Influence of thiol capping on the photoluminescence properties of L‐cysteine‐, mercaptoethanol‐ and mercaptopropionic acid‐capped ZnS nanoparticles

Mercaptoethanol (ME), mercaptopropionic acid (MPA) and L‐cysteine (L‐Cys) having ‐SH functional groups were used as surface passivating agents for the wet chemical synthesis of ZnS nanoparticles. The effect of the thiol group on the optical and photoluminescence (PL) properties of ZnS nanoparticles was studied. L‐Cysteine‐capped ZnS nanoparticles showed the highest PL intensity among the studied capping agents, with a PL emission peak at 455 nm. The PL intensity was found to be dependent on the concentration of Zn²⁺ and S^2– precursors. The effect of buffer on the PL intensity of L‐Cys‐capped ZnS nanoparticles was also studied. UV/Vis spectra showed blue shifting of the absorption edge. Copyright © 2015 John Wiley & Sons, Ltd.     

Morphological and functional characterization of human induced pluripotent stem cell‐derived neurons (iCell Neurons) in defined culture systems

Pre‐clinical testing of drug candidates in animal models is expensive, time‐consuming, and often fails to predict drug effects in humans. Industry and academia alike are working to build human‐based in vitro test beds and advanced high throughput screening systems to improve the translation of preclinical results to human drug trials. Human neurons derived from induced pluripotent stems cells (hiPSCs) are readily available for use within these test‐beds and high throughput screens, but there remains a need to robustly evaluate cellular behavior prior to their incorporation in such systems. This study reports on the characterization of one source of commercially available hiPSC‐derived neurons, iCell^® Neurons, for their long‐term viability and functional performance to assess their suitability for integration within advanced in vitro platforms. The purity, morphology, survival, identity, and functional maturation of the cells utilizing different culture substrates and medium combinations were evaluated over 28 days in vitro (DIV). Patch‐clamp electrophysiological data demonstrated increased capacity for repetitive firing of action potentials across all culture conditions. Significant differences in cellular maturity, morphology, and functional performance were observed in the different conditions, highlighting the importance of evaluating different surface types and growth medium compositions for application in specific in vitro protocols. © 2015 American Institute of Chemical Engineers Biotechnol. Prog., 31:1613–1622, 2015     

Dental root size in bats with diets of different hardness

The relationship between tooth roots and diet is relatively unexplored, although a logical relationship between harder diets and increased root surface area (RSA) is suggested. This study addresses the interaction between tooth morphology, diet, and bite force in small mammals, phyllostomid bats. Using micro computed tomography (microCT), tooth root morphology of two fruit‐eating species (Carollia perspicillata and Chiroderma villosum) and two insect‐eating species (Mimon bennettii and Macrotus californicus) was compared. These species did not differ in skull or estimated body size. Food hardness, rather than dietary classification, proved to be the strongest grouping factor, with the two insectivores and the seed‐processing frugivore (C. villosum) having significantly larger RSAs. Bite force was estimated using skull measurements; bite force significantly correlated with tooth RSA but not with body size. Although the three durophagous species did exhibit larger crowns, the area of the occlusal surface did not vary among the four species. There was a linear relationship between root size and crown size, indicating that the roots were not expanded disproportionately; instead the entire tooth was larger in the hard diet species. MicroCT allows the nondestructive quantification of previously difficult‐to‐access tooth morphology; this method shows the potential for tooth roots to provide valuable dietary, behavioral, and ecological information in small mammals. J. Morphol. 276:1065–1074, 2015. © 2015 Wiley Periodicals, Inc.     

Microstructural roughness of skeletal calcite in ophiuroid vertebral ossicles: evidence of wear?

Local areas of roughened skeletal calcite are reported from the otherwise smooth, imperforate skeletal articulations of the ophiuroid vertebral ossicle (Echinodermata: Ophiuroidea). Complementary patterns of roughness on both proximal and distal articulating surfaces suggest local points of wear between adjacent ossicles, presumably caused by repeated rotation of the intervertebral joint. This surface feature is discussed with respect to its possible origin (mechanical action, resorption of skeletal material, experimental artifact) and the functional morphology of the ophiuroid arm skeleton.     

In vitro effects of 2‐methoxyestradiol‐bis‐sulphamate on the non‐tumorigenic MCF‐12A cell line

A priority in recent anti‐cancer drug development has been attaining better side‐effect profiles for potential compounds. To produce highly specific cancer therapies it is necessary to understand both the effects of the proposed compound on cancer and on normal cells comprising the rest of the human body. Thus in vitro evaluation of these compounds against non‐carcinogenic cell lines is of critical importance. One of the most recent developments in experimental anti‐cancer agents is 2‐methoxyestradiol‐bis‐sulphamate (2ME‐BM), a sulphamoylated derivative of 2‐methoxyestradiol. The aim of this study was to evaluate the in vitro effects of 2ME‐BM on cell proliferation, morphology and mechanisms of cell death in the non‐carcinogenic MCF‐12A breast epithelial cell line. The study revealed changes in proliferative capacity, morphology and cell death induction in response to 2ME‐BM exposure (24 h at 0.4 µM). Microscopy showed decreased cell density and cell death‐associated morphology (increased apoptotic characteristics), a slight increase in acidic intracellular vesicles and insignificant ultra‐structural aberrations. Mitotic indices revealed a G₂M‐phase cell cycle block. This was confirmed by flow cytometry, where an increased fraction of abnormal cells and a decrease in cyclin B1 levels were observed. These results evidently demonstrate that the non‐carcinogenic MCF‐12A cell line is less susceptible when compared to 2ME‐BM‐exposed cancer cell lines previously tested. Further in vitro research into the mechanism of this potentially useful compound is warranted. Copyright © 2010 John Wiley & Sons, Ltd.     

Structure,molting, and mineralization of the dorsal ossicle complex in the gastric mill of the blue crab,Callinectes sapidus

This study examined the mesocardiac and urocardiac ossicles in the gastric mill of the blue crab to describe its structure, mineralization, and dynamics throughout the molt cycle, and to assess its possible utility in age determination. Morphologically, the mineralized ossicles are similar to the calcified dorsal carapace having a lamellate structure comprised of sheets of chitin/protein fibrils. Staining with acridine orange showed the same arrangement of an epicuticle, exocuticle, and endocuticle. In much of the mesocardiac and urocardiac ossicles, the endocuticle is very reduced, with the exocuticle predominating; the reverse of the dimensions of the exoskeleton. The lamellate structure of the ossicles was confirmed with scanning electron microscopy; however, elemental mapping by energy‐dispersive analysis of X‐rays revealed that the ossicles are mineralized with calcium phosphate, in contrast to the calcium carbonate biomineral of the exoskeleton. The medial tooth of the urocardiac ossicle is not calcified, but the epicuticle is highly elaborated and impregnated with silica. Histological examination of the ossicles demonstrated that they are molted during ecdysis, so despite the appearance of bands in the mesocardiac ossicle, it is difficult to hypothesize how the bands could represent a record of chronological age. J. Morphol. 276:1358–1367, 2015. © 2015 Wiley Periodicals, Inc.     

Viscoelastic properties of Staphylococcus aureus and Staphylococcus epidermidis mono‐microbial biofilms

The viscoelastic properties of mono‐microbial biofilms produced by ocular and reference staphylococcal strains were investigated. The microorganisms were characterized for their haemolytic activity and agr typing and the biofilms, grown on stainless steel surface under static conditions, were analysed by Confocal Laser Scanning Microscopy. Static and dynamic rheometric tests were carried out to determine the steady‐flow viscosity and the elastic and viscous moduli. The analysed biofilms showed the typical time‐dependent behaviour of viscoelastic materials with considerable elasticity and mechanical stability except for Staphylococcus aureus ATCC 29213 biofilm which showed a very fragile structure. In particular, S. aureus 6ME biofilm was more compact than other staphylococcal biofilms studied with a yield stress ranging between 2 and 3 Pa. The data obtained in this work could represent a starting point for developing new therapeutic strategies against biofilm‐associated infections, such as improving the drug effect by associating an antimicrobial agent with a biofilm viscoelasticity modifier.     

Linear growth in spines from Acanthaster planci (L.) involving growth lines and periodic pigment bands

We report the first observations of a linear growth pattern in aboral spine ossicles of adult Acanthaster planci (L.). This is unlike the spine development of other echinoderms. Growth in aboral spine ossicles of A. planci is essentially by addition of stereom at the base and the spine's growth history is preserved along its length. There are numerous growth lines perpendicular to the long axis of the ossicle. These are clearly evident in longitudinal spine sections and apparently caused by frequent growth episodes. There are periodic pigment bands which are parallel to the growth lines and evident on the surface of the ossicle. Basal growth of the spine ossicle and the nature of the growth lines were confirmed by tetracycline staining. Size/frequency analyses of a population of A. planci from Davies Reef (GBR) found spine ossicle growth, but not body diameter growth, over the six month period between sampling dates. The additional pigment banding in spine ossicles of 4 individuals recaptured after 6 months suggests that pigment bands are laid down seasonally. If pigment band cyclicity is validated, it offers a simple method for ageing adults of A. planci in field populations.     

Structure of the stomach cuticle in adult and larvae of the spider crab Maja brachydactyla (Brachyura,Decapoda)

The stomach of decapods is a complex organ with specialized structures that are delimited by a cuticle. The morphology and ontogeny of the stomach are largely described, but few studies have focused on the morphology of its cuticle. This study examined the morphology of the stomach cuticle of cardiac sacs, gastric mill ossicles, cardio-pyloric valve and pyloric filters, and during various stages (zoea I and II, megalopa, first juvenile, and adult) of the common spider crab Maja brachydactyla using dissection, histology and transmission electron microscopy. The results show that cuticle morphology varies among structures (e.g., cardiac sacs, urocardiac ossicle, cardio-pyloric valve, pyloric filters), within a single structure (e.g., different sides of the urocardiac ossicle) and among different life stages. The cuticle during the larval stages is very thin and the different layers (epicuticle, exocuticle, and endocuticle) are infrequently distinguishable by histology. Major changes during larval development regarding cuticle morphology are observed after the molt to megalopa, including the increment in thickness in the gastric mill ossicles and cardio-pyloric valve, and the disappearance of the long thickened setae of the cardio-pyloric valve. The cuticle of all the stomach structures in the adults is thicker than in larval and juvenile stages. The cuticle varies in thickness, differential staining affinity and morphology of the cuticle layers. The structure–function relationship of the cuticle morphology is discussed.     

Variations in the mechanical properties of Alouatta palliata molar enamel

Teeth have provided insights into many topics including primate diet, paleobiology, and evolution, due to the fact that they are largely composed of inorganic materials and may remain intact long after an animal is deceased. Previous studies have reported that the mechanical properties, chemistry, and microstructure of human enamel vary with location. This study uses nanoindentation to map out the mechanical properties of Alouatta palliata molar enamel on an axial cross‐section of an unworn permanent third molar, a worn permanent first molar, and a worn deciduous first molar. Variations were then correlated with changes in microstructure and chemistry using scanning electron microscopy and electron microprobe techniques. The hardness and Young's modulus varied with location throughout the cross‐sections from the occlusal surface to the dentin‐enamel junction (DEJ), from the buccal to lingual sides, and also from one tooth to another. These changes in mechanical properties correlated with changes in the organic content of the tooth, which was shown to increase from ～6% near the occlusal surface to ～20% just before the DEJ. Compared to human enamel, the Alouatta enamel showed similar microstructures, chemical constituents, and magnitudes of mechanical properties, but showed less variation in hardness and Young's modulus, despite the very different diet of this species. Am J Phys Anthropol 2010. © 2009 Wiley‐Liss, Inc.     

2‐methoxyestradiol inhibits atorvastatin‐induced rounding of human vascular smooth muscle cells

The cardiovascular benefits of statins, including atorvastatin (ATV), have been reported to be gender‐dependent, but the underlying mechanism is unclear. In this study we examine whether estrogen and its metabolite, 2‐methoxyestradiol (2ME), affect the rounding response of human vascular smooth muscle cells (SMCs) induced by ATV. Twenty‐four hour treatment with ATV (10–100 µM) induced rounding of cultured human SMCs. Addition of 2ME (1–20 µM), but not 17β‐estradiol, for 2 h induced re‐spreading of rounded cells. Our further studies showed that the effects of 2ME were mimicked by microtubule‐disrupting drugs and inhibited by taxol. Inhibition of RhoA and ROCK (Rho‐kinase) by C3‐toxin and H‐1152, respectively, blocked 2ME effects. 2ME effects were also blocked by treatment with either actin‐interfering drugs, such as cytochalasin D and jasplakinolide, or myosin inhibitor blebbistatin. ML‐7 and ‐9, the inhibitors for myosin light chain kinase, inhibited 2ME effect as well. ATV treatment induced a decrease of F‐actin content and Thr18/Ser19 dual phosphorylation of myosin regulatory light chain (MRLC), which was rescued by 2ME or mevalonate. The rescue effects of 2ME on F‐actin content and MRLC dual phosphorylation were abolished by taxol or H‐1152. In addition, kinesin Eg5 inhibitor monastrol and dynein inhibitor erythro‐9‐3‐(2‐hydroxynonyl) adenine (EHNA) significantly blocked 2ME effects. Finally, our results revealed that 2ME inhibited the migration of SMCs induced by ATV (0.1 µM) in wound healing assay and Boyden chamber assay. In summary, our data show that 2ME, but not estrogen, inhibits ATV‐induced rounding of human SMCs through induction of microtubule disassembly and activation of the Rho‐ROCK‐actinomyosin pathway. J. Cell. Physiol. 222: 556–564, 2010. © 2009 Wiley‐Liss, Inc.     

Micro‐ to nanostructure and geochemistry of extant crinoidal echinoderm skeletons

This paper reports the results of micro‐ to nanostructural and geochemical analyses of calcitic skeletons from extant deep‐sea stalked crinoids. Fine‐scale (SEM, FESEM, AFM) observations show that the crinoid skeleton is composed of carbonate nanograins, about 20–100 nm in diameter, which are partly separated by what appears to be a few nm thick organic layers. Sub‐micrometre‐scale geochemical mapping of crinoid ossicles using a NanoSIMS ion microprobe, combined with synchrotron high‐spatial‐resolution X‐ray micro‐fluorescence (μ‐XRF) maps and X‐ray absorption near‐edge structure spectroscopy (XANES) show that high Mg concentration in the central region of the stereom bars correlates with the distribution of S‐sulphate, which is often associated with sulphated polysaccharides in biocarbonates. These data are consistent with biomineralization models suggesting a close association between organic components (including sulphated polysaccharides) and Mg ions. Additionally, geochemical analyses (NanoSIMS, energy dispersive spectroscopy) reveal that significant variations in Mg occur at many levels: within a single stereom trabecula, within a single ossicle and within a skeleton of a single animal. Together, these data suggest that physiological factors play an important role in controlling Mg content in crinoid skeletons and that great care should be taken when using their skeletons to reconstruct, for example, palaeotemperatures and Mg/Ca palaeo‐variations of the ocean.     

Tunable multicolor and white‐light upconversion luminescence in Yb3+/Tm3+/Ho3+ tri‐doped NaYF4 micro‐crystals

NaYF₄ micro‐crystals with various concentrations of Yb³⁺/Tm³⁺/Ho³⁺ were prepared successfully via a simple and reproducible hydrothermal route using EDTA as the chelating agent. Their phase structure and surface morphology were studied using powder X‐ray diffraction (XRD) and scanning electron microscopy (SEM). The XRD patterns revealed that all the samples were pure hexagonal phase NaYF₄. SEM images showed that Yb³⁺/Tm³⁺/Ho³⁺ tri‐doped NaYF₄ were hexagonal micro‐prisms. Upconversion photoluminescence spectra of Yb³⁺/Tm³⁺/Ho³⁺ tri‐doped NaYF₄ micro‐crystals with various dopant concentrations under 980 nm excitation with a 665 mW pump power were studied. Tunable multicolor (purple, purplish blue, yellowish green, green) and white light were achieved by simply adjusting the Ho³⁺ concentration in 20%Yb³⁺/1%Tm³⁺/xHo³⁺ tri‐doped NaYF₄ micro‐crystals. Furthermore, white‐light emissions could be obtained using different pump powers in 20%Yb³⁺/1%Tm³⁺/1%Ho³⁺ tri‐doped NaYF₄ micro‐crystals at 980 nm excitation. The pump power‐dependent intensity relationship was studied and relevant energy transfer processes were discussed in detail. The results suggest that Yb³⁺/Tm³⁺/Ho³⁺ tri‐doped NaYF₄ micro‐crystals have potential applications in optoelectronic devices such as photovoltaic, plasma display panel and white‐light‐emitting diodes. Copyright © 2014 John Wiley & Sons, Ltd.     

2‐methoxyestradiol‐mediated anti‐tumor effect increases osteoprotegrin expression in osteosarcoma cells

Osteosarcoma is a bone tumor that frequently develops during adolescence. 2‐Methoxyestradiol (2‐ME), a naturally occurring metabolite of 17β‐estradiol, induces cell cycle arrest and cell death in human osteosarcoma cells. To investigate whether the osteoprotegrin (OPG) protein plays a role in 2‐ME actions, we studied the effect of 2‐ME treatment on OPG gene expression in human osteosarcoma cells. 2‐ME treatment induced OPG gene promoter activity and mRNA levels. Also, Western blot analysis showed that 2‐ME treatment increased OPG protein levels in MG63, KHOS, 143B and LM7 osteosarcoma cells by 3‐, 1.9‐, 2.8‐, and 2.5‐fold, respectively, but did not affect OPG expression in normal bone cells. In addition, increases in OPG protein levels were observed in osteosarcoma cell culture media after 3 days of 2‐ME treatment. The effect of 2‐ME on osteosarcoma cells was ligand‐specific as parent estrogen, 17β‐estradiol and a tumorigenic estrogen metabolite, 16α‐hydroxyestradiol, which do not affect osteosarcoma cell cycle and cell death, had no effect on OPG protein expression. Furthermore, co‐treating osteosarcoma cells with OPG protein did not further enhance 2‐ME‐mediated anti‐tumor effects. OPG‐released in 2‐ME‐treated cultures led to an increase in osteoblastic activity and a decrease in osteoclast number, respectively. These findings suggest that OPG is not directly involved in 2‐ME‐mediated anti‐proliferative effects in osteosarcoma cells, but rather participates in anti‐resorptive functions of 2‐ME in bone tumor environment. J. Cell. Biochem. 109: 950–956, 2010. © 2010 Wiley‐Liss, Inc.     

Foregut morphology and ontogeny of the mud crab Dyspanopeus sayi (Smith, 1869) (Decapoda,Brachyura, Panopeidae)

The morphology of the foregut of the Say's mud crab Dyspanopeus sayi was described in adults and larvae. The ossicle system was illustrated based on a staining method with Alizarin-Red. The gastric teeth and cardio-pyloric valve were dissected and examined using optical and scanning electron microscopy. In the adults, the morphology of ossicles and gastric teeth of D. sayi is very similar to the related species Rhithropanopeus harrisii. The foregut of first zoea (ZI) presented a functional cardio-pyloric valve while the filter press was lacking. The filter press was observed in the pyloric chamber from ZII. The most significant changes in morphology take place after metamorphosis from ZIV to megalopa, including the occurrence of the gastric mill. The organization and morphology of many megalopal foregut ossicles are recognizable in the adult phase, although the morphology of the gastric teeth differs from the morphology of adults. A correlation of gastric mill structures with food preferences and their contribution to the phylogeny are briefly discussed.     

Three-dimensional Raman imagery of precambrian microscopic organisms

Two major difficulties have long hindered studies of ancient permineralized micro‐organisms: (1) accurate documentation of their three‐dimensional morphology at micron‐scale resolution; and (2) direct characterization of their molecular‐structural composition. Here we introduce a technique new to palaeobiology, the three‐dimensional Raman imagery that meets both of these needs, which is a nonintrusive, nondestructive technique that can provide data by which to accurately and objectively characterize, in situ and at micron‐scale resolution, the morphology and molecular‐structural composition of permineralized micro‐organisms and their associated matrices. Application of this new technique can provide information about the morphology, taphonomy and fidelity of preservation of fossil micro‐organisms unavailable by any other means.     

Micro‐CT evaluation of murine fetal skeletal development yields greater morphometric precision over traditional clear‐staining methods

Traditional techniques for quantification of murine fetal skeletal development (gross measurements, clear‐staining) are severely limited by specimen processing, soft tissue presence, diffuse staining, and unclear landmarks between which to make measurements. Nondestructive microcomputed tomography (micro‐CT) imaging is a versatile, well‐documented tool traditionally used to generate high‐resolution 3‐D images and quantify microarchitectural parameters of trabecular bone. Although previously described as a tool for phenotyping fetal murine specimens, micro‐CT has not previously been used to directly measure individual fetal skeletal structures. Imaging murine fetal skeletons using micro‐CT enables the researcher to nondestructively quantify fetal skeletal development parameters including limb length, total bone volume, and average bone mineral density, as well as identify skeletal malformations. Micro‐CT measurement of fetal limb lengths correlates well with traditional clear‐staining methods (83.98% agreement), decreases variability in measurements (average standard errors: 6.28% for micro‐CT and 10.82% for clear‐staining), decreases data acquisition time by eliminating the need for tissue processing, and preserves the intact fixed fetus for further analysis. Use of the rigorous micro‐CT technique to generate 3‐D images for digital measurement enables isolation of skeletal structures based on degree of mineralization (local radiodensity), eliminating the complications of blurred stain boundaries and soft tissue inclusion that accompany clear‐staining and gross measurement techniques. Microcomputed tomography provides a facile, accurate, and nondestructive method for determining the developmental state of the fetal skeleton using not only limb lengths and identification of malformations, but total skeletal bone volume and average skeletal mineral density as well. Birth Defects Res (Part B) 2008. © 2008 Wiley‐Liss, Inc.     

Combining 3D human in vitro methods for a 3Rs evaluation of novel titanium surfaces in orthopaedic applications

In this study, we report on a group of complementary human osteoblast in vitro test methods for the preclinical evaluation of 3D porous titanium surfaces. The surfaces were prepared by additive manufacturing (electron beam melting [EBM]) and plasma spraying, allowing the creation of complex lattice surface geometries. Physical properties of the surfaces were characterized by SEM and profilometry and 3D in vitro cell culture using human osteoblasts. Primary human osteoblast cells were found to elicit greater differences between titanium sample surfaces than an MG63 osteoblast‐like cell line, particularly in terms of cell survival. Surface morphology was associated with higher osteoblast metabolic activity and mineralization on rougher titanium plasma spray coated surfaces than smoother surfaces. Differences in osteoblast survival and metabolic activity on titanium lattice structures were also found, despite analogous surface morphology at the cellular level. 3D confocal microscopy identified osteoblast organization within complex titanium surface geometries, adhesion, spreading, and alignment to the biomaterial strut geometries. Mineralized nodule formation throughout the lattice structures was also observed, and indicative of early markers of bone in‐growth on such materials. Testing methods such as those presented are not traditionally considered by medical device manufacturers, but we suggest have value as an increasingly vital tool in efficiently translating pre‐clinical studies, especially in balance with current regulatory practice, commercial demands, the 3Rs, and the relative merits of in vitro and in vivo studies. Biotechnol. Bioeng. 2016;113: 1586–1599. © 2015 The Authors. Biotechnology and Bioengineering Published by Wiley Periodicals, Inc.     

Influence of fruit availability on macronutrient and energy intake by female chimpanzees

Daily energy intake of adult female mammals is influenced by environmental conditions and physiological requirements, including reproduction. We examined the effects of fruit availability on macronutrient and metabolisable energy (ME) intake by adult female chimpanzees (Pan troglodytes schweinfurthii) of the Kanyawara community in Kibale National Park, Uganda, from January 2014 through June 2015. Drupe fruits were abundant for 4 months, whereas the other 14 months were dominated by fig fruits. The mean daily intake of food (dry matter) and ME did not differ between drupe‐months and fig‐months. However, foraging costs were higher during fig‐months, as indicated by a 20% increase in feeding time. Furthermore, during drupe‐months female chimpanzees ingested more water‐soluble carbohydrates and lipids, and less available protein and neutral detergent fibre. Although ME intake did not differ consistently between drupe‐months and fig‐months, they consumed more on days when ripe fruit dominated the diet than when leaves and pithy stems dominated the diet. Our data suggest that differences in diet quality between drupes and figs can have important effects on frugivore foraging and that they influence net energy gain more by their effects on macronutrient composition or foraging cost than by their direct impact on energy intake.