Development of an ultrasound-guided technique for pudendal nerve block in cat cadavers.

The objective of this prospective experimental cadaveric study was to develop an ultrasound-guided technique to perform an anaesthetic pudendal nerve block in male cats. Fifteen fresh cadavers were used for this trial. A detailed anatomical dissection was performed on one cat in order to scrutinise the pudendal nerve and its ramifications. In a second step, the cadavers of six cats were used to test three different ultrasonographic approaches to the pudendal nerve: the deep dorso-lateral, the superficial dorso-lateral and the median transperineal. Although none of the approaches allowed direct ultrasonographical identification of the pudendal nerve branches, the deep dorso-lateral was found to be the most advantageous one in terms of practicability and ability to identify useful and reliable landmarks. Based on these findings, the deep dorso-lateral approach was selected as technique of choice for tracer injections (0.1 ml 1% methylene blue injected bilaterally) in six cat cadavers distinct from those used for the ultrasonographical study. Anatomical dissection revealed a homogeneous spread of the tracer around the pudendal nerve sensory branches in all six cadavers. Finally, computed tomography was performed in two additional cadavers after injection of 0.3 ml/kg (0.15 ml/kg per each injection sites, left and right) contrast medium through the deep dorso-lateral approach in order to obtain a model of volume distribution applicable to local anaesthetics. Our findings in cat cadavers indicate that ultrasound-guided pudendal nerve block is feasible and could be proposed to provide peri-operative analgesia in clinical patients undergoing perineal urethrostomy.

Ultrasound-guided block of sciatic and femoral nerves: an anatomical study

The sheep is a popular animal model for human biomechanical research involving invasive surgery on the hind limb. These painful procedures can only be ethically justified with the application of adequate analgesia protocols. Regional anaesthesia as an adjunct to general anaesthesia may markedly improve well-being of these experimental animals during the postoperative period due to a higher analgesic efficacy when compared with systemic drugs, and may therefore reduce stress and consequently the severity of such studies. As a first step 14 sheep cadavers were used to establish a new technique for the peripheral blockade of the sciatic and the femoral nerves under sonographic guidance and to evaluate the success rate by determination of the colorization of both nerves after an injection of 0.5 mL of a 0.1% methylene blue solution. First, both nerves were visualized sonographically. Then, methylene blue solution was injected and subsequently the length of colorization was measured by gross anatomical dissection of the target nerves. Twenty-four sciatic nerves were identified sonographically in 12 out of 13 cadavers. In one animal, the nerve could not be ascertained unequivocally and, consequently, nerve colorization failed. Twenty femoral nerves were located by ultrasound in 10 out of 13 cadavers. In three cadavers, signs of autolysis impeded the scan. This study provides a detailed anatomical description of the localization of the sciatic and the femoral nerves and presents an effective and safe yet simple and rapid technique for performing peripheral nerve blocks with a high success rate.

Identification of erosional mechanisms during past glaciations based on a bedrock surface model of the central European Alps

The bedrock topography beneath the Quaternary cover provides an important archive for the identification of erosional processes during past glaciations. Here, we combined stratigraphic investigations of more than 40,000 boreholes with published data to generate a bedrock topography model for the entire plateau north of the Swiss Alps including the valleys within the mountain belt. We compared the bedrock map with data about the pattern of the erosional resistance of Alpine rocks to identify the controls of the lithologic architecture on the location of overdeepenings. We additionally used the bedrock topography map as a basis to calculate the erosional potential of the Alpine glaciers, which was related to the thickness of the LGM ice. We used these calculations to interpret how glaciers, with support by subglacial meltwater under pressure, might have shaped the bedrock topography of the Alps. We found that the erosional resistance of the bedrock lithology mainly explains where overdeepenings in the Alpine valleys and the plateau occur. In particular, in the Alpine valleys, the locations of overdeepenings largely overlap with areas where the underlying bedrock has a low erosional resistance, or where it was shattered by faults. We also found that the assignment of two end-member scenarios of erosion, related to glacial abrasion/plucking in the Alpine valleys, and dissection by subglacial meltwater in the plateau, may be adequate to explain the pattern of overdeepenings in the Alpine realm. This most likely points to the topographic controls on glacial scouring. In the Alps, the flow of LGM and previous glaciers were constrained by valley flanks, while ice flow was mostly divergent on the plateau where valley borders are absent. We suggest that these differences in landscape conditioning might have contributed to the contrasts in the formation of overdeepenings in the Alpine valleys and the plateau.

Non-acute myocardial infarction-related causes of elevated high-sensitive troponin T in the emergency room: a cross-sectional analysis

To systematically investigate putative causes of non-coronary high-sensitive troponin elevations in patients presenting to a tertiary care emergency department. In this cross-sectional analysis, patients who received serial measurements of high-sensitive troponin T between 1 August 2010 and 31 October 2012 at the Department of Emergency Medicine were included. The following putative causes were considered to be associated with non-acute coronary syndrome-related increases in high-sensitive troponin T: acute pulmonary embolism, renal insufficiency, aortic dissection, heart failure, peri-/myocarditis, strenuous exercise, rhabdomyolysis, cardiotoxic chemotherapy, high-frequency ablation therapy, defibrillator shocks, cardiac infiltrative disorders (e.g., amyloidosis), chest trauma, sepsis, shock, exacerbation of chronic obstructive pulmonary disease, and diabetic ketoacidosis. During the study period a total of 1,573 patients received serial measurements of high-sensitive troponin T. Of these, 175 patients were found to have acute coronary syndrome leaving 1,398 patients for inclusion in the study. In 222 (30 %) of patients, no putative cause described in the literature could be attributed to the elevation in high-sensitive troponin T observed. The most commonly encountered mechanism underlying the troponin T elevation was renal insufficiency that was present in 286 patients (57 %), followed by cerebral ischemia in 95 patients (19 %), trauma in 75 patients (15 %) and heart failure in 41 patients (8 %). Non-acute coronary syndrome-associated elevation of high-sensitive troponin T levels is commonly observed in the emergency department. Renal insufficiency and acute cerebral events are the most common conditions associated with high-sensitive troponin T elevation.

Role of NonO-histone interaction in TNFalpha-suppressed prolyl-4-hydroxylase alpha1.

Inflammation is a key process in cardiovascular diseases. The extracellular matrix (ECM) of the vasculature is a major target of inflammatory cytokines, and TNFalpha regulates ECM metabolism by affecting collagen production. In this study, we have examined the pathways mediating TNFalpha-induced suppression of prolyl-4 hydroxylase alpha1 (P4Halpha1), the rate-limiting isoform of P4H responsible for procollagen hydroxylation, maturation, and organization. Using human aortic smooth muscle cells, we found that TNFalpha activated the MKK4-JNK1 pathway, which induced histone (H) 4 lysine 12 acetylation within the TNFalpha response element in the P4Halpha1 promoter. The acetylated-H4 then recruited a transcription factor, NonO, which, in turn, recruited HDACs and induced H3 lysine 9 deacetylation, thereby inhibiting transcription of the P4Halpha1 promoter. Furthermore, we found that TNFalpha oxidized DJ-1, which may be essential for the NonO-P4Halpha1 interaction because treatment with gene specific siRNA to knockout DJ-1 eliminated the TNFalpha-induced NonO-P4Halpha1 interaction and its suppression. Our findings may be relevant to aortic aneurysm and dissection and the stability of the fibrous cap of atherosclerotic plaque in which collagen metabolism is important in arterial remodeling. Defining this cytokine-mediated regulatory pathway may provide novel molecular targets for therapeutic intervention in preventing plaque rupture and acute coronary occlusion.

Mutations in myosin light chain kinase cause familial aortic dissections.

Mutations in smooth muscle cell (SMC)-specific isoforms of α-actin and β-myosin heavy chain, two major components of the SMC contractile unit, cause familial thoracic aortic aneurysms leading to acute aortic dissections (FTAAD). To investigate whether mutations in the kinase that controls SMC contractile function (myosin light chain kinase [MYLK]) cause FTAAD, we sequenced MYLK by using DNA from 193 affected probands from unrelated FTAAD families. One nonsense and four missense variants were identified in MYLK and were not present in matched controls. Two variants, p.R1480X (c.4438C>T) and p.S1759P (c.5275T>C), segregated with aortic dissections in two families with a maximum LOD score of 2.1, providing evidence of linkage of these rare variants to the disease (p = 0.0009). Both families demonstrated a similar phenotype characterized by presentation with an acute aortic dissection with little to no enlargement of the aorta. The p.R1480X mutation leads to a truncated protein lacking the kinase and calmodulin binding domains, and p.S1759P alters amino acids in the α-helix of the calmodulin binding sequence, which disrupts kinase binding to calmodulin and reduces kinase activity in vitro. Furthermore, mice with SMC-specific knockdown of Mylk demonstrate altered gene expression and pathology consistent with medial degeneration of the aorta. Thus, genetic and functional studies support the conclusion that heterozygous loss-of-function mutations in MYLK are associated with aortic dissections.

Severe aortic and arterial aneurysms associated with a TGFBR2 mutation.

BACKGROUND: A 24-year-old man presented with previously diagnosed Marfan's syndrome. Since the age of 9 years, he had undergone eight cardiovascular procedures to treat rapidly progressive aneurysms, dissection and tortuous vascular disease involving the aortic root and arch, the thoracoabdominal aorta, and brachiocephalic, vertebral, internal thoracic and superior mesenteric arteries. Throughout this extensive series of cardiovascular surgical repairs, he recovered without stroke, paraplegia or renal impairment. INVESTIGATIONS: CT scans, arteriogram, genetic mutation screening of transforming growth factor beta receptors 1 and 2. DIAGNOSIS: Diffuse and rapidly progressing vascular disease in a patient who met the diagnostic criteria for Marfan's syndrome, but was later rediagnosed with Loeys-Dietz syndrome. Genetic testing also revealed a de novo mutation in transforming growth factor beta receptor 2. MANAGEMENT: Regular cardiovascular surveillance for aneurysms and dissections, and aggressive surgical treatment of vascular disease.

Genes to blood pressure: The effects of variation in genes of the renin-angiotensin system on the hormonal and renal control of blood pressure

Objective. Essential hypertension affects 25% of the US adult population and is a leading contributor to morbidity and mortality. Because BP is a multifactorial phenotype that resists simple genetic analysis, intermediate phenotypes within the complex network of BP regulatory systems may be more accessible to genetic dissection. The Renin-Angiotensin System (RAS) is known to influence intermediate and long-term blood pressure regulation through alterations in vascular tone and renal sodium and fluid resorption. This dissertation examines associations between renin (REN), angiotensinogen (AGT), angiotensin-converting enzyme (ACE) and angiotensin II type 1 receptor (AT1) gene variation and interindividual differences in plasma hormone levels, renal hemodynamics, and BP homeostasis.^ Methods. A total of 150 unrelated men and 150 unrelated women, between 20.0 and 49.9 years of age and free of acute or chronic illness except for a history of hypertension (11 men and 7 women, all off medications), were studied after one week on a controlled sodium diet. RAS plasma hormone levels, renal hemodynamics and BP were determined prior to and during angiotensin II (Ang II) infusion. Individuals were genotyped by PCR for a variable number tandem repeat (VNTR) polymorphism in REN, and for the following restriction fragment length polymorphisms (RFLP): AGT M235T, ACE I/D, and AT1 A1166C. Associations between clinical measurements and allelic variation were examined using multiple linear regression statistical models.^ Results. Women homozygous for the AT1 1166C allele demonstrated higher intracellular levels of sodium (p = 0.044). Men homozygous for the AGT T235 allele demonstrated a blunted decrement in renal plasma flow in response to Ang II infusion (p = 0.0002). There were no significant associations between RAS gene variation and interindividual variation in RAS plasma hormone levels or BP.^ Conclusions. Rather than identifying new BP controlling genes or alleles, the study paradigm employed in this thesis (i.e., measured genes, controlled environments and interventions) may provide mechanistic insight into how candidate genes affect BP homeostasis. ^

Transcriptional regulation of the human {\it PAX6\/} gene

PAX6, a member of the paired-type homeobox gene family, is expressed in a partially and temporally restricted pattern in the developing central nervous system, and its mutation is responsible for human aniridia (AN) and mouse small eye (Sey). The objective of this study was to characterize the PAX6 gene regulation at the transcriptional level, and thereby gain a better understanding of the molecular basis of the dynamic expression pattern and the diversified function of the human PAX6 gene.^ Initially, we examined the transcriptional regulation of the PAX6 gene by transient transfection assays and identified multiple cis-regulatory elements that function differently in different cell lines. The transcriptional initiation site was identified by RNase protection and primer extension assays. Examination of the genomic DNA sequence indicated that the PAX6 promoter has a TATA like-box (ATATTTT) at $-$26 bp, and two CCAAT-boxes are located at positions $-$70 and $-$100 bp. A 38 bp ply (CA) sequence was located 992 bp upstream from the initiation site. Transient transfection assays in glioblastoma cells and leukemia cells indicate that a 92 bp region was required for basal level PAX6 promoter activity. Gel retardation assays showed that this 92 bp sequence can form four DNA-protein complexes which can be specifically competed by a 31-mer oligonucleotide containing a PAX6 TATA-like sequence or an adenovirus TATA box. The activation of the promoter is positively correlated with the expression of PAX6 transcripts in cells tested.^ Based on the results obtained from the in vitro transfection assays, we did further dissection assay and functional analysis in both cell-culture and transgenic mice. We found that a 5 kb upstream promoter sequence is required for the tissue specific expression in the forebrain region which is consistent with that of the endogenous PAX6 gene. A 267 bp cell-type specific repressor located within the 5 kb fragment was identified and shown to direct forebrain specific expression. The cell-type specific repressor element has been narrowed to a 30 bp region which contains a consensus E-box by in vitro transfection assays. The third regulatory element identified was contained in a 162 bp sequence (+167 to +328) which functions as a midbrain repressor, and it appeared to be required for establishing the normal expression pattern of the PAX6 gene. Finally, a highly conserved 216 bp sequence identified in intron 4 exhibited as a spinal cord specific enhancer. And this 216 bp cis-regulatory element can be used as a marker to trace the differentiation and migration of progenitor cells in the developing spinal cord. These studies show that the concerted action of multiple cis-acting regulatory elements located upstream and downstream of the transcription initiation site determines the tissue specific expression of PAX6 gene. ^

Standardized definitions and clinical endpoints in trials investigating endovascular repair of aortic dissections.

OBJECTIVES Endovascular therapy is a rapidly expanding option for the treatment of patients with aortic dissection (AD) and various studies have been published. These trials, however, are often difficult to interpret and compare because they do not utilize uniform clinical endpoint definitions. METHODS The DEFINE Group is a collaborative effort of an ad hoc multidisciplinary team from various specialties involved in AD therapy in Europe and the United States. DEFINE's goal was to arrive at a broad based consensus for baseline and endpoint definitions in trials for endovascular therapy of various vascular pathologies. In this project, which started in December 2006, the individual team members reviewed the existing pertinent literature. Following this, a series of telephone conferences and face-to-face meetings were held to agree upon definitions. Input was also obtained from regulatory (United States Food and Drug Administration) and industry (device manufacturers with an interest in peripheral endovascular revascularization) stakeholders, respectively. RESULTS These efforts resulted in the present document containing proposed baseline and endpoint definitions for clinical and morphological outcomes. Although the consensus has inevitably included certain arbitrary consensus choices and compromises, adherence to these proposed standard definitions would provide consistency across future trials, thereby facilitating evaluation of clinical effectiveness and safety of various endovascular revascularization techniques. CONCLUSIONS This current document is based on a broad based consensus involving relevant stakeholders from the medical community, industry and regulatory bodies. It is proposed that the consensus document may have value for study design of future clinical trials in endovascular AD therapy as well as for regulatory purposes.

Comparison of stab wound probing versus radiological stab wound channel depiction with contrast medium

BACKGROUND Instillation of contrast medium into stab wounds has shown promising results regarding visibility and assessment of general stab direction with computed tomography. However, the accuracy of this method--and, incidentally also probing of stab wounds--has to our knowledge not previously been examined. Also the effect of bluntness of different stabbing objects on the examination of stab wounds was not considered before this study. METHODS Using a pocket-knife, a steak-knife, and a Phillips screwdriver, nine stab wounds each were inflicted to three pork haunches. The depths of the stab wounds were determined by probing and multislice computed tomography (MSCT) after instillation of a contrast medium (CM) and then compared to those observed by dissection, our internal "gold standard". RESULTS In stab wounds inflicted by knives, MSCT-CM and probing provided results which differed by roughly 10-11% from the dissection results. In screwdriver stabs MSCT-CM showed a deviation of almost 30%, probing over 33%. DISCUSSION MSCT-CM is a possible alternative to layer-by-layer dissection in autopsy cases of knife stab wounds. Probing, although obsolete in post-mortem examinations, is sufficiently accurate in determining the length of a stab wound of a living person. In cases of stab wounds with blunt objects such as screwdrivers, neither MSCT-CM nor probing proved to be sufficiently accurate. CONCLUSION MSCT-CM is a possible alternative to layer-by-layer dissection in autopsy cases of knife stab wounds. Probing, although obsolete in post-mortem examinations, is sufficiently accurate in determining the length of a stab wound of a living person. In cases of stab wounds with blunt objects such as screwdrivers, neither MSCT-CM nor probing proved to be sufficiently accurate.

Pitfalls in post-mortem CT-angiography--intravascular contrast induces post-mortem pericardial effusion

We present a case where multi-phase post-mortem computed tomography angiography (PMCTA) induced a hemorrhagic pericardial effusion during the venous phase of angiography. Post-mortem non-contrast CT (PMCT) suggested the presence of a ruptured aortic dissection. This diagnosis was confirmed by PMCTA after pressure controlled arterial injection of contrast. During the second phase of multi-phase PMCTA the presence of contrast leakage from the inferior cava vein into the pericardial sac was noted. Autopsy confirmed the post-mortem nature of this vascular tear. This case teaches us an important lesson: it underlines the necessity to critically analyze PMCT and PMCTA images in order to distinguish between artifacts, true pathologies and iatrogenic findings. In cases with ambiguous findings such as the case reported here, correlation of imaging findings with autopsy is elementary.

To see or not to see – Ambiguous findings on post-mortem cross-sectional imaging in a case of ruptured abdominal aortic aneurysm

We present a case of a ruptured abdominal aortic aneurysm (AAA) with ambiguous accessory findings on post-mortem computed-tomography (PMCT), post-mortem magnetic resonance (PMMR) imaging, and PMCT-angiography (PMCTA) suggestive of thoracic aortic dissection. The diagnosis of ruptured AAA was confirmed by autopsy; however, there was no aortic dissection. The imaging findings that mimicked the presence of aortic dissection might have been an atypical presentation of post-mortem clotting or sedimentation. This case is an ideal example to illustrate benefits, limitations, and challenges of post-mortem cross-sectional imaging. It serves as a reminder that both, training as well as correlation of imaging findings with autopsy are fundamental to improve our understanding of radiologic findings on post-mortem cross-sectional imaging.

Total surgical aortic arch replacement as a safe strategy to treat complex multisegmental proximal thoracic aortic pathology.

OBJECTIVE To analyse the results after elective open total aortic arch replacement. METHODS We analysed 39 patients (median age 63 years, median logistic EuroSCORE 18.4) who underwent elective open total arch replacement between 2005 and 2012. RESULTS In-hospital mortality was 5.1% (n = 2) and perioperative neurological injury was 12.8% (n = 5). The indication for surgery was degenerative aneurysmal disease in 59% (n = 23) and late aneurysmal formation following previous surgery of type A aortic dissection in 35.9% (n = 14); 5.1% (n = 2) were due to anastomotical aneurysms after prior ascending repair. Fifty-nine percent (n = 23) of the patients had already undergone previous proximal thoracic aortic surgery. In 30.8% (n = 12) of them, a conventional elephant trunk was added to total arch replacement, in 28.2% (n = 11), root replacement was additionally performed. Median hypothermic circulatory arrest time was 42 min (21-54 min). Selective antegrade cerebral perfusion was used in 95% (n = 37) of patients. Median follow-up was 11 months [interquartile range (IQR) 1-20 months]. There was no late death and no need for reoperation during this period. CONCLUSIONS Open total aortic arch replacement shows very satisfying results. The number of patients undergoing total arch replacement as a redo procedure and as a part of a complex multisegmental aortic pathology is high. Future strategies will have to emphasize neurological protection in extensive simultaneous replacement of the aortic arch and adjacent segments.

Casting materials and their application in research and teaching

From a biological point of view, casting refers to filling of anatomical and/or pathological spaces with extraneous material that reproduces a three-dimensional replica of the space. Casting may be accompanied by additional procedures such as corrosion, in which the soft tissue is digested out, leaving a clean cast, or the material may be mixed with radiopaque substances to allow x-ray photography or micro computed topography (µCT) scanning. Alternatively, clearing of the surrounding soft tissue increases transparency and allows visualization of the casted cavities. Combination of casting with tissue fixation allows anatomical dissection and didactic surgical procedures on the tissue. Casting materials fall into three categories namely, aqueous substances (India ink, Prussian blue ink), pliable materials (gelatins, latex, and silicone rubber), or hard materials (methyl methacrylates, polyurethanes, polyesters, and epoxy resins). Casting has proved invaluable in both teaching and research and many phenomenal biological processes have been discovered through casting. The choice of a particular material depends inter alia on the targeted use and the intended subsequent investigative procedures, such as dissection, microscopy, or µCT. The casting material needs to be pliable where anatomical and surgical manipulations are intended, and capillary-passable for ultrastructural investigations.