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Quantitative microcomputed tomography analysis of mineralization within three-dimensional scaffolds in vitro.

Synthetic and naturally derived scaffold biomaterials in combination with osteogenic cells or bioactive factors have the potential to serve as bone graft substitutes. Porous poly(l-lactide-co-dl-lactide) (PLDL) scaffolds with mechanical properties comparable to trabecular bone and an oriented, interconnected porosity designed to enhance internal mass transport were recently developed. In this study, PLDL scaffolds were seeded with rat calvarial or rat stromal cells and cultured up to 8 weeks in media containing osteogenic supplements. Cell-seeded human demineralized trabecular bone matrix (DTBM) scaffolds were included for comparison. All constructs were imaged weekly from 4 to 8 weeks using microcomputed tomography (micro-CT) to nondestructively quantify the amount and distribution of mineralized matrix formation. The total mineralized matrix volume increased with time in culture for all construct groups. DTBM constructs contained significantly more mineralized matrix than PLDL constructs. However, an analysis of the acellular DTBM scaffolds exposed to osteogenic media revealed partial remineralization of the demineralized matrix whereas no mineralization was detected in acellular PLDL scaffolds. Differences in mineral distribution were also evident with cell-mediated mineralization found throughout the PLDL constructs but localized to the periphery of the DTBM constructs for both cell types. Expression of bone marker genes indicating osteoblast differentiation was demonstrated in all groups at 8 weeks using a quantitative reverse transcription polymerase chain reaction. Osteocalcin expression was significantly higher for calvarial cell constructs compared to stromal cell constructs, regardless of the type of scaffold. This study demonstrated that micro-CT imaging may be used to nondestructively and quantitatively monitor mineralization within three-dimensional scaffolds in vitro. PLDL scaffolds with an oriented microarchitecture were shown to support cell attachment, differentiation, and cell-mediated mineralization comparable to natural DTBM scaffolds.

Animals↗

Optimal segmentation of microcomputed tomographic images of porous tissue-engineering scaffolds.

The morphometric properties of the porous tissue-engineering scaffolds play a dominant role in the initial cell attachment and subsequent tissue regeneration. These properties can be derived nondestructively with the use of quantitative analysis of high-resolution microcomputed tomography (microCT) imaging of scaffolds. Accurate segmentation of these acquired images into solid and porous subspaces is critical to the integrity of morphometric analysis. The absence of a single image-processing technique to provide such accurate separability immune to all the intricacies of the acquired data makes this seemingly simple task significantly error prone. Consequently, an optimal segmentation has to be selected by ranking the segmentations produced by a multiplicity of methods. This article proposes a robust, easy-to-implement, unambiguous, signal-processing-based, ground-truth-free, segmentation rating metric that correlates with visual acuity. With the use of this metric it is possible, for the first time, to threshold the data with a wide range of techniques and select automatically the technique that best delineates the acquired image. The proposed solution has been extensively tested on microCT images of scaffolds fabricated with biodegradable poly (propylene fumarate) (PPF) with the use of a solvent casting particulate leaching process. The approaches proposed and the results obtained may have profound implications for accurate image-based characterization of tissue-engineering scaffolds.

Biocompatible Materials↗

Novel approach for quantification of porosity for biomaterial implants using microcomputed tomography (microCT).

Porous bioceramics have been widely investigated in orthopaedic tissue engineering. Attention has been given to manufacturing of a porous bioceramic that mimics the trabecular bone structure for proper bone regeneration. With the advance of biomedical imaging through microcomputed tomography (microCT), this study attempted to quantify the pore structure of different bioceramics. Two bioceramic blocks (BSC and ChronOS) were synthesized by two methods. The specification claimed the porosity of the bioceramic ranged from 40% to 70%. Six blocks of each bioceramic were evaluated by conventional water immersion method and microCT. The pore size and connectivity were evaluated with standardized protocols. By the water immersion method, the porosity of BSC and ChronOS was 60.4% and 74.7%, respectively. The three-dimensional results of microCT showed that BSC porosity was 26.2% and ChronOS was 60.0%. The pore connectivity was evaluated to be 2.6 for BSC and 39.7 for ChronOS. ChronOS had functional pores with 200 microm to 400 microm in diameter (87.8%+/-0.5%), which is significantly more than 52.8%+/-11.5% of pores in BSC (p<0.05). Providing information on the functional pores objectively, the microCT evaluation serves as a good standard for specification of the bioceramic-related implants.

Coated Materials, Biocompatible↗

Reporting results from a clinical pharmacological laboratory using a microcomputer.

Responsibility lies with an analytical pharmacological laboratory to issue each result with an assessment of its validity for the corresponding patient. This is scarcely possible by the usual manual methods of data handling even when complete diagnostic and pharmacokinetic data are available, because of the staff time required. Use of a microcomputer running a suite of True Basic programs showing "artificial intelligence" solves the temporal difficulty and leads to the issue of more incisive, detailed reports.

Clinical Laboratory Information Systems↗

Microcomputer voice-response telephone entry for balanced clinical trial randomization.

Simple randomization of patients to treatment regimens in clinical trials can result in inbalance among stratum categories. Procedures that produce balance may lack randomness. Systems that are not deterministic yet produce balance are often extremely complex and are not easily performed by randomization clerks. A microcomputer-based voice-response system has been implemented that permits clinic staff to easily randomize patients in clinical trials despite the complexity of the algorithm. Data entry is accomplished by the use of Touch-Tone keys. Input prompts and regimen assignment are spoken by a commercially available voice synthesizer.

Computers↗

Risedronate preserves bone architecture in early postmenopausal women in 1 year as measured by three-dimensional microcomputed tomography.

Risedronate reduces the risk of vertebral fractures by up to 70% within the first year of treatment. Increases in bone mineral density or decreases in bone turnover markers explain only a portion of the anti-fracture effect, suggesting that other factors, such as changes in trabecular bone architecture, also play a role. Our objective was to determine the effects of risedronate on bone architecture by analyzing iliac crest bone biopsy specimens using three-dimensional microcomputed tomography (3-D micro CT). Biopsy specimens were obtained at baseline and after 1 year of treatment from women enrolled in a double-blind, placebo-controlled study of risedronate 5 mg daily for the prevention of early postmenopausal bone loss. Trabecular architecture deteriorated in the placebo group (n = 12), as indicated by a 20.3% decrease in bone volume (25.1% vs. 20.0%, P = 0.034), a 13.5% decrease in trabecular number (1.649 vs. 1.426 mm(-1), P = 0.052), a 13.1% increase in trabecular separation (605 vs. 684 microm, P = 0.056), and an 86.2% increase in marrow star volume (3.251 vs. 6.053 mm(3), P = 0.040) compared with baseline values. These changes in architectural parameters occurred in the presence of a concomitant decrease from baseline in lumbar spine bone mineral density (-3.3%, P = 0.002), as measured by dual energy x-ray absorptiometry. There was no statistically significant ( P < 0.05) deterioration in the risedronate-treated group (n = 14) over the 1-year treatment period. Comparing the actual changes between the two groups, the placebo group experienced decreases in bone volume (placebo, -5.1%; risedronate, +3.5%; P = 0.011), trabecular thickness (placebo, -20 microm; risedronate, +23 microm; P = 0.032), and trabecular number (placebo, -0.223 mm(-1); risedronate, +0.099 mm(-1); P = 0.010), and increases in percent plate (placebo, +2.79%; risedronate, -3.23%; P = 0.018), trabecular separation (placebo, +79 microm; risedronate, -46 microm; P = 0.010) and marrow star volume (placebo, +2.80 mm(3); risedronate, -2.08mm(3); P = 0.036), compared with the risedronate group. These data demonstrate that trabecular architecture deteriorated significantly in this cohort of early postmenopausal women, and that this deterioration was prevented by risedronate. Although there is no direct link in this study between fracture and preservation of architecture, it is reasonable to infer that the preservation of bone architecture may play a role in risedronate's anti-fracture efficacy.

Absorptiometry, Photon↗

Quantification of trabecular bone structure using magnetic resonance imaging at 3 Tesla--calibration studies using microcomputed tomography as a standard of reference.

The purpose of this study is to use high-resolution magnetic resonance (MR) imaging at 3 Tesla (3T) to quantify trabecular bone structure in vitro using femoral head specimens, and to correlate the calculated structure measures with those that were determined using microcomputed tomography (microCT), the standard of reference. Fifteen cylindrical cores were obtained from fresh femoral heads after total hip arthroplasty. MR images were obtained at 3T using a transmit-receive wrist coil. High-resolution coronal images were acquired using a modified three-dimensional (3D) fast-gradient echo sequence. From these data sets two-dimensional (2D) structural parameters analogous to bone histomorphometry were derived by using both mean intercept length (MIL) methods based on the plate model and the more recent model-assumption free 3D distance-transformation (DT) methods. The parameters measured by the 2D plate model-based MIL method and the DT method included apparent (App). BV/TV (bone volume/total volume), App. Tb.Th (trabecular thickness), App. Tb.Sp (trabecular separation), and App. Tb.N (trabecular number). Identical regions of interest were analyzed in the MR images and the microCT data sets, and similar structure measures were derived. The means and standard deviations of the parameters over all slices were calculated and MR-derived measures were correlated with those derived from the microCT data sets using linear regression analyses. Structure measures were overestimated with MRI, for example, the mean App. BV/TV was 0.45 for MRI and 0.20 for microT, and the slope of the graph was 1.45. App. Tb.Th was overestimated by a factor of 1.9, whereas App. Tb.Sp was underestimated; Tb.N showed the smallest effect. Correlations between the individual parameters were excellent (App. BV/TV, r2 = 0.82; App. Tb.Sp, r2 = 0.84; App. Tb.N, r2 = 0.81), except for App.Tb.Th (r2 = 0.67). The results of this study show that trabecular bone structure measures may be obtained using 3T MR imaging. These measures, although higher than the standard of reference, show a highly significant correlation with true structure measures obtained by microCT.

Aged↗

Prediction of cortical bone porosity in vitro by microcomputed tomography.

The high importance of intracortical porosity for mechanical strength of cortical bone has been established. The contribution of other parameters of microstructure such as osteon dimensions for strength is in discussion. The aim of this study was to evaluate the predictive value of microcomputed tomography (mCT) for porosity and other microstructural parameters of cortical bone in cortical bone biopsies. Femoral cortical bone specimens from the middiaphysis of 24 patients were harvested during the procedure of total hip replacement at the location where normally one hole (Ø 4.5 mm) for the relief of the intramedullary pressure is placed. In vitro intracortical porosity and bone mineral density (BMD) measurements by mCT were compared with structural parameters assessed in histological sections of the same specimens. A strong correlation was found between intracortical porosity measured by mCT and histological porosity (r = 0.95, P <0.0001). Porosity measured by mCT was also a strong predictor for other parameters describing dimensions of porous structures. BMD?1 was associated with osteonal area (r = -0.76, P <0.0001). We consider the measurement of porosity by mCT as a very potent procedure for assessing intracortical porosity and parameters related to porous structures of cortical bone nondestructively in vitro.

Aged↗

A morphometric comparison of trabecular structure of human ilium between microcomputed tomography and conventional histomorphometry.

Recently, an imaging technique using microcomputed tomography (micro-CT) has emerged as a method for nondestructively assessing the microarchitecture of unprocessed surgical bone biopsy specimens. Using micro-CT, two-dimensional (2D) axial images were obtained from undecalcified transiliac bone biopsies which were taken from 15 patients with various metabolic bone diseases. Total area, bone area, and bone perimeter were determined, from which the bone volume (BV/TV), trabecular thickness (Tb.Th), trabecular number (Tb.N), and trabecular separation (Tb.Sp) were calculated semiautomatically and instantaneously. To evaluate the validity of this technique as a useful tool, the results were compared with those obtained from conventional histomorphometry. There were significant correlations between the two techniques for all parameters, with correlation coefficients ranging from 0.759 (Tb.N, P < 0.005) to 0.949 (BV/TV, P < 0.0001). Different resolutions seem to lead to major differences in perimeter values measured by the two methods. These factors may explain why the correlation coefficients of Tb.N and Tb.Th estimated from the perimeter and area is lower than that of BV/TV. Our results show that the micro-CT based on 2D images is a useful tool for imaging and nondestructively quantifying the microarchitecture of trabecular bone in unprocessed surgical bone specimens.

Adult↗

[Cardio-pulmonary vascular system. Three-dimensional quantitative evaluation by microcomputed tomography].

In recent years microcomputed tomography (microCT) has become more and more important in basic research. Now commercial microCT scanners are available. Thus, it is very likely that this new, accurate and promising method for three-dimensional and non-destructive quantitative evaluation of intact tissues including vessels will be applied more frequently. The review provides a survey of the basic technology of microCT and its current use for high resolution three-dimensional morphometric and functional analysis within the cardio-pulmonary vascular system.

Aorta, Thoracic↗

Experimental microcomputed tomography study of the 3D microangioarchitecture of tumors.

Differences between soft X-ray imaging and microcomputed tomography (micro-CT) in the microangiographic depiction of small vessels in tumors were compared to evaluate the tumors' 3D microangioarchitecture and the progress of growth-related neovascularization. VX2 carcinomas transplanted to the auricles of 24 rabbits randomly assigned to three groups were examined after 1, 3, and 7 days. Eight rabbits without transplants were the controls. Barium sulfate was injected into the auricular artery, and conventional soft X-ray images and micro-CT microangiograms of auricle specimens were studied. Micro-CT detected vessels with diameters of less than 50 microm, and imaging from several angles clearly showed the network of tumor vessels. Moreover, micro-CT confirmed the 3D process of growth-related neovascularization. There were no significant differences between the present findings and those of our previously published microscopic study. Micro-CT should prove useful for evaluating the 3D microarchitecture of tumors and for clear imaging of tumor angiogenesis.

Angiography↗

Nucleic acid sequence analysis software for microcomputers.

It is clear that a computer-aided data control system is required for even small laboratories generating nucleic acid data. While the molecular biologist at many universities and large research institutions has access to mainframe computers and nucleic acid sequence analysis software, many find it more convenient to perform sequence analysis on microcomputers that are typically located within the investigator's laboratory and totally dedicated to sequence storage and analysis, in essence giving the investigator more personal control of analysis activities than is sometimes possible with shared mini- or mainframe computers. New programs are being written and released at an increasing rate to perform increasingly more complex and specialized analyses using small computer-based systems. This trend will undoubtedly continue, fueled by the need to manage the ever increasing quantity of sequence data.

Base Sequence↗

H-EXCRETION: a program for microcomputers to aid in calculation of acid or base excretion.

H-EXCRETION is a program written in BASIC language for microcomputers and designed to facilitate the calculation of excretion for acid or base from a tissue into the surrounding media. The program is designed in such a way to allow considerable variability in different experimental parameters. Five examples of calculation of H+ excretion in the urinary bladder of the toad Bufo marinus are presented.

Acid-Base Equilibrium↗

A 'hidden line' algorithm for 3D-reconstruction from serial sections--an extension of the NEUREC program package for a microcomputer.

An algorithm is described to generate pictures in arbitrary perspective from serially sectioned biological material. Only those parts of the object which lie on a direct line of sight to the observer are included into the computed paper-plot, while hidden lines are omitted. The program was written for an inexpensive Apple II+ microcomputer as an extension of an earlier program library (NEUREC) for three-dimensional reconstruction. Application examples of neuronal and cardiac tissues are presented.

Animals↗

Long term stability of rat sleep quantified by microcomputer analysis.

An inexpensive microcomputer system is described for the direct recording of electrographic data from animals. Using this system, electrographic data can be recorded continuously on a polygraph and simultaneously quantitated by the computer, for days or weeks. Our system quantifies the amounts of delta waves, spindle bursts, hippocampal RSA activity and movement spikes for 15 sec epochs. These electrographic data are stored by the computer and subsequently can be used to score sleep stages. We find that the computer reliably counts waves; using these data, it can then score sleep stages off-line with about 90% accuracy. With this system, we find that the minutes/hour of both SWS and REM are remarkably stable from day to day. The absolute number of delta waves, spindle bursts, trains of RSA and movement counts/hour also remain stable from day to day. This type of system should find significant application in situations where quantitation of longterm effects of drugs, diets and other environmental inputs on sleep states or EEG wave bands are of interest.

Animals↗

Microcomputer-controlled plotting of environmental health data.

A general microcomputer program has been developed which facilitates the preparation of finished plots of geographical and temporal variations in such data as: the concentrations of suspended airborne particulate matter and of metallic and organic components detected in the particulate; the incidences of several kinds of respiratory illnesses and symptoms; epidemiological information obtained from physicians, hospitals, pharmacies, and schools. Examples of plots produced by the program, detailed explanations of how it works and a complete program listing are included.

Air Pollutants↗

Kinetic analysis of bacterial clearance in mice using the ESTRIPc and KINET microcomputer programs.

Two BASIC microcomputer programs, ESTRIPc and KINET, were used to analyze the kinetics of bacterial clearance from the blood and mesenteric lymph nodes of mice. Because of the similarities between the clearance of bacteria and the clearance of drugs from tissue, blood pharmacokinetic techniques were applied to the analysis of bacterial clearance data. The ESTRIPc program, developed for pharmacokinetic analysis and modified for the study of bacterial clearance, was employed to fit the experimental data of bacterial survival versus time to a polyexponential equation with 1, 2, or 3 terms. The KINET program, written specifically for kinetic analysis of bacterial clearance, uses the biexponential equation constants derived with ESTRIPc to calculate half-life values, rate constants, and other useful kinetic parameters. The combined use of these programs permits precise comparisons of the clearance rates of different bacterial species from the blood or tissues of experimental animals.

Animals↗