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Sequential changes of gap junctions (GJs), tight junctions (TJs) and desmosomes (DSs) between hepatocytes during restorative proliferation were studied in rats after a single intraperitoneal administration of 200 mg/kg thioacetamide (TAA). Antibody against connexin 32 was used to demonstrate GJs; simultaneously the changes in TJs and DSs were studied using antibodies against 7H6 protein and desmoplakins. Propidium iodide and bromodeoxyuridine were used to recognize necrotic and proliferative cells. GJs were evenly distributed in early necrotic hepatocytes at 16 h after TAA treatment, then disappeared from necrotic and surrounding cells at 24 h. At 48 h, GJs had disappeared completely from hepatocytes in whole liver lobules, while many hepatocytes were heavily labelled with BrdU. At 72 h, GJs reappeared, firstly in perinecrotic areas. At 96 h after treatment, when the injured areas had disappeared and restorative proliferation ceased, GJs were distributed evenly throughout the lobules. Immunohistochemical observation of GJs in centrilobular, perinecrotic and periportal areas after TAA-induced hepatic necrosis was confirmed by counting the number of connexin-32-positive spots in the respective areas. TJs and DSs disappeared from necrotic cells at 24 h, but then increased between 24 and 48 h in perinecrotic areas, though the increased intensity of these junctions was more evident at 48 h. At 72 h, localization of TJs and DSs returned to normal. These results suggest that during the course of acute hepatic injury, GJs (cell-cell communication) behave differently from other intercellular junctions.

Quantitative analysis of arterial dimensions from high frequency intravascular ultrasound images (30 MHz) may be hampered by strong blood scattering. Replacement of blood by saline is one method to provide a clear view of the arterial lumen; another method is that of temporal averaging of successive ultrasound images. The accuracy of this latter method was tested by comparing the lumen area measurements on the temporal-averaged image, with the data of the same cross-section obtained from the single-frame and saline-filled images. The mean lumen area measured on the temporal-averaged images was similar to that measured on the single-frame images (mean difference: -0.02 +2- 1.16 mm2; p = ns). The mean lumen area of the saline-filled images was 8% larger than the values obtained from the temporal-averaged and single-frame images (mean difference: -1.14 +/- 0.85 mm2, p < 0.05), probably due to the difference in sound velocity between saline and blood. Intraobserver variation in the averaging method were 2.4 times smaller than the measurements of the single-frame images and close to the data obtained by saline injection (variation coefficient: single-frame: 8.8%; temporal-averaged: 3.6%; saline-filled: 2.9%). It is concluded that analysis from temporal-averaged images is more efficient, enabling accurate and reproducible measurement of the luminal dimensions from images containing blood scattering echoes. This technique is suitable to replace the laborious saline injection method and facilitates off-line quantitative analysis.

We examined the role of the ether a go-go (eag) gene in modulation of K+ currents and the possibility of its protein product Eag as a subunit in the heteromultimeric assembly of K+ channels by voltage-clamp analysis of larval muscle membrane currents. Previous DNA sequence studies indicate that the eag gene codes for a polypeptide homologous to, but distinct from, the Shaker (Sh) K+ channel subunits (Warmke et al., 1991), and electrophysiological recordings revealed allele-specific effects of eag on four identified K+ currents in Drosophila larval muscles (Zhong and Wu, 1991). Further studies of eag alleles indicated that none of the eag mutations, including alleles producing truncated mRNA messages, eliminate any of the four K+ currents, and that the mutational effects exhibit strong temperature dependence. We found that both W7, an antagonist of Ca2+/calmodulin, and cGMP analogs modulated K+ currents and that their actions were altered or even abolished by eag mutations. These results suggest a role of eag in modulation of K+ currents that may subserve integration of signals at a converging site of the two independent modulatory pathways. The Sh locus is known to encode certain subunits of the IA channel in larval muscle. The existence of multiple eag and Sh alleles enabled an independent test of the idea of Eag as a K+ channel subunit by studying IA in different double-mutant combinations. An array of allele-specific interaction between eag and Sh was observed, which reflects a close association between the Sh and eag subunits within the IA channel. Taken together, our data strengthen the possibility that the eag locus provides a subunit common to different K+ channels. The role of the eag subunit for modulating channels, as opposed to that of Sh subunits required for gating, selectivity, and conductance of the channel, suggest a combinatorial genetic framework for generating diversified K+ channels.

The cAMP concentration in Drosophila is increased by mutations of the dunce (dnc) gene and decreased by mutations of the rutabaga (rut) gene. Such mutants provide a unique means for exploring the role of cAMP in functional and developmental regulation of membrane currents. Four distinct K+ currents have been identified in Drosophila larval muscle fibers, i.e. the voltage-activated transient IA and delayed IK and the Ca(2+)-activated fast ICF and slow ICS. Results from our voltage-clamp studies indicated that both IA and IK were increased in dnc alleles. Normal muscle fibers treated with dibutyryl-cAMP showed a similar increase of IA, but no significant effect on IK. In contrast to the dnc alleles, the rut mutations appeared to enhance ICS greatly while leaving the amplitude of other currents largely unchanged. In addition, the dibutyryl-cAMP-induced increase in IA was not observed in rut fibers. Caffeine and W7, which are known to interfere with several second messenger pathways, also modulated K+ currents in larval muscle fibers. The currents in dnc and rut fibers showed strikingly altered responses to caffeine and W7. The results demonstrate that the various K+ currents in Drosophila muscles are affected by altered cAMP cascades in the mutants. The fact that not all dnc and rut mutant defects can be mimicked or reversed by acute application of cAMP suggests that long-term modulation of K+ currents by cAMP may involve mechanisms distinct from the short-term effect of cAMP.

Forensic applications of DNA typing data require the estimation of the frequencies of all observed alleles, which is currently done by a fixed set of groupings (binning) of alleles in a database. Recently it's validity has been questioned on the ground that when a DNA fragment size is close to a bin boundary, the frequencies of all adjacent bins should be added. On the contrary, the current forensic database indicates that when the match window of a DNA fragment overlaps 2 bins, it is enough to consider the bin with the larger frequency, and this never underestimates the frequency within the match interval with the current choice of fixed-bin widths. On average, the current fixed-bin procedure yields an allele frequency at least 2-fold higher than that of a floating-bin.

The tight junction is an essential element of the intercellular junctional complex; yet its protein composition is not fully understood. At present, only three proteins, ZO-1 (Stevenson, B. R., J. D. Siliciano, M. S. Mooseker, and D. A. Goodenough. 1986. J. Cell Biol. 103:755-766), cingulin (Citi, S., H. Sabanay, R. Jakes, B. Geiger, and J. Kendrick-Jones. 1988. Nature (Lond.). 333:272-275) and ZO-2 (Gumbiner, B., T. Lowenkopf, and D. Apatira. 1991. Proc. Natl. Acad. Sci. USA. 88:3460-3464) are known to be associated with the tight junction. We have generated a monoclonal antibody (7H6) against a bile canaliculus-rich membrane fraction prepared from rat liver. This 7H6 antigen was preferentially localized by immunofluorescence at the junctional complex regions of hepatocytes and other epithelia, and 7H6-affiliated gold particles were shown electron microscopically to localize at the periphery of tight junctions. Immunoblot analysis of a bile canaliculus-rich fraction of rat liver using 7H6, anti-ZO-1 antibody (R26.4C), and anti-cingulin antibody revealed that 7H6 reacted selectively with a 155-kD protein, whereas R26.4C reacted only with a 225-kD protein. Anti-cingulin antibody reacted solely with 140 and 108-kD proteins, indicating that the protein recognized by 7H6 is immunologically different from ZO-1 and cingulin. Immunoprecipitation of detergent extracts obtained from metabolically labeled MDCK cells with R26.4C coprecipitated a 160-kD protein, which corresponds to ZO-2, with ZO-1. However, 7H6 did not react with the 160-kD protein. These results strongly suggest that the 7H6 antibody recognizes a novel tight junction-associated protein different from ZO-1, cingulin and ZO-2.

Background: Intravascular ultrasound was used to assess the immediate effect of balloon angioplasty on the superficial femoral artery., Methods and Results: In 16 consecutive patients, corresponding ultrasonic cross sections (n = 72) before and after balloon angioplasty were qualitatively and quantitatively analyzed. The qualitative data were compared with angiographic findings. Before intervention, the angiographically demonstrated obstructive lesions were confirmed by intravascular ultrasound. Ultrasound enabled discrimination between soft (n = 43) and hard (n = 29) lesions, as well as between eccentric (n = 57) and concentric (n = 15) lesions. After balloon angioplasty, the presence of a dissection assessed angiographically in 14 patients was confirmed by intravascular ultrasound. Additional morphological information provided by ultrasound included plaque rupture in 14 patients and internal lamina rupture in six patients. Quantitative ultrasound data revealed an increase in free lumen area from 9.7 +/- 4.7 to 18.3 +/- 7.0 mm2 (p less than or equal to 0.01), an increase in minimal lumen diameter from 2.8 +/- 0.7 to 3.6 +/- 1.2 mm (p less than or equal to 0.01), and an increase in media-bounded area from 21.7 +/- 5.4 to 28.3 +/- 5.8 mm2 (p less than or equal to 0.01). The lesion area for the majority of cases (n = 32) remained unchanged (13.0 +/- 4.9 mm2 versus 12.9 +/- 4.6 mm2), or the lesion disappeared partially (from 9.1 +/- 0.9 to 4.3 +/- 1.4 mm2, n = 4, p less than or equal to 0.01) or totally (from 10.1 +/- 4.2 to 0 mm2, n = 6). Stretching of the arterial wall was further evidenced by medial thinning from 0.55 +/- 0.19 to 0.34 +/- 0.11 mm (p less than or equal to 0.01)., Conclusions: Luminal enlargement by balloon dilatation is achieved primarily by overstretching the arterial wall, with the lesion volume remaining practically unchanged. Overstretching is accompanied almost always by dissection and plaque rupture and occasionally by an internal lamina rupture.

The spatial distribution of capillaries in the optic chiasma was studied quantitatively in 51 fetuses, 8 newborns and 4 babies to reveal that the capillary density was lower in the chiasmal median portion, which was therefore a weak point of blood supply. This finding may be a contributing factor for bitemporal hemianopsia due to neoplasms from the sella turcica.

beta-Sitosterol and eupatilin were isolated and identified from Artemisia argyi. Both of them showed marked inhibition of platelet aggregation.

Activity-dependent synaptic plasticity has been implicated in the refinement and modification of neural circuits during development and learning. Previous studies show that activity-induced facilitation and potentiation are disrupted at larval neuromuscular junctions in the memory mutants dunce (dnc) and rutabaga (rut) of Drosophila. The diminished learning-memory capacity and synaptic transmission plasticity have been associated with altered cAMP levels since dnc affects the cAMP-specific phosphodiesterase and rut affects adenylate cyclase. In this study, the morphology of larval motor axon terminals was examined by anti-HRP immunohistochemistry. It was found that the numbers of terminal varicosities and branches were increased in dnc mutants, which have elevated cAMP concentrations. Such increase was suppressed in dnc rut double mutants by rut mutations, which reduce cAMP synthesis. More profuse projections of larval motor axons have also been reported in double-mutant combinations of ether à go-go (eag) and Shaker (Sh) alleles, which display greatly enhanced nerve activity as a result of reduction in different K+ currents. Therefore, we examined combinations of dnc and rut with eag and Sh mutations to explore the possible relation between activity- and cAMP-induced morphological changes. We found that the expanded projections in dnc were further enhanced in double mutants of dnc with either eag or Sh, an effect that could again be suppressed by rut. The results provide evidence for altered plasticity of synaptic morphology in memory mutants dnc and rut and suggest a role of cAMP cascade in mediating activity-dependent synaptic plasticity.

Galactosyl-neoglycoalbumin (NGA) can bind with the asialoglycoproteins receptor specifically on the surface of mammal's liver. We prepared NGA in different carbohydrate density and made some animal and clinical tests with technetium-99m labeled NGA. The maximum absorption of 99mTc-NGA-21 was 82.6% in liver. We also got the satisfactory radiographs of liver. NGA was synthesized by the covalent coupling of a carbohydrate bifunctional reagent, 2-imino-2-methoxyethyl-1-thio-galactose (4), to human serum albumin. The cyanomethyl-1-thio-glycoside of beta-D-galactose (3) was prepared from the pseudothiourea derivative (2) which was synthesized by acetylation, bromination and substitution of galactose (3) could be converted to (4) by treatment with sodium methoxide.

Voltage-clamp analysis of Drosophila larval muscle revealed that ether à go-go (eag) mutations affected all identified potassium currents, including those specifically eliminated by mutations in the Shaker or slowpoke gene. Together with DNA sequence analysis, the results suggest that the eag locus encodes a subunit common to different potassium channels. Thus, combinatorial assembly of polypeptides from different genes may contribute to potassium channel diversity.

The tritiated alpha-isobutylhydroxybenzyl alcohol (3H-G018) was found to be able to bind benzodiazepine (BZ) receptor on the rat brain membrane. The 125I labeled G-018 also had the similar binding activity with this receptor. In the present investigation, we observed that gastrodigenin and its derivatives inhibited the 125I-G018 binding of BZ receptor competitively, but no inhibition was observed with gastrodin. The results suggested that Gastrodigenin was bound to BZ receptor, and otherwise, gastrodin would have no direct interaction with BZ receptor. Probably, it might metabolize into gastrodigenin in vivo, and then got through the blood brain barrier and bound to BZ receptor, which mediated its pharmacological effects on the central nervous system.

Synaptic transmission was examined in Drosophila mutants deficient in memory function. These mutants, dunce and rutabaga, are defective in different steps of the cyclic adenosine 3',5'-monophosphate (cAMP) cascade. In both dunce and rutabaga larvae, voltage-clamp analysis of neuromuscular transmission revealed impaired synaptic facilitation and post-tetanic potentiation as well as abnormal responses to direct application of dibutyryl cAMP. In addition, the calcium dependence of transmitter release was shifted in dunce. The results suggest that the cAMP cascade plays a role in synaptic facilitation and potentiation and indicate that synaptic plasticity is altered in Drosophila memory mutants.

An intravascular ultrasonic imaging device (40 MHz) was used to obtain in vitro ultrasonic images and matching histologic cross-sections, derived from human vascular specimens. The feasibility of assessing vessel wall morphology as well as the ability to accurately document plaque thickness was determined. Based on the echogenicity of the arterial media, intravascular ultrasound could distinguish muscular arteries from elastic arteries, veins, and bypass grafts. The hypoechoic media only present in the muscular type of artery proved to be an essential landmark to document superimposed atherosclerosis. Plaque thickness calculated in these arteries showed close relationship with the corresponding histologic cross-section. Using real-time in vivo intravascular imaging (30 MHz), the morphology of the vessels interrogated was studied. The dynamic change of the arterial wall, as well as the outcome after intervention, is discussed.

Several novel bioactive components isolated from Chinese medicinal plants will be presented. These include novel maytansinoid tumor inhibitors, some new ent-kaurane and rosane diterpenoids from Mallotus anomalus Meer et Chun (Euphorbiaceae), as well as novel insecticide, stemona alkaloids from Stemona parviflora C. H. Wright (Stemonaceae). Both are native plants of Hainan island, China. 2D NMR techniques such as mono and hetero-COSY, NOESY, COLOC as well as 1H-NMR line broadening effect were utilized for structure elucidation. The separation techniques, structure elucidations and bioassay results will be reported.

This study investigated the accuracy and reproducibility of a computer-aided method for quantification of intravascular ultrasound. The computer analysis system was developed on an IBM compatible PC/AT equipped with a framegrabber. The quantitative assessment of lumen area, lesion area and percent area obstruction was performed by tracing the boundaries of the free lumen and original lumen. Accuracy of the analysis system was tested in a phantom study. Echographic measurements of lumen and lesion area derived from 16 arterial specimens were compared with data obtained by histology. The differences in lesion area measurements between histology and ultrasound were minimal (mean +/- SD: -0.27 +/- 1.79 mm2, p greater than 0.05). Lumen area measurements from histology were significantly smaller than those with ultrasound due to mechanical deformation of histologic specimens (-5.38 +/- 5.09 mm2, p less than 0.05). For comparison with angiography, 18 ultrasound cross-sections were obtained in vivo from 8 healthy peripheral arteries. Luminal areas obtained by angiography were similar to those by ultrasound (-0.52 +/- 5.15 mm2, p greater than 0.05). Finally, intra- and interobserver variability of our quantitative method was evaluated in measurements of 100 in vivo ultrasound images. The results showed that variations in lumen area measurements were low (5%) whereas variations in lesion area and percent area obstruction were relatively high (13%, 10%, respectively). Results of this study indicate that our quantitative method provides accurate and reproducible measurements of lumen and lesion area. Thus, intravascular ultrasound can be used for clinical investigation, including assessment of vascular stenosis and evaluation of therapeutic intervention.

An anatomical and electrophysiological study of Drosophila mutants has been made to determine the effect of altered electrical activity on the development and maintenance of larval neuromuscular junctions. We examined motor axon terminals of (1) hyperexcitable mutants Shaker (Sh), ether a go-go (eag), Hyperkinetic (Hk), and Duplication of para+ (Dp para+); and (2) mutants with reduced excitability, no action potential (napts) and paralytic (parats 1). Nerve terminals innervating larval body-wall muscles were visualized by using anti-HRP immunocytochemistry, which specifically stains neurons in insect species. In wild-type larvae, motor axon terminals were distributed in a stereotypic fashion. However, in combinations of eag and Sh alleles, the basic pattern of innervation was altered. There was an increase in both the number of higher-order axonal branches over the muscles and the number of varicosities on the neurites. A similar phenomenon was found in the double mutant Hk eag and, to a lesser extent, in Dp para+ and Dp para+ Sh mutants. It is known that at permissive temperature the napts, but not parats 1, mutation decreases excitability of larval motor axons and suppresses the behavioral phenotypes of Sh, eag, and Hk. In the mutant napts (reared at permissive temperature), a slight decrease in the extent of branching was observed. Yet, when combined with eag Sh, napts completely reversed the morphological abnormality in eag Sh mutants. No such reversion was observed in parats 1 eag Sh mutants. The endogenous patterns of electrical activity at the neuromuscular junction were analyzed by extracellular recordings in a semi-intact larval preparation. Recordings from wild-type body-wall muscles revealed rhythmic bursts of spikes. In eag Sh mutants, this rhythmic activity was accompanied by or superimposed on periods of strong tonic activity. This abnormal pattern of activity could be partially suppressed by napts in combination with eag Sh.

It is known that gamma-aminobutyric acid (GABA) plays an important role in the pathophysiology of epilepsy. Under physiological conditions it is very difficult for GABA to cross the blood-brain barrier. It is likely that the esters of GABA, Which easily enter the brain, will have useful therapeutic properties. With this in mind, we have designed and synthesized 8 derivatives (IVa to IVh) of gamma-aminobutyric acid with an imine to link a lipophilic carrier in order to facilitate the passage of gamma-aminobutyric acid to cross the blood-brain barrier. All the eight synthesized title compounds were evaluated as anticonvulsants in mice via the Antiepileptic Drug Development Program of NIH. The compounds IVb and IVf showed anticonvulsant activity.

Xifukang is a compound preparation of Chinese herbs consisting of Hanbane grugs mainly. Since 1987, the significant efficiency have been obtained in treatment of 53 patients suffering from silicosis by self-control study. The results indicated that the clinical manifestations including dyspnea, cough, sputum production, chest pain, weakness, etc. were markedly improved (P less than 0.01) and measurements of pulmonary function (FVC, FEV and MVV) significant enhanced (P less than 0.01). After treatment 20 cases roentgenogram exam showed that the lung's clarity and the limited emphysema were improved, the silicolic nodule and mass-mergence opacity of 3 cases lessened in some degree. By discussing the effect of Xifukang which might lower the collagen protein content of experimental silicosis of Wistar rats and improve pathomorphous. The authors concluded that the therapeutic mechanism of Xifukang could be the actions of this remedy on promoting blood circulation to eliminate blood stasis dredging microcirculation, increasing ventilation/perfusion (VA/Q), protecting dust-cells, resisting fibrosis, regulating immune function, enhancing lung clearance, postponing and preventing development of silicosis.

Compounds were isolated from the aqueous extract of the leaves of Crataegus pinnatifida var. major and seven of them were identified as 2-(4-hydroxybenzyl)malic acid, quercetin, hyperin, vitexin, rhamnosylvitexin, diethylamine hydrochloride and sorbitol respectively by UV, IR, NMR, MS, some chemical techniques by comparison with the standard spectra of known compounds.

A malignant astrocytoma cell line TJ-8510 derived from temporoparietal astrocytoma Grade III was established. At present, it has survived for more than 34 months passaging through 220 generations. According to its features in cell kinetics, cytogenetics, morphology, biochemistry and heterotransplantation, it was still identified as malignant astrocytoma. This cell line may be useful in further research on glioma.

Two unusual cases with coronary artery fistula were reported. The diagnoses were first made by color Doppler flow imaging and confirmed by selective coronary angiography and operation. In the first case, color Doppler revealed a continuous flow signal within the dilated left coronary artery and a turbulent jet within the right atrium. In the second case, color Doppler showed turbulent flow signals which could be traced from the proximal left coronary artery, along the left ventricular posterior wall and into the left ventricle. The high velocities at the site of drainage of fistula were recorded by the continuous-wave Doppler and the calculated pressure gradient confirmed that the abnormal flow was from the systemic circulation. We are of the opinion that color Doppler flow imaging is a reliable technique for detecting coronary artery fistula and the combination of color Doppler with continuous-wave Doppler is essential for evaluating the hemodynamics of the shunt flow.

The blockade of K+ channels and enhancement of neuromuscular transmission by dendrotoxin (DTX), a convulsant peptide from mamba snake venom, were examined in normal and mutant larval preparations of Drosophila. Two-microelectrode voltage-clamp experiments showed that DTX reduced the transient K+ current, IA, in muscle membrane. This effect was suppressed by raising the Mg2+ concentration or by lowering the temperature. The interaction of DTX with Mg2+ was further analyzed at a low cation concentration, at which DTX reduced both IA and the delayed rectifier IK. These results were correlated with the action of DTX on the neuromuscular junction. Its facilitatory effect on excitatory junctional potentials (EJPs) was relatively mild but the effect was drastically enhanced when combined with certain mutations and K+ channel blocking drugs, leading to repetitive or prolonged giant EJPs. Only the mutations or drugs that reduced IK or the Ca2(+)-dependent K+ current, ICF, could yield these synergistic effects with DTX. In contrast, the abnormal EJPs caused by the mutation or drug that blocked IA were not further enhanced by DTX, indicating that DTX also affects IA at the neuromuscular junction. Thus, the A-type K+ channels in muscle and nerve terminals appeared very similar in their sensitivity to the specific toxin, drugs and mutations examined here.