Heinz Augsburger

Isotopic signatures of iron in body tissues as a potential biomarker for iron metabolism

Previous studies using MC-ICP-MS for high precision isotope ratio analysis revealed that human blood is enriched in light iron isotopes relative to dietary iron. Moreover, distinct differences in blood iron isotopic patterns between individual subjects can be observed. Discrimination of heavy iron isotopes during uptake of dietary iron in the gastrointestinal tract may explain this observation. This hypothesis was now tested in the pig model. Samples of tissues relevant to iron metabolism were screened for iron isotopic patterns using MC-ICP-MS, revealing considerable isotope fractionation during intestinal uptake on one hand and during distribution between body tissues on the other hand. Light iron isotopes are enriched in the gastrointestinal mucosa relative to feed, with isotopic patterns reflecting major iron absorption sites in mammals along the gut. While heavy iron isotopes are enriched in the liver, spleen and bone marrow, light isotopes are enriched in red blood cells. Well-established links between iron absorption efficiency and iron distribution between functional compartments and stores point to blood iron isotope signatures as a novel type of compound biomarker of element metabolism.

Morphology and stereology of the female canine urethra correlated with the urethral pressure profile

Perfusion-fixed urethrae of five sexually intact nulliparous female beagles in anestrus, between 2 and 3 years of age, were morphologically and morphometrically investigated. Absolute and relative volumes of different tissue components were stereologically estimated per urethral quarter. A discrete smooth muscle sphincter could neither be identified in the bladder neck nor in the urethra. However, the circularly arranged smooth muscle fibers in the proximal half of the urethra were regarded as being capable of sphincteric function. The connective tissue, composed of collagenous and longitudinally orientated numerous elastic fibers, accounted for the most predominant constituent. It comprised 77.3 to 78.2% of the total volume in the proximal three urethral quarters. The vascular plexus was most prominently developed in the proximal two urethral quarters, and its relative and absolute volumes significantly exceeded those of the third quarter. The discovered direct connections between small arteries and the sinusoids of the vascular plexus emphasize its importance as a major contributory component to urinary continence. The striated musculus urethralis was essentially confined to the distal half of the urethra and most strongly developed in the fourth quarter, where it surrounded the urethra transversely at its ventral and lateral aspects. The caudal muscle fibers continued dorsally to the vagina. A transitional epithelium bounded the urethral lumen. Its superficial cell layer progressively flattened towards the external urethral orifice, changing into a stratified squamous lining as also seen in the vestibulum. Recorded pressures, or pressure changes, could neither be correlated with volumes nor with morphology of particular tissue elements in the different urethral quarters.

Stereological Analysis of the Urethra in Sexually Intact and Spayed Female Dogs

The perfusion-fixed urethrae of 5 sexually intact nulliparous and 5 ovariohysterectomized beagles were analyzed stereologically. Different urethral tissue components were estimated stereologically in each of the three proximal urethral quarters and compared between the two groups of dogs. Considerable individual variability within all groups was observed. In all the three urethral quarters considered, average smooth muscle and connective tissue volumes were generally lower in spayed than in intact animals, both in absolute and relative terms. However, only the difference between the mean relative volumes of urethral quarter I was statistically significant. In contrast, mean absolute and relative volumes of the vascular plexus were generally greater in the ovariohysterectomized dogs as compared to the animals of the other group--deviations in statistical relevance concerning the first urethral quarter were noted. Mean epithelial and striated muscle volumes did not vary essentially between the spayed and intact group of dogs. Despite the fact that all findings were not always statistically meaningful, our research nevertheless demonstrates that ovariohysterectomy decreases smooth muscle mass and connective tissue; it also elevates vascular plexus volumes in the urethra of spayed animals. Moreover, our results imply a general weakening of the urethral wall as a sequel to ovariohysterectomy, and this, in turn, could be seen to affect the closure mechanism of the urethra.

Morphological, histochemical and stereological analysis of the female canine M. urethralis

The M. urethralis was morphologically investigated in ten medium-sized female dogs of different breeds and age, as well as histochemically and stereologically analysed in a homogeneous group of five female beagles. Macroscopically, the muscle was essentially confined to the distal third and most strongly developed in the fourth quarter of the urethra. Here, it surrounded the urethra transversely at the ventral and lateral aspects, passing with its caudal fibres dorsally onto the vagina. The muscle fibres were assembled in groups of different sizes and usually separated by thick connective tissue septae. Based on the myofibrillar actomyosin ATPase (mATPase) reaction, type I and two main subtype II fibres could be differentiated. Type II fibres were, however, indistinguishable by their metabolic enzyme activities since both subclasses displayed oxidative-glycolytic properties. The subtype II fibres containing the more acidlabile mATPase activity were classified as IIA, whereas the other main subtype was designated IIS (subtype) and considered as peculiar to the dog. In addition, the investigation revealed a rare fibre type exhibiting the histochemical profile of IIC fibres. There was no evidence of classical glycolytic IIB fibres. The M. urethralis was composed of 24% type I and 76% type II fibres with an average diameter of 34.9 and 28.5 μm, respectively. Connective tissue constituted 52.8% of the total muscle volume. Due to the predominance of fast twitch II fibres, the urethral muscle is not designed to maintain a high tone over long periods of time. Its primary role is to function rapidly and intermittently guaranteeing urinary continence during stress situations when other continence factors are overburdened.

Influence of ovariectomy on the canine striated external urethral sphincter (M. urethralis): a stereological analysis of slow and fast twitch fibres

Abstract Slow and fast twitch fibres were stereologically analysed in the morphologically defined and most strongly developed ventral and ventrolateral region of the external urethral sphincter (M. urethralis) using five sexually intact anestrous and five ovariectomized nulliparous beagles. The estimated mean total cross-sectional area of the investigated part of the muscle consisted of 4.2u2009mm2 (15.1%) type I fibre, 10.0u2009mm2 (32.1%) type II fibre, and 16.3u2009mm2 (52.8%) connective tissue in the control group. The corresponding absolute mean value of type I fibres (3.7u2009mm2/13.5%) was statistically lower in ovariectomized animals. No significant difference between groups was observed in the relative number of transverse profiles of type I and II fibres; type I fibres comprised 23.8% and type II 76.2% of all muscle fibres in the sexually intact group, but 21.8% and 78.2% in the ovariectomy group, respectively. The ovariectomized dogs exhibited a statistically significant lower type I and II fibre number and a concomitant slightly larger mean single profile area (diameter) of fibre type II compared with the control animals. The significantly reduced number and decreased total cross-sectional area of the fatigue-resistant type I fibres in ovariectomized dogs suggest a predominant weakening of the fibre type I portion of the M. urethralis as consequence of ovariectomy. The effect could be mediated by sex hormonal factors and may contribute to the development of postspaying urinary incontinence in female dogs.

Immunohistochemical analysis of collagen types I, III, IV and α-actin in the urethra of sexually intact and ovariectomized beagles

Urinary incontinence is a widespread problem in both postmenopausal women and ovariectomized dogs. The objective of this study was to investigate the influence of ovariectomy on the immunoreactivity and the distribution pattern of collagens I, III, IV and α-actin in the canine urethra. The immunohistochemical results were evaluated in five sexually intact and five ovariectomized beagles. The immunostaining of both collagens I and III delineated urethral connective tissue fibres and co-localized within in the fibres of both groups. The basement membranes of smooth muscle cells and sinusoids showed marked type IV collagen expression, whereas only faint immunoreactivity was present at the urothelial–stromal interface. No differences could be detected in the expression or distribution of the assessed collagen types and actin between ovariectomized and control animals. In conclusion, ovariectomy does not appear to have an effect on urethral collagens I, III, IV and smooth muscle actin in the dog, as ascertained by immunohistochemistry.

Ultrastructural alterations in mammalian parathyroid glands induced by fixation

The influence of fixation methods, buffers and ions on the ultrastructure of parathyroid cells was studied in dogs, cats, rats and mice. Parathyroids fixed by immersion showed 3 chief cell variants referred to as cells in active, intermediate and resting stages, multinucleated syncytial cells, atrophic cells and, only in 1 feline parathyroid, a few oxyphil cells. Parathyroid glands fixed by perfusion, however, consisted only of 1 cell type. Satisfactory preservation was achieved by perfusion with 2.5% glutaraldehyde in 0.1 M Na cacodylate containing 0.25 mM CaCl2 and 0.5 mM MgCl2, and postfixation with 1% OsO4 in 0.1 M s-collidine containing 0.5 mM CaCl2 and 1.0 mM MgCl2. Good preservation was also obtained using Na phosphate during prefixation and postfixation. Other combinations of buffers led to shrinkage, dilation of rough endoplasmic reticulum cisternae, disruption of membranes or loss of matrix and secretory granules. The results demonstrate that the variants of parathyroid chief cells, multinucleated syncytial cells and atrophic cells arise during fixation.

Computed tomography measurements of thoracic structures in 26 clinically normal goats.

The present study was performed to provide computed tomographic (CT) reference values for structures in the thorax of 26 clinically normal Saanen goats. Animals were anesthetized, positioned in sternal recumbency and transverse images with a reconstructed 1.5mm slice thickness were obtained by use of a 40-slice CT scanner. Absolute and relative measurements of the trachea, heart, cranial vena cava, thoracic aorta, caudal vena cava, right and left principal bronchus, right and left caudal lobar bronchus and the concomitant branch of the right and left pulmonary artery and vein, large caudal mediastinal lymph node and lung density were performed with dedicated software. Minimal to moderate interstitial or bronchopneumonia was incidentally found on CT in 24 animals. In conclusion, CT images obtained in this study can be used as a reference for the evaluation of thoracic diseases in goats.

Immunohistochemical expression of collagen types I, III, IV and α-actin in the uterine horns of nulliparous and multiparous beagles

Collagen and smooth muscle cells play essential roles in the remodelling of uterine tissue during pregnancy and involution. To investigate the immunoreactivity and distribution pattern of collagen types I, III, IV and smooth muscle alpha-actin resulting from these processes, two homogenous groups of nulliparous and multiparous beagles were evaluated by immunohistochemistry. Immunostaining patterns of collagens I and III delineated the uterine connective tissue fibers and revealed their dual presence within fibers of both beagle groups. Collagen III staining, in particular, was more pronounced and especially evident in superficial fiber sections. The numerous, large arteries in the myometrial stratum vasculare of multiparous uteri exhibited a highly thickened intima, which distinctly expressed type I and III collagens. Intense collagen IV immunolabeling was discernable in the basement membranes of vascular endothelia and smooth muscle cells. Staining of the basement membranes of the luminal and glandular epithelia, conversely, was either absent or very weak. No difference in the immunoreactivity and distribution of the assessed collagens and actin could be detected between nulliparous and multiparous dogs. Overall, and with the exception of sclerotic arteries, immunohistochemical analysis revealed that the expression of uterine collagens and actin does not change in the uterus of multiparous beagles, even after seven elapsed pregnancies.

Differences of morphometrical parameters in hind limb muscle fibres between ovarectomized and sexually intact female dogs

Slow and fast twitch fibres of the Mm. tibialis cranialis, semitendinosus and sartorius of seven sexually intact and seven ovarectomized female beagles were histochemically and morphometrically analysed. Along with type I and type IIA fibres, another main type II fibre (IIS), which seems to be peculiar to the dog, was found in the Mm. semitendinosus and tibialis cranialis. Type I fibers comprised 26% and type II fibres 74% of all recorded muscle fibres in the M. tibialis cranialis, 29% (type I) and 71% (type II) in the M. semitendinosus and 51% (type I) and 49% (type II) in the M. sartorius, respectively. The average single profile area and the corresponding mean diameter of fibre types I and II in the investigated hind limb muscles were generally larger in ovarectomized than in sexually intact animals. This was more evident in type II than in type I fibres. However only the type II fibres of the M. tibialis cranialis and sartorius exhibited a statistically significant increase in diameter (p < 0.01 and p < 0.05, respectively). Accordingly, the mean density (number of fibres/mm2) of both fibre types in the hind limb muscles of spayed dogs was generally reduced. Again, this reduction attained statistical relevance in the type I and II fibres of the tibialis cranialis. In addition, the fibre densities of type I in the semitendinosus and type II in the sartorius muscles were also significantly reduced in ovarectomized dogs. In conclusion, ovarectomized beagles showed a generally increased mean diameter of the investigated type I and II hind limb muscle fibres and a concomitant decreased average fibre density of the respective types when compared to sexually intact animals.