiveness of platelet and platelet-derived products as augmented graft therapies in hair restoration surgeries and to compare the follicular regeneration rate of follicles transplanted in the presence of platelet lysate (PL) versus activated PRP (AA-PRP). METHODS: PL was administered to the frontal scalp of three male AGA patients. Three treatment zones measuring 4 cm-2 were mapped in the midline scalp region of each patient and equal number of follicular grafts were placed in each box along with PL, AA-PRP, or normal saline. The transplanted follicular grafts of a fourth patient were placed solely with PL. Hair checks in which the surface area of hair coverage was quantified were performed at follow-up appointments ranging from 3.5 to 7 months post-surgery. In these appointments, the number of follicular units with hairs measuring 50 mm or more were counted to determine the percentage of graft hair regeneration. Growth factor (GF) concentrations [vascular endothelial growth factor (VEGF), transforming growth factor beta 1 (TGF-ß1), PDGF-BB, IGF-1] in PL and AA-PRP were also measured for an independent subject set. RESULTS: Follicular regeneration in transplanted grafts was found to be superior for those placed with PL rather than AA-PRP or saline at all follow-up dates. Specifically, at 3.5 months post-op, 89%±9%, 74%±7%, and 57%±10% of follicular units had regenerated hair in the PL, AA-PRP, and saline treatment zones, respectively. At 4 months post-op, 99%, 75%, and 71% of follicle regeneration had occurred in the PL, AA-PRP, and saline treatment areas, respectively. Impressively, when PL was injected alone, the patient experienced a 50% increase in follicular unit density and a 122% increase in hair density 7 months post-injection. When GF concentrations were measured, PL generated from a 30-min sonication of PRP was found to have significantly higher levels of VEGF, PDGF-BB, and TGF-ß1 than AA-PRP. CONCLUSIONS: PRP remains a promising hair loss therapy and should be evaluated further for use not only as an independent therapeutic tool, but also as a treatment to augment surgical procedures. PL in particular affords an effective and efficacious therapeutic product given that the lysate may be obtained by mechanical rather than chemical means. Ultrasonic waves provide sufficient energy to rupture platelet cell walls, and centrifugation may be used to separate the lysate from cell fragments prior to delivery.

Evaluation of Not-Activated and Activated PRP in Hair Loss Treatment: Role of Growth Factor and Cytokine Concentrations Obtained by Different Collection Systems.

Platelet rich plasma (PRP) was tested as a potential therapy for androgenetic alopecia (AGA) through two different clinical protocols in which one population (18 participants) received half-head treatment with autologous non-activated PRP (A-PRP) produced by CPunT Preparation System (Biomed Device, Modena, Italy) and the other half-head with placebo, and a second separated population in which all participants (n = 6, 3 participants per group) received treatment with calcium-activated PRP (AA-PRP) produced from one of two different PRP collection devices (Regen Blood Cell Therapy or Arthrex Angel System). For the A-PRP study, three treatments were administered over 30-day intervals. Trichoscan analysis of patients, three months post-treatment, showed a clinical improvement in the number of hairs in the target area (36 ± 3 hairs) and in total hair density (65±  5 hair cm2), whereas negligible improvements in hair count (1.1±  1.4 hairs) and density (1.9 ± 10.2 hair cm2) were seen in the region of the scalp that received placebo. Microscopic evaluation conducted two weeks after treatment showed also an increase in epidermal thickness, Ki67+ keratinocytes, and in the number of follicles. The AA-PRP treatment groups received a singular set of injections, and six months after the treatments were administered, notable differences in clinical outcomes were obtained from the two PRP collection devices (+90 ± 6 hair cm2 versus -73 ± 30 hair cm2 hair densities, Regen versus Arthrex). Growth factor concentrations in AA-PRP prepared from the two collection devices did not differ significantly upon calcium activation.

Bactericidal Hydrogels via Surface Functionalization with Cecropin A.

The immobilization of antimicrobial peptides (AMPs) to surfaces, enabling their utilization in biosensor and antibacterial/antifouling coating applications, is typically performed using rigid, solid support materials such as glass or gold and may require lengthy, temperamental protocols. Here, we employ a hydrogel immobilization platform to afford facile fabrication and surface functionalization while offering improved biocompatibility for evaluating the influence of linker length, surface density, and AMP conjugation site on retained peptide activity. Rapid, interfacial photo-polymerization using the radical-mediated thiol-ene addition mechanism was used to generate cross-linked, polymeric coatings bearing residual thiol moieties on prefabricated poly(ethylene glycol) (PEG)-based hydrogel supports. The photo-polymerized coatings were 60 µm thick and contained 0.55 nmol of unreacted free thiols, corresponding to a concentration of 410 µM, for use as cecropin A (CPA) immobilization handles via thiol-maleimide conjugation, where the CPA-bound maleimide moiety was localized at either the carboxyl terminus or midsequence between Ala22 and Gly23. Surface presentation of the thiol handles was controlled by varying the thiolated PEG monomer (PEGSH) used in the photo-polymerizable formulation. Bactericidal activity of CPA functionalized hydrogels against E. coli K235 indicated that CPA immobilized at the carboxyl terminus killed 94 ± 6% of the inoculated pathogens when coatings were prepared with high molecular weight PEGSH and 99 ± 1% when prepared with low molecular weight PEGSH. E. coli cell death demonstrated a stronger dependence on peptide concentration than PEG linker length or degree of thiol functionalization, with activity ranging from 34 ± 13% to 99 ± 1% bacterial cells killed as the prefunctionalization thiol concentration in the coatings was increased from 90 to 990 µM. Finally, the immobilization site on the surface-bound CPA strongly affected antibacterial activity; when midsequence modified CPA was bound to a hydrogel coating bearing 990 µM thiol, only 20 ± 4% of the E. coli population was killed.

Thiol-ene functionalized siloxanes for use as elastomeric dental impression materials.

OBJECTIVES: Thiol- and allyl-functionalized siloxane oligomers are synthesized and evaluated for use as a radical-mediated, rapid set elastomeric dental impression material. Thiol-ene siloxane formulations are crosslinked using a redox-initiated polymerization scheme, and the mechanical properties of the thiol-ene network are manipulated through the incorporation of varying degrees of plasticizer and kaolin filler. Formulations with medium and light body consistencies are further evaluated for their ability to accurately replicate features on both the gross and microscopic levels. We hypothesize that thiol-ene functionalized siloxane systems will exhibit faster setting times and greater detail reproduction than commercially available polyvinylsiloxane (PVS) materials of comparable consistencies. METHODS: Thiol-ene functionalized siloxane mixtures formulated with varying levels of redox initiators, plasticizer, and kaolin filler are made and evaluated for their polymerization speed (FTIR), consistency (ISO4823.9.2), and surface energy (goniometer). Feature replication is evaluated quantitatively by SEM. The Tg, storage modulus, and creep behavior are determined by DMA. RESULTS: Increasing redox initiation rate increases the polymerization rate but at high levels also limits working time. Combining 0.86 wt% oxidizing agent with up to 5 wt% plasticizer gave a working time of 3 min and a setting time of 2 min. The selected medium and light body thiol-ene formulations also achieved greater qualitative detail reproduction than the commercial material and reproduced micrometer patterns with 98% accuracy. SIGNIFICANCE: Improving detail reproduction and setting speed is a primary focus of dental impression material design and synthesis. Radical-mediated polymerizations, particularly thiol-ene reactions, are recognized for their speed, reduced shrinkage, and 'click' nature.

Redox Initiation of Bulk Thiol-Ene Polymerizations.

The unique formation-structure-property attributes and reaction behavior of the thiol-ene "click" reaction have been explored extensively for photochemically and thermally initiated reactions but have been much less explored for redox initiation. Therefore, the objective of this work is to characterize fully the impact of the initiation system, monomer structure, degree of functionalization, and inhibitor level on the redox-mediated thiol-ene polymerization rate and behavior. Moreover, this study confirms the ability of redox initiation to achieve full conversion of desired thiol-ene "click" products for small molecules in solution. For the multifunctional thiol-ene systems, polymerization rate was shown to be comparable to photo- and thermally initiated systems, but with the additional advantages of unlimited depth of cure and mild reaction conditions. Additionally, the network properties of the redox-initiated thiol-ene systems were on par with a photocured material formulated with identical monomers and radical initiating potential. Lastly, control over the polymerization rate and preceding induction period was garnered from the concentration of inhibitor included in the reaction mixture. The mechanism of action of quinone inhibition in redox-mediated thiol-ene polymerizations is shown to depend on both the presence of an aniline reducing agent and the concentration of inhibitor, with quinone concentrations in great excess of oxidizing agent concentrations actually leading to heightened polymerization rates when aniline is present.

Synthesis and Characterization of Thiol-Ene Functionalized Siloxanes and Evaluation of their Crosslinked Network Properties.

Three types of linear thiol-functionalized siloxane oligomers and three types of ene-functionalized oligomers were synthesized and subsequently photopolymerized. Within each type of thiol-functionalized oligomer, the ratio of mercaptan repeat units to non-reactive phenyl repeat units was varied to manipulate both the crosslink density and the degree of secondary interactions through pi-pi stacking. Similarly, the repeat units of the three ene-functionalized oligomers are composed of allyl-functional monomers, benzene-functional monomers, and octyl-functional monomers in varying ratios of benzene:octyl but with a constant fraction of allyl moieties. The structural composition of the siloxane oligomers plays a pivotal role in the observed material properties of networks formed through thiol-ene photopolymerization. Networks with a high concentration of thiol functionalities exhibit higher rubbery moduli, ultimate strengths, and Young's moduli than networks with lower thiol concentrations. Moreover, the concentration of functionalities capable of participating in secondary interactions via hydrogen bonding or pi-pi stacking directly impacts the network glass transition temperature and elasticity. The combination of low crosslink density and high secondary interactions produces networks with the greatest toughness. Finally, the fraction of octyl repeats correlates with the hydrophobic nature of the network.

Comparison of manufacturer-recommended exposure durations with those determined using biaxial flexure strength and scraped composite thickness among a variety of light-curing units.

STATEMENT OF THE PROBLEM: Manufacturer-recommended exposure durations for light-curing units are often understated and might not have true clinical relevance. PURPOSE: To compare composite depths of cure among exposure durations provided by the manufacturer and those obtained when optimizing exposure duration for biaxial flexural strength or for composite compule-scraping tests when using different light-curing units. METHODS/MATERIALS: A hybrid composite (Prodigy, A3, Kerr, Orange, CA, USA) was exposed to different light-curing units (all manufactured by Kerr Demetron) (conventional quartz-tungsten-halogen [QTH], conventional blue light-emitting diode [LED(CONV)] or a high-intensity blue LED light [LED(HIGH)]) for various amounts of time, including that recommended by the manufacturer for the given light. A test model was designed in which 0.5-mm thick composite discs were stacked between Mylar sheets to a total composite thickness of 3.0 mm. The top of each stack was exposed to the different lights for a variety of exposures at a 2-mm distance. Twenty-four hours later, the stacks were disassembled, and the individual discs from each 0.5-mm thick increment were tested for biaxial flexure strength. Ten discs were made for each exposure duration from each light. Statistical analysis (analysis of variance, Dunnett-Hsu post hoc test, alpha = 0.05) was used to identify the exposure duration needed for the flexural strength at a 2.5-mm depth (manufacturer-recommended thickness) to be similar to that at the topmost 0.5-mm thick increment. Compules of the same composite were modified to form cylinders in which their contents were forced to one end and photopolymerized (at a 2-mm distance) for a variety of exposure durations using the same light units mentioned above (N = 5). Twenty-four hours later, compule contents were extruded, and the unpolymerized residue was removed using hand scraping with a plastic spatula. The thickness of the resulting specimen was measured, and was plotted as a function of exposure duration for each light. Regression analysis was applied to generate the mathematical correlation between exposure duration and resulting composite scraped thickness. Manual line-drawing methods were used on that generated plot to determine the major inflection in the exposure-thickness relationship that changed, and the exposure time correlated to that inflection point was considered the optimal exposure duration from this method. RESULTS: Manufacturer-recommended exposures for a 2.5-mm thick composite increment from the lights used were: QTH 20 seconds; LED(CONV) 10 seconds; and LED(HIGH) 5 seconds. Flexural strength and scraped composite compule thickness values markedly changed with increase in exposure duration and differed among the lights. Exposure durations needed to provide similar flexural strength at 2.5 mm as that of the topmost increment were: QTH 30 seconds; LED(CONV) 15 seconds; and LED(HIGH) 20 seconds. Exposure durations derived from inflection points of the scraping plots provided optimal exposure duration values of: QTH 25 seconds; LED(CONV) 15 seconds; and LED(HIGH) 17 seconds. CONCLUSIONS: In all cases, use of manufacturer-recommended exposure duration provided a lower flexural strength or scraped composite thickness than did longer exposures used. Exposure durations using the simple scraping method correlated very well with those of the much more sophisticated biaxial test. CLINICAL SIGNIFICANCE: No one can provide a clinician with the optimal exposure duration to use for a given light and a specific lot, shade, and brand of composite. Instead, manufacturers offer a single exposure that is meant to be used for all clinical scenarios and operating conditions. The results of this test indicate that manufacturer-recommended exposures proved inadequate to optimize the flexural strength of the recommended increment of composite, but longer exposures were required. The exposure durations determined from the much more simplified composite compule-scrape test proved to match those found to optimize biaxial flexure testing for each light used. Clinicians can thus adapt this very simple in-office scraping test to develop their own customized exposure guide, providing them with exact exposure durations that will optimize composite properties, thus eliminating the guesswork from this most important aspect of chairside dentistry.