The most commonly performed procedure for treating coronary artery stenosis is percutaneous transluminal coronary angioplasty (PTCA) and, where the vessel lumen is severely narrowed, coronary artery bypass grafting (CABG). In PTCA, regions of atherosclerotic plaques are disrupted, and the vessel lumen increased by inflating a balloon catheter. In CABG an autologous saphenous vein into coronary artery interposition graft is performed in order to bypass occluded regions of epicardial coronary arteries. Both interventions cause varying degrees of vascular damage and the long-term efficacy of these procedures is limited by a high incidence of neointimal formation and subsequent vascular restenosis (Bach et al. 1994; Bryan & Angelini, 1994).

The endothelium-derived constrictor peptide, endothelin-1 (ET-1) (Yanagisawa et al. 1988), also possesses mitogenic activity on vascular smooth muscle cells (Hirata et al. 1989) and has been suggested as playing a role in atherosclerosis (Dashwood et al. 1993; Zeiher et al. 1994) and intimal hyperplasia (Dashwood et al. 1993; Douglas et al. 1994).

In mice a molecular motor of the myosin V class (designated myosin Va) is known to be the product of the dilute locus, where a mutation prevents melanosome transport in melanocytes. There is conflicting evidence about whether it has a role in dendrite outgrowth. We investigated its role by transiently transfecting antisense oligonucleotides to inhibit its expression in a melanocyte cell line. We demonstrated mRNA and protein expression of myosin Va in 3 mouse melanocyte lines and 1 human melanoma cell line, using RT-PCR and immunoblotting. Two splice variants were found in human cells whilst only the longer transcript, containing an additional exon, was present in mouse melanocyte lines. The shorter variant was detected in other mouse tissues. Myosin Va protein levels were similar in 3 melanocyte lines with differing amounts of pigmentation, indicating that expression of myosin Va is not tightly coupled to expression of melanin. Immunocytochemistry showed 2 types of myosin Va localisation. A punctate pattern of staining concentrated in the perinuclear region was indicative of organelle association, and the observation of occasional linear punctate staining aligned with F-actin bundles supported the idea that myosin Va has a role in transporting melanosomes along actin filaments. Staining was also intense at tips of dendrites and at sites of dendrite-cell contact, consistent with a possible role in dendrite growth. Transient transfection of antisense phosphorothioate oligodeoxynucleotides targeted against myosin Va mRNA reduced expression of myosin Va protein in cultured mouse melan-a melanocytes by over 70% 20 h after transfection whereas a control (shuffled sequence) oligonucleotide did not. Upon trypsinisation and replating these cells the capacity of the transfected cells to extend new dendrites was reduced in the cells containing the specific antisense oligonucleotides but unaffected by the control oligonucleotide. Image analysis confirmed that the effect of transfection on morphology was statistically significant (P<0.01). In contrast when cells were not trypsinised and replated following transfection so that previously existing dendrites could persist, the normal dendritic morphology continued to be observed. We conclude that in addition to its involvement in melanosome transport, myosin Va has a role in the extension of new dendrites by melanocytes but not in maintenance of pre-existing dendrites.

A method is described for reconstructing the 3-dimensional coordinates of anatomical landmarks from multiple video images. This method consists of 2 stages: (1) separate reconstructions of landmark coordinates visible in dorsal and ventral orientations using Direct Linear Transformation (Abdel-Aziz & Karara, 1972), and (2) least-squares alignment of the partial reconstructions using landmarks present in both. The repeatability of the overall reconstructions and of individual landmarks were assessed by performing multiple independent reconstructions of landmark coordinates for 2 canid specimens. Results indicate that the repeatability of interlandmark distances based on stereophotogrammetric reconstructions from standard-resolution video images is comparable to that of caliper measurement, and slightly superior to that of electronic 3-dimensional digitisation. At least 4 images should be available per landmark for optimal repeatability, but additional images allow only a slight increase in repeatability.

An antiserum against secretory vesicles from human seminal fluid (prostasomes) was used to study the localisation and distribution of the respective antigen(s) during prenatal development and pubertal maturation of the human prostate. The crude antiserum stained both secretory and membrane proteins in the adult prostate and other glands, such as pancreas and parotid gland. An immunoaffinity purified fraction from the antiserum selectively reacted with the apical plasma membrane of prostatic epithelium adluminal cells, recognizing a 100 kDa antigen (PMS). Even in the earliest stages of embryonic prostate specimens studied, the adluminal plasma membrane of the epithelial cells from developing glandular anlagen reacted strongly. The occurrence of PMS immunoreactivity in prostatic anlagen was directly correlated with lumen formation. As the antigen is an androgen-independently synthesised membrane protein of the prostate, it may possibly be used as a marker of cell polarity in the normal and pathologically altered prostate.

Segmental duplications (also termed ‘fenestrations’) of the fetal anterior cerebral artery are a relatively rare anomaly. Their occurrence was studied in 200 fetuses at 20–40 wk gestational age. Duplications were found in 21 of the 200 fetuses, i.e. 10.5%. In 2 fetuses, i.e. 1%, the duplications were bilateral. In 1 case, i.e. 0.5%, 2 duplications were found on the left anterior cerebral artery. There were 15 segmental duplications on the left anterior cerebral artery, i.e. 62.5% of all duplications, and 9 on the right, i.e. 37.5%. The duplications were located in the distal part of the A1 segment in 45.7%, at the level of the anterior communicating artery (A1–A2) in 37.7%, and in the initial part of the A2 segment in 16.6%. The duplications had 4 different forms namely oviform, fissured, triangular and punctate.

In the early stages of embryonic development, many growth-promoting molecules must be provided by the maternal system. The molecules involved in growth processes may be either hormones or growth factors, or molecules that interact with such factors. The pregnancy related hormone, prolactin (PRL, MW 23 kDa) has been implicated in the control of embryonic growth. The growth-promoting potential of PRL and its species-specificity was investigated by culturing 9.5 d rat embryos in vitro for 48 h in depleted serum in the presence and absence of PRL from 3 different species. The growth-supporting capacity of the serum was reduced by removal of low molecular weight molecules by prolonged filtration of the serum using filters with a molecular weight exclusion of 30 kDa. This method provided a ‘semidefined’ medium (retenate) in which embryonic growth and development was significantly reduced, demonstrating that the low molecular weight fraction of serum may contain some growth-promoting factors. Addition of PRL (0.4–25.6 ng/ml) from different species (human, sheep and rat) to retenate significantly improved embryonic growth and development, suggesting that the developing embryo may utilise PRL. Amongst PRLs, rat PRL was found to be active at much lower concentrations than either of the other molecules, and human PRL had more effect in low concentrations than sheep PRL suggesting a species-specificity for this hormone. It may be that the PRL receptors of the rat embryos have greater affinity for the rat hormone as different responses for hormones from different species have been shown. These findings suggest that embryos may be able to utilise maternally derived PRL during organogenesis.

This study was undertaken to investigate the morphological characteristics and lectin affinity of the testicular lamina propria in healthy boars and in unilateral and bilateral abdominal cryptorchid boars. The lamina propria of scrotal testes from healthy boars and unilateral cryptorchid boars was constituted by an innermost noncellular layer, the basal lamina, and by 2 layers of peritubular cells, each separated by a fibrous layer. The noncellular layers contained collagen fibres and glycoconjugates with abundant N-acetylgalactosamine, galactose, fucose, N-acetylglucosamine and neuraminic acid residues. The inner peritubular cell layer was composed of myoid cells, the outer layer of fibroblasts. In the abdominal testes of unilateral and bilateral cryptorchid boars, the lamina propria of nondegenerating and degenerating seminiferous tubules appeared thickened due to an increased content of collagen fibres and glycoconjugates. Glycoconjugates showed decreased amounts of fucose, neuraminic acid and galactose, and increased amounts of N-acetylglucosamine residues. The basal lamina formed infoldings toward the seminiferous epithelium and contained small cells. Both inner and outer peritubular cells were fibroblasts of immature appearance. In degenerated seminiferous tubules of bilateral cryptorchid boars, the lamina propria was composed of a thickened and collagenised basal lamina, without peritubular cells and with a low content of glycoconjugates. In scrotal testes, therefore, the lamina propria was implicated in tubular contractility and in mediating the communication and the substrate diffusion between seminiferous tubules and interstitial tissue. Cryptorchidism induced morphological and histochemical alterations in the lamina propria of abdominal testes, which may be linked to evidence from other studies of lack of tubular contractility and defective cell–cell communication and substrate diffusion. The severity of these anomalies correlated with the severity of Sertoli cell alterations.

The voltage-gated ‘glial’ sodium channel NaG belongs to a distinct molecular class within the multi-gene family of mammalian sodium channels. Originally found in central and peripheral glia, NaG has since been detected in neurons in rat dorsal root ganglia (DRG) and may play a role in Schwann cell-axon interactions. We have studied the presence of NaG-like immunoreactivity in the intact and injured human peripheral nervous system using a specific affinity-purified antibody. Nerve fibres in normal and injured peripheral nerves and normal skin exhibited intense NaG-immunoreactivity. Numerous NaG-immunoreactive nerve fibres surrounded neuronal cell bodies within postmortem control DRG, and in DRG avulsed from the spinal cord (i.e. after traumatic central axotomy). There were no significant differences in the pattern of NaG immunostaining between control and avulsed DRG, or with delay after injury. Generally, the neuronal cell bodies were only very weakly immunoreactive to NaG, indicating that the NaG immunoreactivity was predominantly in Schwann cells/myelin. In accord, we demonstrated NaG immunostaining in cultured human and rat Schwann cells, and in distal nerve after wallerian degeneration. NaG thus appears to be a useful new marker for Schwann cells in the human PNS, and a role in neuropathy deserves investigation.

The purpose of the present study was to analyse the replicability of the scoring of discontinuous traits. This was assessed on a sample of 100 skulls from the Frassetto collection (Dipartimento di Biologia Evoluzionistica Sperimentale of Bologna University) analysed through intraobserver comparisons: the discontinuous traits were determined on the same skulls and by the same observer on 3 separate occasions. The scoring was also assessed through interobserver comparisons: 3 different observers performed an independent survey on the same skulls. The results show that there were no significant differences in the discontinuous trait frequencies between the 3 different scorings by the same observer, but there were sometimes significant differences between different observers. Caution should thus be taken in applying the frequencies of these traits to population research. After an indispensable control of material conditions (subject age included), consideration must be given to standardisation procedures between observers, otherwise this may be an additional source of variability in cranial discontinuous trait scoring.

Morphological variations of the brachial plexus and variants in the distribution of the anterior division of the middle trunk are relatively frequent. Two of the rarest anomalies occurred in the left brachial plexus of a 62-y-old Japanese male, 1 of 104 plexuses dissected between 1996 and 1997 at Kanazawa University Faculty of Medicine. The superior trunk of the brachial plexus was formed by the anterior primary division of C5 and C6 and a thin branch (0.5 mm in diameter) from C4, the middle trunk by the C7, and the inferior trunk by C8 and T1 (Fig.). We could not determine whether there was a branch derived from T2 to T1, since the subject had died of lung carcinoma. The entire anterior division of the middle trunk crossed the axillary artery and joined the medial root of the median nerve which was the continuation of the medial cord after the cord branched off the ulnar nerve. The lateral cord pierced coracobrachialis and divided into the musculocutaneous nerve and the lateral root of the median nerve just after emerging from the muscle, finally joining the medial root of the median nerve superficial to the brachial artery ∼115 mm distal to the lower border of latissimus dorsi to form the median nerve. The musculocutaneous nerve gave rise to the nerves to biceps brachii, brachialis, and the long head of biceps brachii and finally continued as the lateral cutaneous nerve of the forearm. The branch to coracobrachialis had already been cut and its course could not be traced.

Ultrasound of the calcaneus may be used as a cheap, ionising radiation-free and easy to use indicator of skeletal status, and hence of osteoporotic fracture risk. At present ultrasound is not widely used as it suffers from high precision errors. As ultrasound parameters are determined in part by bone mineral density (BMD), an increase in the accuracy and precision of BMD measurements should reduce the precision error associated with ultrasound measurements. The aim of this study was to define an anatomical site on the calcaneus at which accurate and precise measurements of BMD can be made. Ten dry calcanei and 10 cadaveric feet were scanned using a DXA scanner; 9 anatomically defined regions (1 cm2) were selected in the posterior part of the calcaneus for analysis. The centre of region 1 was positioned halfway along the line joining the anterior border of the calcaneal tubercle and the peak of the posterior superior tubercle, and the remaining 8 regions were placed around this central area. The BMD in these 9 regions was compared with the whole bone BMD and the variability of BMD within each of the 9 regions was measured. The reproducibility of the technique was assessed by taking 10 repeated measurements of 2 bone and 2 cadaveric specimens, each specimen being removed and repositioned between measurements. Region 1 was found to be the most representative of total BMD in cadaveric feet. This region also showed the least variability of BMD and consistently gave the lowest coefficients of variation in the reproducibility study both in the bone and the cadaveric specimens. This region is hence the most suitable site on the calcaneus for measuring absolute values of and changes in BMD. The surface position of region 1 was found to be consistently 5/9 along the line at 45° to the vertical, from the lateral malleolus to the heel. The identification of the surface location of region 1 relative to anatomical landmarks of the foot has enabled the same anatomical site to be measured in all subjects. This allows meaningful intersubject comparisons to be made. Preliminary data suggest that precision errors using ultrasound are also reduced when measurements are taken at this region of the calcaneus. The reduction in the precision error of ultrasound assessment of skeletal status may provide a cheap and safe way to identify individuals at risk from osteoporotic fracture.

The organisation of the stromal cell compartment in the mouse lymph node was studied by light and electron microscopy after tissue impregnation by the zinc iodide-osmium (ZIO) method. Fibroblastic reticular cells (FRCs) represented the main stromal cell population. These cells were located both in the cortical region and in the medulla and exhibited various configurations. In the cortex, FRCs were fusiform in shape and came into close proximity with the floor of the subcapsular sinus. In the medulla, the FRCs were shaped like irregular dendritic cells which formed a complex 3-dimensional network. The FRCs surrounded vascular structures such as capillaries and/or high endothelial venules; in these instances they were organised in a discontinuous sheath-like fashion around the vessel wall. By light and electron microscopy, FRCs have been observed to come in close spatial relationship with a number of cells in the lymph node, including sinus endothelial cells, the endothelium of high endothelial venules and capillaries, various types of lymphocytes, follicular dendritic cells and interdigitating cells. These microanatomical features are consistent with the proposal that FRCs may be involved in the communicative networks between the different lymph node compartments. In particular, the FRCs may be involved in the transport of molecules from the sinus compartment to the high endothelial venules or to the distinct cell populations in the lymphoid parenchyma.

To investigate the effect of maternal adrenocortical hormones on the development of fetal pancreatic islet cells, pregnant rats were adrenalectomised on d 6 of gestation. On d 12–16 the growth patterns of fetal insulin-producing B cells, glucagon-producing A cells, and somatostatin-producing D cells were observed histometrically. Maternal adrenalectomy resulted in growth retardation of fetal B cells on d 12–15. Maternal corticosterone therapy prevented this retardation. Maternal adrenalectomy, however, did not affect the developmental patterns of A and D cells. By Western blotting and immunohistochemistry, glucocorticoid receptors were demonstrated to be present in the islet cells from d 12 to d 15. These results suggest that maternal adrenocortical hormones, glucocorticoids in particular, maintain the early development of fetal pancreatic B cells through their specific intracellular glucocorticoid receptor.

Secretory granule formation in pancreatic acinar cells is known to involve massive membrane flow. In previous studies we have undertaken morphometry of the regranulation mechanism in these cells and in mast cells as a model for cellular membrane movement. In our current work, electron micrographs of pancreatic acinar cells from ICR mice were taken at several time points after extensive degranulation induced by pilocarpine injection in order to investigate the volume changes of rough endoplasmic reticulum (RER), nucleus, mitochondria and autophagosomes. At 2–4 h after stimulation, when the pancreatic cells demonstrated a complete loss of granules, this was accompanied by an increased proportion of autophagosomal activity. This change primarily reflected a greatly increased proportion of profiles retaining autophagic vacuoles containing recognisable cytoplasmic structures such as mitochondria, granule profiles and fragments of RER. The mitochondrial structures reached a significant maximal size 4 h following injection (before degranulation 0.178±0.028 μm3; at 4 h peak value, 0.535±0.109 μm3). Nucleus size showed an early volume increase approaching a maximum value 2 h following degranulation. The regranulation span was thus divided into 3 stages. The first was the membrane remodelling stage (0–2 h). During this period the volume of the RER and secretory granules was greatly decreased. At the intermediate stage (2–4 h) a significant increase of the synthesis zone was observed within the nucleus. The volume of the mitochondria was increasing. At the last step, the major finding was a significant granule accumulation in parallel with an active Golgi zone.

The expression of nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d) and nitric oxide synthase (NOS) in spinal ventral horn neurons was studied in the guinea pig after right knee joint immobilisation (RKJI). At 1 wk after RKJI, neurons in the ipsilateral ventral horn from L4 to S1 segments showed a moderate reactivity for NADPH-d staining. At 2 wk, NADPH-d labelled neurons were also observed in the contralateral ventral horn. Ipsilateral NOS immunoreactive cells were not detectable until wk 2. The intensity of NADPH-d and NOS labelled neurons in the bilateral ventral horns was sustained, peaking at the 4th wk after RKJI. In guinea pigs subjected to 4 wk of RKJI and subsequently released from the immobilisation for 2 and 4 wk, NADPH-d and NOS reactivity in ventral horn neurons diminished. The expression of NADPH-d positive neurons differed from that of NOS labelled neurons in terms of time interval, cell number and staining intensity, the latter being later, fewer and weaker. It is suggested that the induction and upregulation of NADPH-d and NOS are attributable to reduced activity of muscles acting on the knee joint after RKJI; the changes are reversible. It is speculated that increased levels of NO production are involved in protective mechanisms against possible neuronal degeneration as a consequence of target dysfunction.

Previous mouse models have indicated that Otx1 and Otx2 play an important role in brain and sense organ development and, together with the Drosophila orthodenticle (otd) gene, they share a high degree of reciprocal functional equivalence. Interestingly, mouse models replacing the same region of the Otx2 locus with Otx1, otd or lacZ genes have revealed the existence of a differential post-transcriptional control between the visceral endoderm (VE) and epiblast cells. Indeed Otx1, otd or lacZ mRNA were transcribed in both tissues but translated only in the VE. Embryos lacking OTX1 or OTD proteins in the epiblast and derived tissues, such as the neuroectoderm and axial mesendoderm (AME), fail to maintain the anterior identity and result in a headless phenotype. This finding leads us to hypothesise that, during evolution, the specification of the vertebrate-type brain may have required epiblast cells to translate Otx2 mRNA in order to establish maintenance properties. The establishment of this regulatory control might have been reflected into a remarkable reorganisation of the rostral CNS architecture and might have represented an important event in the evolution of the vertebrate head. Current data suggest that the Otx2 replaced region and in particular the 3′ untranslated region (UTR), may contain regulatory element(s) necessary to translate and/or stabilise Otx2 mRNA in epiblast and its derivatives.

Intralobular lymphatics in the guinea pig pancreas were demonstrated enzyme-histochemically showing the extent, distribution and fine structure by combined light and transmission electron microscopy. 5′-nucleotidase(5′-Nase)-positive lymphatic vessels were present throughout the pancreas. Intralobular lymphatics among the acini were comparatively rare and generally independent of the blood capillaries, pancreatic ducts and acini. These lymphatics revealed the usual structural features, such as typical intercellular junctions and very tenuous vascular walls without continuous basal laminae. Fine precipitates of the cerium-based reaction product for 5′-Nase activity were found to be associated with cell membranes of the lymphatic endothelium and pinocytotic vesicles. Lymphatics were not closely related to the endocrine islets, although alkaline phosphatase(ALPase)-positive blood capillaries were well developed. Collecting lymphatic vessels with valves with weaker 5′-Nase activity were also detected in the interlobular connective tissue. ALPase activity, absent in the lymphatics, was positive in the blood capillaries, suggesting that it is also a useful way of demonstrating, histochemically, the blood capillaries in the guinea pig pancreas.

Variations in the arterial pattern of the upper limb are common and have been reported by several investigators (Fuss et al. 1985; Poteat, 1986; Tountas & Bergman, 1993; Rodriguez-Baeza et al. 1995). These variations are often associated with anomalies in the arrangement of the nerves of the brachial plexus (Miller, 1939; Lengele & Dhem, 1989). The presence of a superficial brachial artery (Schwyzer & De Garis, 1935; Skopakoff, 1959; Fuss et al. 1985) and the usual pattern of its branching in the upper arm or forearm have also been reported (McCormack et al. 1953; Keen, 1961; Karlson & Niechalev, 1982; Lippert & Pabst, 1985; Rodriguez-Baeza et al. 1995). The great variability of this arterial pattern may be attributed to the failure of regression of some paths of the embryonic arterial trunks (Tountas & Bergman, 1993; Rodriguez-Baeza et al. 1995).

The aim of the present report is to describe the concomitant appearance of 3 unusual variations in the same upper limb of a male cadaver. In this arm: (1) a superficial brachial artery terminated its course by dividing into 3 branches at the cubital fossa; (2) the definitive brachial artery had an unusual origin; and (3) the ulnar nerve was abnormally formed from 2 roots. This novel variation is compared with other anatomical variations in the arterial supply of the upper limb. In a series of routine dissections of 100 embalmed human cadavers the following variations were observed in the right upper limb of a male subject.

The structure and 3-dimensional pattern of the intraparenchymal microvessels in the brain of the lizard, Podarcis sicula, were studied by a combination of light and transmission electron microscopy as well as scanning electron microscopy of vascular corrosion casts. The angioarchitecture pattern consists of narrow hairpin-shaped microvascular loops of different length originating from the meningeal surface. In each loop, descending and ascending vessels are closely apposed to one another throughout their length and are connected by a narrow U-shaped terminal loop at their tips. The 2 limbs of the vessel pairs show a slightly different diameter but lack other structural differences. While some paired vessels give rise to a secondary hairpin-shaped loop with 2 possible branching patterns, there are no anastomotic intraparenchymal connections with analogous neighbouring structures. The cerebral vascular pattern of Podarcis sicula resembles that found in a few representatives of other vertebrate classes. All cerebral vessels structurally appear to be capillaries. Also the observations carried out on semithin and thin sections strongly support the capillary loop model in the Podarcis brain vasculature and, in accordance with studies carried out on various vertebrates, the general submicroscopic features of the brain capillary wall suggest the presence of an endothelial type blood-brain barrier.