Age and gender-dependent bone density changes of the human skull disclosed by high-resolution flat-panel computed tomography.

INTRODUCTION: The objective of this article was to estimate the age at death in forensic or anthropologic applications based on human skull investigation. Sex-dependent differences were analyzed. METHODS: Digital, high-resolution, flat-panel-based volumetric computed tomography (eXplore Locus Ultra scanner) images (165,920) of 244 European human skulls-163 males, 81 females-were analyzed according to their radiological bone density, based on Hounsfield units (H) that are directly related to the x-ray attenuation of the scanned material. Data were collected by the Department of Forensic Medicine at the University Hospital Giessen and Marburg during 2007 and 2008. Correlation analysis was used for data description. RESULTS: Human skull density estimates are widely scattered as a function of age for both sexes. Male skull bone density remains constant during lifetime, whereas female skull bone density decays slowly from approximately 20 years onwards. CONCLUSIONS: Bone density decay only theoretically provides a new method to determine age at death for adult females. Due to the scattering of the data, an accuracy of approximately ±18 years is found at a confidence interval of 75%, which is, unfortunately, of limited practical interest. We found new sex differences of bone density decay in the skull that are potentially of relevance for the general understanding of bone degradation processes.

High-resolution flat-panel volumetric CT images show no correlation between human age and sagittal suture obliteration--independent of sex.

This study investigated whether digital, high-resolution CT-images of the internal human sagittal suture structure include information that enables a novel method of age at death (aad) determination. To accomplish this, coronal, flat-panel-based volumetric computed tomography (eXplore Locus Ultra scanner) images were automatically analyzed by a software implementation of an algorithm that determines user independent whether a suture is open or closed. 29,205 images of the local vicinity of the sagittal suture of 164 males and 85 females of European descent were investigated separately for both sexes. We used conditional probabilities and a chi(2)-test to investigate whether there is a correlation between aad and suture obliteration or not. The computer-aided analysis enables us to handle huge volumes of data that could not otherwise be analyzed within a reasonable time frame. The implemented algorithm ensured a strongly reproducible, reliable, accurate, and fast differentiation between closed and open sutures. The evaluation of various statistical parameters suggests that there is no reason to assume a correlation between age and suture closure--with equal findings for both sexes. Therefore, we conclude that determination of aad based on the evaluation of sagittal suture obliteration is not possible. This agrees well - thus unsatisfactorily - with the recent literature.

Ossification degrees of cranial sutures determined with flat-panel computed tomography: narrowing the age estimate with extrema.

Since Broca's time (1824-1880), ossification of the neurocranial sutures has been used as a characteristic of age. Current approaches include the visual macroscopic examination of ecto and endocranial sutures. The evaluation of the cross-section of sutures usually necessitates the destruction of the neurocranium. In a nondestructive alternative approach that was tested within the context of the "Digital Forensic Osteology" project that ran in cooperation with the Virtopsy-Project, it emerged that the resolution of conventional multi-slice computed tomography data sets was not high enough to image sutures. Thus for the experiments presented here, the eXplore Locus Ultra flat-panel computed tomography scanner from GE Healthcare was used. Calottes were scanned during autopsy and then immediately returned to the corpse. So far, the skullcaps of 221 individuals have been scanned. The cross-sections of 14 suture segments could be assessed for seven previously defined stages of ossification. In a converse step, the 14 highest and lowest age estimate values corresponding to the individual stages of suture closure found were estimated for each calotte. The obtained ranges narrowing down the age estimate were evaluated with statistics. A mean value of 43.31 years for the range of narrowed age estimates shows that this method can be a useful aid in estimating age. The results of intra- and inter-observer tests showed good overall agreement between the findings of three observers. This method is suitable for a nondestructive age estimation and can be used for the entire calotte.

Estimating age by assessing the ossification degree of cranial sutures with the aid of Flat-Panel-CT.

Estimating age from the skeletons of adults is difficult. The accuracy decreases for old age. Ossification of the neurocranial sutures has been used as an age characteristic since Broca's time (1824-1880). Although the sutures traverse the bone, current approaches only encompass the ecto- and endocranial sutures. Evaluating the cross-section of sutures necessitates the destruction of the neurocranium. In the context of the "Digital Forensic Osteology" project that ran in cooperation with the Virtopsy-Project it emerged that the resolution of conventional MSCT-data sets was not high enough to image sutures. The eXplore Locus Ultra (eLU) Flat-Panel-CT-Scanner from GE was used for these experiments. During autopsy, the skullcap was scanned and then immediately returned to the corpse. To date, the skullcaps of 221 individuals have been scanned. The data sets were reconstructed in 3D. The cross sections of the sutures of 14 segments could be evaluated in the cross-sectional view. Seven stages of ossification were defined and each segment was assessed for these. Several regression formulae for age estimation were developed from the results. This examination method is a suitable means for non-invasively evaluating the ossification degree of cranial sutures in the entire cross-section and for the entire calvarium. An increase in the number of examined individuals in this ongoing project and a look at extreme values will further heighten the conclusiveness of the results.

Monitoring of experimental rat lung transplants by high-resolution flat-panel volumetric computer tomography (fpVCT).

OBJECTIVE: Noninvasive assessment of experimental lung transplants with high resolution would be favorable to exclude technical failure and to follow up graft outcome in the living animal. Here we describe a flat-panel Volumetric Computed Tomography (fpVCT) technique using a prototype scanner. METHODS: Lung transplantation was performed in allogeneic as well as in corresponding syngeneic rat strain combinations. At different time points post-transplantation, fpVCT was performed. RESULTS: Lung transplants can be visualized in the living rat with high-spatial resolution. FpVCT allows a detailed analysis of the lung and the bronchi. Infiltrates developing during rejection episodes can be diagnosed and follow-up studies can easily be performed. CONCLUSIONS: With fpVCT it is possible to control the technical success of the surgical procedure. Graft rejection can be visualized individually in the living animal noninvasively, which is highly advantageous for studying the pathogenesis of chronic rejection or to monitor new therapies.

Evaluation of angiogenesis using micro-computed tomography in a xenograft mouse model of lung cancer.

Quantitative evaluation of lung tumor angiogenesis using immunohistochemical techniques has been limited by difficulties in generating reproducible data. To analyze intrapulmonary tumor angiogenesis, we used high-resolution micro-computed tomography (micro-CT) of lung tumors of mice inoculated with mouse Lewis lung carcinoma (LLC1) or human adenocarcinoma (A549) cell lines. The lung vasculature was filled with the radiopaque silicone rubber, Microfil, through the jugular vein (in vivo application) or pulmonary artery (ex vivo application). In addition, human adenocarcinoma lung tumor-bearing mice treated site-specifically with humanized monoclonal antibody (bevacizumab) against vascular endothelial growth factor. Quantitative analysis of lung tumor microvessels imaged with micro-CT showed that more vessels (mainly small, <0.02 mm(2)) were filled using the in vivo (5.4%) compared with the ex vivo (2.1%) method. Furthermore, bevacizumab-treated lung tumor-bearing mice showed significantly reduced lung tumor volume and lung tumor angiogenesis compared with untreated mice as assessed by micro-CT. Interestingly, microvascularization of mainly the smaller vessels (<0.02 mm(2)) was reduced after bevacizumab treatment. This observation with micro-CT was nicely correlated with immunohistochemical measurement of microvessels. Therefore, micro-CT is a novel method for investigating lung tumor angiogenesis, and this might be considered as an additional complementary tool for precise quantification of angiogenesis.

Treatment with keratinocyte growth factor does not improve lung allograft survival in the rat.

PURPOSE: Lung allografts are threatened by primary graft dysfunction, infections, and rejection. Novel therapies protecting pulmonary allografts are badly needed. Keratinocyte growth factor (KGF) protects the lung against a variety of injurious stimuli and exerts anti-inflammatory effects. The aim of the study was to test the potential of recombinant truncated KGF (DeltaN23-KGF, palifermin) to attenuate pulmonary allograft rejection. MATERIALS AND METHODS: Intratracheal instillation of 5 mg/kg DeltaN23-KGF was performed twice in donor rats on days 3 and 2 before explantation of the lung. In control animals, an equivalent volume of vehicle was instilled. Left lungs were transplanted in the fully allogeneic Dark Agouti to Lewis rat strain combination and in the less stringent Fischer 344 to Wistar Kyoto combination. Allograft recipients were additionally treated with DeltaN23-KGF post-transplantation. Graft outcome, leukocytic infiltration, and major histocompatibility complex (MHC) class II antigen expression was analyzed. RESULTS: In both rat strain combinations, DeltaN23-KGF treatment did not improve pulmonary allograft outcome. Graft infiltration by macrophages and T lymphocytes remained unchanged. In addition, we demonstrated that MHC class II antigens were more abundant in KGF-treated allografts compared to control-treated grafts, which probably results in an increased alloreactivity. CONCLUSION: In conclusion, intratracheal DeltaN23-KGF treatment is not effective to prevent acute pulmonary allograft rejection.

Characterization of a murine model of monocrotaline pyrrole-induced acute lung injury.

BACKGROUND: New animal models of chronic pulmonary hypertension in mice are needed. The injection of monocrotaline is an established model of pulmonary hypertension in rats. The aim of this study was to establish a murine model of pulmonary hypertension by injection of the active metabolite, monocrotaline pyrrole. METHODS: Survival studies, computed tomographic scanning, histology, bronchoalveolar lavage were performed, and arterial blood gases and hemodynamics were measured in animals which received an intravenous injection of different doses of monocrotaline pyrrole. RESULTS: Monocrotaline pyrrole induced pulmonary hypertension in Sprague Dawley rats. When injected into mice, monocrotaline pyrrole induced dose-dependant mortality in C57Bl6/N and BALB/c mice (dose range 6-15 mg/kg bodyweight). At a dose of 10 mg/kg bodyweight, mice developed a typical early-phase acute lung injury, characterized by lung edema, neutrophil influx, hypoxemia and reduced lung compliance. In the late phase, monocrotaline pyrrole injection resulted in limited lung fibrosis and no obvious pulmonary hypertension. CONCLUSION: Monocrotaline and monocrotaline pyrrole pneumotoxicity substantially differs between the animal species.

High resolution imaging of craniofacial bone specimens by flat-panel volumetric computed tomography.

INTRODUCTION: The prototype flat-panel volumetric computed tomograph (fpvCT) provides a new 3D imaging technology with detailed high resolution by using large-area flat-panel X-ray detectors. The object of this study was to evaluate the benefit of high resolution imaging using the experimental fpvCT to visualise different types of human craniofacial bone pathology. The study proved the feasibility of performing an intraoperative evaluation of free margins in bone malignancies using fpvCT. MATERIAL AND METHODS: In this study, 35 bone specimens of various pathological types were examined by fpvCT. fpvCT data were compared with pre-operative multislice clinical CT images as well as with post-operative histological findings. RESULTS: Bone tumours can be visualised with their specific pathological architecture and infiltration structure faster and more precisely by fpvCT than by multislice CT. The analysis of the resection margins supports the surgical procedure intraoperatively, especially when an immediate reconstruction with bone transplantation is carried out. DISCUSSION: The fpvCT has a superior image quality when compared with clinical CT systems. The imaging of the bone structure itself has been shown to be useful for the interpretation of osseous resection borders. Furthermore, it can facilitate the diagnosis of tumour progression, especially in areas that are difficult to access, such as the base of the skull.

A combination hybrid-based vaccination/adoptive cellular therapy to prevent tumor growth by involvement of T cells.

Cancer immunotherapy with dendritic cell-tumor cell fusion hybrids induces polyclonal stimulation against a variety of tumor antigens, including unknown antigens. Hybrid cells can prime CTLs, which subsequently develop antitumor responses. The aim of this study was to enhance the known antitumor effect of hybrid vaccination (HC-Vacc) and hybrid-primed adoptive T-cell therapy (HC-ACT) using the poorly immunogenic Lewis lung carcinoma (LLC1) model. The strategy used was a combination of a double HC-Vacc alternating with HC-ACT (HC-Vacc/ACT). Using flat-panel volumetric computer tomography and immunohistochemistry, we showed a significant retardation of tumor growth (85%). In addition, a significant delay in tumor development, a reduction in the number of pulmonary metastases, and increased survival times were observed. Furthermore, the tumors displayed significant morphologic changes and increased apoptosis, as shown by up-regulation of gene expression of the proapoptotic markers Fas, caspase-8, and caspase-3. The residual tumor masses seen in the HC-Vacc/ACT-treated mice were infiltrated with CD4+ and CD8+ lymphocytes and showed elevated IFNgamma expression. Moreover, splenic enlargement observed in HC-Vacc/ACT-treated mice reflected the increased functionality of T cells, as also indicated by increased expression of markers for CTL activation, differentiation, and proliferation (Cd28, Icosl, Tnfrsf13, and Tnfsf14). Our findings indicate that the combination therapy of dendritic cell-tumor cell HC-Vacc/ACT is a very effective and a promising immunotherapeutic regimen against poorly immunogenic carcinomas.

Non-invasive screening of lung nodules in mice comparing a novel volumetric computed tomography with a clinical multislice CT.

In vivo imaging of small animal models will play an increasingly important role in cancer research, as new imaging systems that employ non-invasive protocols and offer high-resolution capability become available. A flat-panel volumetric computed tomograph (fpvCT) was evaluated to determine if minimally invasive protocols can be used to provide the spatial resolution required for lung imaging in small animals. The detection of small pulmonary nodules in a Lewis carcinoma model was investigated, and fpvCT was compared with a multislice computed tomograph (MSCT). Five C57/BL6 mice with Lewis lung carcinoma were monitored with both modalities over two weeks. Sensitivity of the systems was measured by comparing the results with histology, and the incidence of first visualization of the tumors in the two systems was determined. Compared to MSCT, fpvCT proved its superior sensitivity in detection of lung nodules. Due to its isotropic resolution and a significant reduction of partial volume effects, early detection and reasonable determination of growth in very small tumors was only possible with fpvCT. fpvCT is a high-resolution imaging system that proved its ability to perform in vivo monitoring of a pulmonary lung tumor model in mice. This permits longitudinal investigations in small animals for cancer research.

Potential applications of flat-panel volumetric CT in morphologic and functional small animal imaging.

Noninvasive radiologic imaging has recently gained considerable interest in basic and preclinical research for monitoring disease progression and therapeutic efficacy. In this report, we introduce flat-panel volumetric computed tomography (fpVCT) as a powerful new tool for noninvasive imaging of different organ systems in preclinical research. The three-dimensional visualization that is achieved by isotropic high-resolution datasets is illustrated for the skeleton, chest, abdominal organs, and brain of mice. The high image quality of chest scans enables the visualization of small lung nodules in an orthotopic lung cancer model and the reliable imaging of therapy side effects such as lung fibrosis. Using contrast-enhanced scans, fpVCT displayed the vascular trees of the brain, liver, and kidney down to the subsegmental level. Functional application of fpVCT in dynamic contrast-enhanced scans of the rat brain delivered physiologically reliable data of perfusion and tissue blood volume. Beyond scanning of small animal models as demonstrated here, fpVCT provides the ability to image animals up to the size of primates.

Analysis of tumor vessel supply in Lewis lung carcinoma in mice by fluorescent microsphere distribution and imaging with micro- and flat-panel computed tomography.

In lung carcinomas the blood supply varies depending on tumor type and stage and can develop from pulmonary or bronchial circulation, or both. To examine this in vivo, primary bronchogenic Lewis lung carcinoma cells were intratracheally instilled in C57BL/6 mice. Within 7 days, histological examinations showed progressive tumor growth at the peripheral parenchymal region. The relative contribution of tumor blood supply via the pulmonary and systemic arteries was studied in detail using fluorescent microspheres (10 microm). When compared to healthy lung parenchyma (13:1), Lewis lung carcinoma tumor tissue (52:1) showed a fourfold increase in pulmonary to systemic microspheres, indicating that the pulmonary arteries are the predominant tumor-feeding vessels. After filling the vessels with a vascular cast, the microanatomy of vessels being derived from the pulmonary artery was visualized with micro computed tomography. Flat-panel volumetric computed tomography provided longitudinal visualization of tissue bridges between the growing tumor and the pulmonary vasculature. In this model of peripheral parenchymal malignancy, new imaging techniques allowed effective visualization of lung tumor growth and vascularization in living mice, demonstrating a pulmonary blood supply for lung tumors.

Flat-panel volumetric computed tomography: a new method for visualizing fine bone detail in living mice.

In this report, we present a new noninvasive 3-dimensional (3D) imaging technology for in vivo monitoring of the skeletal development of mice: flat-panel volumetric Computed Tomography (fpvCT). Long-term investigations of 4 mice are presented, with up to 14 scans of each mouse from postnatal day 0 to 86. Examinations of a newborn and an adult mouse, performed with fpvCT and clinical multislice CT (MSCT), demonstrate the superior image quality of high-resolution fpvCT.

Identification of embolic stroke patterns by diffusion-weighted MRI in clinically defined lacunar stroke syndromes.

BACKGROUND: A number of clinical syndromes describing the presentation of deep brain infarcts are called lacunar syndromes resulting from small vessel occlusion (SVO). To verify the reliability of the clinical diagnosis "lacunar syndrome," the value was investigated with diffusion-weighted MRI (DWI). METHODS AND RESULTS: A total of 73 patients (mean age 66 years; range 35 to 83 years) with sudden onset of a classical lacunar syndrome were enrolled. On the basis of the DWI findings, patients were divided into 3 groups: group 1, single subcortical lesion (<15-mm lesion; 43 patients; 59%); group 2, large (> or =15 mm) or scattered lesions in 1 vascular territory (16 patients; 22%); and group 3, multiple lesions in multiple vascular territories (14 patients; 19%). A stroke mechanism other than SVO could be identified in 17 (23%) patients. Cardiac work-up revealed a cardiac embolic source in 8 patients (11%). Duplex sonography revealed symptomatic stenosis in 9 patients (12%). Based on the work-up information, 29 patients (40%) were found to have a potential cause of stroke other than SVO. A significant correlation with >1 single lesion on DWI-MRI and a clinical proven embolic source was observed (P=0.002). In 9 patients with MRI suspicious for a pathomechanism other than SVO, no embolic source was found. CONCLUSIONS: The use of DWI-MRI improves the accuracy of the subtype diagnosis of stroke. Inaccuracy has to be expected in approximately one third if lacunar diagnosis is based on clinical and computed tomography findings. Most of these "false-positive" cases are attributable to large artery or cardiogenic embolic stroke.

Volumetric computed tomography (VCT): a new technology for noninvasive, high-resolution monitoring of tumor angiogenesis.

Volumetric computed tomography (VCT) is a technology in which area detectors are used for imaging large volumes of a subject with isotropic imaging resolution. We are experimenting with a prototype VCT scanner that uses flat-panel X-ray detectors and is designed for high-resolution three-dimensional (3D) imaging. Using this technique, we have demonstrated microangiography of xeno-transplanted skin squamous cell carcinomas in nude mice. VCT shows the vessel architecture of tumors and animals with greater detail and plasticity than has previously been achieved, and is superior to contrast-enhanced magnetic resonance (MR) angiography. VCT and MR images correlate well for larger tumor vessels, which are tracked from their origin on 3D reconstructions of VCT images. When compared with histology, small tumor vessels with a diameter as small as 50 microm were clearly visualized. Furthermore, imaging small vessel networks inside the tumor tissue improved discrimination of vital and necrotic regions. Thus, VCT substantially improves imaging of vascularization in tumors and offers a promising tool for preclinical studies of tumor angiogenesis and antiangiogenic therapies.

Interferon-beta 1b leads to a short-term increase of soluble but long-term stabilisation of cell surface bound adhesion molecules in multiple sclerosis.

Adhesion molecules (AMs) are believed to regulate the transmigration of blood leukocytes across the blood-brain barrier (BBB), which is an essential step in the pathogenesis of multiple sclerosis (MS). Previous studies have investigated changes of the soluble forms of AM during interferon-beta1b (IFN-beta1b) treatment in MS patients. In this study, we analysed the influence of IFN-beta1b treatment on the cell surface bound forms of the AMs cICAM-1 and cICAM-3 on blood mononuclear cells (MNC). Sixty-eight patients with relapsing-remitting MS were enrolled in this open study; thirty of them were treated with IFN-beta1b. Blood samples were collected every three months over a period of 18 months. The expression levels of cell surface bound forms of AM on blood MNC were measured by two colour flow cytometry analysis. sVCAM-1, sICAM-1 and sICAM-3 were determined by ELISA. We found a short-term induction effect on the serum concentrations of sICAM-1 and sVCAM-1 after three months of IFN-beta1b treatment. The expression levels of cell surface bound AMs on blood MNC remained stable during treatment. Untreated MS patients, however, showed a continuous decrease in the expression of cell surface bound AM expression over 18 months. Stabilisation of the expression of cell surface bound AMs on blood MNC may indicate the beneficial effects of IFN-beta1b therapy in MS patients.