Radiation exposure to anaesthetists during endovascular procedures.

Medical radiation exposure increases the likelihood of cataract formation. A personal dosimeter was attached to the left temple of 77 anaesthetists during 45 endovascular aortic aneurysm repairs and 32 interventional neuroradiology procedures. Compared with interventional neuroradiology, the median (IQR [range]) total radiation dose emitted by fluoroscopic equipment was significantly lower during endovascular aortic aneurysm repair (4175 (3127-5091 [644-9761]) mGy than interventional neuroradiology (1420 (613-2424 [165-10,840]) mGy, p < 0.001). However, radiation exposure to the anaesthetist's temple was significantly greater during endovascular aortic aneurysm repair (15 (6-41 [1-109]) µSv) than interventional neuroradiology (4 (2-8 [0-67]) µSv, p < 0.001). These data suggest that anaesthetists at our institution would have to deliver anaesthesia for ~1300 endovascular aortic aneurysm repairs and ~5000 interventional neuroradiology cases annually to exceed the general occupational limits, and ~10,000 endovascular aortic aneurysm repairs and ~37,500 interventional neuroradiology cases to exceed the ocular exposure limits recommended by the International Commission on Radiological Protection. Nevertheless, anaesthetists should be aware of the risk of ocular radiation exposure, and reduce this by limiting the time of exposure, increasing the distance from the source of radiation, and shielding.

Excimer fluorescence as a tool for monitoring protein domain dynamics applied to actin conformation changes based on circulary polarized fluorescence spectroscopy.

Fluorescence-detected circular dichroism (FDCD) was introduced into the study of protein conformation changes. Actin was used as a model protein which undergoes dynamic conformation changes as it polymerizes. Actin labeled with N-(1-pyrene)iodoacetamide (PIA) showed monomer fluorescence peak at 386 and 410 nm, and excimer fluorescence peak at around 480 nm. Excimer was formed by PIA-dimers labeled to different sites of amino acid residues. New information concerned with actin structural changes were monitored by fluorescence emission spectra excited with left- and right-circulary polarized light at 355 nm. FDCD intensities were shown as the difference in the fluorescence emission DeltaF, where DeltaF=(F (L)-F (R))/(F (L)+F (R)) denoting F (L) and F (R) as emissions obtained by excitation with left- and right-circulary polarized light. When solvent conditions of PIA-actin were changed by addition of NaCl, TFE, or ATP, DeltaF showed sensitive responses to these compounds. From the analysis of DeltaF (M) and DeltaF (E) which represent the peaks of DeltaF at the monomer- and excimer-emission band, the information concerned with the actin intrastructural changes were obtained. This method based on monitoring the excimer fluorescence with FDCD could be used for other proteins to extract finer structural changes that cannot be detected by the normal fluorescence spectroscopy.

Discordant iodine-123 metaiodobenzylguanidine uptake area reflects recovery time dispersion in acute myocardial infarction.

lodine-123 metaiodobenzylguanidine (MIBG) uptake was reported to be reduced compared to Tl-201 (Tl) in acute myocardial infarction (AMI). Within such an area, degrees of both sympathetic neural function and ischemic myocardial cell damage are considered to be greatly dispersed. These kinds of damage were reported to effect reporalization time in myocardial cells, and we evaluated our hypothesis that extension of the discordant MIBG uptake area correlates with recovery time (RT) dispersion and relate ventricular arrhythmias in AMI. MIBG and Tl images were obtained in AMI patients. Regional Tl or MIBG uptake was estimated in 9 segments of SPECT by using four-point scoring. The total score was the sum of scores in 9 SPECT segments. ATI-MIBG was calculated by subtracting the total MIBG score from the total Tl score. Corrected RT (RTc) was measured as a signal-averaged ECG. RTc dispersion was defined as the difference between maximal and minimal RTc. The patients were assigned to two groups (group A; < or = Lown 4a, group B; > or = Lown 4b) according to the results of 24-hour Holter monitoring. A positive correlation between RTc dispersion and ATI-MIBG was found. ATI-MIBG and RTc dispersion in group B were greater than those in group A. These results suggested that ATI-MIBG could be used to predict the development of malignant ventricular arrhythmias.

Neuroprotective effect of immunosuppressant FK506 in transient focal ischemia in rat: therapeutic time window for FK506 in transient focal ischemia.

Tacrolimus (FK506), an immunosuppressant currently used in clinic, is known to have neuroprotective properties. However, effects in focal ischemia are shown only in a endothelin induced middle cerebral artery (MCA) occlusion model or with filament technique at a relatively high dose. We have previously shown that FK506 had significant protective effects at a low dose of 0.3 mg kg(-1) when administered immediately after ischemia. In this study, we explored the therapeutic time window of FK506 at this low dose, in a transient focal ischemia model using filament technique. Male Sprague-Dawley rats were subjected to 2 h MCA occlusion and subsequent reperfusion. They received FK506 or vehicle (0.3 mg kg(-1)) i.v. at 30, 60 or 120 min after induction of ischemia, and were decapitated 24 h after ischemia. FK506 injected at 30 and 60 min significantly reduced cortical infarction volume (FK506 vs. vehicle; 30 min: 95 +/- 33 mm3 vs. 170 +/- 62 mm3, p < 0.05; 60 min: 93 +/- 45 mm3, vs. 168 +/- 35 mm3, p < 0.05, respectively). FK506 was ineffective when given at 120 min after ischemia. FK506 had no effect on edema formation, nor on the infarct volume in striatum. The therapeutic time window for this low dose of FK506 given i.v. is between 60 and 120 min in this model.

High-voltage electron microscopy in neurocytology.

The function of the living matter from biomolecules to the whole body of the organism is based on the structure. Consequently, structural information is essential for the understanding of the function of living organisms. The biological structures from biomolecules to cells and tissues are intimately related to each other, and changing their morphological, biochemical, and physiological properties dynamically according to the developmental and functional status of the organism. The molecular dynamics of living matter should be related to the function of whole body. We employed HVEM stereoscopy and morphometry for the purpose of combining the structural information at nanometer level with those of the micrometer level. Novel aspects of three-dimensional organization of neuronal and glial cell processes have been presented. This structural information together with morphometrical data could contribute to the elucidation of brain function.

Relation between myocardial response to dobutamine stress and sympathetic nerve activation in patients with idiopathic dilated cardiomyopathy: a comparison of 123I-MIBG scintigraphic and echocardiographic data.

It is likely that a close association exists between findings obtained by two methods: dobutamine stress echocardiography and 123I-MIBG scintigraphy. Both of these methods are associated with beta-adrenergic receptor mechanisms. This study was conducted to demonstrate the relation between myocardial response to dobutamine stress and sympathetic nerve release of norepinephrine in the failing heart. In 12 patients with heart failure due to idiopathic dilated cardiomyopathy, the myocardial effects of dobutamine stress were evaluated by low-dose dobutamine stress echocardiography: and sympathetic nerve function was evaluated by scintigraphic imaging with iodine-123 [123I] meta-iodobenzylguanidine (MIBG), an analogue of norepinephrine. Echocardiography provided quantitative assessment of wall motion and left ventricular dilation; radiotracer studies with 123I-MIBG provided quantitative assessment of the heart-to-mediastinum (H/M) uptake ratio and washout rate. Results showed that H/M correlated with baseline wall motion (r = 0.682, p = 0.0146), wall motion after dobutamine stress (r = 0.758, p = 0.0043), the change in wall motion (r = 0.667, p = 0.0178), and with left ventricular diastolic diameter (r = 0.837, p = 0.0007). In addition, the 123I-MIBG washout rate correlated with baseline wall motion (r = 0.608, p = 0.0360), wall motion after dobutamine stress (r = 0.703, p = 0.0107), and with the change in wall motion (r = 0.664, p = 0.0185). Wall motion, especially in the myocardial response to dobutamine stress, is related to sympathetic nerve activity in heart failure.

[Reverse redistribution of technetium-99m-tetrofosmin after acute coronary revascularization in patients with myocardial infarction].

This study qualified the severity and localization of reverse redistribution of technetium-99 m (Tc)-tetrofosmin rest imaging. Both Tc-tetrofosmin and thallium-201 (Tl) rest imaging with early images and delayed images were obtained in the subacute phase of myocardial infarction in 21 patients with first anterior myocardial infarction and with successful transluminal angioplasty (including stenting). Relative myocardial uptake (%uptake), degree of reverse redistribution (%), and washout rate (%) were evaluated quantitatively in 6 left ventricular segments (inferoseptal, anteroseptal, anterior, anterolateral, lateral, inferolateral and inferior) by circumferential profile analysis. The percentage reverse redistribution in the infarct area was larger in Tc-tetrofosmin imaging than in Tl imaging (p = 0.013). Reverse redistribution was most prominent in the anterior wall (anterior > anteroseptal > inferoseptal, p = 0.020). This suggests that infradiaphragmatic scatter is unlikely as the mechanism of reverse redistribution. The washout rate of Tc-tetrofosmin in the infarct area (reverse redistribution area) was higher than that in the normal area (non-reverse redistribution area), and was also higher than the washout rate of Tl imaging in the infarct area. The %uptake of delayed images in the infarct area was larger in Tc-tetrofosmin than that in Tl imaging, whereas %uptake of early images did not differ. The percentage reverse redistribution did not correlate with the degree of collateral circulation and the residual stenosis. In conclusion, reverse redistribution of Tc-tetrofosmin was more prominent in the infarct area, and this was due to the relatively lower uptake of reverse redistribution of Tc-tetrofosmin than delayed Tl images.

Increased function of the voltage-dependent calcium channels, without increase of Ca2+ release from the sarcoplasmic reticulum in the arterioles of spontaneous hypertensive rats.

It has been reported that the increased function of the voltage-dependent calcium channels (VDCC) in the artery is involved in the increase of peripheral resistance in hypertension, and that the sarcoplasmic reticulum (SR) in the artery plays an important role in preventing the development of hypertension via a buffering effect. However, no reports have described the role of VDCC and SR in resistance arterioles in the development or maintenance of hypertension. We investigated the function of VDCC and of SR in the cremaster arterioles of spontaneous hypertensive rats (SHR) and age-matched Wistar Kyoto rats (WKY). The changes in diameter and the intracellular calcium ion concentration ([Ca2+]i) in the microdissected arterioles, using fluorescent dyes, were measured with videomicroscopy. The KCl concentration-response curves were analyzed in 4- to 5- and 7- to 8-week-old SHR and WKY. The changes in the vascular diameter and [Ca2+]i in response to ryanodine, an alpha-1 adrenoceptor, and angiotensin-II stimulation were compared between the 7- to 8-week-old SHR and WKY. We found an increase in the Ca2+ influx by VDCC in the early hypertensive stage, but not in prehypertensive SHR. However, after the onset of hypertension, there were no significant differences from WKY in the SR function mediated by Ca2+-induced Ca2+ release or inositol 1,4,5-trisphosphate-induced Ca2+ release. In conclusion, an increased influx of Ca2+ in the cell membrane, without a buffering effect of SR, was associated with progression of hypertension in the cremaster arterioles of SHR.

Clinical usefulness of iodine 123-labeled fatty acid imaging in patients with acute myocardial infarction.

BACKGROUND: Iodine 123-labeled 15-(p-iodophenyl)-3R,S-methylpentadecanoic acid (BMIPP) has recently been developed, since normal myocardium metabolizes free fatty acids. This study investigated the clinical usefulness of BMIPP imaging in patients with acute myocardial infarction (MI), particularly in the detection of stunned myocardium in patients who underwent acute coronary revascularization. METHODS: The subjects were 41 patients with acute MI who had undergone emergency coronary revascularization. Both BMIPP and thallium-201 images at rest were obtained during the subacute phase. The myocardial distribution of radiotracers was quantified by generating circumferential count-distribution profile analysis. Initial 201Tl imaging, delayed 201Tl imaging, and BMIPP imaging were performed, and the mean count densities in the infarct region (initial 201Tl images [TL1], delayed 201Tl images [TL2], and BMIPP images in the infarct region [BM], respectively) were obtained. The differences between mean count densities (TL1-BM: BM subtracted from TL1; TL2-BM: BM subtracted from TL2) were also calculated. RESULTS: BM showed a higher correlation with wall motion data by echocardiography (WM) in the acute phase than other nuclear imaging tests, whereas TL2 showed the highest correlation with WM in the chronic phase. Acute to chronic WM improvement showed a good correlation with TL2-BM. CONCLUSION: Single photon emission computed tomography imaging with BMIPP is a candidate for providing the "memory image" of ischemic damage, whereas TL2 reflects all viable tissue. The mismatch between the tracers can serve as an indicator of myocardial stunning.

A specific Thy-1 molecular epitope expressed on rat mesangial cells.

An experimental model of proliferative glomerulonephritis induced by antibodies against the Thy-1 antigen has been established and used to study the pathological sequence from mesangiolysis to mesangial proliferation. However, the functional role of the Thy-1 molecule distributed on rat glomerular mesangial cells remains unknown, so the present study was undertaken to determine the precise subcellular localization of Thy-1 molecules in vitro using two anti-Thy-1 monoclonal antibodies, 1-22-3 and OX-7. Immunofluorescence and immunoelectron microscopic studies in combination with laser scanning analysis showed that the localization of 1-22-3 and OX-7 bound to cultured rat mesangial cells differed, particularly when cocultured with vascular endothelial cells. An epitope recognized by 1-22-3 was concentrated specifically on the mesangial cell surfaces facing the neighboring endothelial cells. In contrast, OX-7 bound to mesangial cell surfaces and extracellular parts in a diffuse pattern independent of contact with endothelial cells. This finding is consistent with our previous ultrastructural study in which the reactivity of 1-22-3 with normal kidney tissue was limited to mesangial cell surfaces facing endothelial cells. These results led us to conclude that the specific Thy-1 molecular epitope recognized by 1-22-3 is associated with points anchoring mesangial and endothelial cells, where this anti-Thy-1 antibody binds after injection in vivo, resulting in mesangial cell detachment from the vascular capillary wall, mesangiolysis and mesangial cell dysfunction. We believe that the critical epitope detected by 1-22-3 in this study plays an important role in mesangial cell function and injury.

Changes of intermediate filaments in cultured human glioma cells with various growth factors and cytokines using high voltage immunoelectron microscopy.

Morphological and immunocytological changes of intermediate filaments of cultured human malignant glioma cells were studied by adding various growth factors or cytokines using stereoscopic high voltage electron microscopy operated at 1,000 kV. The gold-colloid immuno-cytochemical method was used to stain GFAP and vimentin. Growth rate of tumor cells increased when EGF, TGF-alpha, and PDGF administered and decreased when FGF, TNF, and CLN-IgG administered. Morphological changes of cells were not remarkable when EGF, PDGF, IL-1, and FGF were administered. The cytoplalsmic organellaes were damaged after administrating TNF and CLN-IgG to cells.

Comparative accuracy of various Tl-201 reinjection imaging protocols to detect myocardial viability.

UNLABELLED: The conventional exercise-3 hours-redistribution thallium-201 [201Tl] imaging protocol has been recognized to be suboptimal for reliable detection of myocardial viability. Although 201Tl rest-injection after exercise has improved detection of viable myocardium, it is still underestimated in some patients. The present study was designed to compare detection of viable myocardium in five separate imaging steps: step 1: initial-exercise imaging, step 2: delayed-exercise imaging, step 3: Tl-201 reinjection imaging after delayed-exercise imaging, step 4: separate day rest-injection imaging, and step 5: separate day delayed-rest imaging. The study group consisted of 22 patients scheduled for coronary revascularization (either percutaneous transluminal coronary angioplasty or coronary bypass surgery). Pre- and postintervention echocardiographic wall motion and thickness served as independent markers of myocardial viability. RESULTS: Accuracy in identifying myocardial viability gradually improved incrementally from 201Tl imaging step 1 to step 5. The positive predictive value, negative predictive value and overall accuracy were best for the separate day delayed-rest study (step 5) at 90%, 33% and 78%, respectively. Myocardial segments had fixed defects on separate day delayed-rest 201Tl imaging (step 5), but nevertheless echocardiographic evidence of myocardial viability indicated less severe defects than segments judged nonviable by echocardiography (p = 0.021). The overall accuracy of separate day delayed-rest imaging (step 5) in predicting viability improved to 88% when segments with moderate or mild defects were considered viable. In conclusion, the most reliable predictor of myocardial viability with 201Tl imaging is defect severity on separate day delayed-rest images.

Messenger RNA for the guanine nucleotide-binding regulatory protein (G protein) is reduced in the acute ischemic myocardium.

It has been reported that the function of the guanine-binding regulatory protein (G protein), especially the alpha subunit of the stimulatory G protein (Gs alpha), in myocardium is decreased with acute ischemia. However, it is unclear whether this decrease is due to transcriptional or post-transcriptional changes. Moreover, no studies have examined the distribution of G protein mRNA in ischemic myocardium using in situ hybridization. The purpose of this study was to explore alterations in mRNA of G proteins (Gs and Gi) in ischemic hearts using in situ hybridization. We measured the levels of mRNA for Gs alpha and Gi alpha in ischemic and non-ischemic myocardium by in situ hybridization using a radioisotope imaging system. We compare these mRNA levels in ischemic and non-ischemic myocardium with Northern blot analysis and the protein levels of G proteins by Western blot analysis. The mRNA for Gs alpha and Gi alpha was distributed diffusely in normal hearts. Levels of mRNA detected by in situ hybridization were substantially reduced by acute ischemia, and these results were confirmed by Northern and Western blot analysis. These results suggest that decreased levels of mRNA and protein for G proteins may underlie the impaired function of the receptor--G protein--adenylate cyclase system in ischemic myocardium. In addition, quantitative evaluation of mRNA is possible by in situ hybridization and correlates well with Northern analysis.

Three-dimensional organization of neuronal and glial processes: high voltage electron microscopy.

Neurons and glia cells in the mammalian central nervous system have many complicated processes. They are too fine for light microscopic study and too complicated and widely spread for thin section electron microscopy. High-voltage electron microscopic (HVEM) stereo observation of thick Golgi preparation provides detailed 3-D images of their processes. Three-dimensional fine structures of astocytic processes in the neuropile and on the surface of neuronal somata, and those of the ruffed cell axon initial segment and thorny excrescences of CA3 pyramidal cell dendrites, are elucidated with the aid of HVEM stereoscopy of thick Golgi preparations. In addition, some results obtained by 3-D morphometrical analysis of dendritic spines using HVEM stereo images are shown. The examples presented here clearly show the usefulness of high-voltage electron microscope stereo observation of thick specimens for detailed morphological and morphometric study of the central nervous system.

Localization of protein kinase C alpha, beta and gamma subspecies in sensory axon terminals of the rat muscle spindle.

The localization of protein kinase C (PKC) alpha, beta and gamma subspecies in sensory axon terminals of muscle spindles in the plantar lumbrical muscles of rat was investigated by light and electron microscopic immunocytochemistry using monoclonal and polyclonal antibodies. Immunoreactivity for these subspecies was detected specifically in sensory axon terminals which wound spirally around the intrafusal muscle fibres of the muscle spindle. Immunostaining was found to be stronger with polyclonal than with monoclonal antibodies. By electron microscopy, immunoreactivity for alpha, beta and gamma subspecies was almost diffusely distributed in the cytoplasm of the axon terminal, and the overall pattern of distribution of immunoreactivity was similar for all three subspecies. In the cases of alpha and beta subspecies, some intensely immunostained regions were found in the cytoplasm, but no definite subcellular structures corresponding to such regions could be identified. Considering that PKC plays a crucial role in the regulation of ion channels, it is suggested that PKC might be involved in the control of mechanoelectric transduction in sensory axon terminals.

Adaptation of hybridoma cells to higher ammonia concentration.

Using two mouse-mouse hybridoma cell lines, the response to ammonia step and serial changes was investigated in batch and continuous cultures with serum-free medium. The inhibitory effect of ammonia on cell growth depended on the cultivation mode, and differed markedly between cell lines. The cell line, 4C10B6 producing IgG monoclonal antibody against Pseudomonas, showed a high adaptation ability to ammonia. The 4C10B6 cells could grow under ammonia concentration as high as 21 mmol/l NH4Cl with a viability of 80% in the continuous culture with serial increase in ammonia concentration. Whereas, in the batch culture with ammonia step change the cell growth completely ceased at 12 mmol/l NH4Cl. The other cell line, TO-405 producing IgG monoclonal antibody against hepatitis B surface antigen, could not adapt to ammonia, and the cell growth did not occur at 9 mmol/l NH4Cl even under the ammonia serial change.

Three-dimensional morphometrical study of dendritic spines of the granule cell in the rat dentate gyrus with HVEM stereo images.

Number, length, and diameters of dendritic spines of the granule cell in the dorsal leaf of the rat dentate gyrus were measured by using high-voltage electron microscope stereo images of 5-micron-thick Golgi preparations with the aid of a three-dimensional image analyzer system. Spine densities of 2.02 +/- 0.28, 2.28 +/- 0.33, and 3.36 +/- 0.35 per 1 micron at distal, middle, and proximal portions of the dendrite were obtained. These values were about 1.6-fold of the previous light microscopical report. Mean three-dimensional spine length were 1.244 +/- 0.506 micron, 1.262 +/- 0.563 micron, and 1.254 +/- 0.584 micron at distal, middle, and proximal portions, respectively, which were about 1.4 times longer than those measured in two dimensions. By using measured morphometrical parameters of spines such as lengths, diameters, and population densities, total spine surface areas of 2.401 micron 2, 2.806 micron 2, and 4.180 micron 2 per 1 micron of the dendrite at distal, middle, and proximal portions, respectively, were obtained. The total surface area of dendrite was about doubled by the addition of the spines at each dendritic portion. The advantageous features and the problems of the present method are discussed.

High-voltage electron microscopy of E-PTA-stained synaptic junctions in the rat frontal cortex.

The E-PTA-stained synaptic junctions in the adult rat frontal cortex were examined with high-voltage electron microscopy (HVEM). Perforated whole synaptic junctions were clearly shown in the stereo image. The E-PTA staining procedure provides a useful marker for studies of the 3-dimensional structure of synaptic junctions by means of HVEM.