Amelogenin Induces Biomimetic Mineralization at Specific pH

2003 ◽  
Vol 774 ◽  
Author(s):  
Stefan Habelitz ◽  
Dustin Ford ◽  
Sally J. Marshall ◽  
Pamela K. DenBesten ◽  
Mehdi Balooch ◽  
...  
Keyword(s):  
Composite Material ◽  
Dental Enamel ◽  
Biomimetic Mineralization ◽  
Force Microscopy ◽  
Atomic Force ◽  
Human Enamel ◽  
Glass Ceramic Substrate ◽  
Chemical Conditions ◽  
Supersaturated Solutions

AbstractAmelogenin proteins are assumed to control the calcification of dental enamel with a nanoscale precision that facilitates the formation of fibrous apatite crystals organized in a remarkable microstucture. In this study, recombinant full-length human amelogenin induced protein-guided mineralization and the formation of an enamel-like composite material at specific physical-chemical conditions as observed by atomic force microscopy (AFM). Amelogenin bound specifically to fluoroapatite crystals (FAP) of a glass-ceramic substrate at Ca2+ and PO43- concentrations similar to in-vivo conditions and at pH 8. Layers up to 400 nm high, containing elongated crystals, formed on the (001)-planes of FAP within 24h in supersaturated solutions. In contrast, (hk0)-faces grew by only 10-30 nm, but showed nanospheres aligned parallel to the c-axis of FAP. At pHs different from 8, proteins bound non-specifically to substrate and layers on FAP reached only 5-15 nm thickness. Micro-Raman spectroscopy and AFM revealed the formation of a composite material that resembled a structure and composition comparable to human enamel. These observations suggest that certain conditions are required to activate amelogenin to control and promote crystal growth of apatite along the c-axis and to synthesize an enamel-like material.

Rapid, Refined, and Robust Method for Expression, Purification, and Characterization of Recombinant Human Amyloid-beta M1-42

2019 ◽  
Author(s):  
Priya Prakash ◽  
Travis Lantz ◽  
Krupal P. Jethava ◽  
Gaurav Chopra
Keyword(s):  
Amyloid Beta ◽  
Western Blots ◽  
Force Microscopy ◽  
E Coli ◽  
Atomic Force ◽  
Set Up ◽  
Short Time

Amyloid plaques found in the brains of Alzheimer’s disease (AD) patients primarily consists of amyloid beta 1-42 (Ab42). Commercially, Ab42 is synthetized using peptide synthesizers. We describe a robust methodology for expression of recombinant human Ab(M1-42) in Rosetta(DE3)pLysS and BL21(DE3)pLysS competent E. coli with refined and rapid analytical purification techniques. The peptide is isolated and purified from the transformed cells using an optimized set-up for reverse-phase HPLC protocol, using commonly available C18 columns, yielding high amounts of peptide (~15-20 mg per 1 L culture) in a short time. The recombinant Ab(M1-42) forms characteristic aggregates similar to synthetic Ab42 aggregates as verified by western blots and atomic force microscopy to warrant future biological use. Our rapid, refined, and robust technique to purify human Ab(M1-42) can be used to synthesize chemical probes for several downstream in vitro and in vivo assays to facilitate AD research.

scholarly journals Biomechanics of Neutrophil Tethers

Life ◽  
2021 ◽  
Vol 11 (6) ◽  
pp. 515
Author(s):  
Andrea Cugno ◽  
Alex Marki ◽  
Klaus Ley
Keyword(s):  
Cell Activation ◽  
Optical Trap ◽  
Biomechanical Properties ◽  
Force Microscopy ◽  
Advantages And Disadvantages ◽  
Atomic Force ◽  
Microfluidic Flow ◽  
Biomembrane Force Probe ◽  
Membrane Reservoir

Leukocytes, including neutrophils, which are propelled by blood flow, can roll on inflamed endothelium using transient bonds between selectins and their ligands, and integrins and their ligands. When such receptor–ligand bonds last long enough, the leukocyte microvilli become extended and eventually form thin, 20 m long tethers. Tether formation can be observed in blood vessels in vivo and in microfluidic flow chambers. Tethers can also be extracted using micropipette aspiration, biomembrane force probe, optical trap, or atomic force microscopy approaches. Here, we review the biomechanical properties of leukocyte tethers as gleaned from such measurements and discuss the advantages and disadvantages of each approach. We also review and discuss viscoelastic models that describe the dependence of tether formation on time, force, rate of loading, and cell activation. We close by emphasizing the need to combine experimental observations with quantitative models and computer simulations to understand how tether formation is affected by membrane tension, membrane reservoir, and interactions of the membrane with the cytoskeleton.

scholarly journals Silane-Coating Strategy for Titanium Functionalization Does Not Impair Osteogenesis In Vivo

Materials ◽  
2021 ◽  
Vol 14 (7) ◽  
pp. 1814
Author(s):  
Plinio Mendes Senna ◽  
Carlos Fernando de Almeida Barros Mourão ◽  
Rafael Coutinho Mello-Machado ◽  
Kayvon Javid ◽  
Pietro Montemezzi ◽  
...  
Keyword(s):  
Bone Formation ◽  
Photoelectron Spectroscopy ◽  
Titanium Surface ◽  
Rabbit Model ◽  
Biologically Active ◽  
Force Microscopy ◽  
Atomic Force ◽  
Silane Coating ◽  
Titanium Screws

Silane-coating strategy has been used to bind biological compounds to the titanium surface, thereby making implant devices biologically active. However, it has not been determined if the presence of the silane coating itself is biocompatible to osseointegration. The aim of the present study was to evaluate if silane-coating affects bone formation on titanium using a rabbit model. For this, titanium screw implants (3.75 by 6 mm) were hydroxylated in a solution of H2SO4/30% H2O2 for 4 h before silane-coating with 3-aminopropyltriethoxysilane (APTES). A parallel set of titanium screws underwent only the hydroxylation process to present similar acid-etched topography as a control. The presence of the silane on the surface was checked by x-ray photoelectron spectroscopy (XPS), with scanning electron microscopy (SEM) and atomic force microscopy (AFM). A total of 40 titanium screws were implanted in the tibia of ten New Zealand rabbits in order to evaluate bone-to-implant contact (BIC) after 3 weeks and 6 weeks of healing. Silane-coated surface presented higher nitrogen content in the XPS analysis, while micro- and nano-topography of the surface remained unaffected. No difference between the groups was observed after 3 and 6 weeks of healing (p > 0.05, independent t-test), although an increase in BIC occurred over time. These results indicate that silanization of a titanium surface with APTES did not impair the bone formation, indicating that this can be a reliable tool to anchor osteogenic molecules on the surface of implant devices.

Nanoscale Characterisation of Salivary Pellicle

2004 ◽  
Vol 841 ◽  
Author(s):  
Michelle E. Dickinson ◽  
Adrian B. Mann
Keyword(s):  
Dental Enamel ◽  
Tooth Surface ◽  
Surface Adhesion ◽  
Human Enamel ◽  
Oral Physiology ◽  
Salivary Pellicle ◽  
Initial Adsorption ◽  
Dietary Agents

ABSTRACTSalivary pellicle is an organic biofilm formed by the physisorption of proteins and carbohydrates onto the surface of dental enamel exposed to the oral environment. The pellicle has several key roles in oral physiology including lubrication and reduction of friction between teeth during mastication, as well as chemical protection of the enamel against acidic solutions. However, pellicle proteins are known to react with dietary compounds to cause extrinsic staining on the tooth surface.In this study, nanoindentation and AFM have been used in vitro to examine the acquired salivary pellicle formed in vivo on dental enamel. The mechanical properties, growth, structure and morphology of pellicle grown in vivo on human enamel surfaces have been analysed. In addition, the effects of dietary agents such as polyphenols on the pellicle's morphology and properties have been studied.It was found that initial adsorption of proteins on the enamel surface occurred within 30 seconds of exposure to the oral cavity, with full growth achieved within 2 hours. Differences in the properties of the pellicles such as surface adhesion, and time dependent effects due to polyphenol interaction were measured using nanoindentation. It was seen that the polyphenol interaction has a significant effect on these properties. These results suggest that the stained pellicle is mechanically stiffer, but also less viscous and more fluid like. This could explain why traditional tooth brushing techniques do not efficiently remove this layer.

Diabetes increases stiffness of live cardiomyocytes measured by atomic force microscopy nanoindentation

2014 ◽  
Vol 307 (10) ◽  
pp. C910-C919 ◽  
Author(s):  
Juan C. Benech ◽  
Nicolás Benech ◽  
Ana I. Zambrana ◽  
Inés Rauschert ◽  
Verónica Bervejillo ◽  
...  
Keyword(s):  
Atomic Force Microscopy ◽  
Physiological Condition ◽  
Diabetic Mice ◽  
Cell Nuclei ◽  
Buffer Solution ◽  
Isolated Cardiomyocytes ◽  
Force Microscopy ◽  
Atomic Force ◽  
Myocardial Fibers

Stiffness of live cardiomyocytes isolated from control and diabetic mice was measured using the atomic force microscopy nanoindentation method. Type 1 diabetes was induced in mice by streptozotocin administration. Histological images of myocardium from mice that were diabetic for 3 mo showed disorderly lineup of myocardial cells, irregularly sized cell nuclei, and fragmented and disordered myocardial fibers with interstitial collagen accumulation. Phalloidin-stained cardiomyocytes isolated from diabetic mice showed altered (i.e., more irregular and diffuse) actin filament organization compared with cardiomyocytes from control mice. Sarco/endoplasmic reticulum Ca2+-ATPase (SERCA2a) pump expression was reduced in homogenates obtained from the left ventricle of diabetic animals compared with age-matched controls. The apparent elastic modulus (AEM) for live control or diabetic isolated cardiomyocytes was measured using the atomic force microscopy nanoindentation method in Tyrode buffer solution containing 1.8 mM Ca2+ and 5.4 mM KCl (physiological condition), 100 nM Ca2+ and 5.4 mM KCl (low extracellular Ca2+ condition), or 1.8 mM Ca2+ and 140 mM KCl (contraction condition). In the physiological condition, the mean AEM was 112% higher for live diabetic than control isolated cardiomyocytes (91 ± 14 vs. 43 ± 7 kPa). The AEM was also significantly higher in diabetic than control cardiomyocytes in the low extracellular Ca2+ and contraction conditions. These findings suggest that the material properties of live cardiomyocytes were affected by diabetes, resulting in stiffer cells, which very likely contribute to high diastolic LV stiffness, which has been observed in vivo in some diabetes mellitus patients.

scholarly journals In vivo dynamics of the cortical actin network revealed by fast-scanning atomic force microscopy

Microscopy ◽  
2017 ◽  
Vol 66 (4) ◽  
pp. 272-282 ◽  
Author(s):  
Yanshu Zhang ◽  
Aiko Yoshida ◽  
Nobuaki Sakai ◽  
Yoshitsugu Uekusa ◽  
Masahiro Kumeta ◽  
...  
Keyword(s):  
Atomic Force Microscopy ◽  
Actin Network ◽  
Cortical Actin ◽  
Force Microscopy ◽  
Atomic Force

Dental enamel roughness with different acid etching times: Atomic force microscopy study

2012 ◽  
Vol 1 (3) ◽  
pp. 187 ◽  
Author(s):  
OrlandoMotohiro Tanaka ◽  
ElisaSouza Camargo ◽  
BrunoBochnia Cerci ◽  
Odilon Guariza-Filho ◽  
LucimaraStolz Roman
Keyword(s):  
Atomic Force Microscopy ◽  
Dental Enamel ◽  
Microscopy Study ◽  
Acid Etching ◽  
Atomic Force Microscopy Study ◽  
Force Microscopy ◽  
Atomic Force ◽  
Enamel Roughness

scholarly journals A New Sugarcane Cystatin Strongly Binds to Dental Enamel and Reduces Erosion

2017 ◽  
Vol 96 (9) ◽  
pp. 1051-1057 ◽  
Author(s):  
A.C. Santiago ◽  
Z.N. Khan ◽  
M.C. Miguel ◽  
C.C. Gironda ◽  
A. Soares-Costa ◽  
...  
Keyword(s):  
Potent Inhibitor ◽  
Surface Hardness ◽  
Dental Enamel ◽  
Reversible Inhibitors ◽  
Force Microscopy ◽  
Atomic Force ◽  
Enamel Erosion ◽  
Acquired Enamel Pellicle

Cystatin B was recently identified as an acid-resistant protein in acquired enamel pellicle; it could therefore be included in oral products to protect against caries and erosion. However, human recombinant cystatin is very expensive, and alternatives to its use are necessary. Phytocystatins are reversible inhibitors of cysteine peptidases that are found naturally in plants. In plants, they have several biological and physiological functions, such as the regulation of endogenous processes, defense against pathogens, and response to abiotic stress. Previous studies performed by our research group have reported high inhibitory activity and potential agricultural and medical applications of several sugarcane cystatins, including CaneCPI-1, CaneCPI-2, CaneCPI-3, and CaneCPI-4. In the present study, we report the characterization of a novel sugarcane cystatin, named CaneCPI-5. This cystatin was efficiently expressed in Escherichia coli, and inhibitory assays demonstrated that it was a potent inhibitor of human cathepsins B, K, and L ( Ki = 6.87, 0.49, and 0.34 nM, respectively). The ability of CaneCPI-5 to bind to dental enamel was evaluated using atomic force microscopy. Its capacity to protect against initial enamel erosion was also tested in vitro via changes in surface hardness. CaneCPI-5 showed a very large force of interaction with enamel (e.g., compared with mucin and casein) and significantly reduced initial enamel erosion. These results suggest that the inclusion of CaneCPIs in dental products might confer protection against enamel erosion.

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