Comparison of in vitro and in vivo 44Ca labeling of bone by scanning ion microprobe

1990 ◽  
Vol 259 (4) ◽  
pp. E586-E592 ◽  
Author(s):  
D. A. Bushinsky ◽  
J. M. Chabala ◽  
R. Levi-Setti
Keyword(s):  
Secondary Ion Mass Spectrometry ◽  
Bone Mineralization ◽  
Total Body Weight ◽  
Elemental Distribution ◽  
Ion Microprobe ◽  
Distribution Maps ◽  
Total Body ◽  
Secondary Ion

To determine whether Ca incorporation from medium into cultured bone represents normal mineralization, we labeled some neonatal mouse calvariae in vitro and others in vivo with the stable isotope 44Ca and compared surface label localization with a scanning ion microprobe utilizing secondary ion mass spectrometry. To label in vitro, we incubated live calvariae in medium containing 40Ca or 44Ca for 3 h. Compared with a 44Ca/40Ca ratio of 0.020 with 1 mM 40Ca, the ratio with 1 mM 44Ca was 0.135 and with 2 mM 44Ca was 0.556. Erosion revealed a marked decrease in 44Ca/40Ca with depth. To label in vivo, we subcutaneously injected 40Ca or 44Ca into mice equal to a percentage of their total body weight and dissected the calvariae 24 h later. Compared with a 44Ca/40Ca ratio of 0.021 with 2% 40Ca, the ratio with 2% 44Ca was 0.120 and with 6% 44Ca was 0.205. Erosion revealed only a slight decrease in 44Ca/40Ca with depth. Elemental distribution maps of in vivo labeled samples show broad deposition of 44Ca, whereas maps of in vitro labeled bones show 44Ca preferentially localized at the surface in contact with the medium. Thus calvariae can be labeled with 44Ca both in vitro and in vivo. However, the differing patterns of isotope localization under the conditions of this study indicate that in vitro Ca deposition differs from normal in vivo bone mineralization.

ADAMTS5 promotes murine adipogenesis and visceral adipose tissue expansion

2016 ◽  
Vol 116 (10) ◽  
pp. 694-704 ◽  
Author(s):  
Dries Bauters ◽  
Ilse Scroyen ◽  
Rebecca Deprez-Poulain ◽  
H. Roger Lijnen
Keyword(s):  
Adipose Tissue ◽  
De Novo ◽  
Tissue Expansion ◽  
Total Body Weight ◽  
Stable Gene ◽  
Enhanced Expression ◽  
Total Body ◽  
A Disintegrin And Metalloproteinase

SummaryEnhanced expression of the aggrecanase ADAMTS5 (A Disintegrin And Metalloproteinase with Thrombospondin type 1 motifs; member 5) has been observed in adipose tissue (AT) of obese rodents. Here, we have investigated the role of ADAMTS5 in adipogenesis, AT expansion and associated angiogenesis. In vitro differentiation of precursor cells into mature adipocytes was studied using murine embryonic fibroblasts (MEF) derived from wild-type (Adamts5 +/+) and ADAMTS5 deficient (Adamts5 -/-) mice, or 3T3-F442A preadipocytes with stable gene silencing of Adamts5. De novo adipogenesis was monitored by injection of 3T3-F442A cells with or without Adamts5 knockdown in Nude mice. Furthermore, Adamts5+/+ and Adamts5/- mice were kept on a high-fat diet (HFD) to monitor AT development. Adamts5-/- MEF, as well as 3T3-F442A preadipocytes with Adamts5 knockdown, showed significantly reduced differentiation as compared to control cells. In mice, de novo formed fat pads arising from 3T3-F442A cells with Adamts5 knockdown were significantly smaller as compared to controls. After 15 or 25 weeks on HFD, total body weight and subcutaneous AT weight were similar for Adamts5++/+ and Adamts5-/- mice, but visceral/gonadal fat mass was significantly lower for Adamts5-/-mice. These data were confirmed by magnetic resonance imaging. In addition, the blood vessel density in adipose tissue was higher for Adamts5-/- mice kept on HFD. In conclusion, our data support the concept that ADAMTS5 promotes adipogenesis in vitro and in vivo, as well as development of visceral AT and associated angiogenesis in mice kept on HFD.

Aspects of high-resolution imaging with a scanning ion microprobe

1986 ◽  
Vol 44 ◽  
pp. 750-751
Author(s):  
R. Levi-Setti ◽  
J. M. Chabala ◽  
Y. L. Wang
Keyword(s):  
Metal Ion ◽  
Secondary Ion Mass Spectrometry ◽  
Chromatic Aberration ◽  
Ion Source ◽  
Ion Microprobe ◽  
Emission Pattern ◽  
Intrinsic Resolution ◽  
Resolution Imaging ◽  
20 Nm ◽  
Secondary Ion

We have shown the feasibility of 20 nm lateral resolution in both topographic and elemental imaging using probes of this size from a liquid metal ion source (LMIS) scanning ion microprobe (SIM). This performance, which approaches the intrinsic resolution limits of secondary ion mass spectrometry (SIMS), was attained by limiting the size of the beam defining aperture (5μm) to subtend a semiangle at the source of 0.16 mr. The ensuing probe current, in our chromatic-aberration limited optical system, was 1.6 pA with Ga+ or In+ sources. Although unique applications of such low current probes have been demonstrated,) the stringent alignment requirements which they imposed made their routine use impractical. For instance, the occasional tendency of the LMIS to shift its emission pattern caused severe misalignment problems.

Imaging of Al-Li-Cu alloys with a Scanning Ion Microprobe

1990 ◽  
Vol 48 (2) ◽  
pp. 314-315
Author(s):  
K.K. Soni ◽  
D.B. Williams ◽  
J.M. Chabala ◽  
R. Levi-Setti ◽  
D.E. Newbury
Keyword(s):  
Mass Spectrometry ◽  
Secondary Ion Mass Spectrometry ◽  
Equilibrium Phase ◽  
Icosahedral Symmetry ◽  
Ion Microprobe ◽  
X Ray ◽  
Cu Alloys ◽  
Two Phases ◽  
The University ◽  
Secondary Ion

In contrast to the inability of x-ray microanalysis to detect Li, secondary ion mass spectrometry (SIMS) generates a very strong Li+ signal. The latter’s potential was recently exploited by Williams et al. in the study of binary Al-Li alloys. The present study of Al-Li-Cu was done using the high resolution scanning ion microprobe (SIM) at the University of Chicago (UC). The UC SIM employs a 40 keV, ∼70 nm diameter Ga+ probe extracted from a liquid Ga source, which is scanned over areas smaller than 160×160 μm2 using a 512×512 raster. During this experiment, the sample was held at 2 × 10-8 torr.In the Al-Li-Cu system, two phases of major importance are T1 and T2, with nominal compositions of Al2LiCu and Al6Li3Cu respectively. In commercial alloys, T1 develops a plate-like structure with a thickness <∼2 nm and is therefore inaccessible to conventional microanalytical techniques. T2 is the equilibrium phase with apparent icosahedral symmetry and its presence is undesirable in industrial alloys.

Preparation of cultured astrocytes for x-ray microanalysis

1989 ◽  
Vol 47 ◽  
pp. 988-989
Author(s):  
N.K.R. Smith ◽  
K.E. Hunter ◽  
P. Mobley ◽  
L.P. Felpel
Keyword(s):  
Cultured Cells ◽  
Elemental Content ◽  
Elemental Distribution ◽  
Sprague Dawley ◽  
X Ray ◽  
Cultured Astrocytes ◽  
Freeze Dried ◽  
Ultimate Objective

Electron probe energy dispersive x-ray microanalysis (XRMA) offers a powerful tool for the determination of intracellular elemental content of biological tissue. However, preparation of the tissue specimen , particularly excitable central nervous system (CNS) tissue , for XRMA is rather difficult, as dissection of a sample from the intact organism frequently results in artefacts in elemental distribution. To circumvent the problems inherent in the in vivo preparation, we turned to an in vitro preparation of astrocytes grown in tissue culture. However, preparations of in vitro samples offer a new and unique set of problems. Generally, cultured cells, growing in monolayer, must be harvested by either mechanical or enzymatic procedures, resulting in variable degrees of damage to the cells and compromised intracel1ular elemental distribution. The ultimate objective is to process and analyze unperturbed cells. With the objective of sparing others from some of the same efforts, we are reporting the considerable difficulties we have encountered in attempting to prepare astrocytes for XRMA.Tissue cultures of astrocytes from newborn C57 mice or Sprague Dawley rats were prepared and cultured by standard techniques, usually in T25 flasks, except as noted differently on Cytodex beads or on gelatin. After different preparative procedures, all samples were frozen on brass pins in liquid propane, stored in liquid nitrogen, cryosectioned (0.1 μm), freeze dried, and microanalyzed as previously reported.

scholarly journals Matrix Vesicles: Role in Bone Mineralization and Potential Use as Therapeutics

2021 ◽  
Vol 14 (4) ◽  
pp. 289
Author(s):  
Sana Ansari ◽  
Bregje W. M. de de Wildt ◽  
Michelle A. M. Vis ◽  
Carolina E. de de Korte ◽  
Keita Ito ◽  
...  
Keyword(s):  
Bone Formation ◽  
Bone Regeneration ◽  
Bone Cells ◽  
Bone Mineralization ◽  
Recipient Cell ◽  
Organic Matrix ◽  
Matrix Vesicles ◽  
Matrix Mineralization

Bone is a complex organ maintained by three main cell types: osteoblasts, osteoclasts, and osteocytes. During bone formation, osteoblasts deposit a mineralized organic matrix. Evidence shows that bone cells release extracellular vesicles (EVs): nano-sized bilayer vesicles, which are involved in intercellular communication by delivering their cargoes through protein–ligand interactions or fusion to the plasma membrane of the recipient cell. Osteoblasts shed a subset of EVs known as matrix vesicles (MtVs), which contain phosphatases, calcium, and inorganic phosphate. These vesicles are believed to have a major role in matrix mineralization, and they feature bone-targeting and osteo-inductive properties. Understanding their contribution in bone formation and mineralization could help to target bone pathologies or bone regeneration using novel approaches such as stimulating MtV secretion in vivo, or the administration of in vitro or biomimetically produced MtVs. This review attempts to discuss the role of MtVs in biomineralization and their potential application for bone pathologies and bone regeneration.

scholarly journals LOSS OF PROTEINPOLYSACCHARIDES AT SITES WHERE BONE MINERALIZATION IS INITIATED

1972 ◽  
Vol 20 (4) ◽  
pp. 279-292 ◽  
Author(s):  
D. BAYLINK ◽  
J. WERGEDAL ◽  
E. THOMPSON
Keyword(s):  
Calcium Concentration ◽  
Bone Mineralization ◽  
Bone Matrix ◽  
Indirect Evidence ◽  
Experimental Conditions ◽  
Acid Polysaccharide ◽  
Major Acid ◽  
Tibial Cortex

In both ground sections and demineralized frozen sections of the rat tibial cortex, osteoid but not mature bone matrix stained for proteinpolysaccharides with the Alcian Blue and toluidine blue techniques. The loss of proteinpolysaccharide staining occurred precisely at the mineralizing front, which was identified by in vivo lead or procion markers, not only in normal animals but also in animals in which osteoid width was either increasing or decreasing. In vitro, both proteases and saccharidases abolished proteinpolysaccharide staining of osteoid. Critical electrolyte concentration and other procedures indicated that the major acid polysaccharide component in osteoid is chondroitin sulfate. Consistent with these findings, electron microprobe analyses revealed that sulfur concentration was high in osteoid but dropped abruptly as calcium concentration increased at the mineralizing front. The precise synchronization between loss of proteinpolysaccharides and onset of mineralization under various experimental conditions provides strong indirect evidence that the loss of these macromolecules is somehow involved in initiation of mineralization in bone.

N tau-methylhistidine release: contributions of rat skeletal muscle, GI tract, and skin

1982 ◽  
Vol 243 (4) ◽  
pp. E293-E297 ◽  
Author(s):  
S. J. Wassner ◽  
J. B. Li
Keyword(s):  
Skeletal Muscle ◽  
Gastrointestinal Tract ◽  
Soft Tissue ◽  
Fractional Catabolic Rate ◽  
Gi Tract ◽  
Rat Skeletal Muscle ◽  
Catabolic Rate ◽  
Total Body

The relative contributions of skeletal muscle, gastrointestinal tract, and skin to urinary N tau-methylhistidine (MH) excretion were estimated during in vitro studies using the rat hemicorpus preparation. After 0.5 h of perfusion, MH release into the perfusate was linear for 3 h and averaged 29.8 nmol . h-1 . 100 g hemicorpus-1. In vivo, 24-h urinary MH excretion averaged 37.3 nmol . h-1 . 100 g body wt-1. The ratio of soft tissue to skin weight is equal (3.2:1) in the whole rat and in the hemicorpus. The gastrointestinal tract released 16.0 nmol . h-1 . 100 g body wt-1 or approximately 41% of the total urinary MH excretion. Preparations perfused with or without skin showed modest differences in the rate of MH release that were not statistically significant. Skeletal muscle contains 89.8% of total body MH content, whereas gastrointestinal tract and skin contain 3.8 and 6.4%, respectively. Gastrointestinal tract actomyosin turns over rapidly with a fractional catabolic rate of 24%/day versus 1.4%/day for skeletal muscle actomyosin.

Changes in serum triiodothyronine kinetics and hepatic type I 5'-deiodinase activity of cold-exposed swine

1994 ◽  
Vol 266 (5) ◽  
pp. E786-E795 ◽  
Author(s):  
H. L. Reed ◽  
M. Quesada ◽  
R. L. Hesslink ◽  
M. M. D'Alesandro ◽  
M. T. Hays ◽  
...  
Keyword(s):  
Elevated Serum ◽  
Total Body Weight ◽  
Weight Percent ◽  
Type I ◽  
Free T4 ◽  
Gland Weight ◽  
Total Triiodothyronine ◽  
Total Thyroxine

Swine exposed to cold air have elevated serum values of total triiodothyronine (TT3) and free T3 (FT3). To characterize the mechanism of these increases, we measured in vivo kinetic parameters after a bolus intravenous injection of 125I-labeled T3 by use of both multicompartmental (MC) and noncompartmental (NC) methods and in vitro hepatic type I iodothyronine 5'-deiodinase (5'D-I) activity. Ten ad libitum-fed 5-mo-old boars were divided into two groups, living for 25 days in either control (22 degrees C) or cold (4 degrees C) conditions. Cold-exposed animals consumed 50% more calories than control animals but showed no difference in total body weight, percent body fat, or plasma volume. Thyroid gland weight was increased 86% (P < 0.004), as was serum total thyroxine (TT4) (48%), free T4 (FT4) (61%), TT3 (103%), and FT3 (107%), whereas serum thyrotropin (TSH) was not different in cold-exposed compared with control animals. The T3 plasma clearance rate was similar between groups when both MC and NC techniques were used. However, T3 plasma appearance rate (PAR) was elevated in cold-treated animals 110% over controls by MC (P < 0.001) and 83% by NC methods (P < 0.001). The animal total hormone pool of T3 was increased 76% (MC) and 53% (NC) compared with control (P < 0.01). The Michaelis constant of hepatic 5'D-I was not different between groups, but the maximum enzyme velocity increased (106%; P < 0.02). Therefore cold exposure for 25 days is associated with increased energy intake, thyroid size, T3 PAR, and hepatic 5'D-I activity with little change in serum TSH.

scholarly journals Potential of gas-assisted time-of-flight secondary ion mass spectrometry for improving the elemental characterization of complex metal-based systems

2020 ◽  
Vol 35 (12) ◽  
pp. 2997-3006
Author(s):  
Agnieszka Priebe ◽  
Tianle Xie ◽  
Laszlo Pethö ◽  
Johann Michler
Keyword(s):  
Mass Spectrometry ◽  
Chemical Analysis ◽  
Secondary Ion Mass Spectrometry ◽  
High Sensitivity ◽  
Elemental Distribution ◽  
Tof Sims ◽  
Molecular Information ◽  
Secondary Ion ◽  
Elemental Characterization

Enhancing the spatial resolution of TOF-SIMS, which provides 3D elemental distribution in combination with high sensitivity and molecular information, is currently one of the hottest topics in the field of chemical analysis at the nanoscale.

In vivo and in vitro trimethadione oxidation activity of the liver from various animal species including mouse, hamster, rat, rabbit, dog, monkey and human

1999 ◽  
Vol 18 (1) ◽  
pp. 12-16 ◽  
Author(s):  
E Tanaka ◽  
A Ishikawa ◽  
T Horie
Keyword(s):  
Animal Species ◽  
In Vitro Study ◽  
Metabolic Clearance ◽  
Oxidation Activity ◽  
In Vitro Characterization ◽  
Demethylase Activity ◽  
Total Body

Trimethadione (TMO) has the properties required of a probe drug for the evaluation of hepatic drug-oxidizing capacity and, in this study, we have summarized the in vivo and in vitro metabolism of TMO in various animal species including mouse, hamster, rat, rabbit, dog, monkey and human. In the in vivo study, the plasma TMO level was measured after intravenous or oral (human) administration of TMO at a dose of 4 mg/kg to various animal species. The rate of TMO metabolic clearance in these animal species in vivo was in the order mouse > hamster >rat>rabbit>dog>monkey>human. In the in vitro study, species differences were observed in the cytochrome P450 (P450) content and drug-oxidizing enzyme activity. The content of P450 was monkey> mouse>dog>rabbit>hamster>rat>human. On the other hand, TMO N-demethylation was in the order mouse >hamster >rat >rabbit>dog>monkey>human. There was a good correlation between the mean total body clearance of TMO ( in vivo)andthemeanTMON-demethylase activity ( in vitro) (y=1.7×+0.11, r=0.965, P<0.001). These results show that TMO is a probe agent with metabolic and pharmacokinetic characteristics making it attractive for the in vivo and in vitro characterization of metabolic activity in various animal species.

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