Chromatin-associated HMG-17 is a major regulator of homeodomain transcription factor activity modulated by Wnt/beta-catenin signaling.

Homeodomain (HD) transcriptional activities are tightly regulated during embryogenesis and require protein interactions for their spatial and temporal activation. The chromatin-associated high mobility group protein (HMG-17) is associated with transcriptionally active chromatin, however its role in regulating gene expression is unclear. This report reveals a unique strategy in which, HMG-17 acts as a molecular switch regulating HD transcriptional activity. The switch utilizes the Wnt/beta-catenin signaling pathway and adds to the diverse functions of beta-catenin. A high-affinity HMG-17 interaction with the PITX2 HD protein inhibits PITX2 DNA-binding activity. The HMG-17/PITX2 inactive complex is concentrated to specific nuclear regions primed for active transcription. beta-Catenin forms a ternary complex with PITX2/HMG-17 to switch it from a repressor to an activator complex. Without beta-catenin, HMG-17 can physically remove PITX2 from DNA to inhibit its transcriptional activity. The PITX2/HMG-17 regulatory complex acts independently of promoter targets and is a general mechanism for the control of HD transcriptional activity. HMG-17 is developmentally regulated and its unique role during embryogenesis is revealed by the early embryonic lethality of HMG-17 homozygous mice. This mechanism provides a new role for canonical Wnt/beta-catenin signaling in regulating HD transcriptional activity during development using HMG-17 as a molecular switch.

Atrial structure and plasma ANF levels in rats with chronic diabetes mellitus.

The effect of streptozocin-induced diabetes (STZ-D) on right atrial structure was investigated in male Wistar rats. STZ (55 mg/kg) or saline (1 ml/kg) was administered by intravenous injection 12 wk before the experimental studies. Tissue was sampled from four regions of the atrium, processed, and embedded in plastic. Quantitative stereological analysis indicated that in STZ-D rats, there was a significant diminution in size of the musculi pectinati (muscular ridges), which form a network making up the wall of the atrium. In addition, within the muscular ridges, there was a significant reduction in the relative proportion of cardiocytes within the cardiac tissue. The rest of the cardiac tissue consisted of interstitial regions, connective tissue, and blood vessels, which correspondingly increased. This suggests there was some form of cardiomyopathy. When atrial granularity was determined relative to cardiocyte volume density, a significant decrease (54%) was found in tissue from STZ-D rats. The blood pressure of conscious STZ-D rats was significantly lower than control rats, whereas right atrial pressure was not different. The level of resting plasma immunoreactive atrial natriuretic factor (ANF) in conscious STZ-D rats (98 +/- 5 pg/ml) was significantly higher than in control rats (52 +/- 7 pg/ml). The decreased atrial granularity could be related to the higher resting plasma ANF levels, suggesting a more rapid turnover or increased synthesis bypassing storage in the granular form.

Coexistence of diabetes and hypertension results in unique structural alterations in the renal artery in rats beyond that found with diabetes alone.

Numerous investigators have presented evidence of increased mortality in patients with diabetes mellitus due to cardiovascular disease. It is still unclear as to the reasons why there is a predisposition to vascular pathology that in the advanced state leads to atherosclerosis. Our hypotheses were: (1) The condition of diabetes mellitus in a streptozocin animal model may show vascular changes similar to early pathology in macrovessels and (2) since the model is normotensive, inducing hypertension will result in early atherogenic pathology. We carried out a quantitative analysis of the renal artery using light and electron microscopy to test the hypotheses. Male Wistar rats had diabetes mellitus induced using streptozocin and 1 week later half of the diabetic animals had hypertension induced with deoxycorticosterone acetate (DOCA). Samples were taken following 7 weeks of diabetes, or 7 weeks of diabetes with DOCA administration during the final 6 weeks. The renal artery from the diabetic group did not have any differences in wall or luminal dimensions from control, but did have proportionately more extracellular matrix than smooth muscle in the tunica media. This is evidence of structural change, in a large supply artery, as a manifestation of diabetes mellitus, similar to that seen in vascular disease. Vessels from the control/hypertensive had a significantly thickened tunica media as did the diabetic/hypertensive over control values. The latter also had proportionately even greater significant elevation of the extracellular matrix compared with either the diabetic or control/hypertensive. In addition, only the diabetic/hypertensive group showed marked subendothelial invasion of macrophage type cells and deposits of various shapes and densities. We have, therefore, demonstrated significant vascular alteration due to the diabetic condition in this animal models and also shown that with hypertension and diabetes combined, the early vascular pathology is exacerbated.

Efficacy and specificity of pharmacological therapies for behavioral disorders in persons with mental retardation.

This review assesses the efficacy and specificity of psychotropic medications used to control aberrant behavior in persons with mental retardation. It is concluded that neuroleptics, the most widely used psychotropic agents in this population, suppress aberrant behavior, but do so by suppressing behavior generally. An exception to this conclusion is that it may be possible to selectively suppress stereotyped behavior with neuroleptics. In addition, the empirical evidence indicates that, in some persons with mental retardation, opioid antagonists and methylphenidate are useful therapies for self-injurious behavior and hyperactivity, respectively. Lithium and beta-blockers are potentially useful for treating aggression.

Intraluminal endothelium-covered bridges in chronic fat-fed balloon-injured Yucatan miniswine.

The Yucatan miniswine has been recommended as an animal model of advanced atherosclerosis. Atherosclerotic plaques developed in this model demonstrate foam cells, widespread fibrosis, and calcification, features suggestive of human atherosclerosis. We have observed the occurrence of intraluminal projections that appear peculiar to this animal model. Forty-three miniswine, weighing between 20 and 30 kg, were rendered atherosclerotic with a combination of balloon endothelial injury of the aortoiliac segments and dietary supplementation with 2% cholesterol and 15% lard. Endothelial injury was created by retrograde balloon catheter injury of the aorta and both external iliac arteries via cutdowns on the femoral arteries. Serum cholesterol prior to starting the diet and at 1, 2, and 6 weeks following initiation of the diet was 2.0 +/- 0.4, 11.6 +/- 4.0, 15.9 +/- 5.0, and 16.4 +/- 4.2 mM, respectively (p < .0001, ANOVA). Angiographically significant lesions were apparent in 33 of 37 (89%) animals (occlusion 20/37, stenosis 17/37) at 6 weeks postinjury. In three of six (50%) animals followed up to 16 weeks postinjury, trabecular areas were seen in the external iliac arteries on angiography. Light and electron microscopy demonstrated that these areas were covered with normal endothelium and projected into the lumen or bridged with the adjacent arterial wall. Foam cells and calcification were not seen in these lesions. This finding is not typical of human atherosclerosis and appears peculiar to this type of animal model.

Changes in size and shape of smooth muscle cells from the portal vein of spontaneously hypertensive rats: an ultrastructural analysis.

The portal vein was investigated in male spontaneously hypertensive rats (SHR), from Wistar-Kyoto (WKY) and Wistar strains, in animal 16-20 weeks old. In SHR, the inner circular smooth muscle was unchanged, but the outer longitudinal layer showed marked alterations in shape and size, readily observed in three-dimensional micrographs using scanning electron microscopy. The cells in both Wistar and WKY were elongate and tubular with little variation along their lengths and with a relatively smooth sarcolemma. This applied to both the inner and outer layers of smooth muscle. In contrast, the smooth muscle cells from SHR in the outer layer varied considerably in thickness along their lengths, and had very irregular outlines with numerous pits or depressions of varying sizes. In addition, the cells frequently had major forks or branches. The vasa vasorum running through the muscle layer, fibroblasts and nerve bundles were also identified. Sectioned material (transmission electron microscopy) showed a change in shape and hypertrophy of the smooth muscle cells from the portal vein of SHR, and also demonstrated a significant increase in paracellular connective tissue in the outer layer of smooth muscle. Such major morphological alterations in the outer layer of smooth muscle in the portal vein from SHR could have profound effects on functional studies.

Trophic interactions between rat nerves and blood vessels in denervated peripheral arteries and in anterior eye chamber transplants.

This investigation was undertaken to examine trophic interrelationships between nerves and arteries in male Wistar rats. Two approaches were used. (1) Surgical denervation of two peripheral muscular arteries in the thigh (the superficial epigastric and saphenous) was carried out on young animals (5-20 days old). (2) Arteries from young adults, either with a high density of innervation in situ (the tail artery), or virtually uninnervated (the femoral artery), were transplanted into intact or sympathectomized anterior eye chambers of adult rat hosts. In the denervation experiments, the maximum length of time before reinnervation occurred was 15 days postoperatively. The only evidence of morphological change in the vessel wall was in the external elastic lamina that became irregular and laminated. Reinnervation followed the typical developmental sequence, and was accelerated in the younger animals and by a double lesion. Translocating the proximal part of the nerve carrying the vasomotor innervation indicated that sprouting was directional toward the muscular arteries, bypassing an artery with very sparse innervation. The transplant experiments into the anterior eye chamber showed that only an artery densely innervated in situ (the tail artery) could induce reinnervation by iridean nerve sprouting. The tail artery, in the chamber lacking adrenergic innervation of the iris, became reinnervated by terminals with small agranular vesicles. These vesicles were part of Schwann cell complexes, at a similar relative density, occupying the same position in the vessel wall, as the ingrowing nerves in the fully innervated iris. The latter also had a proportion of terminals with the small clear vesicles. A small population of large granular vesicles could also be found in both types of terminals. Therefore, tissue normally having only sympathetic innervation cannot be assumed to be completely noninnervated when transplanted into a sympathectomized anterior eye chamber. The denervation and transplant experiments described here demonstrated the presence of trophic interactions between nerves and arteries, but also revealed a heterogeneity of response between vessels with very high and extremely low levels of innervation in situ.

Hypertensive structural changes in blood vessels: do endothelial cells hold the key?

In recent decades, the complexity of the endothelium and its major role in maintaining or altering blood vessel architecture are being revealed. In contrast, the vascular smooth muscle cell previously received the most attention. I suggest support of the hypothesis that the endothelium is the key to vascular disease. An altered endothelium in diabetes mellitus likewise is likely to be pivotal in vascular complications that develop. We have demonstrated that adherent monocytes, indicators of altered endothelium, occur in deoxycorticosterone acetate induced hypertension in male Wistar rats. The coronary artery and thoracic aorta were investigated using transmission electron microscopy. Details of hypertensive changes were revealed as well as early atherogenic pathology in the absence of dietary modifications. Scanning electron microscopy of thoracic aorta showed details of the luminal endothelial surface and adherent monocyte-macrophages in hypertensive animals. There were two cell types: numerous typical monocytes with upstream tails, and larger cells that may have been free grazing macrophages or macrophages that had returned to the circulation. Debris and amorphous material were particularly evident in vessels from hypertensive animals. Monocytes squeezed between intact endothelial plasma membranes (as seen in section), and were found as subendothelial foam cells and phagocytosing macrophages. The endothelial adherence of monocytes to the aortas from diabetic animals was significantly (p less than 0.05) elevated over that found in controls (but not different from control-hypertensive or diabetic-hypertensive animals) supporting the concept of altered endothelium in diabetes.

Development of adrenergic innervation in rat peripheral vessels: a fluorescence microscopic study.

The postnatal development of adrenergic innervation was followed in peripheral blood vessels of Wistar rats. The femoral vessels and their branches, the superficial epigastric and saphenous vessels, and the tail artery, were investigated from birth to maturity. The proximal ends of the vessels were studied with fluorescence microscopy after the catecholamine was converted to a fluorophore by hot formaldehyde vapour, and ultrastructural morphology confirmed that the nerve varicosities mainly contained small vesicles with dense cores, typical of adrenergic innervation. Further confirmation was obtained with reserpine pre-pretreatment, the sodium borohydride specificity test, and experiments to alter the non-specific fluorescence of elastin. The nerves in the arteries were immediately adjacent to the external elastic lamina, and they retained this position throughout postnatal development. Of the three muscular arteries, the development of innervation was earlier and more intense in the saphenous and superficial epigastric arteries than in the tail artery. However, the tail artery surpassed the other two both in the total number of nerves and in the density of innervation per unit area beyond 12 days of age, and maintained this lead to maturity. The superficial epigastric artery had the smallest total number of nerves but had a greater density of innervation than the saphenous. The femoral artery did not develop any appreciable innervation. The femoral vein demonstrated the greatest amount of fluorescence of any of the veins, the others having considerably less innervation than their companion arteries.

Arterial wall and smooth muscle cell development in young Wistar rats and the effects of surgical denervation.

Development of the muscular saphenous artery and the effect of surgical denervation on normal development was investigated in young rats at 3 and 6 weeks of age. During this interval, the weight and blood pressures (systolic, diastolic, and mean) of the animals increased significantly. The tunica media of the artery and the lumen increased significantly with age, but the proportion of smooth muscle cell to paracellular matrix did not alter. Computer-assisted three-dimensional reconstructions were used to investigate the smooth muscle cells. They increased significantly in length, volume, and angle of orientation within the vessel wall with age but maintained an approximate surface area-to-volume ratio. The cells in any one vessel tended to be oriented in either a clockwise or counterclockwise direction. The size of the nucleus also increased significantly in length and volume with age, but an approximate surface area-to-volume ratio and a constant nucleocytoplasmic ratio were maintained. The nuclei tended to be eccentrically located, with less than half of all nuclei wholly within the middle third of the cell. Surgical denervation at 10 days of age resulted in abnormalities of growth in vessel dimensions, thinner tunica media at 3 weeks (denervated 11 days previously), and smaller lumen at 6 weeks (denervated 32 days previously). Elevated amounts of paracellular matrix occurred in both age groups, but denervation did not alter smooth muscle cell size. In the 3-week-old animals, denervation resulted in smooth muscle cells with hypertrophied nuclei. This may account for the increase in growth of the tunica media between 3 and 6 weeks of age in the denervated artery.(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of postnatal growth and denervation on characteristics of the saphenous artery in SHR and WKY rats between 3 and 6 weeks of age: an in vitro study.

Thigh vessels of spontaneously hypertensive (SHR) and Wistar-Kyoto (WKY) rats were unilaterally surgically denervated at 10 days of age by femoral nerve section. Denervated and contralateral control segments of saphenous arteries from 3- and 6-week-old rats were mounted in a small vessel myograph for study. Both strains showed growth changes in blood pressure, but there was no significant difference between WKY and SHR. Both strains also had significant growth changes in vessel dimensions and the in vitro measurements suggested that SHR vessels had a thicker wall. Denervation did not affect vessel size. Transmural nerve stimulation indicated loss of innervation due to the surgical procedure. In the denervated vessels, both norepinephrine (NE) and 5-hydroxytryptamine (5-HT) dose response curves were shifted to the left, indicating a postjunctional increase in sensitivity. Maximum tension developed was in the order K+ greater than 5-HT greater than NE. In comparing the two strains, vessels from 6-week-old SHR were less sensitive to 5-HT. Relaxation to acetylcholine was significantly decreased in denervated arteries from WKY, whereas in SHR the significant decrease occurred only at 3 weeks. Denervated vessels from both rat strains at 3 weeks showed greater relaxation to beta-receptor activation, but not at 6 weeks of age. Therefore, the absence of functional innervation resulted in altered function of the saphenous artery wall.

Improved ultrastructural detail in tissues fixed with potassium permanganate.

An improved method has been developed for fixation with potassium permanganate. Although this is one of the methods widely used to preserve the dense cores of adrenergic storage vesicles, fixation of other tissue components is usually poor. The main differences from previously reported methods using potassium permanganate are the use of a physiological saline as the vehicle for all solutions, and, following this, very rapid dehydration before infiltration with plastic. Cellular and intercellular details of tissue ultrastructure may, in general, be evaluated as satisfactorily as with conventional fixatives, with the exception of certain protein elements associated with ribosome, microtubule, and myofilament organization. Nerve endings with agranular or clear vesicles may be distinguished from adrenergic endings since the dense cores of the vesicles of the latter are preserved by this method.

The rat-tail artery maintained in culture: an experimental model.

The rat-tail artery was maintained in vitro for 2 weeks to investigate its suitability as an experimental model. The criteria were that (a) it should retain the overall histological organization with normal ultrastructural appearance of the smooth-muscle cells; (b) stored neurotransmitter which could be activated by experimental treatment should be absent; and (c) smooth-muscle ion transport mechanisms should fall within normal range. Vessels were maintained in Falcon tissue-culture dishes in Dulbecco's modified Eagle's medium. Either 2% or no serum supplement was found to be more suitable than 10% serum due to the high rate of cell proliferation induced by the latter. Light and electron microscopy of cross sections of the vessels indicated that the overall normal vessel architecture was retained, and the ultrastructural features predicted normal function. There were no discernible differences dependent on the length (up to 8- to 10-cm lengths) of the cultured vessel. Preliminary experiments with fluorescent microscopy showed that stored neurotransmitter in the nerves of the vessel wall was no longer present after 48 hr. Ultrastructural examination revealed that storage vesicles in vitro lost their dense cores, representing noradrenalin, between 41 and 48 hr in culture. Normal ion transport mechanisms were retained in the smooth-muscle cells of the arteries in vitro for up to 2 weeks when tested with ion-specific electrodes. Morphological and physiological evidence support the suitability of the rat-tail artery as a model for experimental testing of vascular tissues.

Some effects of the ionophore X-537A on the isolated rat tail artery.

The effects of micromolar concentrations of the ionophore X-537A (RO 2-2985) were studied using isolated preparations of the rat tail artery. The ionophore causes complete release of catecholamines from adrenergic nerves, which is the sole cause of the transient contractile response. The amines are released by a nonexocytotic process which seems to be related to the ability of X-537A to act as an efficient transmembrane carrier of Na+, k+, and H+. The ionophore also causes an almost complete and irreversible loss of the cocaine-sensitive component of metaraminol uptake by the tissue. X-537A dissipates the transmembrane concentration gradients of Na and K in the smooth muscle component of the preparation. This effect is unrelated to the release of endogenous catecholamines, and it can also be observed after the Na pump has been inhibited with ouabain. It is fully reversible, though not readily, and it can be induced repeatedly. In catecholamine-depleted strips, X-537A dissipates the transmembrane Na+ and K+ gradients without causing any change in tension. Stimulation of the rate of O2 consumption by X-537A in catecholamine-depleted tissue is reversible, and it is unaffected by ouabain and (or) removal of external Ca2+.

A teaching model to illustrate the variation in size and shape of the maxillary sinus.

A technique to produce teaching models of the maxillary sinus is described. Dental impression material is injected into the maxillary sinus of a cadaver and subsequently dissected out. Using an impression and mould technique in a series of steps, a stone model of the maxillary sinus is produced. The shape, size and configuration of the model can be seen and the dimensions measured. A radiopaque impression material can be used so that the radiographic outline of the maxillary sinus on standard radiographic projections can be shown. Models of both dentate and edentulous patients can be used to show anatomical variation and the relationship to teeth and are useful in teaching both gross and radiographic anatomy. Since 10 out of 11 specimens had 3 walls leading to the apex from a 4-sided base, it would appear that the classical description of a 4-sided pyramidal shape to the maxillary sinus is relatively uncommon.

The mandibular lingual foramen: a consistent arterial foramen in the middle of the mandible.

The lingual foramen in the midline of the mandible causes confusion in terminology, incidence of occurrence and contents. A survey of 314 dried mandibles showed the foramen to be present in 311 specimens (99.04%). Sectioning of cadaver specimens illustrated a canal traversing the bone to approximately 50% of the buccolingual dimension of the mandible. The contents of the foramen were found to be an artery, which was an anastomosis of the sublingual branches of the right and left lingual arteries. Wire markers were placed in the foramen and the genial tubercles were covered with lead foil to illustrate the radiographic relationship between them. The radio-opacity peripheral to the foramen as seen on a radiograph is produced by the wall of the canal and not the genial tubercles as previously reported. While the foramen is not seen on many radiographs of the lower incisor region, this can be accounted for by a change in orientation of the x-ray beam. A pilot study revealed an incidence of 49% of the lingual foramen on periapical radiographs of the mandibular incisor region in an adult population, the previous reported incidence being 28%.

Relationship between atrial granularity and release of atrial natriuretic factor in rats with diabetes mellitus.

We examined the effect of a 25% blood volume expansion on the release of atrial natriuretic factor (ANF) in conscious Wistar rats (13-15 wk old) injected 6 wk earlier with streptozotocin (55 mg/kg iv, diabetic) or saline (1 ml/kg iv, control). The diabetic rats demonstrated a significant (P less than 0.05) resting hypotension (132 +/- 2/91 +/- 1 mmHg, systolic/diastolic) and bradycardia (340 +/- 5 beats/min) compared with the controls (145 +/- 2/98 +/- 2 mmHg, 377 +/- 8 beats/min). Resting plasma immunoreactive (IR) ANF levels were significantly (P less than 0.05) elevated in the diabetic rats (control: 72 +/- 4 pg/ml; diabetic: 87 +/- 4), although resting right atrial pressures were not different (control: 6.0 +/- 0.8 cmH2O; diabetic: 5.2 +/- 0.6). Volume expansion with donor blood from similarly treated animals significantly (P less than 0.05) elevated IR ANF levels in both groups, but the increase in the saline-injected group (+527 +/- 80 pg/ml) was significantly (P less than 0.05) greater than that of the streptozotocin-injected group (+323 +/- 45 pg/ml). Both groups showed similar elevations in right atrial pressure (control: +1.8 +/- 0.3 cmH2O; diabetic: +1.6 +/- 0.4). Morphological examination of tissue taken from right atria demonstrated no difference in cardiocyte volume percent per unit of tissue but a significant (P less than 0.05) reduction in the relative frequency of occurrence of atrial granules in the diabetic group. The cause of the reduction in atrial granularity in these animals is as yet unknown.

Determining light dose for photodynamic therapy of atherosclerotic lesions in the Yucatan miniswine.

PURPOSE: To determine the light dose required for photodynamic therapy of atherosclerotic lesions in the miniswine. METHODS: Aortic atherosclerosis was created in seven Yucatan miniswine by a combination of balloon endothelial injury and 2% cholesterol and 15% lard for 7 weeks. Six animals received the photosensitizer Photofrin 2.5 mg/kg, while an additional swine received no drug. After 24 hours, the abdominal aorta was exposed and the aorta opened longitudinally in each animal. Three 1-cm spots were illuminated with energy densities of 60, 120, and 240 J/cm2 from an argon-pumped dye laser tuned to 630 nm with a laser output of 1 W. Four weeks later, the animals were killed, abdominal aortae removed, and intimal thickness determined by morphometry. RESULTS: The percentage intimal thickness (mean +/- SD) was 36.7 +/- 27.1, 9.1 +/- 5.0, and 6.4 +/- 8.1 for the three energy densities, respectively. Although both 120 and 240 J/cm2 energy densities produced significant (p < 0.05) reduction in atheroma, considerable damage to the underlying media was also observed in the 240 J/cm2 group. CONCLUSIONS: A Photofrin dose of 2.5 mg/kg and 120 J/cm2 light are necessary for adequate ablation of atheroma while avoiding extensive medial damage.

Dosage and timing of Photofrin for photodynamic therapy of intimal hyperplasia.

Photodynamic therapy has been recommended as a method of preventing intimal hyperplasia. The purpose of this study was to determine the dose and timing of Photofrin porfimer sodium needed to achieve a 3:1 or higher ratio between injured and control arteries after balloon endothelial injury. New Zealand White rabbits were anesthetized and their right femoral artery surgically exposed. A 4Fr Fogarty balloon catheter was passed retrograde into the lower abdominal aorta, inflated and pulled distally into the external iliac artery six times. All rabbits received heparin 100 IU/kg. Arteriotomies were closed and the animals recovered. Rabbits (n = 5 per group) were given intravenous Photofrin at a dose and time according to the following scheme: group I, 5.0 mg/kg immediately after balloon injury; group II, 2.5 mg/kg immediately after injury; group III, 5.0 mg/kg after 1 week; group IV, 5.0 mg/kg after 2 weeks; or group V, 2.5 mg/kg after 2 weeks. Animals were killed 24h after drug administration and the aortoiliac segments removed for spectrophotofluorometric determination of Photofrin levels from injured and control segments. Mean(s.d.) ratios of injured: control arteries for groups I to V were 4.8 (2.6), 2.8 (1.2), 3.0 (1.0), 1.4 (0.3) and 1.0 (0.0) respectively. This ratio was significantly higher for group I rabbits compared with groups IV and V (P < 0.01, ANOVA). Fluorescence and light microscopy showed that Photofrin was localized primarily in the tunica media, and that the drug must be administered before significant intimal hyperplasia occurs.(ABSTRACT TRUNCATED AT 250 WORDS)