Tsg101 is upregulated in a subset of invasive human breast cancers and its targeted overexpression in transgenic mice reveals weak oncogenic properties for mammary cancer initiation.

Previous studies reported that the Tumor Susceptibility Gene 101 (TSG101) is upregulated in selected human malignancies, and the expression of exogenous Tsg101 was suggested to transform immortalized fibroblasts in culture. To date, the potential oncogenic properties of Tsg101 have not been examined in vivo owing to the lack of appropriate model systems. In this study, we show that Tsg101 is highly expressed in a subset of invasive human breast cancers. Based on this observation, we generated the first transgenic mouse model with a targeted overexpression of Tsg101 in the developing mammary gland to test whether exogenous Tsg101 is capable of initiating tumorigenesis. Normal functionality of exogenous Tsg101 was tested by rescuing the survival of Tsg101-deficient mammary epithelial cells in conditional knockout mice. The overexpression of Tsg101 resulted in increased phosphorylation of the epidermal growth factor receptor and downstream activation of MAP kinases. Despite an increase in the activation of these signal transducers, the mammary gland of females expressing exogenous Tsg101 developed normally throughout the reproductive cycle. In aging females, the overexpression of Tsg101 seemed to increase the susceptibility of mammary epithelia toward malignant transformation. However, owing to the long latency of tumor formation and the sporadic occurrence of bona fide mammary cancers, we conclude that the Tsg101 protein has only weak oncogenic properties. Instead of cancer initiation, it is therefore likely that Tsg101 plays a more predominant role in the progression of a subset of spontaneously arising breast cancers.

The relationship of the subarachnoid injection of blood and blood fractions with cardiac rate change and arrhythmias.

The relationship of subarachnoid hemorrhage and cardiac arrhythmias was studied utilizing a Sprague-Dawley rat model. A total of 30 male animals were divided into five groups and given subarachnoid injections of either blood, blood fractions, or control substances. Blood pressure, intracranial pressure, serum electrolytes, arterial blood gases, hypothalamic multiple unit activity and an electrocardiogram were concurrently monitored. Cardiac arrhythmias were graded on a 0 to 4 + objective scale. Control parameter values were similar for all animals. Arrhythmias, hypotension, and decreased hypothalamic multiple unit activity were seen with infusion of whole blood and packed red blood cells. Packed red blood cells were statistically demonstrated to have the most potent arrhythmogenic effect. Cardiac histopathology revealed myocardial contraction band lesions most predominant in the packed red blood cell group. In addition, significant QT interval prolongation was observed after subarachnoid injection of either whole blood or packed red blood cells. These findings indicate that packed red blood cells, or a component thereof, may play an important role in the etiology of immediate (i.e. acute) post subarachnoid hemorrhage induced cardiac arrhythmias.

Acute cardiotoxic effects of cocaine and a hyperadrenergic state in anesthetized dogs.

The purpose of this study was to determine whether a subthreshold dose of cocaine (one that is well-tolerated in a resting state with no significant hemodynamic changes) causes cardiotoxic responses when superimposed upon a subthreshold infusion of norepinephrine. Adult mongrel dogs were anesthetized, instrumented and assigned to one of five treatment groups: saline control, norepinephrine infusion (0.75 microgram/kg/min, i.v. for 60 min); cocaine (5.0 mg/kg, i.v. bolus); combination of cocaine (0.5 mg/kg) or (5.0 mg/kg) with the last 45 min of norepinephrine infusion. The combination groups exhibited increased blood pressure and contractility (dP/dt), and a greater variety and frequency of arrhythmias. Cocaine did not increase plasma catecholamine levels beyond norepinephrine alone. There were significantly more contraction band lesions/mm2 with cocaine in combination with norepinephrine than in any of the individual treatment groups. These data indicate that, even in anesthetized animals, a well-tolerated dose of cocaine may become toxic when superimposed on an excited or hyperadrenergic state with elevated levels of norepinephrine.

Measurement of blood flow to osseous tissue in dogs using the radiolabelled microsphere method.

1. The purpose of this study was to examine bone blood flow in various intra- and extra-oral sites. 2. The radiolabelled microsphere method was utilized to assess osseous blood flow in the following regions of 10 dogs: rib, long bone, and anterior and posterior regions of the maxilla and mandible. 3. Samples of cancellous and cortical bone were also obtained from each of these regions with the exception of the maxilla and the anterior mandible. 4. Mean blood flow ranged from 3.71 +/- 0.81 (SE) ml min.-1 100 g-1 in the mandibular posterior cortical bone to 22.7 +/- 4.66 ml min-1 100 g-1 in the cancellous rib samples. 5. Blood flow to the cancellous tissue of the rib was significantly greater (P < 0.05) than the other tissues with the exception of maxillary posterior bone and cortical rib. 6. Results from this study indicate that blood flow to the maxillary posterior bone is relatively high, but blood flow in other intraoral osseous sites is significantly less than that of cancellous rib bone.

Effects of systemic and topical nicotine on pulpal blood flow in dogs.

It has been suggested that nicotine exerts cardiovascular effects which are similar to stimulation of the sympathoadrenal system. If this observation is true, nicotine administration would decrease pulpal blood flow and possible alter the pulpal response to injury. The purpose of this study was to measure pulpal blood flow in dogs using the radiolabelled microsphere method following topical or systemic administration of nicotine for 28 days. Thirteen dogs were divided into three groups. Group one received topical nicotine (8 mg nicotine/kg/day) combined with orabase which was applied in two equal doses to the mandibular anterior gingiva. Group two received systemic nicotine (2.5 mg/kg/day) delivered by osmotic pumps implanted subcutaneously in the back of each animals' neck. Group three were controls, and these animals received either topical orabase twice daily applied to the mandibular anterior gingiva or saline via osmotic pumps. Results indicated pulpal blood flow increased from Day 0 to Day 28 in both nicotine treated groups. Group one (topical nicotine) exhibited a mean increase in blood flow of 21.8 ml/min/100 g, while group two exhibited a mean increase of 50.1 ml/min/100 g. Group three, the control animals, exhibited a mean decrease in pulpal blood flow of 22.1 ml/min/100 g over the 28-day interval. These changes were not statistically significant (p > 0.05).

Application of the MRFIT smoking cessation program to a healthy, mixed-sex sample.

The Multiple Risk Factor Intervention Trial (MRFIT) included a smoking cessation program that was highly successful (40.3% abstinence prevalence rate at 48-month follow-up) when used with other interventions for a male, middle-aged population at high risk for coronary heart disease (CHD). Our study employed the MRFIT cessation program alone with a mixed-sex, mixed-age, healthy population. We wished to determine its effectiveness when applied in a manner similar to other smoking cessation programs. Fifty-six subjects participated in a 10-week intervention followed by maintenance or extended intervention programs. The 52% abstinence prevalence rate at the end of the 10-week intervention dropped to 32% after four months, 25% at eight months, 25% at 12 months, and 27% at 16 months. The higher cessation rates of the original MRFIT study may be related to motivation and other characteristics of the high-risk population and to the combination of the smoking component with other interventions for CHD, rather than to the characteristics of the smoking intervention itself. Although the MRFIT program is comprehensive and includes vigorous maintenance activities, it is also expensive and may not be cost-effective or as desirable as programs with slightly lower cessation rates.

Effects of topical and systemic nicotine on gingival blood flow in dogs.

It has been suggested that due to its vasoconstrictive action, nicotine may have a deleterious effect on the periodontium. This study examined the effects of topical and systemic nicotine administration on gingival blood flow. Eighteen young adult dogs were divided into three groups receiving the following treatments for 28 days; topical nicotine in orabase, systemic nicotine via osmotic mini-pumps, and topical orabase or systemic saline via osmotic mini-pumps. Blood flow to the gingiva was measured (at days 0 and 28) by the radiolabeled microsphere method. Blood flow was consistently increased from day 0 to day 28 in the nicotine-treated animals. Comparison of days 0 and 28 blood-flow values demonstrated a statistically significant change (p less than 0.05) in the anterior regions of the topical-nicotine group as compared with the control group. The increased flow may be a reflection of the mode of nicotine delivery and timing of the blood-flow determination procedures.

Effects of chronic non-ketotic diabetes and aging on myocardial function and fatty acid oxidation.

The effects of aging and chronic non-ketotic diabetes on contractile properties, oxygen consumption, palmitate oxidation and morphology were studied in isolated, perfused working hearts of 2, 9, 12 and 22 month old rats. The heart rate, coronary flow, and oxygen consumption were no different among the 9, 12 and 22 month control and diabetic hearts. Cardiac work was not depressed in control hearts until 22 months of age. Depression of cardiac output due to aging in the control hearts progressed in stages. The superimposition of chronic diabetes in the 9, 12 and 22 month rats did not further depress the cardiac work or cardiac output. [1-14C] palmitate oxidation in the 2 and 9 month control hearts was higher than that of the 12 and 22 month controls. Chronic diabetes did not affect fatty acid oxidation in the 9 and 12 month rats compared to their controls, but was diminished in the 22 month diabetic rat heart. These results suggest that impairments in the contractile properties of the isolated hearts of the chronically diabetic, senescent rats were primarily due to aging.

Cardioprotective effects of diltiazem when given before, during or delayed after infusion of norepinephrine in anesthetized dogs.

Catecholamine excess has been shown to produce 2 distinct forms of irreversible myocardial necrosis termed contraction band lesions. Calcium channel blocking agents provided a variable protective effect from these contraction band lesions. The purpose of this study was to determine the temporal responses of the most effective of these blocking agents, diltiazem, when given before, simultaneous with or after an initial exposure to a necrogenic infusion of norepinephrine (NE). Forty-one adult mongrel dogs were anesthetized with sodium pentobarbital (32 mg/kg) and infused with saline solution or NE (4 micrograms/kg/min) for 60 minutes or diltiazem at a rate of 20 micrograms/kg/min for the first 5 minutes and 10 micrograms/kg/min for the remaining 70 minutes. Diltiazem was infused as pretreatment 15 minutes before continued infusion with NE for 60 minutes, simultaneously infused with NE for 60 minutes or delayed 30 minutes after the start of NE infusion. Diltiazem alone exhibited no significant effect on hemodynamics, but pretreatment with diltiazem was able to moderate the rapid NE-induced increases in heart rate. NE infusion produced significant numbers of the 2 forms of contraction band lesions: (1) paradiscal contraction band lesions involving a small portion of the cell adjacent to the disc, and (2) holocytic contraction band lesions involving the entire cell. Diltiazem reduced the number of contraction band lesions, particularly the holocytic contraction band lesions, provided diltiazem was available before the insult and massive influx of calcium with a pharmacologic dose of NE. Although the exact mechanism of diltiazem's cardioprotective properties is not known, the timing of drug administration does appear to affect the degree of protection.

Blood flow and epithelial thickness in different regions of feline oral mucosa and skin.

The relationship between epithelial thickness and blood flow was examined in 6 mucosal and 3 skin regions of the cat. Blood flow to these tissues was determined using the radiolabelled microsphere method. From histologic sections the proportion of the tissue biopsy occupied by epithelium and the average epithelial thickness were calculated. The oral tissues had a significantly higher blood flow than the skin regions (p less than 0.05). In terms of epithelial thickness, the tissues could be divided into 4 groups (p less than 0.05). These were: a) palate; b) gingival regions and dorsum of the tongue; c) lip and buccal mucosa; d) all skin regions. When epithelial thickness was related to blood flow there was a significant positive correlation (p less than 0.005) indicating that a thicker epithelium is associated with a higher blood flow. This finding may reflect the greater metabolic demands of the thicker epithelia.

Determination of plasma catecholamines in dogs after experimental gingival retraction with epinephrine-impregnated cord.

Blood catecholamine levels were determined by high-performance liquid chromatography after placement of the cord around two teeth in a random sequence of the following treatments: (1) untreated cord to intact gingival crevice (C), (2) epinephrine cord to intact crevice (E1), and (3) epinephrine cord to a crevice disrupted with a tapered diamond bur to simulate a subgingival procedure such as in crown preparation (E2). Each cord was left in place for 30 min while blood was drawn from the external jugular vein at 0, 5, 10, 20 and 30 min, and at 10 and 30 min after removal of the cord. Catecholamine values were compared to baseline (time 0) measurements. Treatment C resulted in no significant change in blood catecholamines. Treatment E1 caused a significant increase (p less than 0.01) in blood epinephrine levels, reaching a 641 per cent increase at 30 min. Treatment E2 produced a highly significant increase (p less than 0.001) in blood epinephrine with a greater than 5000 per cent increase 10 min after cord removal. Norepinephrine and dopamine levels were not significantly altered by any of the treatments.

Protective effects of slow channel calcium antagonists on noradrenaline induced myocardial necrosis.

Catecholamine excess results in two distinct forms of coagulative myocytolysis, apparently due to increased membrane permeability followed by a large influx of calcium. To determine if three slow channel calcium antagonists, verapamil, nifedipine, and diltiazem, could reduce the calcium overload and prevent the development of noradrenaline induced acute myocardial contraction band lesions, 48 adult mongrel dogs in eight groups (n = 6) were continuously infused with saline alone, noradrenaline alone (4 micrograms X kg-1 X min-1), nifedipine (1 microgram X kg-1 X min-1), or other calcium blockers (10 micrograms X kg-1 X min-1) with saline or noradrenaline. After 15 minutes of pretreatment with a calcium antagonist, the antagonists were simultaneously infused with either saline or noradrenaline for 60 minutes. Nifedipine increased heart rate to the same degree as noradrenaline alone, whereas verapamil and diltiazem significantly suppressed the noradrenaline induced increases in heart rate. All three calcium antagonists reduced the increases in blood pressure and frequency of ventricular arrhythmias seen with noradrenaline alone. Only nifedipine produced a moderate increase in contractility (dP/dtmax) within 5 min and a pronounced synergistic increase when combined with noradrenaline. The effect of the other antagonists with noradrenaline was no different than the effect with noradrenaline alone. Contraction band lesions in the hearts of dogs in the saline and saline plus calcium antagonist groups were rare. The group receiving noradrenaline alone showed large numbers of the two predominant lesions: small paradiscal contraction band lesions and large holocytic contraction band lesions.(ABSTRACT TRUNCATED AT 250 WORDS)

Experimental catecholamine-induced myocardial necrosis. II. Temporal development of isoproterenol-induced contraction band lesions correlated with ECG, hemodynamic and biochemical changes.

Catecholamines have been shown to produce irreversible contraction band lesions of myocardial cells. However, little is known about the temporal appearance and correlation of the acute form of coagulative myocytolysis with ECG, hemodynamic and biochemical parameters. Groups of adult mongrel dogs were anesthetized with sodium pentobarbital, infused continuously with isoproterenol (2.5 micrograms/kg/min) and killed after periods of 0, 5, 15, 30, or 60 min. There were two predominant myocardial patterns: 'paradiscal' and 'holocytic' contraction band lesions. Either type of lesion was non-existent or rare in the control hearts. The small 'paradiscal' contraction band lesions were present as early as 5 min of isoproterenol infusion, particularly in the inner myocardial layer. The large 'holocytic' contraction band lesions were present by 15 min, however, they were not produced in any significant numbers before 30 min. Both types of contraction band lesions continued to accumulate up to 60 min. ST segment depression was the predominant ECG change. This occurred as early as 5 min when heart rate, blood pressure and dP/dt values had also significantly changed. The high-energy phosphates, phosphocreatine and ATP, started declining as early as 5 min. Furthermore, these phosphates and lactate were distributed in transmural gradients across the left ventricular wall with the greatest change in the endocardial third. This was also the site of the largest accumulation of each type of contraction band lesion. While the lesions correlated with certain biochemical and hemodynamic changes, the underlying pathophysiology is more complex than ischemia or high-energy phosphate depletion alone.

Experimental catecholamine-induced myocardial necrosis. I. Morphology, quantification and regional distribution of acute contraction band lesions.

Acute histologic and ultrastructural changes, quantification and regional distribution (transmural, circumferential, transaxial) of damaged myocells from anesthetized, open-chested dogs following one hour of intravenous infusions of saline or increasing doses of isoproterenol (0.1, 1.0, 2.5 micrograms/kg/min) or norepinephrine (4.0 micrograms/kg/min) were investigated. Two predominant subsets of acute contraction band lesions were produced: 'paradiscal' involving aggregation of less than 15 sarcomeres adjacent to the intercalated disc and 'holocytic' involving coagulation of groups of adjoining sarcomeres into transverse bands interspersed with areas of myofibrillar rhexis throughout the cell. Both lesions were distributed as isolated cells or as small foci of myocells surrounded by normal myocardium. Quantification of 'paradiscal' and 'holocytic' contraction band lesions/mm2 of area was used as an index of the severity of catecholamine-induced necrosis. Numbers of 'paradiscal' myocells increased with increasing doses of isoproterenol, while 'holocytic' myocells were not present in any significant numbers until 1.0 microgram/kg/min) and increased further at 2.5 micrograms/kg/min. 'Paradiscal' myocells with both isoproterenol (2.5 micrograms/kg/min) and norepinephrine were distributed with the greatest number in the inner third of the free wall. This gradient was not significant for 'holocytic' lesions. There was, generally, no significant difference in distribution of either type of lesion around the circumference. However, transaxially there was a higher frequency of 'paradiscal' lesions at the apex with norepinephrine. The lesions were identical and present in comparable numbers at both the highest dose of isoproterenol and with norepinephrine. Thus, these two catecholamines result in a similar cardiotoxicity, each with two predominant subsets of lesions, despite their hemodynamic diversities.

Physiologic changes during a marathon, with special reference to magnesium.

In a single case study of a moderately trained, healthy man, physiologic changes during a marathon are reported. Blood was drawn prior to the race, at 1 hour and 2 hours into the race, at the end of the race, and after 1 hour of recovery. By 1 hour into the race, norepinephrine, epinephrine, and dopamine had increased nearly nine-fold, two-fold and five-fold, respectively. After 1 hour of recovery, epinephrine had returned to the pre-race value but norepinephrine and dopamine were still elevated. Cortisol increased gradually and was more than doubled by the end of the race. It was still elevated after 1 hour of recovery. White blood cells gradually increased, reaching their maximum value at the end of the race; a four-to-five-fold increase. Thromboxane B2, which had an inverse relationship to serum magnesium, was below the pre-race value for the first 2 hours but increased nine-fold by the end of the race. Serum magnesium increased from 1.44 meq/l to 1.68 meq/l at 2 hours into the marathon, dropped to 1.07 meq/l by the end of the race, and returned to its pre-race value by 1 hour of recovery. The decrease in serum magnesium at the end of the race may be associated with increased plasma free fatty acid levels.

Attenuation of myocardial acidosis by propranolol during ischaemic arrest and reperfusion: evidence with 31P nuclear magnetic resonance.

31P nuclear magnetic resonance (NMR) spectroscopy was used to ascertain whether propranolol could reduce the development of myocardial acidosis during periods of ischaemic arrest and were studied. Cardiac pH progressively declined during ischaemia from a normal 6.97 +/- 0.02 (n = 23) to 6.09 +/- 0.04 or 5.96 +/- 0.04, respectively. Normalisation of pH following reperfusion occurred only in the 35 min ischaemic hearts. Propranolol (1 mg. litre-1) given prior to arrest significantly reduced the magnitude of developing acidosis regardless of the length of ischaemia. Furthermore, it aided in the normalisation of intramyocardial pH upon reperfusion in both groups. Propranolol significantly reduced the magnitude of phosphocreatine (PCr loss normally seen during ischaemic arrest alone, but it did not protect against depletion of ATP. Restoration of PCr reperfusion was virtually complete in all cases, while transient increases in ATP were seen only in those hearts protected by propranolol. In summary, this NMR study demonstrated the first direct evidence that a significant component of the myocardial acidosis caused by global ischaemia and arrest can be blocked by propranolol.