Anti-inflammatory activity of the sesquiterpene lactone diacethylpiptocarphol in dextransulfate sodium-induced colitis in mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Sesquiterpene lactones are organic compounds derived mainly from plants that exhibit anti-inflammatory and antitumor activities being one of the key mechanism of action of NF-kB pathway and synthesis of cytokines such as IL-1 and TNF- α. AIM OF THE STUDY: The overall objective of the present study was to evaluate the anti-inflammatory action of a sesquiterpene lactone diacethylpiptocarphol (DPC) from Vernonia scorpioides (Lam.) Pers. and parthenolide (PTH) in Balb-c mice with DSS-induced colitis. MATERIALS AND METHODS: The anti-inflammatory effects of Intraperitonial administration of DPC (5 mg/kg/day) were evaluated in Balb/c mice with DSS-induced colitis, and further the body weight measurement, TNF-α and TGF-ß level was determined. RESULTS: After intraperitoneal treatment for one week, DSS-induced colitis was significantly reduced in mice treated with either of both sesquiterpenes lactones, as witnessed by reduced cellular infiltration, tissue damage, TNF-α production, and enhanced production of TGF-ß. CONCLUSIONS: Sesquiterpene lactone DPC, isolated from Vernonia scorpioides showed anti-inflammatory activity, in this experimental model of colitis the sesquiterpene lactones DPC and PTH exhibit equal anti-inflammatory activity.

Blastocyst production after intracytoplasmic sperm injection with semen from a stallion with testicular degeneration.

In horse breeding, intracytoplasmic sperm injection (ICSI) has gained interest to obtain offspring from subfertile individuals. This paper presents a case report of a stallion with severe testicular degeneration. Semen analysis showed very low motility and 83.5% of detached heads. Histology of a testicular biopsy showed severely decreased spermatogenesis, while transmission electron microscopy of the sperm cells revealed no significant abnormalities. A total of 39 oocytes were fertilized by ICSI with frozen-thawed spermatozoa of this stallion: 25 oocytes with intact spermatozoa and 24 with detached heads. When using intact sperm cells, 8 out of the 25 oocytes cleaved, and 1 developed to the blastocyst stage 9 days after ICSI. None of the oocytes injected with a detached sperm head cleaved. Studies on the paternal influence on ICSI outcome are limited in the horse and further research is needed to define which stallion factors may influence ICSI results. Here, we report the possibility to produce a blastocyst by ICSI of a stallion suffering from testicular degeneration with a poor spermiogram, as long as an intact sperm cell containing a centriole is selected.

Revisiting the segmental organization of the human spinal cord.

In classic anatomic atlases, the spinal cord is standardly represented in its anatomical form with symmetrically emerging anterior and posterior roots, which at the level of the intervertebral foramen combine into the spinal nerves. The parts of the cord delimited by the boundaries of the roots are called segments or myelomeres. Associated with their regular repetitive appearance is the notion that the cord is segmentally organized. This segmental view is reinforced by clinical practice. Spinal cord roots innervate specific body parts. The level of cord trauma is diagnosed by the de-innervation symptoms of these parts. However, systemically, the case for a segmentally organized cord is not so clear. To date, developmental and genetic research points to a regionally rather than a segmentally organized cord. In the present study, to what degree the fila radicularia are segmentally implanted along the cord was investigated. The research hypothesis was that if the fila radicularia were non-segmentally implanted at the cord surface, it would be unlikely that the internal neuron stratum would be segmented. The visual segmented aspect of the myelomeres would then be the consequence of the necessary bundling of axons towards the vertebral foramen as the only exits of the vertebral canal, rather than of an underlying segment organization of the cord itself. To investigate the research hypothesis, the fila radicularia in the cervical-upper thoracic part of five spinal cords were detached from their spinal nerves and dissected in detail. The principal research question was if the fila radicularia are separated from their spinal nerves and dissected from their connective tissues up to the cord, would it be possible to reconstruct the original spinal segments from the morphology and interspaces of the fila? The dissections revealed that the anterior fila radicularia emerge from the cord at regular regionally modulated interspaces without systematic segmental delineations. The posterior fila radicularia are somewhat more segmentally implanted, but the pattern is individually inconsistent. The posterior and anterior roots have notable morphological differences, and hypotheses are presented to help explain these. The macroscopic observations are consistent with a regionally but not a segmentally organized cord. This conclusion was visually summarized in photographs of spinal cords with ipsilateral intact roots and contralateral individually dissected fila radicularia. It was suggested that this dual view of the spinal cord be added to the standard anatomic textbooks to counterbalance the current possibly biased view of a segmented cord.

Postmortem pump-driven reperfusion of the vascular system of porcine lungs: towards a new model for surgical training.

PURPOSE: The objective of this experiment is to establish a continuous postmortem circulation in the vascular system of porcine lungs and to evaluate the pulmonary distribution of the perfusate. This research is performed in the bigger scope of a revascularization project of Thiel embalmed specimens. This technique enables teaching anatomy, practicing surgical procedures and doing research under lifelike circumstances. METHODS: After cannulation of the pulmonary trunk and the left atrium, the vascular system was flushed with paraffinum perliquidum (PP) through a heart-lung machine. A continuous circulation was then established using red PP, during which perfusion parameters were measured. The distribution of contrast-containing PP in the pulmonary circulation was visualized on computed tomography. Finally, the amount of leak from the vascular system was calculated. RESULTS: A reperfusion of the vascular system was initiated for 37 min. The flow rate ranged between 80 and 130 ml/min throughout the experiment with acceptable perfusion pressures (range: 37-78 mm Hg). Computed tomography imaging and 3D reconstruction revealed a diffuse vascular distribution of PP and a decreasing vascularization ratio in cranial direction. A self-limiting leak (i.e. 66.8% of the circulating volume) towards the tracheobronchial tree due to vessel rupture was also measured. CONCLUSIONS: PP enables circulation in an isolated porcine lung model with an acceptable pressure-flow relationship resulting in an excellent recruitment of the vascular system. Despite these promising results, rupture of vessel walls may cause leaks. Further exploration of the perfusion capacities of PP in other organs is necessary. Eventually, this could lead to the development of reperfused Thiel embalmed human bodies, which have several applications.

The calcium-sensing receptor as a regulator of cellular fate in normal and pathological conditions.

The calcium-sensing receptor (CaSR) belongs to the evolutionarily conserved family of plasma membrane G protein-coupled receptors (GPCRs). Early studies identified an essential role for the CaSR in systemic calcium homeostasis through its ability to sense small changes in circulating calcium concentration and to couple this information to intracellular signaling pathways that influence parathyroid hormone secretion. However, the presence of CaSR protein in tissues is not directly involved in regulating mineral ion homeostasis points to a role for the CaSR in other cellular functions including the control of cellular proliferation, differentiation and apoptosis. This position at the crossroads of cellular fate designates the CaSR as an interesting study subject is likely to be involved in a variety of previously unconsidered human pathologies, including cancer, atherosclerosis and Alzheimer's disease. Here, we will review the recent discoveries regarding the relevance of CaSR signaling in development and disease. Furthermore, we will discuss the rational for developing and using CaSR-based therapeutics.

Postgraduate cadaver surgery: An educational course which aims at improving surgical skills.

OBJECTIVE: To describe the postgraduate surgical skills training programme of the Flemish Society of Obstetrics and Gynaecology (VVOG*). Laparoscopic surgical techniques and indications have increased substantially during the past two decades. From surgeons it is expected that they keep up with all techniques and should be able to perform all relevant procedures. Learning new procedures in daily practice is hazardous and difficult to achieve. A training experience with cadaver surgery could improve the course and outcome of surgery on patients. We present the objective, structure, and outcome of the endoscopic postgraduate training course. STRUCTURE: The overall objective of the endoscopic postgraduate training course is to refresh anatomical knowledge and improve general gynaecological laparoscopic surgical skills. The VVOG endoscopic training programme is based on black box training, followed by pig surgery. New is the possibility to perform surgical procedures on specifically prepared human cadavers. The course consists of an anatomical teaching session followed by lectures with videotaped procedures on the anatomical exploration of the pelvis, laparoscopic hysterectomy and pelvic lymphadenectomy. During the hands-on session the participant performs the surgical procedures in a controlled, nonthreatening and interactive way under the guidance of an experienced trainer. CONCLUSIONS: All participants provided feedback on their experiences. The evaluation of the workshop revealed that this course is an opportunity to practise and improve clinical laparoscopic skills of gynaecological procedures and anatomy. Attending the course was regarded as of genuine additional value for surgical practice.

Helicobacter heilmannii sp. nov., isolated from feline gastric mucosa.

Three gram-negative, microaerophilic bacteria, strains ASB1(T), ASB2 and ASB3, with a corkscrew-like morphology isolated from the gastric mucosa of cats were studied using a polyphasic taxonomic approach. The isolates grew on biphasic culture plates under microaerobic conditions at 37 °C and exhibited urease, oxidase and catalase activities. They were also able to grow in colonies on dry agar plates. Based on 16S rRNA gene sequence analysis, ASB1(T), ASB2 and ASB3 were identified as members of the genus Helicobacter and showed 98 to 99 % sequence similarity to strains of Helicobacter felis, Helicobacter bizzozeronii, 'Candidatus Helicobacter heilmannii', Helicobacter cynogastricus, Helicobacter baculiformis and Helicobacter salomonis, six related Helicobacter species previously detected in feline or canine gastric mucosa. Sequencing of the partial hsp60 gene demonstrated that ASB1(T), ASB2 and ASB3 constitute a separate taxon among the feline and canine Helicobacter species. The urease gene sequences of ASB1(T), ASB2 and ASB3 showed approximately 91 % similarity to those of 'Candidatus Helicobacter heilmannii'. Protein profiling, the absence of alkaline phosphatase activity and several other biochemical characteristics also allowed strains ASB1(T), ASB2 and ASB3 to be differentiated from other Helicobacter species of feline or canine gastric origin. The results of this polyphasic taxonomic study show that the cultured isolates constitute a new taxon corresponding to 'Candidatus Helicobacter heilmannii', which was previously demonstrated in the stomach of humans, wild felidae, cats and dogs. The name Helicobacter heilmannii sp. nov. is proposed for these isolates; the type strain is ASB1(T) (=DSM 24751 (T) =LMG 26292(T)) [corrected].

In vitro validation and reliability study of electromagnetic skin sensors for evaluation of end range of motion positions of the hip.

There is growing evidence that femoroacetabular impingement (FAI) is a probable risk factor for the development of early osteoarthritis in the nondysplastic hip. As FAI arises with end range of motion activities, measurement errors related to skin movement might be higher than anticipated when using previously reported methods for kinematic evaluation of the hip. We performed an in vitro validation and reliability study of a noninvasive method to define pelvic and femur positions in end range of motion activities of the hip using an electromagnetic tracking device. Motion data, collected from sensors attached to the bone and skin of 11 cadaver hips, were simultaneously obtained and compared in a global reference frame. Motion data were then transposed in the hip joint local coordinate systems. Observer-related variability in locating the anatomical landmarks required to define the local coordinate system and variability of determining the hip joint center was evaluated. Angular root mean square (RMS) differences between the bony and skin sensors averaged 3.2° (SD 3.5°) and 1.8° (SD 2.3°) in the global reference frame for the femur and pelvic sensors, respectively. Angular RMS differences between the bony and skin sensors in the hip joint local coordinate systems ranged at end range of motion and dependent on the motion under investigation from 1.91 to 5.81°. The presented protocol for evaluation of hip motion seems to be suited for the 3-D description of motion relevant to the experimental and clinical evaluation of femoroacetabular impingement.

Muscle fibres crossing the line of incision used in carpal tunnel decompression.

Muscle fibres that cross the proposed line of incision of the flexor retinaculum at carpal tunnel decompression can be a source of confusion, particularly for the less experienced surgeon. We investigated how frequently muscle fibres crossed the line of incision on the palmar surface of the flexor retinaculum at carpal tunnel decompression in 143 hands, and dissected 103 cadaver hands to study the origin and insertion of these muscle fibres. The line of incision was defined as a longitudinal line between thenar and hypothenar eminences along the third web space, with the wrist in neutral radioulnar deviation and the fingers in extension. Muscle fibres crossing the line of incision were absent in 50% of the operated hands, 2-10 mm wide in 39% and more than 10 mm wide in 11%. In the cadaver hands the proportions were similar at 50%, 35% and 15%, respectively. The fibres were extensions of the thenar and hypothenar muscles and did not appear to represent a separate anomalous muscle.

Deltoid muscle volume estimated from ultrasonography: in vitro validation and correlation with isokinetic abduction strength of the shoulder.

Accurate prediction of muscle parameters can be used for customization of biomechanical models. A method is presented to estimate the volume of triangular-shaped muscles from ultrasonographic measurements. The deltoid muscle was used to validate the technique in a cadaver study of 12 shoulders. The relationship between isokinetic abduction strength and estimated deltoid muscle volume of the dominant shoulder of 30 healthy male subjects was analysed. Isokinetic joint moments were quantified using the Biodex System 3. A significant relationship (R = 0.87; p < 0.001) between the calculated deltoid volume and the isokinetic abduction peak torque was established.

Connexin-related signaling in cell death: to live or let die?

Evidence is accumulating that some forms of cell death, like apoptosis, are not only governed by the complex interplay between extracellular and intracellular signals but are also strongly influenced by intercellular communicative networks. The latter is provided by arrays of channels consisting of connexin proteins, with gap junctions directly connecting the cytoplasm of neighboring cells and hemichannels positioned as pores that link the cytoplasm to the extracellular environment. The role of gap junctions in cell death communication has received considerable interest and recently hemichannels have joined in as potentially toxic pores adding their part to the cell death process. However, despite a large body of existing evidence, especially for gap junctions, the exact contribution of the connexin channel family still remains controversial, as both gap junctions and hemichannels may furnish cell death as well as cell survival signals. An additional layer of complexity is formed by the fact that connexin proteins as such, beyond their channel function, may influence the cell death process. We here review the current knowledge on connexins and their channels in cell death and specifically address the molecular mechanisms that underlie connexin-related signaling. We also briefly focus on pannexins, a novel set of connexin-like proteins that have been implicated in cellular responses to pathological insults.

Connexin 43 hemichannels contribute to the propagation of apoptotic cell death in a rat C6 glioma cell model.

Gap junctions (GJs) have been demonstrated to communicate cell death signals from apoptotic to healthy cells, thereby spatially extending apoptosis. Before being incorporated into GJs, hemichannels (hemi-GJs) are normally closed but recent evidence suggests that they can be opened by various messengers and conditions, thereby forming a pore through which molecules can enter or leave the cell potentially leading to cell death. The aim of this study was to determine the contribution of GJs and hemichannels in the communication of apoptosis toward surrounding cells. We induced apoptosis in C6 glioma cells stably transfected with connexin (Cx)43, with cytochrome C (cytC) using in situ electroporation and found that healthy surrounding cells underwent apoptotic transformation. Work with various cell death markers, wild-type (WT) and Cx43-expressing cells, inhibitors of GJs and/or hemichannels, and Cx43 gene silencing showed that GJs contribute to the spread of apoptosis in a zone next to where apoptosis was triggered whereas hemichannels also promoted cell death beyond this area. Buffering cytoplasmic Ca(2+) changes inhibited the spread of apoptosis in both cases. We conclude that Cx43 hemichannels, in concert with their GJ counterparts, play a role in communicating cytC-induced apoptotic cell death messages.

Molecular-based identification of Sarcocystis hominis in Belgian minced beef.

Sarcocystis hominis, one of the three species of Sarcocystis that cause muscular cysts in cattle, is a protozoan parasite that can infect the human intestinal tract. The objective of the present study was to develop a new molecular identification method capable of discriminating among the bovine Sarcocystis species and to apply this tool in combination with stereomicroscopy to determine the presence of Sarcocystis spp. in minced beef in Belgium, with special attention to Sarcocystis hominis. A PCR technique based on the 18S rRNA sequence and by sequencing of the amplicon was highly specific. Sequence analysis of PCR products from thick-walled cysts collected from minced beef in Belgium revealed that S. hominis was present in 97.4% of the samples. Because the consumption of raw minced beef is common in Belgium and certain other European countries, these findings may point to an underestimated risk to public health.

The effect of Helicobacter felis and Helicobacter bizzozeronii on the gastric mucosa in Mongolian gerbils: a sequential pathological study.

In contrast to Helicobacter(H.) pylori, little is known about the pathogenic mechanisms of gastric non-H. pylori Helicobacter species. Mongolian gerbils were inoculated intragastrically with H. felis or H. bizzozeronii and killed at different timepoints post-inoculation (p.i.), stomach tissue being taken for light and transmission electron microscopy (TEM) and polymerase chain reaction (PCR) analysis. Parietal cells (PCs), apoptosis, cell proliferation and nuclear factor-kappaB (NF-kappaB) activation were "visualized" immunohistochemically. Inflammation consisted of neutrophilic granulocytes, mainly in the antrum, and lymphocytic infiltrates around the limiting ridge and throughout the stomach mucosa and submucosa. From day 11 p.i. onwards, H. felis-inoculated animals showed moderate to severe loss of PCs extending from the limiting ridge into the fundus. Apoptotic cells, spiral bacteria, cell proliferation, and NF-kappaB activation were detected at the transition zone between affected and normal PCs. TEM revealed interaction of H. felis flagella with PCs and chief cells. Moreover, H. felis was seen in proximity to, and inside, necrotic cells. At 10 weeks p.i., some H. felis-infected gerbils showed complete loss of fundic glands, and mucous metaplasia of the epithelium. H. bizzozeronii, which made no flagellar contact with epithelial cells, was associated with only mild PC loss. The mechanism by which H. felis induces PC necrosis and apoptosis remains unclear. The observed flagellar contact and NF-kappaB activation may play an important role in H. felis-associated inflammation.

Macrophages use different internalization mechanisms to clear apoptotic and necrotic cells.

The present study characterized two different internalization mechanisms used by macrophages to engulf apoptotic and necrotic cells. Our in vitro phagocytosis assay used a mouse macrophage cell line, and murine L929sAhFas cells that are induced to die in a necrotic way by TNFR1 and heat shock or in an apoptotic way by Fas stimulation. Scanning electron microscopy (SEM) revealed that apoptotic bodies were taken up by macrophages with formation of tight fitting phagosomes, similar to the 'zipper'-like mechanism of phagocytosis, whereas necrotic cells were internalized by a macropinocytotic mechanism involving formation of multiple ruffles directed towards necrotic debris. Two macropinocytosis markers (Lucifer Yellow (LY) and horseradish peroxidase (HRP)) were excluded from the phagosomes containing apoptotic bodies, but they were present inside the macropinosomes containing necrotic material. Wortmannin (phosphatidylinositol 3'-kinase (PI3K) inhibitor) reduced the uptake of apoptotic cells, but the engulfment of necrotic cells remained unaffected. Our data demonstrate that apoptotic and necrotic cells are internalized differently by macrophages.

Gap junctions and the propagation of cell survival and cell death signals.

Gap junctions are a unique type of intercellular channels that connect the cytoplasm of adjoining cells. Each gap junction channel is comprised of two hemichannels or connexons and each connexon is formed by the aggregation of six protein subunits known as connexins. Gap junction channels allow the intercellular passage of small (< 1.5 kDa) molecules and regulate essential processes during development and differentiation. However, their role in cell survival and cell death is poorly understood. We review experimental data that support the hypothesis that gap junction channels may propagate cell death and survival modulating signals. In addition, we explore the hypothesis that hemichannels (or unapposed connexons) might be used as a paracrine conduit to spread factors that modulate the fate of the surrounding cells. Finally, direct signal transduction activity of connexins in cell death and survival pathways is addressed.

Simultaneous occurrence of a thyromediastinal muscle, a truncus bicaroticobrachialis, and a left superior vena cava.

A case is presented of a combination of anatomical anomalies found in a 67-year-old female cadaver during routine dissection by medical students. They include a thyromediastinal muscle, a truncus bicaroticobrachialis, and a left superior vena cava, with complete absence of the right superior vena cava, but with a normal azygos vein opening into the right atrium at the expected site of entry of the superior vena cava. No associated congenital cardiac malformations were found. Clinical implications include the difficulty of heart catheterization through the subclavian veins and misleading images on CT or MRI scans, where the azygos vein could be mistaken for a right superior vena cava.

Therapeutic ultrasound: temperature increase at different depths by different modes in a human cadaver.

A specific increase in temperature is necessary to achieve a temperature-mediated therapeutic impact by ultrasound in rehabilitation. In order to obtain a physical basis for a definite temperature rise at a certain depths in the human body a human cadaver study in situ was conducted. A set of 16 experiments was carried out with an ultrasound frequency of 1 and 3 MHz. For each frequency the pulsed and continuous mode of energy delivery were compared at a variable intensity of 0.5, 1.0, 1.5 and 2.0 W/cm2. The ultrasonic energy was delivered in direct contact to the cadaver at the posterior crural compartment of the leg in a static manner. Temperature was monitored with a 1-minute interval during an insonation of 10 minutes. Results revealed that theoretical physical predictions concerning tissue heating by therapeutic ultrasound could not always be consolidated and that thermal therapeutic effects for deeper conditions are not obvious to be achieved by ultrasonic therapy.

Mitochondrial transmembrane potential changes support the concept of mitochondrial heterogeneity during apoptosis.

Dissipation of mitochondrial membrane potential (DeltaPsi(m)) and release of cytochrome c from mitochondria appear to be key events during apoptosis. The precise relationship (cause or consequence) between both is currently unclear. We previously showed in a model of serum-free cultured granulosa explants that cytochrome c is retained in a subset of respiring mitochondria until late in the apoptotic process. In this study we further investigated the issue of heterogeneity by using the DeltaPsi(m)-sensitive probe CM-H2TMRos in combination with a DNA fluorochrome. Changes of DeltaPsi(m) were assessed qualitatively by epifluorescence microscopy and were quantified using digital imaging microscopy. This approach yielded the following results: (a) CM-H2TMRos staining is a reliable and specific procedure to detect DeltaPsi(m) changes in granulosa cells explants; (b) dissipation of transmembrane potential is an early event during apoptosis preceding nuclear changes but is confined to a subpopulation of mitochondria within an individual cell; (c) in frankly apoptotic cells a few polarized mitochondria can be detected. These findings support the hypothesis that ATP needed for completion of the apoptotic cascade can be generated during apoptosis in a subset of respiring mitochondria and is not necessarily derived from anaerobic glycolysis.

Contribution of progesterone, follicle stimulating hormone and glucocorticoids in survival of serum-free cultured granulosa cell explants.

To investigate the role of progesterone (P4) as a survival factor in quail granulosa cell explants, P4 content was determined under various conditions and correlated with apoptotic indexes (AIs) evaluated by 2',6'-diamidino-2-phenylindole (DAPI)-staining. Analysis of serum-free cultures from 24 to 96 h shows decreased P4 levels in the medium paralleled by increasing AI. Inhibiting apoptosis by gonadotropic support (FSH, 100 ng/ml) stimulates a 3-fold increase of the P4 level in the medium (83.49+/-8.69 vs 26.31+/-1.61 ng/ml in serum-free controls) together with a significant decrease in AI from 8.81+/-1.06% in serum-free controls to 3.50+/-0.72%. Substantial evidence for P4 as an autocrine/paracrine survival factor can be inferred from experiments with aminoglutethimide (AG, 1 mM) and RU486 (20 microM). Blocking P4 synthesis by AG causes a 2-fold increase in apoptosis from 6.08+/-0.67% in serum-free controls to 12.53+/-1.60%. Blocking P4 receptors by RU486 causes a similar increase in AI (3.02+/-0.98% in serum-free controls to 17.07+/-3.20%) and about a 50% decrease in P4. The effect of RU486 could be attenuated by exogenous P4 but not by dexamethasone indicating selective binding of P4 to the progesterone receptor. Dexamethasone treatment promotes survival without affecting P4 levels. In further support of an autocrine/paracrine action for P4 in the granulosa cells, both the A and B form of the avian P4 receptor (PR) are identified in vivo and in vitro by Western blotting. Exogenous administration of P4 only affects survival when endogenous P4 synthesis is blocked or after 48 h of serum-free culture when endogenous P4 production is very low. Because FSH also affects survival when its stimulatory effect on P4 synthesis is blocked by AG (AI decrease from 6.08+/-0.67% in serum-free controls to 1.64+/-0.71% in FSH+AG treated) it is proposed that (1) P4 is an autocrine/paracrine survival factor in the preovulatory granulosa and (2) FSH mediates both P4-dependent and P4-independent survival pathways.