Role of contrast-enhanced ultrasound in the evaluation of post-liver transplant vasculature.

Liver transplantation is a frequently used treatment for patients with end-stage liver disease and ultrasound is often the first-line imaging technique for detection of vascular complications after liver transplant. Although colour Doppler ultrasound is a good screening method for evaluation of post-liver transplant vasculature, it has limitations in evaluating small-calibre vessels and vessels in close proximity. Contrast-enhanced ultrasound (CEUS) has been proposed to overcome these limitations by improving visualisation of post-liver transplant vasculature and reducing the number of false-positive cases, which necessitate unnecessary additional investigations such as computed tomography or angiography. Liver transplant anatomy and the wide array of post-transplant imaging findings on colour Doppler have already been well described but literature on the use of CEUS and its image interpretation remain scarce. This review aims to discuss the indications for CEUS after liver transplant, to demonstrate CEUS technique and familiarise readers with the imaging appearances of post-transplant vascular complications on CEUS.

Governance of automated image analysis and artificial intelligence analytics in healthcare.

The hype over artificial intelligence (AI) has spawned claims that clinicians (particularly radiologists) will become redundant. It is still moot as to whether AI will replace radiologists in day-to-day clinical practice, but more AI applications are expected to be incorporated into the workflows in the foreseeable future. These applications could produce significant ethical and legal issues in healthcare if they cause abrupt disruptions to its contextual integrity and relational dynamics. Sustaining trust and trustworthiness is a key goal of governance, which is necessary to promote collaboration among all stakeholders and to ensure the responsible development and implementation of AI in radiology and other areas of clinical work. In this paper, the nature of AI governance in biomedicine is discussed along with its limitations. It is argued that radiologists must assume a more active role in propelling medicine into the digital age. In this respect, professional responsibilities include inquiring into the clinical and social value of AI, alleviating deficiencies in technical knowledge in order to facilitate ethical evaluation, supporting the recognition, and removal of biases, engaging the "black box" obstacle, and brokering a new social contract on informational use and security. In essence, a much closer integration of ethics, laws, and good practices is needed to ensure that AI governance achieves its normative goals.

Impact of transient acute hypoxia on the developing mouse EEG.

Hypoxemic events are common in sick preterm and term infants and represent the most common cause of seizures in the newborn period. Neonatal seizures often lack clinical correlates and are only recognized by electroencephalogram (EEG). The mechanisms leading from a hypoxic/ischemic insult to acute seizures in neonates remain poorly understood. Further, the effects of hypoxia on EEG at various developmental stages have not been fully characterized in neonatal animals, in part due to technical challenges. We evaluated the impact of hypoxia on neonatal mouse EEG to define periods of increased susceptibility to seizures during postnatal development. Hippocampal and cortical electrodes were implanted stereotaxically in C57BL/6 mice from postnatal age 3 (P3) to P15. Following recovery, EEG recordings were obtained during baseline, acute hypoxia (4% FiO2 for 4min) and reoxygenation. In baseline recordings, maturation of EEG was characterized by the appearance of a more continuous background pattern that replaced alternating high and low amplitude activity. Clinical seizures during hypoxia were observed more frequently in younger animals (100% P3-4, 87.5% P5-6, 93% P7-8, 83% P9-10, 33% P11-12, 17% P15, r(2)=0.81) and also occurred at higher FiO2 in younger animals (11.2±1.1% P3-P6 vs. 8.9±0.8% P7-12, p<0.05). Background attenuation followed the initial hypoxemic seizure; progressive return to baseline during reoxygenation was observed in survivors. Electrographic seizures without clinical manifestations were observed during reoxygenation, again more commonly in younger animals (83% P3-4, 86% P5-6, 75% P7-8, 71% P9-10, 20% P11-12, r(2)=0.82). All P15 animals died with this duration and degree of hypoxia. Post-ictal abnormalities included burst attenuation and post-anoxic myoclonus and were more commonly seen in older animals. In summary, neonatal mice exposed to brief and severe hypoxia followed by rapid reoxygenation reliably develop seizures and the response to hypoxia varies with postnatal age and maturation.

Increased excitability and excitatory synaptic transmission during in vitro ischemia in the neonatal mouse hippocampus.

OBJECTIVE: The present study tested the hypothesis that exposure to in vitro hypoxia-ischemia alters membrane properties and excitability as well as excitatory synaptic transmission of CA1 pyramidal neurons in the neonatal mouse. METHODS: Experiments were conducted in hippocampal slices in P7-P9 C57Bl/6 mice using whole-cell patch clamp in current- and voltage-clamp mode. Passive membrane potential (Vm), input resistance (Rin) and active (action potential (AP) threshold and amplitude) membrane properties of CA1 pyramidal neurons were assessed at baseline, during 10 min in vitro ischemia (oxygen-glucose deprivation (OGD)) and during reoxygenation. Spontaneous and miniature excitatory post-synaptic currents (s and mEPSCs) were studied under similar conditions. RESULTS: OGD caused significant depolarization of CA1 pyramidal neurons as well as decrease in AP threshold and increase in AP amplitude. These changes were blocked by the application of tetrodotoxin (TTX), indicating Na(+) channels' involvement. Following 10 min of reoxygenation, significant membrane hyperpolarization was noted and it was associated with a decrease in Rin. AP threshold and amplitude returned to baseline during that stage. sEPSC and mEPSC frequency increased during both OGD and reoxygenation but their amplitude remained unchanged. Additionally, we found that OGD decreases Ih (hyperpolarization activated current) in CA1 neurons from neonatal mice and this effect persists during reoxygenation. SIGNIFICANCE: These results indicate that in vitro ischemia leads to changes in membrane excitability mediated by sodium and potassium channels. Further, it results in enhanced neurotransmitter release from presynaptic terminals. These changes are likely to represent one of the mechanisms of hypoxia/ischemia-mediated seizures in the neonatal period.

Increased neurosteroid sensitivity of hippocampal GABAA receptors during postnatal development.

To investigate developmental changes in neurosteroid modulation of GABA(A) receptors, whole-cell currents were elicited by applying GABA with allopregnanolone or pregnenolone sulfate (PS) to dentate granule cells (DGCs), acutely isolated from 7-14-day-old and adult rats. GABA evoked larger currents from dentate granule cells acutely isolated from adult rats (adult DGCs) than from neonatal DGCs, due to increased efficacy (1662+/-267 pA in adult DGCs versus 1094+/-198 pA in neonatal DGCs, P=0.004), and current density (0.072+/-0.01 pA/microm(2) in neonatal rat DGCs to 0.178+/-0.02 pA/microm(2) in adult DGCs), but unchanged potency (EC(50) was 18.5+/-2 microm in adult DGCs, and 26.6+/-7.9 microm in neonatal DGCs, P=0.21). Allopregnanolone sensitivity of GABA(A) receptor currents increased during development due to an increased potency (21.1+/-4.7 nM in adult DGCs versus 94.6+/-9 nM in neonatal DGCs, P=0.0002). The potency and efficacy of PS inhibition of GABA(A) receptor currents were remained unchanged during development (13+/-6 microm and 13.2+/-5.9 microm, P=0.71 and 85.5%+/-3.5% and 83.6%+/-0.8%, P=0.29, respectively). To investigate possible mechanism of developmental changes in GABA(A) receptor properties, in situ hybridization for alpha1, alpha4 and gamma2 subunit mRNAs was performed in dentate gyrus of the two age groups. Qualitatively, alpha1 subunit mRNA was expressed at low levels in neonatal rats while it was well expressed in adult rats. The alpha4 and gamma2 subunits were well expressed in the dentate gyrus of adult and neonatal rats. Immunohistochemical staining for alpha1 subunit in hippocampal slices from neonatal and adult rats was examined under confocal laser scanning microscope. This demonstrated that cell bodies and dendrites of granule cells are moderately positive for the alpha1 staining in adult rats but weakly so in neonatal rats. Higher-magnification images demonstrate large number of clusters of alpha1-subunit in the cell bodies of dentate granule cells of adult rat but rare clusters in granule cells of neonatal rats. Maturation of GABA(A) receptors in DGCs is characterized by increased number of GABA(A) receptors that are more sensitive to endogenous neurosteroid allopregnanolone, which might be related to increased expression of alpha1 subunit.

A gain-of-function mutation in the sodium channel gene Scn2a results in seizures and behavioral abnormalities.

The GAL879-881QQQ mutation in the cytoplasmic S4-S5 linker of domain 2 of the rat brain IIA sodium channel (Na(v)1.2) results in slowed inactivation and increased persistent current when expressed in Xenopus oocytes. The neuron-specific enolase promoter was used to direct in vivo expression of the mutated channel in transgenic mice. Three transgenic lines exhibited seizures, and line Q54 was characterized in detail. The seizures in these mice began at two months of age and were accompanied by behavioral arrest and stereotyped repetitive behaviors. Continuous electroencephalogram monitoring detected focal seizure activity in the hippocampus, which in some instances generalized to involve the cortex. Hippocampal CA1 neurons isolated from presymptomatic Q54 mice exhibited increased persistent sodium current which may underlie hyperexcitability in the hippocampus. During the progression of the disorder there was extensive cell loss and gliosis within the hippocampus in areas CA1, CA2, CA3 and the hilus. The lifespan of Q54 mice was shortened and only 25% of the mice survived beyond six months of age. Four independent transgenic lines expressing the wild-type sodium channel were examined and did not exhibit any abnormalities. The transgenic Q54 mice provide a genetic model that will be useful for testing the effect of pharmacological intervention on progression of seizures caused by sodium channel dysfunction. The human ortholog, SCN2A, is a candidate gene for seizure disorders mapped to chromosome 2q22-24.

The association of HLA and immune responses to bovine collagen implants.

The HLA type of patients with various kinds of immune reactions to bovine collagen implants were evaluated to determine a possible genetic basis for such responses. All patients suffering adverse clinical reactions to bovine collagen implants were lacking the HLA-DR4 antigen. All patients who received multiple bovine collagen injections without having adverse clinical reactions were lacking HLA-B5 and HLA-DR5 and had a significantly increased incidence of HLA-DR4. Combinations of histocompatibility antigens may influence immune response to bovine collagen implants.

NMDA receptor activation mediates the loss of GABAergic inhibition induced by recurrent seizures.

Previously we have shown that delivery of rapidly recurring hippocampal seizures (RRHS) to awake rats causes a rapid kindling and that RRHS in urethane-anesthetized rats leads to a progressive lengthening of afterdischarges and diminution of paired pulse inhibition. The present experiments examined the relationship between the changes in afterdischarge durations and inhibition. Pre-treatment before RRHS with the non-competitive NMDA receptor antagonists MK-801 and ketamine blocked afterdischarge lengthening. MK-801 also prevented RRHS-induced changes in paired pulse inhibition. For pharmacodynamic and pharmacokinetic reasons the ability of ketamine to counteract RRHS-induced changes of paired pulse inhibition was not examined. MK-801 also blocked the rightward shift of stimulus intensity vs. population spike curves which RRHS caused. We suggest that RRHS leads to an enduring diminution of GABAergic inhibition and that this accounts, at least in part, for the lengthening of afterdischarges seen with recurrent hippocampal seizures. In addition, NMDA receptor activation appears to play a role in this decrease of the potency of GABAergic inhibition. However, mechanisms which are not dependent on NMDA receptor activation also play a critical role in hippocampal epileptogenesis.

Ketamine controls prolonged status epilepticus.

New treatments are needed to control prolonged status epilepticus given the high failure rate of current therapies. In an animal model of status epilepticus based on electrical stimulation of the hippocampus, rats demonstrate at least 5 five-hours of seizure activity following stimulation. Phenobarbital (70 mg/kg) administered 15 min after stimulation effectively controlled seizures in 66% of animals (n=6). When phenobarbital (70 mg/kg) was administered 60 min after stimulation, seizures were controlled in 25% of animals (n=4). Ketamine (100 mg/kg) administered 15 min after stimulation did not control seizures in any animal (n=4). But when ketamine was administered one hour after stimulation it effectively controlled seizures in all animals (n=4). Increasing doses of ketamine were administered 60 min after stimulation to generate a dose-response curve. The ketamine dose response (fraction of seizure free rats) data were fit to a sigmoid curve to derive an ED(50) of 58 mg/kg. These findings suggest that prolonged status epilepticus becomes refractory to phenobarbital but can be effectively controlled by ketamine. For patients experiencing prolonged status epilepticus that is refractory to phenobarbital, ketamine may be an alternative to general anesthesia.

Recurrent spontaneous hippocampal seizures in the rat as a chronic sequela to limbic status epilepticus.

A period of continuous hippocampal stimulation (CHS) establishes an acute condition of self-sustaining limbic status epilepticus (SSLSE) which is followed by chronic neuropathological changes reminiscent of hippocampal sclerosis encountered in epileptic patients. In the chronic (greater than or equal to 1 month) condition following CHS-induced SSLSE, extended electrographic monitoring in the hippocampus revealed spontaneous recurrent paroxysmal discharges. All 6 animals studied had persistent interictal spiking; 3 had multiple fully developed electrographic seizures. There was a marked diminution of paired pulse inhibition, demonstrated by a protocol known to reflect the potency of inhibition mediated by GABAA receptors. Hippocampal slices from animals that had previously experienced CHS-induced SSLSE demonstrated an increased excitability relative to slices from control animals as evidenced by epileptiform bursting in increased extracellular potassium ([K+]0) and decreased extracellular calcium ([Ca2+]0). These studies establish that CHS-induced SSLSE in rats provides an experimental model with recurrent spontaneous hippocampal seizures. Based on electrophysiological data we suggest that a decrease in GABA-mediated inhibition and/or altered sensitivity to extracellular ions may play roles in the development of such seizures.

Evidence for a chronic loss of inhibition in the hippocampus after kindling: biochemical studies.

Brain tissue from kindled animals prepared 1 month after their last seizure was compared to tissue from matched surgical control animals. Quantitative film autoradiography was used to study muscimol binding in the CA1 region and 3 other brain areas (dentate gyrus, cerebral cortex, and thalamus). The Kd values so obtained were constant from region to region and comparable to those published by others. Bmax values varied; of the 4 regions studied CA1 had the lowest value of Bmax. There were no differences in either Kd or Bmax values in any region studied between kindled and surgical control rats. The release of GABA from nerve terminals was assessed with hippocampal tissue maintained in vitro and perfused with different solutions in which the concentrations of K+ and Ca2+ were varied. This allowed the examination of K+-induced depolarization release and the Ca2+ dependence of this process. K+-induced, Ca2+-dependent release of GABA from hippocampus derived from kindled animals was significantly less than that from hippocampus derived from controls. The biochemical studies reported here provide additional support for the hypothesis that there is a chronic decrease in GABA-mediated inhibition in the hippocampus associated with kindling. The data point to a dysfunction at the presynaptic level, within the GABAergic interneuron, but do not exclude changes at a level postsynaptic to the GABAergic interneuron not detected with the methods employed.

Evidence for a chronic loss of inhibition in the hippocampus after kindling: electrophysiological studies.

Rats were kindled with either of 2 protocols: (1) a rapidly recurring hippocampal seizure (RRHS) paradigm in which 10 sec stimulus trains were delivered every 5 min through hippocampal electrodes; and (2) a traditional approach in which 1 sec stimulus trains were given to the amygdala once daily. Three groups of kindled rats were prepared: (1) one of amygdala-kindled rats that had experienced 9-15 seizures; (2) one of RRHS-kindled rats that had experienced 96 seizures; and (3) one of RRHS-overkindled rats that had experienced 144-336 seizures. After a 1 month seizure-free period, the animals were anesthetized with urethane and measurements were made on the potency of paired pulse inhibition in the CA1 region of the hippocampus. All groups of kindled animals were found to have significantly less paired pulse inhibition than control rats. This decrement was confined to interpulse intervals less than or equal to 70 msec. The amount of inhibition lost correlated with the number of seizure that had occurred. The GABAergic agonist muscimol restored paired pulse inhibition in kindled animals for interpulse intervals less than or equal to 70 msec towards normal values. These results indicate that not only RRHS, but also other modes of kindling, reduced GABAergic inhibition in the CA1 region of the hippocampus and that this diminution was long-lasting, if not permanent.

Photothrombotic brain infarction results in seizure activity in aging Fischer 344 and Sprague Dawley rats.

This study was designed to determine whether photothrombotic brain infarction could result in epileptic seizures in adult animals. Male Fischer 344 (F344) rats at 2, 6, 12, 24, and 30 months of age and male Sprague Dawley (SD) rats at 2 and 6 months of age underwent photothrombotic brain infarction with the photosensitive dye rose bengal by focusing a wide (6 mm) or narrow (3 mm) diameter white light beam on the skull overlying left hemisphere anterior frontal, midfrontal, frontoparietal, or parietal areas. Animals were monitored with video and EEG recordings. Morphological analysis of infarct size was performed with a computer-assisted image analysis system. The primary finding of this study was that epileptic seizures were recorded in post-mature rats 2 months after lesioning the frontoparietal cortex with large photothrombotic infarcts that extended to the cortical-subcortical interface. These seizures were characterized behaviorally by motor arrest, appeared to originate in the periinfarct area, and could be distinguished from inherited spontaneous bilateral cortical discharges by the morphology, frequency, duration, and laterality of the ictal discharges. Small cortical lesions were ineffective in producing seizures except for one animal that demonstrated recurrent prolonged focal discharges unaccompanied by behavioral change. Stage 3 seizures were observed in a small number of mid-aged and aged animals lesioned with large infarcts in anterior frontal and frontoparietal areas. These results suggest that the technique of photothrombosis can be used to produce neocortical infarction as a means to study mechanisms of secondary epileptogenesis.

The chemistry of 9 alpha-hydroxysteroids. 1. Preparation of 9 alpha,17 beta-dihydroxy-17 alpha-ethynylandrost-4-en-3-one.

9 alpha-Hydroxyandrost-4-ene-3,17-dione 1, when allowed to react with dipotassium acetylide in tetrahydrofuran, resulted, after chromatographic separation, in 4-methyl-19-norandrosta-4,9-diene-1,17-dione 2, 4 xi-methyl-19-norandrosta-5(10),9(11)-diene-1,17-dione 3, 4-methyl-17 alpha-ethynyl-17 beta-hydroxy-19-norandrosta-4,9-dien-1-one 4, 4 xi-methyl-17 alpha-ethynyl-17 beta-hydroxy-19-norandrosta-5(10),9(11)-dien- 1-one 5, and 17 alpha-ethynyl-17 beta-hydroxy-9,10-secoandrost-4-ene-3,9-dione 6. Selective protection of delta 4-3-ketone of 9 alpha-hydroxyandrost-4-ene-3,17-dione 1 as its dienol methyl ether 7, and subsequent reaction with lithium acetylide-ethylenediamine followed by acidic hydrolysis, afforded 9 alpha,17 beta-dihydroxy-17 alpha- ethynylandrost-4-en-3-one 8.

Changes in lipids and free fatty acid fractions in adult Haemonchus contortus during incubation in vitro.

Adult Haemonchus contortus (Rud., 1803) has been investigated for its ability to utilize lipids with regard to the total lipids, sterols, free fatty acids, acylglycerols and phospholipids produced during incubation in vitro. All these components exhibit extensive fluctuations, decreasing at some times and increasing at others, thus indicating both biosynthesis and utilization. Also, changes in fatty acid components of total lipids have been analyzed by gas liquid chromatography. These observations indicate that H. contortus is capable of utilizing 16:0, 18:0, 18:1, 18:2, 20:0, 20:un1, 20:un2, 21:0, 21:un1, 21:1, 21:un2, 22:un1, 22:3, 22:?, 22:un2 and 24:3 fatty acids. At the same time, because of large increases in 15:0, 16:0, 18:0, 18:?, 18:1, 18:2, 20:0, 20:un1, 20:un2, 21:0, 21:un1, 21:1 and 21:un2 acids, it is postulated that these acids are synthesized by adult H. contortus.

Amino acid biosynthesis in Haemonchus contortus from C14-labelled precursors, in vitro.

Adult Haemonchus contortus (Nematoda: Trichostrongylidae) was investigated for its ability to utilize various C14-labelled precursors, i.e., glucose, acetate, CO2 and palmitic acid, for amino acid biosynthesis. H. contortus has been demonstrated to be capable of synthesizing essential as well as non-essential amino acids. Label from all the precursors was detected in aspartic acid, lysine, histidine, cystine, cysteine, glutamic acid, proline, arginine, tyrosine, alanine, glycine, serine, valine, methionine, leucine and isoleucine. Glutamic acid, aspartic acid, alanine, glycine and serine were synthesized to a greater extent relative to the other amino acids, regardless of the precursor employed. However, in case of glucose, there was comparatively less incorporation into glycine and serine. Incorporation of C14 into various amino acids is evidence for the operation of tricarboxylic acid cycle. Further, the fact that carbon from palmitic acid appears in amino acids, indicates that adult H. contortus is capable of catabolizing long-chain fatty acids. Possible mechanisms for the involvement of various precursors in amino acids biosynthesis are examined here.

Evaluation of enzyme immunoassays using purified protein derivative (PPD) and its pooled fractions 3 and 4 for diagnosis of pulmonary tuberculosis.

A critical evaluation of two enzyme immunoassays (EIAs) for diagnosis of pulmonary tuberculosis is reported. Purified protein derivative (PPD) or its pooled fractions 3 and 4 were used as antigens for detection of antibodies in sera from 53 patients with active pulmonary tuberculosis and 10 normal healthy individuals. The cut-off point for each EIA was based on the absorbance (mean + 3 SD) of normal sera with the respective antigens. All the normal sera were negative in both the assays. The positivity of tuberculosis patients in either assay was 86.8 per cent. Thus, for serodiagnosis of tuberculosis fractions 3 and 4 of PPD could serve as a good substitute for whole PPD. Sera from 45 leprosy patients were also analysed to assess the specificity of the EIAs. The mean reactivity of tuberculoid leprosy sera was comparable to that of normal sera. The ratio of the mean absorbance of lepromatous leprosy (LL) sera and normal sera was 16.73 with PPD, in comparison to 21.95 for pooled fractions 3 and 4. Out of 10 LL patients 9 (90%) were positive with fractions 3 and 4, in comparison to 10 (100%) with PPD. 71.1 per cent of leprosy patients belonging to different categories were positive in assay based on PPD in comparison to 64.4% in EIA using fractions 3 and 4. The high false positivity of leprosy sera in an assay designed for detection of pulmonary tuberculosis has immense implications in interpretation of results of the assay for diagnostic and epidemiological purposes.

Kinetics of antibody response by Dot-ELISA in rabbits immunized with adult Haemonchus contortus antigen.

Whole adult soluble extract of Haemonchus contortus as an antigen along with Freund's complete adjuvant, was used to immunize rabbits. Antisera from immunized rabbits were collected at intervals of 30, 60, 90, 120, 150 and 180 days. For the detection and titration of anti-H. contortus antibodies in these sera, Dot-ELISA was developed. Sera collected 30 days post-immunization exhibited a titre of 1:5,000 in all the rabbits except one, where a titre of 20,000 was recorded. Later, all the rabbits attained the highest titre of 40,000 at different periods of post-immunizations, which were maintained 150-180 days. These high titre sera can be of immense use in the identification and characterization of immunodominant antigens of adult H. contortus.