Phosphodiesterase-5 (PDE-5) Inhibitors as Therapy for Cerebrovascular Dysfunction in Chronic Traumatic Brain Injury.

Multiple phase III randomized controlled trials (RCTs) for pharmacologic interventions in traumatic brain injury (TBI) have failed despite promising results in experimental models. The heterogeneity of TBI, in terms of pathomechanisms and impacted brain structures, likely contributes to these failures. Biomarkers have been recommended to identify patients with relevant pathology (predictive biomarkers) and confirm target engagement and monitor therapy response (pharmacodynamic biomarkers). Our group focuses on traumatic cerebrovascular injury as an understudied endophenotype of TBI and is validating a predictive and pharmacodynamic imaging biomarker (cerebrovascular reactivity; CVR) in moderate-severe TBI. We aim to extend these studies to milder forms of TBI to determine the optimal dose of sildenafil for maximal improvement in CVR. We will conduct a phase II dose-finding study involving 160 chronic TBI patients (mostly mild) using three doses of sildenafil, a phosphodiesterase-5 (PDE-5) inhibitor. The study measures baseline CVR and evaluates the effect of escalating sildenafil doses on CVR improvement. A 4-week trial of thrice daily sildenafil will assess safety, tolerability, and clinical efficacy. This dual-site 4-year study, funded by the Department of Defense and registered in ClinicalTrials.gov (NCT05782244), plans to launch in June 2023. Biomarker-informed RCTs are essential for developing effective TBI interventions, relying on an understanding of underlying pathomechanisms. Traumatic microvascular injury (TMVI) is an attractive mechanism which can be targeted by vaso-active drugs such as PDE-5 inhibitors. CVR is a potential predictive and pharmacodynamic biomarker for targeted interventions aimed at TMVI. (Trial registration: NCT05782244, ClinicalTrials.gov ).

Cerebrovascular Reactivity Measurement with Functional Near Infrared Spectroscopy.

Cerebrovascular reactivity (CVR) is the capacity of blood vessels in the brain to alter cerebral blood flow (either with dilation or constriction) in response to chemical or physical stimuli. The amount of reactivity in the cerebral microvasculature depends on the integrity of the capacitance vasculature and is the primary function of endothelial cells. CVR is, therefore, an indicator of the microvasculature's physiology and overall health. Imaging methods that can measure CVR are available but can be costly, and require magnetic resonance imaging centers and technical expertise. In this study, we used fNIRS technology to monitor changes of oxyhemoglobin (HbO) and deoxyhemoglobin (HbR) in the cerebral microvasculature to assess the CVR of 15 healthy controls (HC) in response to a vasoactive stimulus (inhaled 5% carbon dioxide or CO2). Our results suggest that this is a promising imaging technology that offers a non-invasive, accurate, portable, and cost-effective method of mapping cortical CVR and associated microvasculature function, resulting from a traumatic brain injury or other conditions associated with cerebral microvasculopathy.

Cerebrovascular Reactivity Measures Are Associated With Post-traumatic Headache Severity in Chronic TBI; A Retrospective Analysis.

OBJECTIVE: To characterize the relationship between persistent post-traumatic headache (pPTH) and traumatic cerebrovascular injury (TCVI) in chronic traumatic brain injury (TBI). Cerebrovascular reactivity (CVR), a measure of the cerebral microvasculature and endothelial cell function, is altered both in individuals with chronic TBI and migraine headache disorder (Amyot et al., 2017; Lee et al., 2019b). The pathophysiologies of pPTH and migraine are believed to be associated with chronic microvascular dysfunction. We therefore hypothesize that TCVI may contribute to the underlying migraine-like mechanism(s) of pPTH. MATERIALS AND METHODS: 22 moderate/severe TBI participants in the chronic stage (>6 months) underwent anatomic and functional magnetic resonance imaging (fMRI) scanning with hypercapnia gas challenge to measure CVR as well as the change in CVR (ΔCVR) after single-dose treatment of a specific phosphodiesterase-5 (PDE-5) inhibitor, sildenafil, which potentiates vasodilation in response to hypercapnia in impaired endothelium, as part of a Phase2a RCT of sildenafil in chronic TBI (NCT01762475). CVR and ΔCVR measures of each participant were compared with the individual's pPTH severity measured by the headache impact test-6 (HIT-6) survey. RESULTS: There was a moderate correlation between HIT-6 and both CVR and ΔCVR scores [Spearman's correlation = -0.50 (p = 0.018) and = 0.46 (p = 0.03), respectively], indicating that a higher headache burden is associated with decreased endothelial function in our chronic TBI population. CONCLUSION: There is a correlation between PTH and CVR in chronic moderate-severe TBI. This relationship suggests that chronic TCVI may underlie the pathobiology of pPTH. Further, our results suggest that novel treatment strategies that target endothelial function and vascular health may be beneficial in refractory pPTH.

Imaging biomarkers of vascular and axonal injury are spatially distinct in chronic traumatic brain injury.

Traumatic Brain Injury (TBI) is associated with both diffuse axonal injury (DAI) and diffuse vascular injury (DVI), which result from inertial shearing forces. These terms are often used interchangeably, but the spatial relationships between DAI and DVI have not been carefully studied. Multimodal magnetic resonance imaging (MRI) can help distinguish these injury mechanisms: diffusion tensor imaging (DTI) provides information about axonal integrity, while arterial spin labeling (ASL) can be used to measure cerebral blood flow (CBF), and the reactivity of the Blood Oxygen Level Dependent (BOLD) signal to a hypercapnia challenge reflects cerebrovascular reactivity (CVR). Subjects with chronic TBI (n = 27) and healthy controls (n = 14) were studied with multimodal MRI. Mean values of mean diffusivity (MD), fractional anisotropy (FA), CBF, and CVR were extracted for pre-determined regions of interest (ROIs). Normalized z-score maps were generated from the pool of healthy controls. Abnormal ROIs in one modality were not predictive of abnormalities in another. Approximately 9-10% of abnormal voxels for CVR and CBF also showed an abnormal voxel value for MD, while only 1% of abnormal CVR and CBF voxels show a concomitant abnormal FA value. These data indicate that DAI and DVI represent two distinct TBI endophenotypes that are spatially independent.

Assessment of cerebrovascular dysfunction after traumatic brain injury with fMRI and fNIRS.

Traumatic cerebral vascular injury (TCVI) is a frequent, but under-recognized, endophenotype of traumatic brain injury (TBI).  It likely contributes to functional deficits after TBI and TBI-related chronic disability, and represents an attractive target for targeted therapeutic interventions. The aim of this prospective study is to assess microvascular injury/dysfunction in chronic TBI by measuring cerebral vascular reactivity (CVR) by 2 methods, functional magnetic resonance imaging (fMRI) and functional Near InfraRed Spectroscopy (fNIRS) imaging, as each has attractive features relevant to clinical utility. 42 subjects (27 chronic TBI, 15 age- and gender-matched non-TBI volunteers) were enrolled and underwent outpatient CVR testing by 2 methods, MRI-BOLD and fNIRS, each with hypercapnia challenge, a neuropsychological testing battery, and symptom survey questionnaires. Chronic TBI subjects showed a significant reduction in global CVR compared to HC (p < 0.0001). Mean CVR measures by fMRI were 0.225 ±â€¯0.014 and 0.183 ±â€¯0.026 %BOLD/mmHg for non-TBI and TBI subjects respectively and 12.3 ±â€¯1.8 and 9.2 ±â€¯1.7 mM/mmHg by fNIRS for non-TBI versus TBI subjects respectively. Global CVR measured by fNIRS imaging correlates with results by MRI-BOLD (R = 0.5). Focal CVR deficits seen on CVR maps by fMRI are also observed in the same areas by fNIRS in the frontal regions. Global CVR is significantly lower in chronic TBI patients and is reliably measured by both fMRI and fNIRS, the former with better spatial and the latter with better temporal resolution.  Both methods show promise as non-invasive measures of CVR function and microvascular integrity after TBI.

Phosphodiesterase-5 inhibition potentiates cerebrovascular reactivity in chronic traumatic brain injury.

BACKGROUND: Traumatic cerebrovascular injury (TCVI), a common consequence of traumatic brain injury (TBI), presents an attractive therapeutic target. Because phosphodiesterase-5 (PDE5) inhibitors potentiate the action of nitric oxide (NO) produced by endothelial cells, they are candidate therapies for TCVI. This study aims to: (1) measure cerebral blood flow (CBF), cerebrovascular reactivity (CVR), and change in CVR after a single dose of sildenafil (ΔCVR) in chronic TBI compared to uninjured controls; (2) examine the safety and tolerability of 8-week sildenafil administration in chronic symptomatic moderate/severe TBI patients; and as an exploratory aim, (3) assess the effect of an 8-week course of sildenafil on chronic TBI symptoms. METHODS: Forty-six subjects (31 chronic TBI, 15 matched healthy volunteers) were enrolled. Baseline CBF and CVR before and after administration of sildenafil were measured. Symptomatic TBI subjects then completed an 8-week double-blind, placebo-controlled, crossover trial of sildenafil. A neuropsychological battery and neurobehavioral symptom questionnaires were administered at each study visit. RESULTS: After a single dose of sildenafil, TBI subjects showed a significant increase in global CVR compared to healthy controls (P < 0.001, d = 0.9). Post-sildenafil CVR maps showed near-normalization of CVR in many regions where baseline CVR was low, predominantly within areas without structural abnormalities. Sildenafil was well tolerated. Clinical Global Impression (CGI) scale showed a trend toward clinical improvement while on sildenafil treatment. FINDINGS: Single-dose sildenafil improves regional CVR deficits in chronic TBI patients. CVR and ΔCVR are potential predictive and pharmacodynamic biomarkers of PDE5 inhibitor therapy for TCVI. Sildenafil is a potential therapy for TCVI.

Vascular Abnormalities within Normal Appearing Tissue in Chronic Traumatic Brain Injury.

Magnetic resonance imaging (MRI) is a powerful tool for visualizing traumatic brain injury(TBI)-related lesions. Trauma-induced encephalomalacia is frequently identified by its hyperintense appearance on fluid-attenuated inversion recovery (FLAIR) sequences. In addition to parenchymal lesions, TBI commonly results in cerebral microvascular injury, but its anatomical relationship to parenchymal encephalomalacia is not well characterized. The current study utilized a multi-modal MRI protocol to assess microstructural tissue integrity (by mean diffusivity [MD] and fractional aniosotropy [FA]) and altered vascular function (by cerebral blood flow [CBF] and cerebral vascular reactivity [CVR]) within regions of visible encephalomalacia and normal appearing tissue in 27 chronic TBI (minimum 6 months post-injury) subjects. Fifteen subjects had visible encephalomalacias whereas 12 did not have evident lesions on MRI. Imaging from 14 age-matched healthy volunteers were used as controls. CBF was assessed by arterial spin labeling (ASL) and CVR by measuring the change in blood-oxygen-level-dependent (BOLD) MRI during a hypercapnia challenge. There was a significant reduction in FA, CBF, and CVR with a complementary increase in MD within regions of FLAIR-visible encephalomalacia (p < 0.05 for all comparisons). In normal-appearing brain regions, only CVR was significantly reduced relative to controls (p < 0.05). These findings indicate that vascular dysfunction represents a TBI endophenotype that is distinct from structural injury detected using conventional MRI, may be present even in the absence of visible structural injury, and persists long after trauma. CVR may serve as a useful diagnostic and pharmacodynamic imaging biomarker of traumatic microvascular injury.

Imaging of Cerebrovascular Function in Chronic Traumatic Brain Injury.

Traumatic cerebrovascular injury (TCVI) is a common pathologic mechanism of traumatic brain injury (TBI) and presents an attractive target for intervention. The aims of this study were to assess cerebral blood flow (CBF) and cerebrovascular reactivity (CVR) using magnetic resonance imaging (MRI) to assess their value as biomarkers of TCVI in chronic TBI, characterize the spatial distribution of TCVI, and assess the relationships between each biomarker and neuropsychological and clinical assessments. Forty-two subjects (27 chronic TBI, 15 age- and gender-matched healthy volunteers) were studied cross-sectionally. CBF was measured by arterial spin labeling and CVR by assessing the MRI-blood oxygen level-dependent signal with hypercapnia challenge. A focused neuropsychological battery adapted from the TBI Common Data Elements and neurobehavioral symptom questionnaires were administered at the time of the imaging session. Chronic TBI subjects showed a significant reduction in mean global, gray matter (GM), and white matter (WM) CVR, compared with healthy volunteers (p < 0.001). Mean GM CVR had the greatest effect size (Cohen's d = 0.9). CVR maps in chronic TBI subjects showed patchy, multifocal CVR deficits. CBF discriminated poorly between TBI subjects and healthy volunteers and did not correlate with CVR. Mean global CVR correlated best with chronic neurobehavioral symptoms among TBI subjects. Global, GM, and WM CVR are reliable and potentially useful biomarkers of TCVI in the chronic stage after moderate-to-severe TBI. CBF is less useful as biomarker of TCVI. CVR correlates best with chronic TBI symptoms. CVR has potential as a predictive and pharmacodynamic biomarker for interventions targeting TCVI.

Prefrontal Hemodynamics in Toddlers at Rest: A Pilot Study of Developmental Variability.

Functional near infrared spectroscopy (fNIRS) is a non-invasive functional neuroimaging modality. Although, it is amenable to use in infants and young children, there is a lack of fNIRS research within the toddler age range. In this study, we used fNIRS to measure cerebral hemodynamics in the prefrontal cortex (PFC) in 18-36 months old toddlers (n = 29) as part of a longitudinal study that enrolled typically-developing toddlers as well as those "at risk" for language and other delays based on presence of early language delays. In these toddlers, we explored two hemodynamic response indices during periods of rest during which time audiovisual children's programming was presented. First, we investigate Lateralization Index, based on differences in oxy-hemoglobin saturation from left and right prefrontal cortex. Then, we measure oxygenation variability (OV) index, based on variability in oxygen saturation at frequencies attributed to cerebral autoregulation. Preliminary findings show that lower cognitive (including language) abilities are associated with fNIRS measures of both lower OV index and more extreme Lateralization index values. These preliminary findings show the feasibility of using fNIRS in toddlers, including those at risk for developmental delay, and lay the groundwork for future studies.

A machine learning approach to identify functional biomarkers in human prefrontal cortex for individuals with traumatic brain injury using functional near-infrared spectroscopy.

BACKGROUND: We have explored the potential prefrontal hemodynamic biomarkers to characterize subjects with Traumatic Brain Injury (TBI) by employing the multivariate machine learning approach and introducing a novel task-related hemodynamic response detection followed by a heuristic search for optimum set of hemodynamic features. To achieve this goal, the hemodynamic response from a group of 31 healthy controls and 30 chronic TBI subjects were recorded as they performed a complexity task. METHODS: To determine the optimum hemodynamic features, we considered 11 features and their combinations in characterizing TBI subjects. We investigated the significance of the features by utilizing a machine learning classification algorithm to score all the possible combinations of features according to their predictive power. RESULTS AND CONCLUSIONS: The identified optimum feature elements resulted in classification accuracy, sensitivity, and specificity of 85%, 85%, and 84%, respectively. Classification improvement was achieved for TBI subject classification through feature combination. It signified the major advantage of the multivariate analysis over the commonly used univariate analysis suggesting that the features that are individually irrelevant in characterizing the data may become relevant when used in combination. We also conducted a spatio-temporal classification to identify regions within the prefrontal cortex (PFC) that contribute in distinguishing between TBI and healthy subjects. As expected, Brodmann areas (BA) 10 within the PFC were isolated as the region that healthy subjects (unlike subjects with TBI), showed major hemodynamic activity in response to the High Complexity task. Overall, our results indicate that identified temporal and spatio-temporal features from PFC's hemodynamic activity are promising biomarkers in classifying subjects with TBI.

Abnormality of low frequency cerebral hemodynamics oscillations in TBI population.

Functional Near Infrared Spectroscopy (fNIRS) can non-invasively capture dynamic cognitive activation and underlying physiological processes by measuring changes in oxy- and deoxy-hemoglobin levels, correlated to brain activation. It is a portable, inexpensive and user-friendly device which is easily adapted to the outpatient setting for the assessment of cognitive functions after Traumatic Brain Injury (TBI). Low frequency oscillations in hemodynamic signal, attributed in the literature to cerebral autoregulation, were assessed using recently introduced metrics, Oxygenation Variability (OV Index), obtained from oxy/deoxy-hemoglobin variations in response to mental tasks for a group of healthy control (HC, n=14) and TBI (n=29). Participants responded to an action complexity judgment task (evaluating the complexity of daily life activities by classifying the number of steps as "few" or "many") with a varying degree of cognitive load to produce brain activation. During the task, we measured blood variations with fNIRS and analyzed OV Index changes. Mean OV indices, corresponding to high complexity tasks, are higher than that of low complexity tasks in the HC group, revealing strong parametric effect (0.039±0.017 for low, 0.057±0.036 for high, p-value=0.069). However, no significant difference has been recorded for the OV indexes for two different loads in the TBI group (0.055±0.033 for low, 0.054±0.035 for high, p=0.9). OV index metrics proves to be sensitive to chronic TBI and can potentially be used to separate subpopulations TBI vs. HC. Noticeable differences in OV index spatial distributions between subpopulations have been observed.

Cerebral Vascular Injury in Traumatic Brain Injury.

Traumatic cerebral vascular injury (TCVI) is a very frequent, if not universal, feature after traumatic brain injury (TBI). It is likely responsible, at least in part, for functional deficits and TBI-related chronic disability. Because there are multiple pharmacologic and non-pharmacologic therapies that promote vascular health, TCVI is an attractive target for therapeutic intervention after TBI. The cerebral microvasculature is a component of the neurovascular unit (NVU) coupling neuronal metabolism with local cerebral blood flow. The NVU participates in the pathogenesis of TBI, either directly from physical trauma or as part of the cascade of secondary injury that occurs after TBI. Pathologically, there is extensive cerebral microvascular injury in humans and experimental animal, identified with either conventional light microscopy or ultrastructural examination. It is seen in acute and chronic TBI, and even described in chronic traumatic encephalopathy (CTE). Non-invasive, physiologic measures of cerebral microvascular function show dysfunction after TBI in humans and experimental animal models of TBI. These include imaging sequences (MRI-ASL), Transcranial Doppler (TCD), and Near InfraRed Spectroscopy (NIRS). Understanding the pathophysiology of TCVI, a relatively under-studied component of TBI, has promise for the development of novel therapies for TBI.

A Review of the Effectiveness of Neuroimaging Modalities for the Detection of Traumatic Brain Injury.

The incidence of traumatic brain injury (TBI) in the United States was 3.5 million cases in 2009, according to the Centers for Disease Control and Prevention. It is a contributing factor in 30.5% of injury-related deaths among civilians. Additionally, since 2000, more than 260,000 service members were diagnosed with TBI, with the vast majority classified as mild or concussive (76%). The objective assessment of TBI via imaging is a critical research gap, both in the military and civilian communities. In 2011, the Department of Defense (DoD) prepared a congressional report summarizing the effectiveness of seven neuroimaging modalities (computed tomography [CT], magnetic resonance imaging [MRI], transcranial Doppler [TCD], positron emission tomography, single photon emission computed tomography, electrophysiologic techniques [magnetoencephalography and electroencephalography], and functional near-infrared spectroscopy) to assess the spectrum of TBI from concussion to coma. For this report, neuroimaging experts identified the most relevant peer-reviewed publications and assessed the quality of the literature for each of these imaging technique in the clinical and research settings. Although CT, MRI, and TCD were determined to be the most useful modalities in the clinical setting, no single imaging modality proved sufficient for all patients due to the heterogeneity of TBI. All imaging modalities reviewed demonstrated the potential to emerge as part of future clinical care. This paper describes and updates the results of the DoD report and also expands on the use of angiography in patients with TBI.

Normative database of judgment of complexity task with functional near infrared spectroscopy--application for TBI.

The ability to assess frontal lobe function in a rapid, objective, and standardized way, without the need for expertise in cognitive test administration might be particularly helpful in mild traumatic brain injury (TBI), where objective measures are needed. Functional near infrared spectroscopy (fNIRS) is a reliable technique to noninvasively measure local hemodynamic changes in brain areas near the head surface. In this paper, we are combining fNIRS and frameless stereotaxy which allowed us to co-register the functional images with previously acquired anatomical MRI volumes. In our experiment, the subjects were asked to perform a task, evaluating the complexity of daily life activities, previously shown with fMRI to activate areas of the anterior frontal cortex. We reconstructed averaged oxyhemoglobin and deoxyhemoglobin data from 20 healthy subjects in a spherical coordinate. The spherical coordinate is a natural representation of surface brain activation projection. Our results show surface activation projected from the medial frontopolar cortex which is consistent with previous fMRI results. With this original technique, we will construct a normative database for a simple cognitive test which can be useful in evaluating cognitive disability such as mild traumatic brain injury.

A hematoma detector-a practical application of instrumental motion as signal in near infra-red imaging.

In this paper we discuss results based on using instrumental motion as a signal rather than treating it as noise in Near Infra-Red (NIR) imaging. As a practical application to demonstrate this approach we show the design of a novel NIR hematoma detection device. The proposed device is based on a simplified single source configuration with a dual separation detector array and uses motion as a signal for detecting changes in blood volume in the dural regions of the head. The rapid triage of hematomas in the emergency room will lead to improved use of more sophisticated/expensive imaging facilities such as CT/MRI units. We present simulation results demonstrating the viability of such a device and initial phantom results from a proof of principle device. The results demonstrate excellent localization of inclusions as well as good quantitative comparisons.

Quantitative principal component model for skin chromophore mapping using multi-spectral images and spatial priors.

We describe a novel reconstruction algorithm based on Principal Component Analysis (PCA) applied to multi-spectral imaging data. Using numerical phantoms, based on a two layered skin model developed previously, we found analytical expressions, which convert qualitative PCA results into quantitative blood volume and oxygenation values, assuming the epidermal thickness to be known. We also evaluate the limits of accuracy of this method when the value of the epidermal thickness is not known. We show that blood volume can reliably be extracted (less than 6% error) even if the assumed thickness deviates 0.04mm from the actual value, whereas the error in blood oxygenation can be as large as 25% for the same deviation in thickness. This PCA based reconstruction was found to extract blood volume and blood oxygenation with less than 8% error, if the underlying structure is known. We then apply the method to in vivo multi-spectral images from a healthy volunteer's lower forearm, complemented by images of the same area using Optical Coherence Tomography (OCT) for measuring the epidermal thickness. Reconstruction of the imaging results using a two layered analytical skin model was compared to PCA based reconstruction results. A point wise correlation was found, showing the proof of principle of using PCA based reconstruction for blood volume and oxygenation extraction.

Principal component model of multispectral data for near real-time skin chromophore mapping.

Multispectral images of skin contain information on the spatial distribution of biological chromophores, such as blood and melanin. From this, parameters such as blood volume and blood oxygenation can be retrieved using reconstruction algorithms. Most such approaches use some form of pixelwise or volumetric reconstruction code. We explore the use of principal component analysis (PCA) of multispectral images to access blood volume and blood oxygenation in near real time. We present data from healthy volunteers under arterial occlusion of the forearm, experiencing ischemia and reactive hyperemia. Using a two-layered analytical skin model, we show reconstruction results of blood volume and oxygenation and compare it to the results obtained from our new spectral analysis based on PCA. We demonstrate that PCA applied to multispectral images gives near equivalent results for skin chromophore mapping and quantification with the advantage of being three orders of magnitude faster than the reconstruction algorithm.

Direct curvature correction for noncontact imaging modalities applied to multispectral imaging.

Noncontact optical imaging of curved objects can result in strong artifacts due to the object's shape, leading to curvature biased intensity distributions. This artifact can mask variations due to the object's optical properties, and makes reconstruction of optical/physiological properties difficult. In this work we demonstrate a curvature correction method that removes this artifact and recovers the underlying data, without the necessity of measuring the object's shape. This method is applicable to many optical imaging modalities that suffer from shape-based intensity biases. By separating the spatially varying data (e.g., physiological changes) from the background signal (dc component), we show that the curvature can be extracted by either averaging or fitting the rows and columns of the images. Numerical simulations show that our method is equivalent to directly removing the curvature, when the object's shape is known, and accurately recovers the underlying data. Experiments on phantoms validate the numerical results and show that for a given image with 16.5% error due to curvature, the method reduces that error to 1.2%. Finally, diffuse multispectral images are acquired on forearms in vivo. We demonstrate the enhancement in image quality on intensity images, and consequently on reconstruction results of blood volume and oxygenation distributions.

Topology of the heterogeneous nature of the extracellular matrix on stochastic modeling of tumor-induced angiogenesis.

We have modeled tumor-induced angiogenesis; our model includes the phenomena of the migratory response of endothelial cells (ECs) to tumor angiogenic factors, and the interaction of ECs with the extracellular matrix (ECM). ECs switch between growth, differentiation, motility, or apoptotic behavior in response to the local topology and composition of the ECM. Assuming the ECM medium as a statistically inhomogeneous medium (some area support sprout growth, some not), we show that the ECM can be a natural barrier to angiogenesis. We study vascular network formation for several ECM distributions and topologies, and we find an analogy with percolation. A threshold exists, under which sprouts cannot reach the tumor. During the growth of the vascular network, a competition exists between the attraction exerted by tumor and the preferred path created by the ECM. We also examined the influence of branching on the tumor vascularization. Branching is a natural phenomenon which helps the tumor become vascularized. By increasing the number of sprouts, the vascular network increases the probability of reaching the tumor, as it can explore more pathways. Our simulations show after two branching events, the vascular network is very likely to reach the tumor.

Use of scaling relations to extract intrinsic fluorescence lifetime of targets embedded in turbid media.

We present a novel method for estimating the intrinsic fluorescence lifetime of deeply embedded localized fluorophores. It is based on scaling relations, characteristic for turbid media. The approach is experimentally substantiated by successfully reconstructing lifetimes for targets at depths up to 14.5 mm. A derived correction factor was determined from the product of the transport-corrected scattering coefficient mu(s) (') and the index of refraction n(r). In addition, data from an array of detectors (> or =2) can be used to estimate mu(s) (')n(r). The suggested algorithm is a promising tool for diagnostic fluorescence, since lifetime can be a sensitive indicator of the fluorophore environment.