Standardized quick en bloc technique for procurement of cadaveric liver grafts for pediatric liver transplantation

This paper describes a quick procedure for cadaveric liver graft retrieval during multiple organ harvesting. The technique is based on minimal preliminary dissection, absence of in situ direct portal perfusion, and en bloc removal of the liver and pancreas, with an aortic patch encompassing the coeliac trunk and superior mesenteric artery. The results of 110 pediatric liver transplantations with 109 organs harvested using this technique are reported. There were no graft harvesting injuries. The liver graft primary nonfunction rate was 4.5% (5/110). The 3-month retransplantation rate was 10%. The actual patient survival rates were 93% at 3 months and 90% at 1 year; actual graft survival rates were 85.5% and 78%, respectively. The technique described was at least as safe as conventional procedures. A major advantage of the procedure is its flexibility, which allows for the easily combined procurement of other organs (whole pancreas and intestine).     

Neurotransmitter release evoked by α-latrotoxin in the smooth muscle of the female pig urethra

Neuronal regulation of smooth muscle tone in the female pig urethra has mainly been studied in vitro using electrical field stimulation (EFS) of nerves. Excitatory control is considered to be exerted by released noradrenaline, whereas inhibitory control is non-adrenergic non-cholinergic (NANC), and mediated by nitric oxide (NO), and an as yet unidentified agent. We investigated the functional and morphological effects of α-latrotoxin (αLTX), a spider neurotoxin believed to cause massive release of vesicle-stored neurotransmitters, on spontaneously developed urethral smooth muscle tone. The effects were compared to those of EFS and high potassium. In the presence of the NO-synthesis inhibitor N^ω-nitro-L-arginine (L-NOARG: 0.3 mM) both αLTX and EFS evoked contractions. After treatment with scopolamine and phentolamine, no contraction was observed, and under these conditions αLTX and EFS induced relaxation. At low frequencies (<12 Hz), the EFS-induced relaxations were rapid, whereas at higher frequencies (>12 Hz), they were biphasic, consisting of a rapid first phase followed by a more long-lasting second phase. L-NOARG abolished the relaxations at low frequencies, as well as the first phase of the biphasic relaxation. The second phase was not affected by treatment with L-NOARG, but 0.1 μM ω-conotoxin GVIA, blocker of N-type voltage-operated calcium- channels (VOCCs), markedly reduced or abolished the response. In the presence of L-NOARG or ω-conotoxin GVIA, the αLTX-induced relaxation was significantly decreased, and the combination of L-NOARG and ω-conotoxin GVIA further reduced or abolished the relaxation. In preparationstreated with tetrodotoxin or scorpion venom, believed to inactivate nerves by acting on sodium channels, αLTX and EFS had no effects. αLTX-induced relaxation was not associated with changes in cyclic GMP or cyclic AMP content. High (80 mM) potassium solution induced a triphasic response of the preparation. A transient relaxation was followed by a restoration of tone, and then by a persistent relaxation. The persistent relaxation was slightly reduced by scorpion venom or L-NOARG, but reduced by 50% by a combination of L-NOARG and ω-conotoxin GVIA. Ultrastructural analysis of the urethral circular smooth muscle layer revealed a moderate amount of nerve profiles supplying the smooth muscle. In control preparations, the nerve profiles contained both small synaptic vesicles and large dense core vesicles. αLTX caused a major loss of both types of vesicle. The present data suggest that αLTX has the ability to release not only adrenergic and cholinergic transmitters, but also NANC mediators of relaxation, including NO, from nerve terminals in the urethra. Received: 13 January 1997 / Accepted: 17 April 1997     

Langendorff and ischemia in immature and neonatal myocardia. Two essential key-words in Today's cardiothoracic research

Background

Oscar Langendorff first published a method of nutrition of the isolated, mammal heart in 1895. Numerous studies have been performed on adult animals since then. With the growing incidence of pediatric heart surgery, however, studies were also done on immature, and neonatal myocardia. This article reviews the experimental efforts undertaken to protect immature and neonatal myocardia during cardiac surgery in comparison to the adult one.

Methods

The Langendorff preparation, used in most of the experiments mentioned above, is based on a retrograde aortic perfusion with physiological buffer. Due to the closure of the aortic valves, the fluid flows through the coronaries. Flow, pressure, rhythm, and biochemical analyses can be evaluated.

Results

Immature, or neonatal hearts have a higher tolerance of ischemia than adult ones. Possibilities of how to protect the myocardium will be briefly reviewed.

Conclusion:

The different energetic metabolism and the age dependent changes in the ultrastructures of immature and neonatal hearts call for a different kind of protection. Again, the Langendorff's method seems to be a successful experimental approach as it allows to study effects independently of the organism, and presents an experimental set-up with minor ethical concerns.     

NANC transmitters in the female pig urethra–localization and modulation of release via α2-adrenoceptors and potassium channels

1. To investigate further the release, localization and identity of a non-nitrergic mediator of smooth muscle relaxation in the female pig urethra, we studied the effects of drugs acting at α₂-adrenoceptors or K⁺ channels, the effects of capsaicin and chemical sympathectomy, and the actions of several transmitter candidates.
2. Electrical field stimulation (EFS; frequencies above 12 Hz) of spontaneously contracted smooth muscle strips from the female pig urethra evoked long-lasting, frequency-dependent relaxations in the presence of prazosin, scopolamine, and N^G-nitro-L-arginine. Treatment with the selective α₂-adrenoceptor agonist UK-14 304 markedly reduced the relaxations evoked by EFS at all frequencies tested (16–30 Hz). The inhibitory effect of UK-14 304 was completely antagonized by the α₂-adrenoceptor antagonist rauwolscine. The muscarinic M₁ receptor antagonist, pirenzepine, or exogenously administered carbachol, did not have any effects on the electrically evoked relaxations.
3. Inhibition of high conductance Ca²⁺ activated K⁺ channels by iberiotoxin or charybdotoxin significantly enhanced the relaxations evoked by EFS at all frequencies. However, inhibition of voltage-sensitive K⁺ channels with 4-aminopyridine or dendrotoxin-1, treatment with the ATP-sensitive K⁺ channel blocker, glibenclamide, or treatment with the high and low conductance Ca²⁺ activated K⁺ channel blockers, tetraethylammonium chloride and apamin, had no effect on the relaxations evoked by EFS.
4. Electrically evoked relaxations were not affected by adrenergic denervation with 6-hydroxydopamine (6-OHDA) at any frequency. However, treatment with 6-OHDA abolished prazosin-sensitive electrically induced contractions, and a long-lasting relaxation was revealed. Treatment with capsaicin, believed to damage selectively a subpopulation of primary afferent fibres, did not affect basal tone or relaxations evoked by EFS.
5. Exogenously applied vasoactive intestinal polypeptide (VIP), pituitary adenylate cyclase-activating peptide (PACAP)-27, PACAP-38, adenosine, ATP and 5-hydroxy-tryptamine caused relaxations of the urethral preparations, whereas prostaglandin E₂ and calcitonin gene-related peptide had no effects. VIP 10-28, α, β-methylene-ATP, reactive blue-2, suramin or indomethacin did not reduce the electrically-evoked relaxations at any frequency. However, the relaxations were slightly reduced by trypsin or α-chymotrypsin.
6. The present results suggest that the release of the unknown mediator in the female pig urethra can be modulated via α₂-adrenoceptors and high conductance Ca²⁺ activated K⁺ channels. Furthermore, the mediator does not appear to be localized to or released from adrenergic or capsaicin-sensitive sensory nerve-endings. The identity of the transmitter remains to be established.

     

Optimization and Application of a Biotinylation Method for Quantification of Plasma Membrane Expression of Transporters in Cells

Quantitative proteomics, using LC-MS/MS, is increasingly used to quantify drug transporters present in tissues and cells. Most of these investigations quantify total transporter expression in the cells by utilizing a total membrane fraction, not only the plasma membrane. Here, we report development and optimization of a biotinylation method to quantify protein expression of transporters in the plasma membrane of cells. The Pierce cell surface isolation protocol was optimized for plasma membrane isolation. Incubation of OATP1B1-expressing CHO cells with 0.78 mg/mL of membrane impermeable biotinylation reagent (sulfo-NHS-SS-biotin) at 37°C for 1 h resulted in optimum isolation of the plasma membrane. Subsequently, the expression of transporters in the plasma membrane as a percent of the total was determined by quantitative proteomics using LC-MS/MS. Mean (±SD) plasma membrane expression of OATP1B1 in plated OATP1B1-expressing CHO, MDCKII, and HEK293 cells was found to be 79.7% (±4.7%), 67.7% (±12.2%), and 65.3% (±6.8%) of total cell OATP1B1 expression. Mean (±SD) plasma membrane expression of OATP1B3 in plated OATP1B3-expressing HEK293 cells, OATP2B1 in plated OATP2B1-expressing MDCKII cells, and sodium/taurocholate co-transporting polypeptide (NTCP) in plated NTCP-expressing CHO cells was 63.2% (±1.6%), 37.1% (±15.7%), and 71.7% (±1.2%), respectively. This method of quantifying transporter protein expression in the plasma membrane will be useful in the future to predict transporter-mediated drug disposition.     

Extended stereotactic brain biopsy in suspected primary central nervous system angiitis: good diagnostic accuracy and high safety

Journal of Neurology - To evaluate the diagnostic accuracy and safety of extended stereotactic brain biopsy (ESBB) in a single center cohort with suspected primary angiitis of the central nervous...     

Severe Preeclampsia and Maternal Self‐Report of Oral Health,Hygiene, and Dental Care

Background: Maternal periodontal disease diagnosed by a detailed oral health examination is associated with preeclampsia. Our objective was to measure the association between maternal self‐report of oral symptoms/problems, oral hygiene practices, and/or dental service use before or during pregnancy and severe preeclampsia. Methods: A written questionnaire was administered to pregnant females at the time of prenatal ultrasound and outcomes were ascertained by chart abstraction. The χ² test compared maternal oral symptoms/problems, hygiene practices, and dental service use between females with severe preeclampsia versus normotensive females. Multivariable logistic regression was used to calculate adjusted odds ratios (aORs) and 95% confidence intervals (CIs) for severe preeclampsia. Results: A total of 48 (10%) of 470 females reported ≥2 oral symptoms/problems in the 6 months before pregnancy and 77 (16%) since pregnancy. Fifty‐one (11%) reported previous periodontal treatment. Twenty‐eight (6%) of 470 developed severe preeclampsia. Females with a history of periodontal treatment were more likely to develop severe preeclampsia (aOR = 3.71; 95% CI = 1.40 to 9.83) than females without a history of periodontal treatment. Self‐reported oral health symptoms/problems, oral hygiene practices, or dental service use before or during pregnancy were not associated with severe preeclampsia when considered in the context of other maternal risk factors. Conclusion: Maternal self‐report of previous periodontal treatment before pregnancy is associated with severe preeclampsia.     

Repair of aortic root in patients with aneurysm or dissection: comparing the outcomes of valve-sparing root replacement with those from the Bentall procedure

Introduction

Management of aortic root aneurysm or dissection has been the subject of much discussion that has led to some modifications. The current trend is a valve-sparing root replacement. We compared the outcome following valve sparing root repair with Bentall procedure.

Methods

We retrospectively evaluated 70 patients who underwent root replacement for aneurysm or dissection and compared the outcomes of valve-sparing root replacement with those of the Bentall procedure from January 2007 to December 2011 at our institution.

Results

Twenty-five patients had valve-sparing aortic root replacement (VSR, including reimplantation or remodeling) (23 males and 2 females), and 45 patients had the Bentall procedure (34 males and 11 females). Patients who underwent a VSR were younger with a mean age of 55.4 ± 14.8 years compared to those who underwent the Bentall procedure with a mean age of 60.6 ± 12.7 (P=ns). The preoperative aortic insufficiency (AI) in the VSR group was moderate in 8 (32%) patients, and severe in 6 (24%). Preoperative creatinine was 1 ± 0.35 mg/dl in the VSR group and 1.1 ± 0.87 mg/dl in the Bentall group. In the VSR group, 3 (12%) patients had emergency surgery; by contrast, in the Bentall group, 8 (17%) patients had emergent surgery. Concomitant coronary artery bypass grafting (excluding coronary reimplantation) was performed in 8 (32%) patients in the VSR group and in 12 (26.6%) patients in the Bentall group (P=0.78); additional valve procedures were performed in 2 (8%) patients in the VSR group and in 11 (24.4%) patients in the Bentall group. The perioperative mortality was 8% (n=2) and 13.3% (n=6), for the VSR and Bentall procedures, respectively (P=0.7, ns). The total duration of intensive care unit stay was 116.6 ± 106 hours for VSR patients and 152.5 ± 218.2 hours for Bentall patients (P=0.5). The overall length of stay in the hospital was 10 ± 8.1 days for VSR and 11 ± 9.52 days for Bentall (P=0.89). The one-year survival was 92% for the VSR group and 79.0% for the Bentall group. The seven-year survival for the VSR group was 92% and 79% for the Bentall group (95% CI [1.215 to 0.1275], P=0.1).

Conclusion

Aortic valve-sparing root replacement can be performed with acceptable morbidity and mortality with a comparable long-term survival to the Bentall procedure.     

Biological Trojan horse: Antigen 43 provides specific bacterial uptake and survival in human neutrophils

Escherichia coli is a versatile pathogen causing millions of infections in humans every year. This bacterium can form multicellular aggregates when it expresses a self-associating protein, antigen 43 (Ag43), on its surface. We have discovered that Ag43-expressing E. coli cells are efficiently taken up by human defense cells, polymorphonuclear neutrophils (PMNs), in an opsonin-independent manner. Surprisingly, the phagocytosed bacteria were not immediately killed but resided as tight aggregates within the PMNs. Our observations indicate that Ag43-mediated uptake and survival in PMNs constitute a mechanism to subvert one of the primary defense mechanisms of the human body.