Biomedical subjects

Joseph D Fenstermacher

Publications and source records attributed to Joseph D Fenstermacher.

14 recordsLinked to original sources

PubMed2006

Role of angiotensin II in retinal leukostasis in the diabetic rat.

To study if the endogenous renin-angiotensin system affects diabetic retinal leukostasis, rats with streptozotocin-induced diabetes were treated with an ACE inhibitor (ramipril), an angiotensin II AT(1) receptor antagonist (losartan) and the Ca channel blocker, (nifedipine). In the diabetic rats, these drug treatments reduced systolic blood pressure by approximately 16 mmHg but did not change blood glucose. After 2 weeks, the rats were examined for retinal leukostasis in vivo with a scanning laser ophthalmoscope (SLO). Retinal leukostasis, which was defined as no movement of arrested leukocytes over 2 min, was markedly higher in diabetic rats than normal controls (P<0.01). Leukostasis was significantly decreased by ramipril and losartan (P<0.01 vs. untreated diabetic rats) but was still higher than normal. Retinal leukostasis after nifedipine treatment was not significantly different than in untreated diabetic rats. The same trend was observed when leukostasis was analyzed on retinal flat mounts with concanavalin A and CD45 immunofluorescence; ramipril and losartan treatment, however, decreased leukostasis to values no different than controls. Retinal leukostasis was lowered by nifedipine (P<0.05, untreated diabetes vs. nifedipine-treated) but was still higher than in normal, ramipril-, or losartan-treated rats. Assays of gene expression of retinal intercellular adhesion molecule (ICAM-1) by semi-quantitative RT-PCR indicated that ICAM-1 mRNA was increased in diabetic rats but was decreased markedly by treatment with losartan or ramipril, and modestly by nifedipine. In summary, suppressing the activity of the endogenous renin-angiotensin system markedly decreases, perhaps even normalizes, the retinal leukostasis that accompanies type I diabetes in rats. These effects seem to be partly independent of blood pressure and to be associated with a decrease in ICAM-1 gene expression. Angiotensin II may, thus, mediate retinal leukostasis in early diabetes.

Angiotensin II↗

PubMed2006

Application of arsenazo III in the preparation and characterization of an albumin-linked, gadolinium-based macromolecular magnetic resonance contrast agent.

A macromolecular magnetic resonance contrast agent (MMCA) was prepared by linking bovine serum albumin (BSA) to gadolinium (Gd) via a chelating agent, diethylenetriaminepentaacetic acid (DTPA). Colorimetric testing with 2,7-bis(o-arsenophenylazo)-1,8-dihydroxynaphthalene-3,6-disulfonic acid (arsenazo III) was performed to check for the appearance of free gadolinium during preparation and to quantify the Gd content in the final product. The complex was purified by dialysis, concentrated by lyophilyzation and characterized by magnetic resonance (MR) proton relaxation times. The resultant product had a molecular weight of about 90 kDa, Gd:BSA ratio of 14:1, and T1 and T2 relaxation times of 128.3 and 48.9 ms, respectively, at a field strength of 7Tesla (T) and at 20% concentration. Contrast enhancement of Gadomer-17 (a dendritic MMCA) and Gd-linked to BSA (Gd-BSA) was sequentially evaluated in a rat brain gliosarcoma model (n = 5) by MR imaging (MRI). Following intravenous injection, the blood concentration of Gadomer-17 fell rapidly, whereas that of Gd-BSA was almost constant for the duration of imaging. The areas of enhancement of both MMCAs were comparable. The spatial distribution of Gd-BSA showed good agreement with Evans blue-tagged albumin. Treatment with dexamethasone decreased Gd-BSA enhancement in the tumor. These results suggest that the arsenazo III method is applicable in preparing Gd-BSA to image brain tumors and their response to treatment. This simple method may also be useful for preparing other gadolinium-linked MMCAs.

Animals↗

PubMed2006

Model selection in magnetic resonance imaging measurements of vascular permeability: Gadomer in a 9L model of rat cerebral tumor.

Vasculature in and around the cerebral tumor exhibits a wide range of permeabilities, from normal capillaries with essentially no blood-brain barrier (BBB) leakage to a tumor vasculature that freely passes even such large molecules as albumin. In measuring BBB permeability by magnetic resonance imaging (MRI), various contrast agents, sampling intervals, and contrast distribution models can be selected, each with its effect on the measurement's outcome. Using Gadomer, a large paramagnetic contrast agent, and MRI measures of T(1) over a 25-min period, BBB permeability was estimated in 15 Fischer rats with day-16 9L cerebral gliomas. Three vascular models were developed: (1) impermeable (normal BBB); (2) moderate influx (leakage without efflux); and (3) fast leakage with bidirectional exchange. For data analysis, these form nested models. Model 1 estimates only vascular plasma volume, v(D), Model 2 (the Patlak graphical approach) v(D) and the influx transfer constant K(i). Model 3 estimates v(D), K(i), and the reverse transfer constant, k(b), through which the extravascular distribution space, v(e), is calculated. For this contrast agent and experimental duration, Model 3 proved the best model, yielding the following central tumor means (+/-s.d.; n = 15): v(D) = 0.07 +/- 0.03 for K(i) = 0.0105 +/- 0.005 min(-1) and v(e) = 0.10 +/- 0.04. Model 2 K(i) estimates were approximately 30% of Model 3, but highly correlated (r = 0.80, P < 0.0003). Sizable inhomogeneity in v(D), K(i), and k(b) appeared within each tumor. We conclude that employing nested models enables accurate assessment of transfer constants among areas where BBB permeability, contrast agent distribution volumes, and signal-to-noise vary.

Animals↗

PubMed2005

9L gliosarcoma cell proliferation and tumor growth in rats are suppressed by N-hydroxy-N'-(4-butyl-2-methylphenol) formamidine (HET0016), a selective inhibitor of CYP4A.

The present study examined the effects of N-hydroxy-N'-(4-butyl-2 methylphenyl) formamidine (HET0016), a selective inhibitor of the formation of 20-hydroxyeicosatrienoic acid (20-HETE) on the growth of 9L rat gliosarcoma cells in vitro and in vivo. After 48 h of incubation, HET0016 reduced the proliferation of 9L in vitro by 55%, and this was associated with a fall in p42/p44 mitogen-activated protein kinase and stress-activated protein kinase/c-Jun NH(2)-terminal kinase phosphorylation and increased apoptosis. HET0016 inhibited epidermal growth factor (EGF) and platelet-derived growth factor (PDGF)-induced proliferation and diminished phosphorylation of PDGF receptors. A stable 20-HETE analog increased 9L cell proliferation. In vivo, chronic administration of HET0016 (10 mg/kg/day i.p.) for 2 weeks reduced the volume of 9L tumors by 80%. This was accompanied by a 4-fold reduction in the mitotic index, a 3- to 4-fold increase in the apoptotic index, and a approximately 50% decrease in vascularization in the tumor. HET0016 treatment increased mean survival time of the animals from 17 to 22 days. Liquid chromatography/mass spectrometry experiments indicated that neither 9L cells grown in vitro nor 9L tumors removed produce 20-HETE when incubated with arachidonic acid. The normal surrounding brain tissue, however, avidly makes 20-HETE, and this activity is selectively inhibited by HET0016. These results suggest that HET0016 may be the prototype of a class of antigrowth compounds that may be efficacious for treating malignant brain tumors. In vivo, it may act in part by inhibiting the formation of 20-HETE by the surrounding tissue. However, the antiproliferative effects of HET0016 on 9L cells in vitro seem unrelated to its ability to inhibit the formation of 20-HETE.

Amidines↗

PubMed2005

In normal rat, intraventricularly administered insulin-like growth factor-1 is rapidly cleared from CSF with limited distribution into brain.

BACKGROUND: Putatively active drugs are often intraventricularly administered to gain direct access to brain and circumvent the blood-brain barrier. A few studies on the normal central nervous system (CNS) have shown, however, that the distribution of materials after intraventricular injections is much more limited than presumed and their exit from cerebrospinal fluid (CSF) is more rapid than generally believed. In this study, we report the intracranial distribution and the clearance from CSF and adjacent CNS tissue of radiolabeled insulin-like growth factor-1 after injection into one lateral ventricle of the normal rat brain. METHODS: Under barbiturate anesthesia, 125I-labeled insulin-like growth factor-1 (IGF-1) was injected into one lateral ventricle of normal Sprague-Dawley rats. The subsequent distribution of IGF-1 through the cerebrospinal fluid (CSF) system and into brain, cerebral blood vessels, and systemic blood was measured over time by gamma counting and quantitative autoradiography (QAR). RESULTS: Within 5 min of infusion, IGF-1 had spread from the infused lateral ventricle into and through the third and fourth ventricles. At this time, 25% of the infused IGF-1 had disappeared from the CSF-brain-meningeal system; the half time of this loss was 12 min. The plasma concentration of cleared IGF-1 was, however, very low from 2 to 9 min and only began to rise markedly after 20 min. This delay between loss and gain plus the lack of radiotracer in the cortical subarachnoid space suggested that much of the IGF-1 was cleared into blood via the cranial and/or spinal nerve roots and their associated lymphatic systems rather than periventricular tissue and arachnoid villi. Less than 10% of the injected radioactivity remained in the CSF-brain system after 180 min. The CSF and arteries and arterioles within the subarachnoid cisterns were labeled with IGF-1 within 10 min. Between 60 and 180 min, most of the radioactivity within the cranium was retained within and around these blood vessels and by periaqueductal gray matter. Tissue profiles at two sites next to ventricular CSF showed that IGF-1 penetrated less than 1.25 mm into brain tissue and appreciable 125I-activity remained at the tissue-ventricular CSF interface after 180 min. CONCLUSION: Our findings suggest that entry of IGF-1 into normal brain parenchyma after lateral ventricle administration is limited by rapid clearance from CSF and brain and slow movement, apparently by diffusion, into the periventricular tissue. Various growth factors and other neuroactive agents have been reported to be neuroprotective within the injured brain after intraventricular administration. It is postulated that the delivery of such factors to neurons and glia in the injured brain may be facilitated by abnormal CSF flow. These several observations suggest that the flow of CSF and entrained solutes may differ considerably between normal and abnormal brain and even among various neuropathologies.

Journal Article↗

PubMed2005

Arterial spin labeling: validity testing and comparison studies.

Arterial spin labeling (ASL) is a potential means of obtaining quantitative images of cerebral blood flow (CBF). However, few validation studies of ASL have been performed in animal models using gold-standard CBF methods. Other methods that use radiolabeled water as a tracer underestimate CBF in high flow states, but this effect has not been evident in ASL studies. In this study the accuracy of ASL measurements of CBF were modeled and experimentally validated, with particular attention paid to high flow rates. The ASL signal as modeled included the contributions from intravascular labeled spins. The modeling demonstrated linearity of the ASL signal with respect to baseline flow, and linearity of ASL signal changes with respect to changes in flow, including high-flow conditions. Validation studies using quantitative autoradiography (QAR) to image flow in a rat model of unilateral cerebral ischemia showed that ASL systematically overestimated CBF by 34%. A similar overestimation was also predicted by modeling. These results indicate that ASL signals are linear with respect to flow (even high flow), but ASL-CBF measurements are systematically overestimated.

Animals↗

PubMed2005

Quantitation and localization of blood-to-brain influx by magnetic resonance imaging and quantitative autoradiography in a model of transient focal ischemia.

The ability of gadolinium-diethylenetriaminepentaacetic acid (Gd-DTPA) enhanced MRI to localize and quantitate blood-brain barrier (BBB) opening was evaluated against quantitative autoradiographic (QAR) imaging of (14)C-alpha-aminoisobutyric acid (AIB) distribution. The blood-to-brain transfer constant (K(i)) for Gd-DTPA was determined by MRI in rats after 3 h of focal cerebral ischemia plus 2.5 h of reperfusion (n = 9), and that of AIB was determined by QAR shortly thereafter. Tissue regions of interest (ROIs) for Gd-DTPA leakage were identified by ISODATA segmentation of pre- and post-Gd-DTPA Look-Locker (L-L) T(1) maps. Patlak plots were constructed using time course of blood and tissue T(1) changes induced by Gd for estimating K(i). Among the nine rats, 14 sizable regions of AIB uptake were found; 13 were also identified by ISODATA segmentation. Although the 13 MRI-ROIs spatially approximated those of AIB uptake, the segmentation sometimes missed small areas of lesser AIB uptake that did not extend through more than 60% of the 2.0-mm-thick slice. Mean K(i)'s of AIB were highly correlated with those of Gd-DTPA across the 13 regions; the group means (+/-SD) were similar for the two tracers (7.1 +/- 3.3 x 10(-3) and 6.8 +/- 3.5 x 10(-3) ml.g(-1) . min(-1), respectively). In most instances, Gd-DTPA MRI accurately localized areas of BBB opening.

Animals↗

PubMed2005

Acute blood-brain barrier opening in experimentally induced focal cerebral ischemia is preferentially identified by quantitative magnetization transfer imaging.

Pathologic changes in brain tissue during and after stroke may lead to injury of the blood-brain barrier (BBB) and subsequent hemorrhagic transformation (HT). In a rat model of HT, the apparent diffusion coefficient of water, cerebral blood flow, relaxation times, T(1) and T(2), and magnetization transfer (MT) related parameters (T(1sat), K(for) and the MT ratio) were repetitively measured during 3 h of focal ischemia and 2 h of reperfusion (n = 8). Areas of BBB opening were identified by sequential assay of the transcapillary influx of Gd-diethylenetriaminepentaacetic acid (Gd-DTPA) by MRI and (14)C-alpha-aminoisobutyric acid (AIB) by quantitative autoradiography. Ischemia-injured regions of interest were identified from the MRI data and divided into those with and without BBB opening. Of the several MRI parameters measured, the T(1sat) in the caudate-putamen and preoptic area during ischemia and the first 2 h of reperfusion correlated best with the regional pattern of BBB opening observed thereafter. These data suggest that an ipsilateral/contralateral T(1sat) ratio > 1.6 demarcates leakage of small molecules such as Gd-DTPA and AIB across the BBB. As to clinical relevance, the quantitation of MT parameters in acute stroke may enable the early detection of areas of BBB opening and potential HT.

Acute Disease↗

PubMed2004

A quantitative method for measuring gene expression magnitude and volume delivered by gene therapy vectors.

This study describes a quantitative method to measure the magnitude and distribution of gene expression following local delivery of an adenoviral vector containing the human sodium iodide symporter (hNIS) reporter gene into the canine prostate. Following systemic administration of Na(99m)TcO(4), autoradiographs of prostate sections depicting hNIS-dependent (99m)TcO(4)(-) uptake were digitized and stacked to produce a three-dimensional reconstruction of gene expression. Frequency histograms reflecting hNIS gene expression magnitude and volume were used to quantify hNIS function. The method demonstrated submillimeter resolution allowing for precise measurements of gene expression magnitude and volume in vivo. The method developed here could be applied to other reporter gene systems in which the readout can be digitized from thin tissue sections.

Adenoviridae↗

PubMed2003

Patlak plots of Gd-DTPA MRI data yield blood-brain transfer constants concordant with those of 14C-sucrose in areas of blood-brain opening.

The blood-to-brain transfer rate constant (K(i)) of Gd-DTPA was determined in MRI studies of a rat model of transient cerebral ischemia. The longitudinal relaxation rate, R(1), was estimated using repeated Look-Locker measurements. A model-independent analysis of deltaR(1), the Patlak plot, produced maps of K(i) for Gd-DTPA and the distribution volume of the mobile protons (V(p)) with intravascular-Gd changed R(1)'s. The K(i)'s of Gd-DTPA were estimated in regions of interest with blood-brain barrier (BBB) opening (regions of interest, ROIs) and compared to those of (14)C-sucrose determined shortly thereafter by quantitative autoradiography. The K(i)'s for both Gd-DTPA and sucrose were much higher than normal within the ROIs (n = 7); linear regression of K(i) for Gd-DTPA vs. K(i) for sucrose yielded a slope of 0.43 +/- 0.11 and r(2) = 0.72 (P = 0.01). Thus, K(i) for Gd-DTPA varied in parallel with, but was less than, K(i) for sucrose. In the ROIs, mean V(p) was 0.071 ml g(-1) and much higher than mean vascular volume estimated by dynamic-contrast-enhancement (0.013 ml g(-1)) or mean V(p) in contralateral brain (0.015 ml g(-1)). This elevated V(p) may reflect increased capillary permeability to water. In conclusion, K(i) can be reliably calculated from Gd-DTPA-MRI data by Patlak plots.

Animals↗

PubMed2003

GENIS: gene expression of sodium iodide symporter for noninvasive imaging of gene therapy vectors and quantification of gene expression in vivo.

With the goal of optimizing adenovirus-mediated suicide gene therapy for prostate cancer, we have developed a method based on the human sodium iodide symporter (hNIS) that allows for noninvasive monitoring of adenoviral vectors and quantification of gene expression magnitude and volume within the prostate. A replication-competent adenovirus (Ad5-yCD/mutTK(SR39)rep-hNIS) coexpressing a therapeutic yeast cytosine deaminase (yCD)/mutant herpes simplex virus thymidine kinase (mutTK(SR39)) fusion gene and the hNIS gene was developed. Ad5-yCD/mutTK(SR39)rep-hNIS and a replication-defective hNIS adenovirus (rAd-CMV-FLhNIS) were injected into contralateral lobes of the dog prostate and hNIS activity was monitored in live animals following administration of Na(99m)TcO(4) using gamma camera scintigraphy. Despite the close proximity of the urinary bladder, (99m)TcO(4)(-) uptake was readily detected in the prostate using viral dose levels (10(10) to 10(12) viral particles) that have been safely administered to humans. Due to its rapid clearance and short physical half-life (6 h), it was possible to obtain daily measurements of (99m)TcO(4)(-) uptake in vivo, allowing for dynamic monitoring of reporter gene expression within the prostate as well as biodistribution throughout the body. High-resolution autoradiography of prostate sections coupled with 3D reconstruction of gene expression demonstrated that the magnitude and volume of gene expression could be quantified with submillimeter resolution. Implementation of the GENIS (gene expression of Na/I symporter) technology in the clinic will facilitate optimization of future human gene therapy trials.

Adenoviridae↗

PubMed2003

Direct comparison of local cerebral blood flow rates measured by MRI arterial spin-tagging and quantitative autoradiography in a rat model of experimental cerebral ischemia.

The present study determined cerebral blood flow (CBF) in the rat using two different magnetic resonance imaging (MRI) arterial spin-tagging (AST) methods and 14C-iodoantipyrine (IAP)-quantitative autoradiography (QAR), a standard but terminal technique used for imaging and quantitating CBF, and compared the resulting data sets to assess the precision and accuracy of the different techniques. Two hours after cerebral ischemia was produced in eight rats via permanent occlusion of one middle cerebral artery (MCA) with an intraluminal suture, MRI-CBF was measured over a 2.0-mm coronal slice using single-coil AST, and tissue magnetization was assessed by either a spin-echo (SE) or a variable tip-angle gradient-echo (VTA-GE) readout. Subsequently ( approximately 2.5 hours after MCA occlusion), CBF was assayed by QAR with the blood flow indicator 14C-IAP, which produced coronal images of local flow rates every 0.4 mm along the rostral-caudal axis. The IAP-QAR images that spanned the 2-mm MRI slice were selected, and regional flow rates (i.e., local CBF [lCBF]) were measured and averaged across this set of images by both the traditional approach, which involved reader interaction and avoidance of sectioning artifacts, and a whole film-scanning technique, which approximated total radioactivity in the entire MRI slice with minimal user bias. After alignment and coregistration, the concordance of the CBF rates generated by the two QAR approaches and the two AST methods was examined for nine regions of interest in each hemisphere. The QAR-lCBF rates were higher with the traditional method of assaying tissue radioactivity than with the MRI-analog approach; although the two sets of rates were highly correlated, the scatter was broad. The flow rates obtained with the whole film-scanning technique were chosen for subsequent comparisons to MRI-CBF results because of the similarity in tissue "sampling" among these three methods. As predicted by previous modeling, "true" flow rates, assumed to be given by QAR-lCBF, tended to be slightly lower than those measured by SE and were appreciably lower than those assessed by VTA-GE. When both the ischemic and contralateral hemispheres were considered together, SE-CBF and VTA-GE-CBF were both highly correlated with QAR-lCBF ( P< 0.001). If evaluated by flow range, however, SE-CBF estimates were more accurate in high-flow (contralateral) areas (CBF > 80 mL. 100 g(-1). min(-1) ), whereas VTA-GE-CBF values were more accurate in low-flow (ipsilateral) areas (CBF < or= 60 mL. 100 g(-1). min(-1) ). Accordingly, the concurrent usage of both AST-MRI methods or the VTA-GE technique alone would be preferred for human studies of stroke.

Animals↗

PubMed2003

Hemorrhagic transformation is related to the duration of occlusion and treatment with tissue plasminogen activator in a nonembolic stroke model.

The availability of reperfusion therapy for acute ischemic stroke patients has made the causes and significance of hemorrhagic transformation an area of intense interest and controversy. Ninety-two male Wistar rats underwent transient middle cerebral artery occlusion (MCAO) of between 1 and 6 h. Forty animals received 10 mg kg-1 of recombinant tissue plasminogen activator (rtPA), infused over 20 min, starting 5 min before reperfusion. At 18-24 h, the animals were sacrificed. The presence of hemorrhagic transformation (HT) on stained sections was recorded and total ischemic lesion area was quantified using image analysis software. Seventeen animals (11 with HT) were subjected to immunohistochemical analysis for detection of endothelial barrier antigen (EBA), quantified in three sections, in eight different fields per section. Chi-squared analysis and logistic regression were used to assess the contribution of rtPA and duration of occlusion to HT development. Nested, repeated measures analyses of variance were performed to assess the changes in EBA caused by ischemia and associated with HT. Fifty-nine animals developed HT that was significantly associated with occlusion duration (p < 0.0001) and ischemic lesion size (p = 0.0007). The presence of rtPA accelerated HT development. Statistically significant side-to-side differences in the presence of EBA were found in the striatum (core of the infarct) of animals with HT (p < 0.001) and without HT (p < 0.001), but only in animals with durations of occlusion of 2 h or more. Duration of occlusion is an important predictor of HT in transient MCAO in the rat and is closely associated with EBA expression.

Animals↗

PubMed2002

Acute postischemic renormalization of the apparent diffusion coefficient of water is not associated with reversal of astrocytic swelling and neuronal shrinkage in rats.

BACKGROUND AND PURPOSE: Initially decreased apparent diffusion coefficient (ADC) values are reversible if reperfusion is rapidly performed after focal brain ischemia. We sought to determine if reperfusion-induced renormalization of initially abnormal values indicates reversal of cellular, morphologic changes that occur during acute ischemia. METHODS: Eighteen rats underwent 30 minutes of middle cerebral artery occlusion (MCAO) without reperfusion (group A, n = 6), with 1.5 hours of reperfusion (group B, n = 6), or with 12 hours of reperfusion (group C, n = 6). Diffusion- and perfusion-weighted MR images were obtained at the end of MCAO and 1.5 and 12 hours after reperfusion. Immediately after the final MR study, the brains were fixed by cardiac perfusion with 4% paraformaldehyde. Neuronal injury was evaluated on hematoxylin-eosin-stained slices, and astrocytic size was determined by the area of glial fibrillary acidic protein (GFAP) plus S-100 expression. RESULTS: In group A in which ADC values decreased significantly, 47 +/-12% of the neurons were slightly shrunken; astrocytes were moderately swollen, and the area expressing GFAP plus S-100 was larger than that in the contralateral hemisphere (117 microm(2) +/- 6 vs 89 microm(2) +/- 2; P <.001). In group B in which ADC had renormalized, most neurons were moderately shrunken, and the frequency of such neurons was greater in group B (92% +/- 2) than in group A (P <.001); astrocytes were markedly swollen, and the area was larger than that in the contralateral hemisphere (123 microm(2) +/- 8 vs 85 microm(2) +/- 4, P <.001). In group C in which a secondary ADC decrease occurred, most neurons (94% +/- 3) were severely shrunken, and some had eosinophilic cytoplasm; astrocytes were disintegrated, and the area of GFAP plus S-100 expression was reduced (78 microm(2) +/- 4 vs 90 microm(2) +/- 5, P <.001). CONCLUSION: Reperfusion-induced acute renormalization of ADC values is not associated with the reversal of neuronal shrinkage and astrocytic swelling that occur during ischemia. Conversely, the morphologic changes of astrocytes and neurons progressively worsen over time, although ADC values show a biphasic change.

Animals↗