Regional cerebral blood flow in cerebrovascular disease studied using N-isopropyl-p-(123I) iodoamphetamine

Twenty-one patients with cerebrovascular disease (5 with hemorrhage, 10 with infarction, 2 with TIA, 4 with motor disturbance of unknown cause) were studied using N-isopropyl-p-(123I)iodoamphetamine (123I-IMP) and single photon emission CT. In 3 of 5 cases with cerebral hemorrhage, perfusion defects were shown in and around the region of hematomas, furthermore, one of the cases with internal capsular hematoma, the perfusion defect extended to the cortical area corresponding to the neurological pathway. In one case with MCA infarction, the perfusion defect was greater and clearer than the low density area on X-ray computed tomography (CT). These results revealed the 123I-IMP study provides physiological information in contrast with X-ray CT which provides anatomical information. In 3 of 9 cases with multiple small deep hemispheric infarctions on X-ray CT, the perfusion to the basal ganglia was suspected to be decreased on 123I-IMP images. However, this visual findings was not definitive and, in fact, the diseased side was not always consistent with the clinical findings. For quantitative analysis, symmetrical regions of interest (ROIs) were constructed both basal ganglia and the ratio of average counts over the ROI to those over the whole slice was calculated. In the small infarction group, the mean +/- S.D. of the values was 0.89 +/- 0.09 in right and 0.89 +/- 0.08 in left. Although the values were not significantly different from those of normal subjects (0.99 +/- 0.02 in right, 0.97 +/- 0.03 in left), they distributed in the range less than normal in 5 of 9 cases. This method was thought to be useful and practical to evaluate the cerebral blood flow in basal ganglia of patients with deep hemispheric infarction.     

Eletroarteriography: A non-invasive technique for blood flow measurement

Peripheral vascular diseases, such as athero- and arteriosclerosis, result in modifications of the blood flow characteristics which give a clinician an indication of the status of these diseases. A non-invasive technique, called electroarteriography (EAG), that permits the measurement and analysis of pertinent hemodynamic information based upon simple voltage measurements taken on the skin surface has been developed. The technique is very similar to that used in electrocardiography (ECG). Verification experiments were conducted on a variety of cardiovascular mechanical models, in rats and with human volunteers. Evidence is presented that supports the postulate that EAG analysis does indeed permit detection of rather subtle changes in peripheral blood flow.     

A simple technique to measure regional cerebral blood flow during intravascular balloon clamping

A case of giant internal carotid ophthalmic aneurysm was presented. In order to clarify whether the patient could tolerate carotid occlusion, a balloon clamping test was performed. before surgery. The cerebral blood flow was measured using early imaging by single photon emission computed tomography (SPECT) with N-isopropyl-(iodine-123)-p-iodoamphetamine (123I-IMP). When the balloon clamping test was performed the tracer was injected, and scanning was performed 35 minutes after removing the catheter. This tracer enabled a "memory of blood flow" during temporary ischemia to determine the character of quick diffusion and slow wash out, that could not be performed by other methods of cerebral blood flow measurement. SPECT with 123I-IMP can simplify the measurement of cerebral blood flow during the balloon clamping test.     

Measurement of regional cerebral blood flow by 133Xe inhalation method in patients with ruptured aneurysm

Regional cerebral blood flow (rCBF) was measured in 20 patients with ruptured aneurysm. rCBF images are obtained by 133Xe inhalation method using a ring type gamma camera (SET-020, Shimazu Co.) with high sensitivity collimator. Examinations are performed at acute stage (1-7 days after onset) and at subacute stage (8-14 days after onset). In acute stage, rCBF in the affected cerebral hemisphere has tendency to be low in 12 patients with angiospasm after the aneurysmal rupture comparing to those in patients without angiospasm. Low rCBF areas in cerebral hemisphere at acute stage were apt to become normal in subacute stage. rCBF measurement by 133Xe inhalation method seemed to be an useful method to evaluate rCBF in patients with ruptured aneurysm.     

Nuclear and DNA-binding proteins in human brain tumors

Nuclear proteins obtained from human brain tumor cell lines by differential salt extraction were subjected to high-resolution two-dimensional electrophoresis. Several hundred spots were detectable in the low salt (0.4 M NaCl) extract using silver staining. These patterns exhibited remarkable differences between the different cell lines we analyzed. A less complex pattern occurred when nuclei were subsequently treated with high salt (2.5 M NaCl/5 M urea). We compared the electropherograms from various human glioblastoma cell lines and found them very similar and even a high degree of similarity occurs between glioblastomas and other human tumor cell lines. Beside these more general observations we detected several proteins at least enriched in human glioblastomas which were totally absent in low grade astrocytomas and nonglial tumors. They could be separated from the bulk of nonspecific proteins by simple modifications of the isoelectric focusing conditions. From these results we conclude that nuclear proteins obtained by sequential salt extraction and separated by two-dimensional techniques may provide tumor specific proteins suitable for antibody production.     

Synthesis, characterization and evaluation of99mTc-L, L-ethylenecysteine dimer to assess regional cerebral blood flow

L,L-ethylenecysteine dimer (L,L-ECD) compound was synthesized with an overall yield 31.2% and a melting point 197–198°C. A systematic study has been carried out on the labelling of L,L-ECD with technetium-99m by ligand exchange via^99mTc-EDTA. The percent labelling yield of L,L-ECD was found ≥95% when 0.75 2.0 mg and 100 μg of L,L-ECD, disodium-EDTA and tin (II) were used, respectively, at pH 5.5. The produced^99mTc-L,L-ECD complex is stable, neutral and lipid soluble. The^99mTc-L,L-ECD complex penetrate the blood-brain barrier following intravenous injection in the tail vein of mice with an initial brain uptake equal to 0.9% at 2 minutes and a slow washout equal to 0.65% after 25 minutes. High uptake in the gastrointestinal tract (GIT) was observed. It reaches up to 38.9% at 2 minutes and increased as time pass reaching 48.3% at 60 minutes. These results indicate that this ligand is suitable for brain imaging and has a tendency to be excreted via liver and GIT due to its lipophilicity.     

Evaluation of gamma camera collimator for regional cerebral blood flow measurements using a 133Xe inhalation method

In the measurements of regional cerebral blood flow (rCBF) using inhalation of 133Xe gas, the activity present is generally limited in lower levels than those of usual brain scintigraphy. Measurements with low count-rate are usually resulted in diminishing the accuracies of results obtained. Therefore, it is necessary to make measurements using a high sensitive collimator for getting as much count-rate as possible when a gamma camera is used. The relationships among sensitivity and structures of multi-parallel collimator were mathematically analyzed. The results of analysis suggested that sensitivity usually increased by using a collimator with holes of reduced height and diameter. A prototype multi-parallel collimator with holes of low height and small diameter was made in our laboratory for testing sensitivity and resolution. The collimator possessing 1141 holes of 6 mm phi in hole diameter, 1.5 cm is hole height and septal thickness of 1 mm lead showed 24 times more sensitive than those of a general all purpose collimator supplied by the manufacturer. However, resolution measured in FWHM was of 9 to 14 mm at the collimator face and of 29 to 38 mm at 5 cm from the face. The results indicated that this collimator was useful enough in rCBF measurements with 133Xe inhalation using a gamma camera. The mathematical analysis however, suggested that optimum collimator for rCBF measurements was approximate 4.5 mm phi in hole diameter and 1.0 cm in hole height.     

Low electroosmotic flow measurement by tilting microchip

A novel method for low electroosmotic flow (EOF) rates measurement by tilting microchip which based upon the hydrostatic pressure conception and sampling zone method is described. Sampling zone could be detected in the tilting microchip but not in non-tilting one due to the hydrostatic pressure driven. The method is fulfilled to calculate low EOF rates by detecting the liquid flow velocity driven by hydrostatic pressure, and difference between the apparent mobility of the migrating analyte in two modes is caused by the effect of hydrostatic pressure. And then the EOF rates in unknown low EOF microchip can be calculated. Different microchannels modified with bovine serum albumin (BSA), myoglobin (MB) and polyvinyl alcohol (PVA) were used to verify the method, the EOF rate value was 1.73+/-0.03, 1.21+/-0.05, 0.34+/-0.04 x 10(-4)cm(2) V(-1)s(-1), respectively. The results obtained by the proposed method were agreed well with conventional methods.     

mTHPC-mediated photodynamic diagnosis of malignant brain tumors.

Radical tumor resection is the basis for the prolonged survival of patients suffering from malignant brain tumors such as glioblastoma multiforme. We have carried out a phase-II study involving 22 patients with malignant brain tumors to assess the feasibility and the effectiveness of the combination of intraoperative photodynamic diagnosis and fluorescence-guided resection (FGR) mediated by the second-generation photosensitizer meta-tetrahydroxyphenylchlorin (mTHPC). In addition, intraoperative photodynamic therapy (PDT) was performed. Several commercially available fluorescence diagnostic systems were investigated for their applicability in clinical practice. We have adapted and optimized a diagnostic system that includes a surgical microscope, an excitation light source (filtered to 370-440 nm), a video camera detection system and a spectrometer for clear identification of the mTHPC fluorescence emission at 652 nm. Especially in regions of faint fluorescence, it turned out to be essential to maximize the spectral information by optimizing and matching the spectral properties of all components, such as excitation source, camera and color filters. To sum up, on the basis of 138 tissue samples derived from 22 tumor specimens, we have been able to achieve a sensitivity of 87.9% and a specificity of 95.7%. This study demonstrates that mTHPC-mediated intraoperative FGR followed by PDT is a highly promising concept in improving the radicality of tumor resection combined with a therapeutic approach.     

Concurrent measurement of unbound genistein in the blood, brain and bile of anesthetized rats using microdialysis and its pharmacokinetic application

Genistein, the major isoflavone in soybeans, has been shown to have a wide range of effects. We used an HPLC-UV combined with microdialysis method to detect unbound genistein in rat blood, brain and bile. Genistein dialysates were eluted with a mobile phase containing acetonitrile-water (40:60, v/v, pH 3.5 adjusted by 0.1% acetic acid). Samples were separated using a phenyl (5 microm) column maintained at ambient temperature. The UV detector wavelength was set at 259 nm. The flow rate was 1.0 m/min. The limit of quantitation for genistein was 50 ng/ml. The in vitro recoveries of genistein were 31 +/- 1, 13 +/- 1 and 59 +/- 4% in microdialysis probes of blood, brain and bile, respectively (n = 4). Inter- and intra-assay accuracy and precision of the analysis were less than 10% in the concentration ranges of 0.05-5.0 microg/ml. A small ratio of genistein penetrates the blood-brain barrier (BBB) and goes through hepatobiliary excretion after genistein administration (10 or 30 mg/kg, i.v.). The brain-to-blood (AUC(brain)/AUC(blood)) and bile-to-blood (AUC(bile)/AUC(blood)) distribution ratios were 0.04 +/- 0.01 and 1.85 +/- 0.42, respectively for the dosage of genistein 30 mg/kg. After co-administration of cyclosporine, a P-glycoprotein (P-gp) inhibitor, the distribution ratios of genistein in brain and bile were not significantly altered. These results suggest that the BBB penetration and hepatobiliary excretion of genistein may not regulated by P-gp.     

Photobleaching-based flow measurement in a commercial capillary electrophoresis chip instrument

For microfluidic analytical instruments, a facile, fast, and accurate instrument test is highly demanded. The test includes the quantitative verification of the relationship between pressure drop and flow velocity for the hydrodynamic pump, between the electric voltage and electroosmotic flow (EOF) for the high-voltage supply, and the chip quality. The key point for the test is the measurement of the flow velocity. However, most currently available velocimetries cannot be directly used without any instrumental modification or adding extra instruments. We applied a recently developed Laser Induced Fluorescence Photobleaching Anemometer (LIFPA) for the instrument test through measuring fluid flow velocity in a microfluidic instrument with optical measurement without any modification and extra instrument. We have successfully used the method to test Caliper HTS 250 System from Caliper Life Sciences (Hopkinton, MA) with its own light source and detector. The experimental result demonstrates that this single-point method of measuring flow velocity can be easily used for accurate test of a microfluidic instrument in less than 10 min at extremely low cost without any modification and extra instrument.     

Systematic errors in the measurement of heat by flow microcalorimetry

It is shown that a number of systematic errors must be considered when performing heat measurements by flow microcalorimetry because the nature of the flow technique is such that substantial heat loss can be incurred. The conventional procedure of electrical calibration is found to be an inadequate correction parameter. Equations to account for the effects of thermal disequilibrium are derived from the basic principles incorporated in the Tian equation. The predicted relationships are tested by simple experiments and shown to be correct. The various correction parameters are measured for a wide range of flow rate and heat input conditions. A composite equation is presented which allows for the correction of heat loss while deconvoluting electrical heat from a heat of reaction. The total heat output rate from a flow calorimeter can be calculated for most experimental conditions by reference to this equation and to the tabulated correction values.     

Determination of thiamine by continuous flow chemiluminescence measurement

The chemiluminescence produced by oxidation of thiamine by ferricyanide in alkaline medium has been investigated by using a simple continuous flow analyser and a procedure developed for the determination of thiamine hydrochloride or nitrate in the range 2.00 x 10(-5)-5.00 x 10(-4)M (equivalent to 6.75-169 mug/ml thiamine hydrochloride) with coefficients of variation <2%. A measurement rate of 112/hr can be obtained. When applied to pharmaceutical formulations, the only interferent among common excipients and coexisting drugs is ascorbic acid. The results obtained for the assay of dosage forms compared well with those obtained by an official method and demonstrated an error <4%.     

Raman spectroscopic imaging for in vivo detection of cerebral brain metastases

We report for the first time a proof-of-concept experiment employing Raman spectroscopy to detect intracerebral tumors in vivo by brain surface mapping. Raman spectroscopy is a non-destructive biophotonic method which probes molecular vibrations. It provides a specific fingerprint of the biochemical composition and structure of tissue without using any labels. Here, the Raman system was coupled to a fiber-optic probe. Metastatic brain tumors were induced by injection of murine melanoma cells into the carotid artery of mice, which led to subcortical and cortical tumor growth within 14 days. Before data acquisition, the cortex was exposed by creating a bony window covered by a calcium fluoride window. Spectral contributions were assigned to proteins, lipids, blood, water, bone, and melanin. Based on the spectral information, Raman images enabled the localization of cortical and subcortical tumor cell aggregates with accuracy of roughly 250 μm. This study demonstrates the prospects of Raman spectroscopy as an intravital tool to detect cerebral pathologies and opens the field for biophotonic imaging of the living brain. Future investigations aim to reduce the exposure time from minutes to seconds and improve the lateral resolution.     

Determination of the migration times of flow measurement markers in CEC

The electroosmotic migration times of fructose, water, and phenol have been measured in several solutions. The electroosmotic flow rate was fundamentally dependent on buffer concentration and on the concentration of additives such as sodium chloride; additives such as methanol and sodium dodecylsulfate did not influence the flow rate, yet tetrabutylammonium bromide had a significant effect.     

Cerebral blood flow imaging in patients with moyamoya disease

The usefulness of IMP-SPECT and rCBF image by 133Xe inhalation method on rCBF in patients with moyamoya disease was studied. Six patients with moyamoya disease were diagnosed by cerebral angiogram, and STA-MCA anastomosis technique and EMS were done to reconstruct the rCBF. Low perfusion areas were detected around the cerebral infarction and hemorrhage and that of anterior and middle cerebral arteries by IMP-SPECT and 133Xe-rCBF image met by X-ray CT. After surgery, 4 out of 6 cases showed the improvement of rCBF by IMP-SPECT and 133Xe-rCBF image, as for clinical symptoms, there were reduction of TIA in 3 cases, and no rehemorrhage in 3 cases. In summary IMP-SPECT and 133Xe-rCBF image may be useful method to evaluate the rCBF in moyamoya disease and the change of rCBF post STA-MCA anastomosis technique and EMS.     

High-throughput biophysical measurement of human red blood cells

This paper reports a microfluidic system for biophysical characterization of red blood cells (RBCs) at a speed of 100-150 cells s(-1). Electrical impedance measurement is made when single RBCs flow through a constriction channel that is marginally smaller than RBCs' diameters. The multiple parameters quantified as mechanical and electrical signatures of each RBC include transit time, impedance amplitude ratio, and impedance phase increase. Histograms, compiled from 84,073 adult RBCs (from 5 adult blood samples) and 82,253 neonatal RBCs (from 5 newborn blood samples), reveal different biophysical properties across samples and between the adult and neonatal RBC populations. In comparison with previously reported microfluidic devices for single RBC biophysical measurement, this system has a higher throughput, higher signal to noise ratio, and the capability of performing multi-parameter measurements.     

Detection of collagens in brain tumors based on FTIR imaging and chemometrics

Fourier transform infrared (FTIR) imaging has been used as a molecular histopathology tool on brain tissue sections after intracranial implantation and development of glioma tumors. Healthy brain tissue (contralateral lobe) as well as solid and diffuse tumor tissues were compared for their collagen contents. IR spectra were extracted from IR images for determining the secondary structure of protein contents and compared to pure product spectra of collagens (types I, III, IV, V, and VI). Multivariate statistical analyses of variance and correspondence factorial analysis were performed to differentiate healthy and tumor brain tissues as well as their classification according to their secondary structure profiles. Secondary structure profiles revealed that no collagen was present in healthy tissues; they are also significantly different from solid and diffuse tumors (p < 0.05). Solid and diffuse tumors could be discriminated with respect to the secondary structure profile of fibrillar and non-fibrillar collagens, respectively. We can thus propose to develop FTIR imaging for histopathology examination of tumors on the basis of collagen contents.