Left ventricular wall resection for aneurysm and akinesia  due to coronary artery disease: fifty consecutive patients.

Left ventricular wall resection with or without vein bypass grafting was performed in 50 consecutive patients with aneurysm or an akinetic segment with poor left ventricular hemodynamics. Ejection fraction averaged 29% and left ventricular end-diastolic pressure averaged 20 mm Hg. Associated valve disease was present in 14 patients (28%). Hospital mortality was 22% (11 patients) and late mortality was 20% (10 patients). The lowest mortality (3%, 1 death) was found in patients who had anterior apical resection with associated vein bypass, in whom prophylactic balloon pumping was used when needed for ejection fractions of less than 30%. A high mortality occurred with associated valve replacement (6 deaths, 86%) and posterior aneurysm. Among 39 patients followed for an average of twenty-four months, 34 (87%) improved one class and 18 (46%) improved two classes of the New York Heart Association Functional Classification.


A better surgical technique for repairing pulmonary arteries after banding.

A technique is presented to eliminate the residual gradient more completely after removing the main pulmonary band at the time of primary repair of ventricular septal defect. The band and underlying pulmonary artery are circumferentially excised, except for a small posterior part, and pulmonary artery is reanastomosed.


Posterior rupture of the interventricular septum after acute myocardial infarction: successful early surgical repair.

A patient with a large posterior ventricular septal defect complicating an acute inferior myocardial infarction is reported. Because of medically intractable biventricular failure, temporary circulatory assistance was initiated using intraaortic balloon pumping. Emergency coronary angiography, ventriculography, and subsequent operation were carried out. Operative repair involved closure of the septal defect with the use of a Dacron patch, infarctectomy, and aortocoronary bypass grafting and resulted in long-term survival of the patient.


Vigorous medical management of acute fulminant hepatitis.

Twenty patients with acute fulminant hepatic failure and stage II, III, or IV hepatic encephalopathy attributable to viral hepatitis were studied to assess the risk factors, as well as the affects of vigorous medical management. These patients were treated according to a protocol that directed aggressive medical management of fluid balance with electrolyte solutions, plasma, and blood; acid-base balance; coagulation defects with fresh frozen plasma; blood replacement as needed; dietary protein elimination; and orally administered neomycin sulfate. Among the 20 patients there were eight survivors (40%). Seven of the 13 patients who were positive for the hepatitis B surface antigen (HB-Ag) survived (54%), while one of the seven patients whoe were negative for HB-Ag survived (14%). The stage of encephalopathy on admission did not correlate with survival. Patients under the age of 40 years had a 43% survival rate, while those over 40 years had a 33% survival rate. Conservative but vigorous medical management may improve survival in fulminant hepatic failure.


Studies with an artificial endocrine pancreas.

Short-term studies with an artificial endocrine pancreas have provided insights into the pathophysiology of diabetes and demonstrated the ability of such a system to restore and maintain glucose homeostasis in human subjects given either their usual diet, a 50 gm oral glucose tolerance test, or a moderate level of physical exercise. Animals were used to demonstrate that a peripheral route of insulin delivery by the artificial pancreas is equivalent to the portal route.


Heart failure. A proposed definition and classification.

Heart failure exists when either the systolic or diastolic operation of the ventricle is impaired to a degree that, despite compensatory mechanisms, the demands of the peripheral organs are not satisfied, the peripheral muscle shortens inadequately, and/or the pulmonary or systemic venous system becomes congested from high filling pressures. Since every pumping system has finite limits and can fail if excessive and prolonged demands are made on it, any definition of failure must take into consideration the degree of stress imposed and whether or not the cardiac response is appropriate or subnormal. When failure is present, it becomes essential to discern whether it involves the intrinsic pumping structures of the heart, the myocardial cells, or, rather, whether failure of one of the other components of the integrated cardiovascular system (for example, valvular dysfunction, ruptured ventricular septum) has occurred.


Lupus nephritis in a neuronal storage disease.

Systemic lupus erythematosus (SLE) with glomerulonephritis occurred in a patient with a long-standing neuronal storage disease and myoclonus. Intravenously administered diazepam was useful in controlling the myoclonus to permit a renal biopsy. The exact relationship between the patient's underlying illness and the development of SLE remains unknown.


Salvage of infected total joint replacements.

Among 424 total joint replacement (362 hip and 62 knee replacements) since 1971, eight deep infections have occurred (two primary and six late) in six patients. Removal of the prosthesis has been necessary in only one patient to date. Our follow-up experience, extending over a maximum period of 20 months, indicates that successful salvage of infected total joint arthroplasties without bone involvement may be accomplished without removal of the prosthesis if there is adequate surgical debridement and proper antibiotic therapy.


Epidermoid splenic cysts.

Five young women were treated by splenectomy for epidermoid cysts of the spleen. They all had vague abdominal symptoms, and had a mass palpable in the left upper quadrant of the abdomen. The hematologic indices were unremarkable. The diagnosis was suggested by routine radiologic examinations (including chest roentgenogram, upper gastrointestinal x-ray series, barium enema, and excretory urogram). It was confirmed in one patient by gray-scale ultrasonography, and in another by selective splenic arteriography. Of the true cysts of the spleen, the epidermoid cyst is the rarest, representing 10% of the benign, nonparaslitic cysts. The symptoms are completely relieved by splenectomy, and this remains the treatment of choice.


Adenocarcinoma in regional enteritis of the small intestine.

An increased risk for regional enteritis patients of small bowel adenocarcinoma to develop has been suspected but unproved. We have analyzed 49 cases reported since 1957 and two additional ones of our own. These have been compared with a current group of small bowel adenocarcinomas not associated with regional enteritis. The Crohn-associated cancers differed from adenocarcinomas not associated with Crohn disease in that (1) mean age at cancer discovery was less (46 vs 64 years), (2) more cancers arose in the ileum (76% vs 27%), (3) diagnosis and cure were less successful, and (4) they occurred more frequently. The 32 cases reported in the past five years were compared with the expected 0.1 to 5 cases. Regional enteritis patients were found to have an increased risk for the development not only of small bowel adenocarcinoma, but one that is more occult and lethal than that in individuals wihtout Crohn disease.


Bilateral catamenial pneumothorax.

Bilateral catamenial pneumothorax occurred in a 32-year-old parous woman. The 41 previously reported cases of catamenial pneumothorax occurred on the right side. This patient was treated with left-sided pleurodesis and, subsequently, with hysterectomy and bilateral salpingo-oophorectomy. The pathologic mechanism of this peculiar syndrome has been attributed to reflux of air via the genitalia and fenestrations in the diaphragm, menstrual shedding of endometrial cells growing in the visceral pleura, or, possibly, aveolar tissue damage secondary to vascular and bronchiolar spasm resulting from dinoprost tromethamine released from menstrual debris. Treatment needs to be tailored to fit the individual patient's specific pelvic and intrathoracic pathology and procreative desires.


Bilateral renal artery stenosis causing acute oliguric renal failure. Report of a case corrected by renovascular surgery.

A 70-year-old woman with chronic hypertension and previously normal renal function had acute oliguric renal failure requiring hemodialysis. Renal arteriograms revealed the presence of bilateral renal artery stenosis and normal-sized kidneys. Nineteen days after admission to hospital, after undergoing nine hemodialysis procedures, surgical revascularization of renal artery stenosis was performed utilizing a single bypass graft of the left renal artery. Postoperatively, an immediate diuresis ensued, with resolution of acute renal failure. It is critically important in the evaluation of patients with anuria, acute renal failure without obvious cause, or impending uremia in patients with chronic stable renal insufficiency, to consider the possibility of renal artery stenosis or thrombosis. Recognition and then surgical correction of significant renal arterial hypoperfusion allows the reasonable potential for reversibility of this important form of acute or progressive renal failure.


Paradoxical embolization. Case report and review of the literature.

A 32-year-old morbidly obese woman had bilateral subclavian artery embolism. After embolectomy, a right heart catheterization showed a patent foramen ovale and a right-to-left shunt. Pulmonary angiography revealed multiple pulmonary emboli. Bilateral lower extremity venography demonstrated left ileofemoral thrombi as the probable source. The patient was successfully treated with an inferior vena caval umbrella and anticoagulant drugs.


Malabsorption and abdominal pain secondary to celiac artery entrapment.

A patient with abdominal pain was found to have severe compression of his celiac artery on abdominal angiography. Preoperative evaluation with base line and provocative xylose absorption studies were compatible with decreased intestinal blood flow. Surgical division of the median arcuate ligament corrected the intraoperatively determined pressure gradient. Postoperative studies at three months demonstrate absence of celiac artery compression on angiography and normal provocative xylose absorption studies. This case lends support to the existence of the median arcuate ligament syndrome.


Primary adenocarcinoma of the appendix masquerading as a bladder tumor.

Appendiceal adenocarcinoma, a rare lesion, is difficult to diagnose preoperatively. The unusual combination of genitourinary symptoms, a fungating adenocarcinoma of the deeper layers of the bladder wall, and an abnormal barium enema resulted in what seems to be the first reported case to be diagnosed preoperatively. Preoperative radiotherapy and radical exenteration of the tumor with ileotransverse colectomy and ureteroileal cutaneous diversion were performed. The patient is well without evidence of recurrence two years later.


Uteroenteric fistula due to advanced extrauterine pregnancy.

A case of an uteroenteric fistula is reported, representing an unusual sequel of a long-standing ectopic pregnancy. A correct diagnosis could have been made preoperatively by visualization of the fragments of the fetal skeleton on abdominal roentgenogram in the area of the fistula.


Type A influenza: postmortem virus isolations from different organs in human lethal cases.

Trachea, lung, liver, spleen, pancreas and brain of 77 heman patients who had died in the course of clinically diagnosed influenza were subjected to virological and histopathological examination. Type A influenza viruses closely related to the virus variants contemporarily in circulation were isolated from 12 of the lethal cases. In 10 of them, virus was demonstrated in organs other than respiratory, most often the brain. Influenza antigen was also demonstrated in brain tissue by immunofluorescence.


[Tissue organization of mesothelium under normal conditions and during reparative regeneration].

The investigation of tissue organization of the mesothelium by non-parametric statistic methods has shown that under conditions of reparative regeneration the principle of organization of the mesothelium as a tissue is preserved. Heteromorphism characteristic of the normal mesothelium is increased at all steps of regeneration, and the degree of the layer organization is decreased. The mitotic activity leading to the reparation of defect develops undulatorily, the periodicity of time does not coincide with the periodicity in space.


[Types of organization of interneuronal contacts in the ventral horn of the cat spinal cord].

The ultrastructure of intermediate neurons of the ventral horn of the spinal cord of cats was studied. It has been established that interneuronal contacts having different structural organization may be grouped according to the principle of different space distribution of pre- and postsynaptic elements. Contacts of the first type realize convergence of different synaptic effects on one post-synaptic structure. In contacts of the second type the synaptic action of the axon fibre or a terminal diverge to different elements of the neuropile. Several synaptic forms of contacts of the second type were revealed. The role of the above structures in the physiological activity of intermediate neurons of the spinal cord is discussed.


[Histochemical changes in regenerating adrenal autotransplants].

The aim of the work was to study certain histochemical changes in the process of regeneration of adrenal autotransplants. The autotransplants regenerated at the expense of the external area of the glomerular zone (the subcapsular zone). Though certain changes in the capsule were observed, in no case the adrenals capsule elements were turned into cortical cells. On the 28th day after the operation the cortical substance regenerated almost completely. The medullary substance failed to restore.


[Ultrastructural organization of chromaffin paraganglia in the ganglia of the solar plexus].

Under study was the ultrastructural organization of chromaffin cells in the nodes of the solar plexus of albino rats. The chromaffin cells form accumulations--paraganglia, bent for capillaries. The chromaffin cells are of small size and have a dense glial capsule. The cytoplasms are rich in mitochondria, Golgi Complexes and vesicles containing catecholamines. The state of cytoplasmic components speaks of asynchronous secretory activity of chromaffin cells. Among catecholamine-containing vesicles three main types can be distinguished, different is size, structure of the content and localization in the perikaryon. Excretion of catecholamines in realized through cytoplasmic processes which, getting free of the glial capsule, come out directly to capillaries. The coordinating influence of the sympathetic nervous system on the endocrinous function of chromaffin paraganglia of the solar plexus is exercised by paraganglionic fibres, whose terminal portions form synaptic contacts on the chromaffin cells.


[Dynamics of morphofunctional changes in the fibroblasts of a focus of inflammation].

Morphometrical, histochemical and electron-microscopic methods were used in order to show that the inflammatory reactions in the traumatized connective tissue were accompanied by massive death of fibroblasts. In surviving cells the protein synthesis increased and the mitotic activity rose. The synthetic and secretory organs in fibroblasts became mostly developed by the 2--5th day. The increased synthesis of proteins was associated with the formation of the connective tissue capsule around the foreign body. As the capsule matured, the synthetic activity and the mitotic activity of fibroblasts decreased. In some cells the destruction phenomena were observed.


[Ocular scale for logarithmic grouping of classes of spherical nuclei and measurement of the diameters of their nucleoli].

The proposed scale is prepared by a photographic method and consists of some vertical lines, each of them being disposed at the distance equal to a nucleus diameter having bordering size between classes of lg mkm2, and of three horizontal lines, forming two patterns designated for measuring diameters of nucleoli. The scale is meant for working with the objective 90 X and the ocular 20 X and makes the measuring of nuclei and nucleoli diameters 2--2,5 times quicker as compared with the screw ocular micrometer with a sufficient coincidence of results, obtained by both methods.


[Morphologic changes in the liver and kidney in ethylene glycol poisoning].

The liver and kidneys of 48 persons who had died of poisoning with ethyleneglycol were investigated at periods from 4 hours to 32 days following the poisoning. In 44 deceased who had died 2--32 days after the poising of acute renal and hepato-renal insufficiency there were revealed lesions characteristic of this intoxication. The structural basis of glycolic nephrosis and hepatosis was hydropic dystrophy of the nephrothelium of proximal and distal tubules and of hepatocytes of the centers of the liver lobules with outcome into colliquative necrosis through ballon dystrophy. Specific lesions of thekidney and liver may be accounted for by a high osmotic activity of ethyleneglycol and products of its metabolism; these were captured by the renal and hepatic epithelium in the process of reabsorption and pinocytosis increasing drastically the osmotic concentration of the intracellular liquid, which was accompanied by rapid inflow of water into the cell and its hydropic dystrophy.


[Pathologo-anatomic and experimental study of the morphology of Q-fever].

The article presents a survey of the literature containing reports on lesions of some human organs in Q-fever and on a possible role of the pathogene--Rickettsia burnetii in the development of these lesions. Pathoanatomical findings were compared with those of the experimental investigations of the Q-rickettsial infectious process. The literature data and the author's own investigations confirm the capability of Rickettsia burnetii of parasitizing in cells of the reticulo-endothelial system. This is an important factor in the pathogenicity of this causative agent and lies at the root of the pathogenesis of Q-rickettsial infection.


Supratentorial arachnoid cysts in adults. A discussion of two cases from a pathophysiologic and surgical perspective.

Large congenital arachnoid cysts are a rare cause of increased intracranial pressure and progressive neurologic deficits in adult life. We describe two patients, diagnosed by computerized axial tomography and surgically treated. The pertinent medical literature was reviewed. The outer cyst membranes were excised, and a communication between cyst and subarachnoid space was established. When possible, the membranes and fluid content of these lesions should be studied. This fluid is best obtained by aspiration prior to dural incision. At present, three etiologic mechanisms of cyst enlargement appear tenable: (1) secretion of fluid by ependymal cells, (2) fluid ingress due to an osmotic gradient, and finally, (3) trapping of fluid by a ball-valve mechanism. Regardless of the reasons why these lesions enlarge, drainage of cyst content into the venous system warrants a clinical consideration.


The relationship of arterial hypertension to intracranial aneurysms.

The rather widely held belief that systemic arterial hypertension is a major factor either in the genesis of saccular aneurysms or in their subsequent rupture is questioned on the basis of our clinical and pathological analysis of 250 patients with aneurysms. Analysis of 150 patients with ruptured and 100 patients with unruptured saccular aneurysms for clinical and morphologic evidence of hypertension revealed no notable excess over an age- and sex-matched control autopsy population. There is no evident association of hypertension with multiplicity of aneurysms, the age at which aneurysms present clinically, or with their rupture. All available data clearly indicate that saccular aneurysms can arise in the absence of fixed arterial hypertension and that they can also rupture in the absence of fixed hypertension.


Pheniramine aminosalicylate overdosage. Reversal of delirium and choreiform movements with tacrine treatment.

We describe a case of central anticholinergic syndrome with choreiform movements after an overdose of pheniramine aminosalicylate. The symptoms and signs of the condition were reversed by the intravenous administration of the cholinesterase-inhibitor tetrahydroaminacrine. We also discuss the concept of adrenergic-cholinergic imbalance in relation to this case and recommend the use of centrally acting anticholinesterase agents in the treatment of specific cases of drug overdose.


Pattern visual evoked responses in multiple sclerosis.

The latency of the first major positive component of the visual evoked response to pattern reversal was measured from a midoccipital electrode for each eye in 30 patients with a definite diagnosis of multiple sclerosis (MS). Their latencies were compared with those of a group of 18 normal subjects and 18 patients with other neurological diseases. Of 26 MS patients with a measurable response, 77% gave values that were outside the ranges of the two control groups. A total of 75% of the MS patients without history or findings of optic neuritis had an abnormal response. The procedure appears to be useful in detecting asymptomatic lesions of the visual pathways in suspected MS patients.


Nasal projection.

The challenge of enduring tip projection is met with the use of a septal cartilage graft between the medial crura. With suturing, a "sandwich" is created composed of the two medial crura and the graft between them. The sandwich can be trimmed as necessary, in accordance with the surgeon's taste requirements for good tip profile and projection. The graft allows much more projection than from the medial crura alone. We describe the Goldman technique of rim incisions, delivery of the lower lateral cartilages, bisection of each, and then creation of the sandwich. The technique has been performed on more than 40 patients, in most of whom the results have remained successful more than six months postoperatively. The results indicate excellent fixation of the projected tip with maintenance of position, strength, and form. The indications for this tip technique are the classical ones for medial crura projection, such as hypoplastic tip cartilages, short columella revision rhinoplasty, and most importantly, the increase of real projection not attained by simple rotation.


Comparative histological study of mini pig skin after chemical peel and dermabrasion.

Although individual studies of the histological changes after chemical peel and dermabrasion have been reported, there has not been a comparative study of these modalities in the same animal mode. Using the mini pig, whose skin most closely resembles human skin, a study was performed of histological changes 24 hours to 16 weeks after dermabrasion and chemical peel. A considerable increase in thickness of the new collagen layer was noted after chemical peel in contrast to that seen in skin after dermabrasion. The skin surface after chemical peel appeared smoother after 16 weeks, suggesting that chemical peel may be a more effective modality for treatment of the fine wrinkles of aging skin.


Atypical ossicle joint lesions in rheumatoid arthritis with sicca syndrome (Sjögren syndrome).

Atypical incudomalleal and incudostapedial joint changes were found in a 55-year-old woman with long-standing rheumatoid arthritis and sicca syndrome (Sjögren syndrome). Available audiograms taken at 37 and 42 years of age demonstrated slight bilateral high-frequency loss of hearing. The ossicular joint changes involved dissolution of disk material together with proliferation of synovial-type elements of the disk and articular surfaces, with formation of pannus-like tissue. There was extensive destruction of cartilage, with cellular collagenous tissue extending along the exposed bone surfaces. Despite strong similarities to rheumatoid arthritis, a definite diagnosis cannot be made in the absence of the inflammatory, lymphocyteplasma cell component.


Experience with gel-filled implants in augmentation mentoplasty.

Many materials have been used for augmentation mentoplasty with varying degrees of success. In our experience with the silicone bag-gel implant in 50 cases over the past 18 months the results have been quite satisfactory and there were no untoward complications.


Thyroid abscess in a 3 1/2-year-old child.

Thyroid abscess is a rare occurrence; there have been only six reported cases in children in the past 20 years and none of these in the otorhinolaryngologic literature. The presentation of suppurative thyroiditis in the 3 1/2-year-old child described here is somewhat atypical, with a rather insidious onset of swelling in the neck. The white blood cell count was elevated; thyroid function, however, was normal. The child also had an abnormal thyroid scan. Rapid improvement ensued with incision and drainage and appropriate antibiotic therapy. A brief discussion and review of the literature is provided regarding manifestations of previously reported cases. The recognition of this entity is important in order to differentiate from other fascial abscesses of the head and neck.


Dysphagia and Forestier disease.

Forestier disease (ankylosing hyperostosis) is a well-recognized and common disease entity. Although these patients rarely have complaints related to their spine, they are seen by the otorhinolaryngologist because of dysphagia, since the osteophytic spurs may impinge on the posterior pharyngeal wall. Since the plain roentgenogram and barium swallow is so specific, biopsy examination of the pharyngeal mass is not necessary.


Blood diseases in old age.

There is an increased incidence of chronic disease in old age, and blood disorder is not uncommon. Some diseases are more frequent in the latter half of life, while others -- such as the hereditary disorders -- are rare and occur only in a mild degree. Diagnosis of blood disease is more difficult in the elderly because symptoms of anaemia such as weakness, shortness of breath and mental disorder are often attributed to old age.


Association of Australian arboviruses with nervous disease in horses.

An outbreak of Murray Valley encephalitis (MVE) occurred in New South Wales during the first five months of 1974. Specimens from 52 horses with nervous disease collected January to May 1974 were examined histopathological or virologically. Although MVE virus was not isolated, 13 horses had serological evidence of recent infection with MVE virus. Another 4 horses had evidence of recent infection with Ross River virus. Two animals had histological evidence of viral infection of the central nervous system. Attempts to experimentally infect 2 horses with a low dose of MVE virus were not successful by intravenous, intramuscular and subcutaneous routes.


Disease and injury in the southern elephant seal.

Disease and injury was recorded in 170 tagged soughern elephant seals (Mirounga leonina) on the Antarctic continent. Trauma due to interspecific predation was the most common disorder. No small seals were observed with lesions attributable to this in the Antarctic population. This finding differs from data recorded in a sub-Antarctic (Macquarie Island) population where seals of all class sizes exhibit lesions due to interspecific trauma. No external parasites were observed in the Antarctic population.


Stabilization of gas cavitation nuclei by surface-active compounds.

Gas bubbles are the primary agent in producing the pathogenic effects of decompression sickness. Numerous experiments indicate that bubbles originate in water, and probably also in man, as pre-existing gas nuclei. This is surprising considering that gas phases larger than 1 micron should rise to the surface of a standing liquid, whereas smaller ones should dissolve rapidly due to surface tension. Several stabilizing mechanisms have been suggested, and each has been refuted on experimental grounds. In this article, we propose a new model that arises out of a systematic study of the earlier theories. We review these theories and conclude that gas cavitation nuclei must be held intact by surface-active skins that are initially permeable. The first quantitative analysis of bubble formation data from supersaturated gelatin is summarized and leads to the further conclusion that skins can become impermeable if the ambient pressure is increased rapidly by a sufficient amount. Our model owes much to Sirotyuk, who "demonstrated experimentally that stabilization of gas bubbles acting as cavitation nuclei in water is always attributable to the presence of surface-active substances in the water".


On the interaction of otolithic and cupular sensations.

A differential equation is proposed for describing how the brain compromises between a sensation of rotation about a particular axis (derived from the semicircular canals) and a sensation of gravity perpendicular to this axis (derived from the otolith organs). It is shown that this differential equation, with suitably chosen and reasonable values of the parameters and initial conditions, will satisfactorily explain the experimental results of Lansberg, Guedry, and Graybiel (7).


Mechanism of head and neck response to -Gx impact acceleration: a math modeling approach.

Mathematical modeling has attained wider acceptance in recent years. In particular, the use of computer programs to simulate the dynamic response of a human in a crash situation has become an attractive alternative to full-scale experimental testing. This paper analyzes data on the dynamic response of the living human head and neck to -Gx impact acceleration, where the motion of the subject's head and neck in the midsagittal plane was monitored with inertial instrumentation and high-speed photography for confirmation. The Calspan "3D Computer Simulator of Motor Vehicle Crash Victims" was used to predict expected responses for the deceleration pulses employed. These estimates were compared to the fully instrumented human test runs. The standard 15-segment and 14-joint representation of the occupant was modified to include two sternoclavicular joints, increasing the articulation in the upper torso. Analysis of the data indicated that muscular activity in the head and neck seemed to be evident and does influence motion of the head, even at relatively high (10-G peak, 530 G/s onset) acceleration levels. Simulation of muscular contraction, using a spring-damper arrangement, improved the results significantly. Additionally, possible limitations to head-to-neck motion, such as ligament restrictions, were also modeled.


Dietary requirements of subjects exposed to hyperbaric He-O2 conditions: effects of various levels of vitamin supplementation.

A 24-d growth study was conducted on rats exposed to ambient air, 11 ATA He-O2, or 21 ATA  He-O2 conditions. The rats were fed either a standard diet or the standard diet supplemented with 25, 50, or 100% increase of all vitamins, or the last with an additional 50 or 100% increase of casein. Fat, as cod liver oil was increased 0.5, 1.0, or 2.0% of the diet as the vitamins were increased. The vitamin and fat-supplemented diets, with or without supplemental casein, were adequate to support normal growth of the rats exposed to 11 ATA but not 21 ATA He-O2 conditions. Urine excretion and water consumption were closely related and varied in accordance with the adequacy of the diet. Feed digestibility was not a limiting factor in determining the growth of the rats under the three environments. The composition of the rat carcasses varied minimally.


Carbon dioxide hazards in general aviation.

There are numerous reports of carbon dioxide fire extinguishers or significant loads of dry ice causing actual or potential carbon dioxide intoxication leading to accidents or incidents, both in the military and in the airlines. To evaluate the significance of the problem in general aviation, dry ice sublimation reported in literature and measured sublimation was theoretically applied to general aviation aircraft, and carbon dioxide from fire extinguishers was measured, both in static and flight condition. The results indicate that the use of carbon dioxide fire extinguishers or the transportation under some conditions of dry ice could produce levels which could cause severe anxiety due to respiratory stimulation or actually produce pilot impairment and contribute to or cause accidents.


ST segment distortions by high-side frequency filtration in direct-writing ECG recorders.

ST segment depression in 12-lead resting and single-lead or multi-lead exercise ECGs is a useful screening criterion for detection of coronary heart disease (CHD). The recommended minimum frequency specification for direct-writing ECG recorders is 0.05-100 Hz (-3 dB). ST segment distortion by unintention low- and high-side frequency filtration could possibly weaken this screening capability. Consequently, one low- and two high-side potential filtration causes of such distortions were investigated in resting and exercise recordings of normal and both J-junction and ST-depressed ECGs. Of 4,914 filtered ST segments, 365 ST category changes were observed. Of these changes, approximately 15% was produced by filtration of the 0.05-0.1 Hz (amplifier function) and 45-100 Hz (60-Hz "noise" filtering) frequency bands. The remaining 85% was attributable to filtration of the 23-45 Hz (stylus overpressure) band. An optional remedy is discussed.


Aseptic necrosis presenting as wrist pain in SLE.

Aseptic necrosis of the wrist in systemic lupus erythematosus (SLE) is not rare. In 156 patients with SLE, aseptic necrosis occurred in 11 patients, of whom 3 (27%) had wrist involvement. Onset of pain was insidious and the symptoms were thought to be related to synovitis due to SLE. An average of 11 months elapsed before aseptic necrosis was correctly diagnosed. An awareness of this possibility is important in the management of any patient with SLE who complains of wrist pain.


Carrageenin-induced arthritis. IV. Rate changes in cartilage matrix proteoglycan synthesis.

A localized inflammatory response was initiated by both single and repeated injections of carrageenin into femorotibial joints. Histologic changes were observed 24 hours after a single intraarticular injection, and an inhibition in the in vitro rate of proteoglycan synthesis was detected 72 hours after the injection. This inhibition was relieved in vitro by the addition of beta-D-xyloside, an exogenous initiator of glycosaminoglycan biosynthesis. Following repeated carrageenin injections, most cells appeared to be dead on histologic examination and no in vitro proteoglycan synthesis could be detected; nor could any stimulation be achieved by adding exyloside.


Connective tissue activation. XI. Stimulation of glycosaminoglycan and DNA formation by a platelet factor.

This report describes a small basic protein found in human platelets which stimulates hyaluronic acid formation, glucose uptake, and lactate formation in several types of human connective tissue cells. In addition, it stimulates in incorporation of [3H] methyl thymidine into DNA in human synovial cell cultures. This platelet factor (connective tissue activating peptide-II, CTAP-III) clearly differs form CTAP-I, found in human lymphocytes, and may play a role as a mediator of the inflammatory process.


Connective tissue synthesis by cultured scleroderma fibroblasts. II. Incorporation of 3h-glucosamine and synthesis of glycosaminoglycans.

Fibroblasts from normal and scleroderma skin, grown tissue culture, were incubated with 3H-glucosamine for 24 hours. No definite trends could be established in 3H-glucosamin incorporation or glycosaminoglycan synthesis. Over 90% of the total 3H activity as well as the glycosaminoglycans synthesized were secreted into the medium. Characterization of glycosaminoglycans showed that in both the medium and cells, hyaluronic acid ins the major glycosaminoglycan in normal and scleroderma fibroblasts. In the medium, hyaluronic acid represented 87% of the total glycosaminoglycans; it was slightly decreased in the cells. Fibroblasts were compared from the upper and lower areas of the affected dermis and an uninvolved dermal area of the same scleroderma patients.


Preliminary criteria for the classification of the acute arthritis of primary gout.

The American Rheumatism Association sub-committe on classification criteria for gout analyzed data from more than 700 patients with gout, pseudogout, rheumatoid arthritis, or septic arthritis. Criteria for classifying a patient as having gout were a) the presence of characteristic urate crystals in the joint fluid, and/or b) a topus proved to contain urate crystals by chemical or polarized light microscopic means, and/or c) the presence of six of the twelve clinical, laboratory, and X-ray phenomena listed in Table 5.


Comparison of assay methods for selective measurement of plasma lipase. The effect of clofibrate on hepatic and lipoprotein lipase in normals and patients with hypertriglyceridemia.

Three different assays for selective measurement of plasma lipoprotein lipase (LPL) and hepatic triglyceride lipase (H-TGL) were compared. These were: an immunochemical method based on enzyme antibody precipitation (IM), a procedure in which both enzymes were separated by affinity chromatography on small heparin--Sepharose columns (HS), and an assay in which one enzyme was inhibited by protamine sulfate (PS). Good correlations were found between the immunochemical and the heparin--Sepharose method, but not between these and the protamine sulfate assay procedure. The IM was then used to evaluate the effect of clofibrate on the two lipolytic enzymes. It was found that both in normals and in patients with Type IV hyperlipoproteinemia, clofibrate treatment leads to a specific increase of plasma LPL while H-TGL activity remains almost unaffected. The magnitude of the LPL response was different in normals and in patients with endogenous hyperlipoproteinemia. Furthermore, in normals the maximal increase of LPL activity was already reached one week after drug treatment was begun, while in hypertriglyceridemic patients, this effect was not evident prior to four weeks of clofibrate treatment.  The marked enzyme increase following clofibrate administration indicates that an increased peripheral removal rate for triglycerides is one major mechanism responsible for the lipid-lowering effect of this drug.


Prevalence of coronary artery disease and peripheral artery disease in patients with different types of primary hyperlipidemia.

The prevalence of coronary artery disease (CAD) and peripheral artery disease (PAD) was studied in 280 (203 males, 77 females) patients with different types of primary hyperlipoproteinemia. In primary hyperbetalipoproteinemia the prevalence of CAD (45% for Type IIa and 47% for Type IIb) is significatly higher than that in the other types of hyperlipoproteinemia (38% for Type IV and 17% for Type V). On the other hand, PAD prevalence is much higher in hypertriglyceridemia (21% in Type IIb and 20% in Type V) than in hypercholesterolemia alone (9% in Type IIa). These results suggest ths atherosclerotic complications are concerned. Moreover, the high frequency of PAD found in hypertriglyceridemia can be related to the high occurrence of diabetes in these patients. The effects of other major risk factors of atherosclerosis (smoking and hypertension) were also evaluated. Our results indicate that the association of hypercholestolemia and hypertension is more dangerous than the co-occurence of hypercholesterolemia and smoking.


The treatment of hypercholesterolemia with beta-pyridylcarbinol. Part 5. Report on 16 cases with severe hypercholesterolemia treated for 12 years.

A group of 78 patients with severe hypercholesterolemia (-X = 464 mg/dl) and symptoms of vascular disease of the heart, the extremities, or the brain, started a beta-pyridylcarbinol treatment with an average daily dosage of 1.2 g in 1964. In 1976 we could re-examine 12 patients, still on the same therapy. No myocardial infarction has occurred in this group since 1973, only 2 patients have had more attacks of angina pectoris that 1964. In contrast patients discontinuing therapy or replacing beta-pyridylcarbinol by other hypolipidemic drugs had a higher mortality.


Experimental atherosclerosis of the heart valves in rats following a long-term atherogenic regimen.

Male and female rats kept on a long-term atherogenic regimen for periods up to 15 months developed marked atherosclerotic changes of cardiac valves in 100% of cases. The degree and the topography of valvular atherosclerotic plaques corresponded widely to that seen in human atherosclerotic valvular disease. The regular topography of atherosclerotic valvular lesions was believed to be primarily due to differences in blood pressure as occurring during cardiac performance. Histologically, there was a striking contrast between the uniform composition of atheromatous plaques as found on the valves and the variability in composition of aortic atheromas.


Inhibition of platelet adhesion to rabbit aorta by sulphinpyrazone and acetylsalicylic acid.

A technique for the isolation of rabbit thoracic aorta in a form in which it can withstand normal pressure and a new in vitro system in which it was tested is described. Segments of aorta were selectively damaged with a balloon catheter by the Baumgartneer technique and supensions of washed platelets, labelled with 51Cr, were perfused through them. More platelets adhered to the damaged surface than to the undamaged surface. Suophinpyrazone was more effective than aspirin in inhibiting platelet adherence to both the damaged and undamaged surface.


Prenatal diagnosis and gonadal findings in X/XXX mosaicism.

Prenatal diagnosis of a case of X/XXX mosaicism is presented. In spite of the fact that over 50% of the cells cultured from both ovaries were trisomic for the X chromosome, fetal öocytes were rarely found. This case illustrates that the presence of a triple-X cell line, even in a relatively high percentage of ovarian cells, does not necessarily protect the ovary from 'aöogenesis'. This observation might prove useful in the counselling of future cases involving the prenatal detection of sex chromosome mosaicism.


Recessive form of Freeman-Sheldon's syndrome or 'whistling face',.

Freeman-Sheldon's syndrome is a rare genetic disease inherited as an autosomal dominant trait in some families but showing sporadic appearance in the majority of the reported cases. In the present paper we report a family having two affected children from normal consanguineous parents suggesting that Freeman-Sheldon's syndrome may be heterogeneous from the genetic point of view.


Assessment of response to therapy in advanced breast cancer.

A system is proposed by the UICC for assessing response to treatment of advanced breast cancers.


Interaction of hyperthermia and the hypoxic cell sensitizer Ro-07-0582 on the EMT6 mouse tumour.

The combination of hyperthermia and the hypoxic cell radiosensitizer Ro-07-0582 has been investigated on the EMT6 tumour implanted into the legs of BALB/c mice. Treatments at a drug dose of 1 mg/g over a range of waterbath temperatures from 37 to 45 degrees C are described. The surviving clonogenic fraction following treatment was assayed in vitro. Measurements of intra-tumour temperature have been made, and shown to be better correlated with the cytocidal effect on the tumour than the waterbath temperature. No significant effect of Ro-07-0582 was observed at 37 degrees C. However, marked cytotoxicity due to the drug was seen at intra-tumour temperatures above 42-5 degrees C for 1 h. These were in addition to the cytocidal effect of the hyperthermia. The results are discussed in relation to the distribution of temperatures and hypoxic cell populations throughout the tumour.


Effect of hyperthermia on differential cytotoxicity of a hypoxic cell radiosensitizer, Ro-07-0582, on mammalian cells in vitro.

There is now evidence that several classes of nitro compounds which have been used as radiosensitizers also function as cytotoxic agents specific for hypoxic cells. The 2-nitroimidazole, Ro-07-0582, (1-(2-nitroimidazol-1-yl)-3-methoxy-2-propanol) is a compound of this type, and its effectiveness as a cytotoxic agent is dependent on drug concentration, contact time and temperature. In vitro, Ro-07-0582 in air at 37 degrees C does not cause loss of cell viability at concentrations up to 2 mM, even when in contact for several days. In contrast, hypoxic cells do not tolerate much lower concentrations of drugs, even if the contact time is only a few hours. When the temperature is raised above 37 degrees C, there is a pronounced increase in the slope of the survival curves; for example, at 41 degrees C (for 1 mM Ro-07-0582, (200 microng/ml), the slope changes by a factor of 2-0 relative to that for 37 degrees C. For cells in air at 41 degrees C, as at 37 degrees C, there is no toxic effect at the concentration of drug tested. In the absence of drug, there is no cytotoxic effect of hyperthermia alone under these conditions. These results are discussed in terms of Arrhenius parameters.


Dephenylation of N-phenyl-2-naphthylamine in dogs and its possible oncogenic implications.

N-Dephenylation of N-phenyl-2-naphthylamine (PBNA) is strictly limited in dogs, and a 5 mg/kg dose gives 0-10 microng of urinary 2-naphthylamine (BNA), which does not appear to undergo further metabolism. Neither 2-naphthylhydroxylamine (BNHA) nor 2-amino-1-naphthylsulphate were detected in the urine of treated animals. Urinary output of BNA varies markedly between dogs, and at different times in the same animal. The extent of PBNA N-dephenylation is unaltered by chronic administration. Calculations based on Druckery and Küpfmüller's equation (1948) and present data indicate that, for dogs to form BNA tumours through exposure to a relatively high dose-level of PBNA, the period of daily dosing would occupy, or even exceed, the normal life-span. The carcinogenic risk of PBNA to human subjects is discussed.


Avascular and vascular phases of tumour growth in the chick embryo.

The chick embryo was used to study the relationship between the onset of tumour neovascularization and tumour growth. Walker 256 carcinosarcoma was implanted on the chrioallantoic membrane (CAM) of about 600 embryos aged 5-16 days. Tumour diameter and changes in the CAM vasculature in response to the implants were recorded daily. Representative tumours were examined by light microscopy of Epon-embedded tissue and autoradiography after injection of [3H]-thymidine. Tumours remained avascular for 72 h, after which they were penetrated by new blood vessels and began a phase of rapid growth. The rate of growth during this vascular phase was greatest for implants on 5- and 6-day-old embryos and decreased the later the day of implantation. The time of onset of tumour angiogenesis appears to be independent of the immunological state of the chick embryo, although the rate of growth after vascularization may be modified by the onset of immunity. This study suggests that the avascular and vascular phases of tumour growth are separable, and that the avascular tumour population lives under the growth constraints which limit the size of a tumour spheroid growing in soft agar or aqueous humour.


Adjustment and war bereavement - some considerations.

Forty-seven parents from 25 families who had lost a son in the War of Attrition of 1969-1970 were interviewed in the course of this study. The interviews took place 2-3 years after the bereavement. The purpose of the study was an assessment of personal adjustment of bereaved parents, and an examination of background and behavioural characteristics which might differentiate between parents making a better and less good adjustment to the loss of a son. To this end data were collected via semistructured interviews. Non-objective variables: religiosity, social interpersonal relationships and adjustment were rated by independent judges. Reliability of interjudge agreement ranged from 0-67 to 1-00. Hypotheses were tested using correlation matrices. The hypothesis stating a positive relationship of high religiosity and good adjustment was not confirmed. The results suggest a very significant relationship between good adjustment and level of education (r = 0-668, alpha less than 0-01). Level of education was responsible for most of the variance and thus made impossible an answer regarding the relationship between religiosity and recovery after bereavement. To elucidate religiosity's function in adjustment, control of education level would be necessary. While this was not done in the present study, a further research plan was suggested. The hypothesis stating a positive correlational relationship between adjustment and interpersonal skills as reflected in social contacts and marital relations was confirmed (r = 0-735, r = 0-573, alpha less than 0-01). The findings of this study suggest that individuals who have suffered a severe stress, may gain strength, encouragement and compensation when their life style includes higher educational level and satisfying interactions with other people.


Loss: a central theme in psychotherapy.

As an essential aim of psychotherapy is to change in order to develop and grow the patient has to give up some mal-adaptive patterns of behaviour and out-of-date values and belief systems. The inevitable losses involved in this process are discussed in detail. The losses the patient has to face when entering therapy are considered first, followed by consideration of the losses he encounters when during therapy he has to change his attitudes towards himself and others, including the giving up of infanitle wishes and illusions and of value systems originally imposed upon him from outside sources. The losses and disappointments he experiences in the therapeutic relationship itself are discussed, and attention is focused on the way in which what is at first experienced as a loss can become the starting point of new developments and lead to a more positive acceptance of his psychic and physical functioning. To achieve these aims the therapist himself needs to learn to adopt a similar, positive attitude towards the losses and limitations involved in his own life experience and therapeutic work.


Attitudes of psychiatric patients to staff roles and treatment methods: a replication and extension.

A questionnaire dealing with attitudes toward staff roles and treatment methods was administered to a group of 191 psychiatric patients and their responses subjected to a rotated principal components analysis. The first three components accounted for the same proportion of the overall variance as did those obtained in an earlier study. The first components, emphasizing high regard for physical treatments and formal interpersonal relationships, were essentially the same in both studies. The second and third components, however, showed important differences; our second component dealt with the psychological and social aspects of patienthood, and our third reflected a critical attitude toward doctors. Scores on the first component were significantly associated with age, score on the Hysteroid-Obsessoid Questionnaire and social class. Women scored higher than men on the second component and scores on the third were related to total hostility scores on the Hostility and Direction of Hostility Questionnaire and to schizophrenia. Differences between diagnostic groups on the first component could be attributed almost entirely to the groups' mean ages.


The multiple techniques approach to behavioural psychotherapy: a retrospective evaluation of effectiveness and an examination of prognostic indicators.

A retrospective evaluation of the multiple techniques approach to behaviour therapy is described. The effectiveness of the approach seems to compare favourably with a single-technique approach as far as can be ascertained. The observed improvement can be attributed to some aspect of the treatment package and not to 'spontaneous remission' factors. A prospective trial would not seem valuable, providing the problems of replication can be solved. An examination of prognostic indicators reveals a range of variables associated with outcome. Amongst the diagnostic groups, the 'social phobics' and 'anxiety states' did poorly. A number of general adjustment variables also seemed important, particularly marriage, and social and vocational adjustment. The implications of these findings for the nature and deployment of services and briefly discussed.


Hierarchies of personality deviance and personal illness.

Three hundred and twenty-five psychiatric patients were allocated to classes within the hierarchy of personal illness by means of the Delusions-Symptoms-States Inventory. They were then given the Personality Deviance Scales. The results showed that the classes ranked in the same hierarchical order as on the DSSI on extrapunitiveness and intropunitiveness, but not on dominance. Maladjustive personality deviance, as statistically defined, was 3 1/2 times as frequent among patients as among non-patients. Whereas symptom measures had previously been shown to change considerably after one month, personality measures did not. Longer follow-up periods are needed before it can be decided whether personality measures contribute substantially to prediction of type of illness or whether they are determined, at least in part, by the type of illness. In the latter event, personality measures might still prove useful in providing a more fundamental estimate of long-term clinical improvement than symptom measures alone if they were found to change more slowly or only with more intensive clincial effort.


Thumbsucking and falling asleep.

A review of the studies on the aetiology of habitual thumbsucking reveals either contradictory or inconclusive results. In this study carried out in Turkey, 50 thumbsuckers, 50 non-thumbsuckers, 250 school children and 312 'problem' children were investigated through interviews, questionnaires and other clinical techniques with their mothers. Among variables studied were aspects of feeding, onset and incidence of thumbsucking, strength of sucking drive, sex distribution, educational level and occupation of mothers, parental attitudes toward physical contact with children, mother-child relationships, and particular forms of falling asleep. It was found that thumbsucking was aetiologically more related to ways of falling asleep than to other factors. An attempt was made to explain the social, psychological and physiological basis of the aetiological significance of the falling asleep-stage in habitual thumbsucking. These findings now permit predictive longitudinal investigations to test this accuracy.


Vitrectomy technique in anterior segment surgery.

Instruments designed primarily for vitreous surgery were used in 74 anterior segment operations. In each case the instrument was introduced into the eye through a limbal incision. This preliminary report shows that this new approach offers several advantages over conventional methods in selected cases. It is of particular value in the management of presenile and complicated cataracts and in the removal of capsulo-lenticular remnants and pupillary membranes. A larger number of cases and longer follow-up are necessary to define further the specific indications and limitations of this technique.


Immunoglobulins in tear in trachoma patients.

Tear immunoglobulin concentrations have been measured in 100 healthy people and 62 patients in different stages of trachoma. In healthy people the average IgA level was 27-8 mg/100 ml. There was no significant difference in the IgA level in various age groups and between the sexes. IgG was detected in 92 samples, and it was less than 1 mg/100 ml. IgM in tears was detected in only one sample. IgD was not detected in any specimen. In tracoma cases, the mean IgA level was found to be significantly lower (22-0 mg/100 ml) than in healthy people. There was no significant difference in IgA level between different stages of trachoma, IgG, IgD, and IgM could not be detected in any sample from the trachoma cases.


Histopathological changes in malignant melanomas of the choroid after cobalt plaque therapy.

A series of 23 eyes removed after cobalt plaque therapy for malignant melanoma of the choroid is presented, and the histological changes are described with particular reference to the presence or absence of necrosis in the tumours. In 17 of the 23 eyes no evidence of tumour necrosis was found, though all showed evidence of radiation changes in the adjacent tissues. Necrosis was found in 6 cases but was prominent in only 2, one of which was the only example of a mixed-cell tumour, the other 22 being spindle-cell tumours. Enucleation became necessary on average at about 32 months after treatment (23 out of 100 treated eyes) because of obvious failure of treatment with evidence of continued growth, or because of radiational complications leading to a painful blind eye. The 5-year survival rate was 86% compared with about 50% when the treatment is enculeation.


Sulphydryl group modification of aspartate aminotransferase with 3-bromo-1,1,1-trifluoropropanone during catalysis.

After protection of cysteine-45 and -82 with iodoacetamide or N-ethylmaleimide, and in the presence of saturating concentrations of substrates, the supernatant isozyme of pig heart aspartate transaminase has been covalently modified at cysteine-390 with 3-bromo-1,1,1-trifluoropropanone. The modified enzyme retains 60-70% of the initial specific activity and is similar to native enzyme in pH and temperature stability. After tagging cysteine-390 with the fluorinated compound, the enzyme retains  substrate and inhibitor binding abilities; as shown by direct spectrophotometric titration of the active-site chromophores. The 19F NMR spectrum of the modified enzyme has been obtained by a Fourier transform NMR method. Although the transaminase is a dimeric enzyme, 19F bound at each subunit's cysteine-390 gives rise to only a single 19F resonance upfield from that of trifluoroacetic acid. The fact that the chemical shifts of the 19F probe differ in native and guanidine hydrochloride (Gdn-HCl) denatured enzyme is interpreted as the effect of the native protein groups on the probe. The discordance between the changes induced by varying concentrations of Gdn-HCl on the 19F resonance parameters, on the one hand, and the changes in enzyme activity and prosthetic group absorbance, on the other, suggests that, in aspartate transaminase, cysteine-390 lies in an environment dissimilar from that of the active-site components.


Active site specific inactivation of chymotrypsin by cyclohexyl isocyanate formed during degradation of the carcinostatic 1-(2-chloroethyl)-3-cyclohexyl-1-nitrosourea.

Prolonged incubation of 1-(2-chloroethyl)-3-([1-14C]cyclohexyl)-1-nitrosourea with chymotrypsin resulted in covalent modification and concomitant inactivation of chymotrypsin via degradation of the nitrosourea to form cyclohexyl isocyanate. Cyclohexyl isocyanate was shown to be an active-site-specific inactivator of chymotrypsin. A cyclohexyl isocyanate to enzyme molar ratio of 0.63 was required to produce 50% enzyme inactivation, thus demonstrating the high specificity of inactivation. At 2.38 X 10(-4) M chymotrypsin this near stoichiometric inactivation was not significantly affected by the presence of 1, 5, and 10 mM L-lysine. Degradation of an excess of 1-(2-chloroethyl)-3-([1-14C]-cyclohexyl)-1-nitrosourea in the presence of enzyme yielded 1.11 +/- 0.07 mol of covalently bound [14C]cyclohexyl moiety per mol of enzyme inactivated. Short-term incubation demonstrated that the nitrosourea neither inhibited nor protected the enzyme from cyclohexyl isocyanate inactivation. Treatment of chymotrypsin with less than stoichiometric amounts of cyclohexyl isocyanate or titration of the active-site serine with phenylmethanesulfonyl fluoride followed by in situ degradation of excess 1-(2-chloroethyl)-3-([1-14C]cyclohexyl)-1-nitrosourea resulted in a decreased amount of covalently bound 14C proportional to the extent of inactivation by these reagents prior to 14C labeling. These results strongly suggest that cyclohexyl isocyanate, whether added directly or generated by CCNU degradation, reacted almost exclusively with the active site of the enzyme. The extent of this inactivation indicates that 70% of the CCNU degraded in such a manner as to form cyclohexyl isocyanate.


Significance of surface potential in interaction of 8-anilino-1-naphthalenesulfonate with mitochondria: fluorescence intensity and zeta-potential.

The 8-anilino-1-naphthalenesulfonate (Ans) fluorescence in nonenergized and energized mitochondria was measured at various concentrations of Ans and KCl. Under the same experimental conditions, the zeta potential was determined from measurements of the electrophoretic mobility of mitochondria. The fluorescence intensity under various conditions was represented quantitatively in terms of the Langmuir adsorption isotherm where the electrostatic interaction acting between Ans and mitochondria was properly taken into account. The values of qN (q) proportionally constant related to quantum yield of Ans: N, the maximum number of the adsorption site) and deltaG (nonelectrical part of the free-energy change due to the binding of Ans to mitochondria) were constant irrespective of difference in energy state and in ionic strength in media. It was concluded that changes in the Ans fluorescence in mitochondria are mainly attributed to changes in the surface potential of mitochondria.


Cyanogen bromide treatment of methionine-containing compounds.

The preparation of a series of X-Met-Gly-OEt and X-Met-Phe-OMe and their treatment with CNBr in either 70% or 97-100% formic acid at 25 degrees C are described where X is methanesulfonyl (mesyl), p-nitrobenzyloxycarbonyl, phthaloyl, trifluoroacetyl, acetyl, formyl, or tert-butyloxycarbonyl. Total cleavage of the peptide esters was found with mesyl-, p-nitrobenzyloxycarbonyl-, phthaloyl-, and trifluoroacetylmethionyl derivatives which indicated the suitability of these derivatives as amino protecting groups in peptide synthesis. Treatment of the acetylmethionyl peptide esters with CNBr in 70 and 97-100% formic acid resulted in 92 and 98% cleavage, respectively. With formylmethionyl peptide esters, about 85-95% cleavage was estimated when either 70 or 97-100% formic acid was used as the solvent. With the tert-butyloxycarbonylmethionyl derivatives, CNBr treatment in 70% formic acid resulted in about 93% cleavage of peptides, while treatment in 97-100% formic acid led to only 30-33% release of C-terminal amino acid esters. Quantitative cleavage of the carbonylbis(methionyl peptide esters) was observed. The reaction of CNBr with N-terminal methionyl derivatives containing free alpha-amino groups revealed that free methionine was quantitatively converted to homoserine lactone, whereas methionine ethyl ester and methionyl peptides (Met-Gly and Met-Phe) disappeared from the reaction mixture in 70% formic acid with only partial splitting of the ester (16%) or peptide bond (45%).


Purification of rat liver and mouse ascites DNA-dependent RNA polymerase I.

Three forms of RNA polymerase were assayed in nuclei and nucleoli isolated from rat liver and from Krebs II ascites cells. Assays of rat liver nuclei in the absence of exogenous DNA showed polymerase I accounted for 72% of the total activity, polymerase II for 17%, and polymerase III for 11%. The total activity in ascites nuclei was similar but the ratios of polymerase activities were different: polymerase I, 53%; polymerase II, 41%; and polymerase III, 6%. These values may reflect differences in the transcriptional activity of the nuclei. After isolation of nucleoli, both rat liver and ascites polymerase I accounted for 85% of enzyme activity. When exogenous calf-thymus DNA was added to nucleoli, there was a greater than 50% increase in activity suggesting that less than one-half of the polymerase I present was bound to endogenous template. Polymerase I was solubilized from either rat liver or ascites nucleoli by sonication at high ionic strength and subsequently purified by ion filtration, phosphocellulose, sucrose gradient centrifugation, and DNA-cellulose chromatography. The essentially homogenous ascites enzyme had a specific activity of 86 units/mg when assayed with native calf-thymus DNA and of 876 units/mg when assayed with poly(deoxycytidylic acid). Electrophoresis of the enzyme in sodium dodecyl sulfate indicated the presence of six subunits with molecular weights of 205 000, 125 000, 51 000, 44 000, 26 000 and 16 000. After the same purification procedure, the rat liver enzyme had a similar specific activity (98 units/mg) on native calf thymus and 362 units/mg on poly(deoxycytidylic acid).


Nucleotide sequences at the N6-methyladenosine sites of HeLa cell messenger ribonucleic acid.

Borate gel chromatography was used to separate internal oligonucleotides containing N6-methyladenosine (m6A) from methylated 5'-terminal oligonucleotides of HeLa cell polyadenylylated mRNA. N6-Methyladenosine occurs primarily in two sequences, -G-m6A-C (70%) and -A-m6A-C-(30%). The nucleoside immediately following cytidine may be uridine, cytidine, or adenosine, while guanosine as well as other nucleosides occupy subsequent positions. Each of the four positions preceding the -(G or A)-m6A-C- sequence may be occupied by a pyrimidine or a purine ribonucleoside. Since on a random basis all possible sequences containing -(G or A)-A-C-(U or C or A)- could occur once per 43 nucleotides whereas there is only one m6A residue per thousand nucleotides, then either (1) not all potential sites are methylated, (2) there are multiple unique sequences perhaps methylated by several different enzymes, or (3) there are other unrecognized discriminating factors. The possibility that methylation of adenosine occurs exclusively in the region close to the 5' terminus of the mRNA was considered. However, such a localization was excluded since the majority of m6A residues were not found in 4 to 6S 5'-terminal fragments isolated by borate gel chromatography.


Fractionation of purified nucleosomes on the basis of aggregation properties.

Monomeric nucleosomes from micrococcal nuclease digest of rat liver nuclei have been purified on Sepharose columns in the presence of divalent cations and 0.6 M NaCl. The particles contain histones H2A, H2B, H3, H4, and a complement of nonhistone chromosomal proteins. In 0.6 M NaCl, the nucleosome mixture sedimented at 10 S; however, when the NaCl was removed approximately 30% of the particles aggregated and precipitated, and the remaining soluble fraction sedimented at 11 S. The aggregation phenomenon was divalent cation-dependent and reversible. Characterization of the macromolecular components of the subfractions of nucleosomes showed that the subfractions differed in composition of species of histone H3 as well as of several nonhistone chromosomal proteins but not in the size of the DNA fragment present. The aggregation properties of the isolated nucleosomes showed similarities to the divalent cation-dependent differences in the extent of chromatin condensation in the intact eukaryote nucleus.


Amino acid sequence studies on the alpha chain of human fibrinogen. Characterization of 11 cyanogen bromide fragments.

The alpha chain of human fibrinogen consists of 600 +/- 25 amino acid residues, 10-11 of which are methionines. In this regard, we have identified and characterized 11 cyanogen bromide peptide fragments of 2, 3, 26, 28, 28, 37, 51, 56, 60 +/- 5, 64 +/- 5, and 260 +/- 20 residues, respectively. The sequences of five of these and a portion of a sixth have been reported previously. We now report the complete amino acid sequences of another of these fragments (56 residues), partial sequences for four others, and a preliminary characterization of the largest fragment. In a companion study (Doolittle, R. F., Cassman, K. G., Cottrell, B. A., Friezner, S. J., and Takagi, T. (1977), Biochemistry 16 (following paper in this issue)), we have obtained key overlap sequences from plasmic digests of fibrinogen which allow all but one of these cyanogen bromide peptides to be arranged in order. The sequences of some of these newly reported fragments have revealed an internal homology in the alpha chain, as well as structural similarities to the corresponding portions of the beta and gamma chains.


Ultracentrifugal studies of human luteinizing hormone and its subunits: dependence on protein concentration and ionic strength.

Purified human pituitary luteinizing hormone (hLH) and its alpha and beta subunits have been examined by physical methods. Freshly prepared hLH showed three closely adjacent bands on electrophoresis in polyacrylamide gels under alkaline conditions, but on standing even in the freeze-dried condition additional bands appeared. alpha and beta subunits gave bands which were quite different from the main hLH bands but comparable with the additional bands, as well as traces of hLH bands. hLH was investigated at three ionic strengths (0.1, 0.2, 0.5) at pH 5.9 +/- 0.1. Sedimentation velocity experiments demonstrated a complex system of association-dissociation which was further investigated by sedimentation equilibrium. Association occurred at the higher protein concentrations at each ionic strength, but to a significantly higher level at I = 0.5. Only at I = 0.1, pH 5.8, was there a clear indication over a range of protein concentration of the occurrence of a species of molecular weight 32 000 +/- 2000, in fair agreement only with the sum of the molecular weights of the alpha and beta subunits. At higher ionic strengths, there were indications of dissociation at low protein concentration (less than 0.10 g/100 mL) as well as association at higher values (greater than 0.20 g/100 mL). In view of the occurrence of molecular weights less than 28 000 at I = 0.2, HLH was treated in terms of a monomer of molecular weight 14 000, and some evidence was obtained for tetramer formation (4M = M4). At higher ionic strength this model does not apply and it is thought that indefinite association may be occurring to some extent also. The alpha subunit gave indications of association from its sedimentation coefficient vs. concentration plot, and sedimentation equilibrium (at pH 5.9 I = 0.1) demonstrated molecular weights increasing with increasing concentration. Evidence for tetramer formation also was obtained. The beta subunit, in spite of an apparently simple sedimentation coefficient-concentration curve, showed molecular weights varying from well below 14 000 to beyond 20 000. There is evidence to suggest that the isolated alpha and beta subunits, even on standing as dry solid, are not stable but give rise to lower molecular weight products. Aged freeze-dried hLH did not show such impurity.


Comparison of hemerythrins from four species of sipunculids by optical absorption, circular dichroism, fluorescence emission, and resonance Raman spectroscopy.

Resonance Raman, optical absorption, circular dichroic, and fluorescence emission spectroscopy of hemerythrins from four species of sipunculids (Phascolopsis gouldii, Phascolosoma agassizii, Themiste dyscritium, and Themiste pyroides) reveals no major differences in their active site or tertiary structures. This precludes any change in iron ligands or coodination geometry and makes it unlikely that the active-site structures of P. gouldii and T. dyscritum hemerythrins could be as disparate as indicated by present crystallographic interpretations (Stenkamp, R. E., Sieker, L. C., and Jensen, L. H. (1976), Proc. Natl. Acad. Sci. U.S.A. 73, 349; Klotz, I. M., Klippenstein, G. L., and Hendrickson, W. A. (1976), Science 192, 335). Resonance Raman enhancement profiles of the stretching modes involving coordinated dioxygen maximize with excitation at approximately 525 nm, and correspond to the circular dichroic (CD) transition at approximately 520 nm. For coordinated azide modes in metazidohemerythrins these profiles maximize with excitation at approximately 505 nm corresponding to the 500-nm CD transition. Hemerythrins also possess another resonance Raman peak at approximately 510 cm-1 which show maximum intensity enhancement at approximately 530 nm and this vibration is most likely associated with a permanent iron ligand.


In vitro reconstitution of calf brain microtubules: effects of solution variables.

The effects of solution variables on the in vitro reconstitution of calf brain tubulin, purified by the method of Weisenberg et al. (Weisenberg, R. C., Borisy, G. G., and Taylor, E. W. (1968), Biochemistry 7, 4466-4479; Weisenberg, R. C., and Timasheff, S. N. (1970), Biochemistry 9, 4110-4116), as modified by Lee et al. (Lee, J. C., Frigon, R. P., and Timasheff, S. N. (1973), J. Biol. Chem. 248, 7253-7262), were investigated at pH 7.0. Reconstitution of microtubules was successful in a variety of buffer systems, the free energy of the propagation step of microtubule formation being little dependent on the buffer. Microtubule formation is promoted by magnesium ions and guanosine triphosphate, but inhibited by calcium ions. The dependence of the apparent association constant for microtubule formation on ligand concentration was analyzed by the linked function theory of Wyman (Wyman, J. (1964), Adv. Protein Chem. 19, 224-286), leading to the conclusion that the formation of a tubulin-tubulin contact involves the binding of one additional magnesium ion per tubulin dimer. Microtubule formation is also accompanied by the apparent binding of one additional proton and the release of water molecules, as suggested by the thermodynamic parameters determined. The reaction is entropy driven with an apparent heat capacity change, deltaCp, of -1500 +/- 500 cal/deg-mol. The enhancement of tubulin reassembly by glycerol is most likely due to nonspecific protein-solvent general thermodynamic interactions.


Relationship between inhibitor binding by chloroplasts and inhibition of photosynthetic electron transport.

The binding of radioactively labelled atrazin, metribuzin and phenmedipham by broken chloroplasts was studied. From the double-reciprocal plots (bound vs. free inhibitors) a high affinity binding reaction is graphically isolated which is related to the inhibition of photosynthetic electron transport. It is concluded that the specific binding sites correspond to the electron carrier molecules which are attacked by the inhibitors. The relative concentration of specific binding sites is 1 per 300-500 chlorophyll molecules. The binding of the labelled substances is competitively inhibited by each of the indicated unlabelled substances, by DCMU and by several pyridazinone derivatives. These results suggest that triazines, triazinones, pyridazinones, biscarbamates and phenylureas interfere with the same electron carrier of the photosynthetic electron transport chain, according to the same molecular mechanism.


Energy-dependent exchange of adenine nucleotides on chloroplast coupling factor (CF1).

1. [14C]ADP is incorporated into washed broken chloroplasts in the light. The bound labelled nucleotides which cannot be removed by washing are almost exclusively related to coupling factor CF1. [14C]ADP binding exhibits a monophasic concentration curve with a Km of 2 micronM. 2. By illumination of the chloroplasts, previously incorporated labelled nucleotides are released. A fast release is obtained in the presence of unlabelled ADP and ATP, indicating an energy-dependent exchange. A slow and incomplete release is induced by light in the absence of unlabelled adenine nucleotides. Obviously, under those conditions, an adenine nucleotide depleted CF1 conformation is established. 3. Re-binding of [14C]ADP by depleted membranes is an energy-independent process. Even after solubilization of adenylate-depleted CF1, [14C]ADP is incorporated into the protein. By re-binding of ADP in the dark, CF1 is converted to a non-exchangeable form. 4. Energy-dependent adenine nucleotide exchange on CF1 is suggested to include three different conformational states of the enzyme: (1) a stable, non-exchangeable form which contains firmly bound nucleotides, is converted to (2), an unstable form containing loosely bound adenine nucleotides. This conformation allows adenylate exchange; it is in equilibrium with (3) a metastable, adenylate-depleted form. The transition from state (1) to state (2) is the energy-requiring step.


Flash induced fluorescence kinetics in chloroplasts in the 20 microseconds-100 s time range in the presence of 3(3,4-dichlorophenyl)-1,1-dimethylurea. Effects of hydroxylamine.

Flash induced variations of the fluorescence yield have been studied at 2 degrees C over a long time range (at 1 microseconds and from 20 microseconds to 3 min) in chloroplasts in the presence of 3(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU) with or without addition of hydroxylamine. 1. In the presence of DCMU, a polyphasic rise is observed. A first fast rise (less than 1 microseconds) is ascribed to the transfer of the positive charge from the primary Photosystem II donor Chl to a secondary donor Y. Two hypotheses are proposed to interpret the existence of the subsequent slower increase (t1/2 approximately equal to 70 microseconds) which then follows the initial fast rise. 2. The effects or various hydroxylamine concentrations have been studied with a sufficient incubation time to inactivate the secondary donors Y and Z. This inactivation leads to a complete inhibition of the ability to emit oxygen. Under these conditions, the initial fast rise (less than 1 microseconds) is inhibited as shown by den Haan, G.A., Duysens, L. N. M. and Egberts, D. J. N. (1974) Biochim. Biophs. Acta 368, 409-421, and the oxidized Chl+ is reduced by an auxiliary donor D. The slow fluorescence rise observed after destruction of Y and Z has a similar kinetic behaviour to that observed in the presence of DCMU only and is polyphasic. In the presence of 10 mM hydroxylamine, the constant rate of the back reaction k1 between Chl+ and the primary acceptor Q- is estimated to be approx. (135 microseconds)-1 while the transfer of the positive charge from Chl+ to D has a rate constant k2 of approx. (105 microseconds)-1. 3. In the presence of hydroxylamine concentrations higher than 10 mM, there appears a rise in the 1-20 microseconds range ascribed to a direct reduction of oxidized Chl+ by hydroxylamine. 4. In chloroplasts treated with 10 mM hydroxylamine for 15 min and washed afterwards, the rate constant k3 of the back reaction between D+ and Q- is estimated to be approx. (100 ms)-1 which leads to a value of about 700 for the equilibrium constant between Chl and D. Hydroxylamine added under these conditions is able to reduce D+. The rate constant k4 of this reduction is estimated to be (350 ms)-1 in 0.1 mM hydroxylamine.


Time-resolved fluorescence spectroscopy of spinach chloroplast.

Picosecond fluorescent kinetics and time-resolved spectra of spinach chloroplast were measured at room temperature and low temperatures. The measurement is conducted with 530 nm excitation at an average intensity of 2-10(14) photons/cm2, pluse and at a pulse separation of 6 ns for the 100 pulses used. The 685 nm fluorescent kinetics was found to decay with two components, a fast component with a 56 ps lifetime, and a slow component with a 220 ps lifetime. The 730 nm fluorescent kinetics at room temperature is a single exponential decay with a 100 ps lifetime. The 730 nm fluorescence lifetime was found to increase by a factor of 6 when the temperature was lowered from room temperature to 90 K, while the 685 and 695 nm fluorescent kinetics were unchanged. The time-resolved spectra data obtained within 10 ps after excitation is consistent with the kinetic data reported here. A two-level fluorescence scheme is proposed to explain the kinetics. The effect of excitation with high light intensity and multiple pulses is discussed.


Distribution of excitation energy between photosystem I and photosystem II in red algae. II. Kinetics of the transition between state 1 and state 2.

The kinetics of chlorophyll fluorescence were measured during the state 1-state 2 and the state 2-state 1 transition in some members of the Florideae (Halymenia latifolia, Phycodrys rubens and Delesseria sanguinea) and in Porphyra umbilicalis as a member of the Bangiales. With the members of the Florideae it was possible to induce 70-80% of the maximum transition in direction to state 1 (obtained by us in longtime experiments) by an illumination of only a few (2-5) seconds with medium intensities of light 1. A complete transition back could be induced by a similar short illumination with light 2. The transition process itself is slow with values of tau1/2 in the range of 10 to 20 s and occurs in the dark with nearly the same speed as in continuous light 1 or light 2. The state 1-state 2 transient of chlorophyll fluorescence is kinetically clearly different from the dark-light transient phenomenon. In dark times longer than 1 min Halymenia (starting from state 1) slowly approaches state 2. This decay of state 1 is slower by a factor of about five in Halymenia than in Porphyra. This may explain, why in Halymenia, but not in Porphyra, the whole transition process can be resolved experimentally into a fast light reaction and a slow dark reaction. The results are discussed on the basis of a model which assumes the rearrangement of charges in the plane of the thylakoid membrane as a primary inducing event in the state 1-state 2 transition. This may lead to a redistribution of excitation energy among Photosystems I and II by a slow dark process, which may include a conformation change of some protein(s).


The protein-mediated net transfer of phosphatidylinositol in model systems.

The phospholipid monolayer technique has been used to study the transfer activity of the phospholipid exchange protein from beef brain. In measuring the transfer between a monolayer consisting of equimolar amounts of phosphatidylcholine and phosphatidylinositol and liposomes consisting of 98 mol% phosphatidylcholine and 2 mol% phosphatidylinositol, the beef brain protein demonstrates an 8-fold higher transfer activity for phosphatidylinositol than for phosphatidylcholine. Under similar conditions the phosphatidylcholine exchange protein from beef liver showed a great preference for phosphatidylcholine. Phosphatidylcholine liposomes devoid of phosphatidylinositol still functioned as receptors of phosphatidylinositol when the beef brain exchange protein was present. This indicates that this protein can catalyse a net transfer of phosphatidylinopsitol. Binding of both phosphatidylinositol and phosphatidylcholine to the beef brain protein was shown.


Asymmetric binding of cytochrome b5 to the membrane of human erythrocyte ghosts.

The intact, amphipatic form of cytochrome b5 could bind to unsealed ghosts, but not to resealed ghosts, suggesting that the cytochrome could bind only to the inner (cytoplasmic) surface of the ghost membrane. This was further confirmed by the finding that the cytochrome could bind to closed, inside-out vesicles prepared from the ghosts. This asymmetric binding was not due to the exclusive localization of sialic acid and sugar chains on the outer surface of the ghosts membrane, because the cytochrome could not bind to ghosts even after enzymatic removal of these components. Although liposomes consisting of phosphatidylcholine or both phosphatidylcholine and sphingomyelin could effectively bind the cytochrome, this binding capacity was progressively decreased as increasing amount of cholesterol was included in the composition of phosphatidylcholine liposomes. Removal of cholesterol from resealed ghosts by incubation with egg phosphatidylcholine liposomes resulted in the binding of cytochrome b5 to the outer surface of the treated ghosts. The possibility is discussed that the asymmetric binding is due to preferential localization of cholesterol in the outer leaflet of the lipid bilayer that constitutes the ghost membrane.


Glycopeptide fractions prepared from purified central and peripheral rat myelin.

Myelin was purified from rat brain and sciatic nerve after in vivo labeling with [3H]fucose and [14C]glucosamine to provide a radioactive marker for glycoproteins. The glycoproteins in the isolated myelin were digested exhaustively with pronase, and glycopeptides were isolated from the digest by gel filtration on Bio-Gel P-10. The glycopeptides from brain myelin separated into large and small molecular weight fractions, whereas the glycopeptides of sciatic nerve myelin eluted as a single symmetrical peak. The large and small glycopeptide fractions from central myelin and the single glycopeptide fraction from peripheral myelin were analyzed for carbohydrate by colorimetric and gas liquid chromatographic techniques. The glycopeptides from brain myelin contained 2.4 micrograms of neutral sugar and 0.59 micrograms of sialic acid per mg total myelin protein, whereas sciatic nerve myelin glycopeptides contained 10 micrograms of neutral sugar and 3.8 micrograms of sialic acid per mg total protein. Similarly, the gas-liquid chromatographic analyses showed that the glycopeptides from peripheral myelin contained 4- to 7-fold more of each individual per mg total myelin protein than those from central myelin. Most of the sialic acid and galactose in the glycopeptides from central myelin were in the large molecular weight fraction, and the small molecular weight glycopeptides contained primarily mannose and N-acetylglucosamine. The considerably higher content of glycoprotein-carbohydrate in peripheral myelin supports the results of gel electrophoretic studies, which indicate that the major protein in peripheral myelin is glycosylated while the glycoproteins in purified central myelin are quantitatively minor components.


Potassium release from submandibular salivary gland in vitro.

Rat submandibular gland slices, incubated in continuously-gassed Krebs-Ringer bicarbonate buffer, were shown to release K+ in response to alpha-adrenergic and muscarinic cholinergic stimulation. The system employed the specific alpha-, beta-adrenergic and cholinergic receptor-blocking agents phentolamine, propranolol and atropine, respectively, in combination with the agonists L-epinephrine and carbamylcholine both of which required the presence of Ca2+ for their effect. The introduction of Ca2+ into the cell via the ionophore A23187, with all neurotransmitter receptors blocked, resulted in K+ release. Ouabain also allowed extensive K+ release which was in addition to, and hence independent of, that elicited by epinephrine and carbamylcholine. Ethacrynic acid, a potent inhibitor of salivary secretion in vivo, had no influence on K+ movement. K+ was released by both physalaemin and an eledoisin-related peptide independently of normal neurotransmitter receptors. The activity of the eledoisin-related peptide did not require the presence of extracellular Ca2+. The implication of cyclic GMP at some stage of K+ release was suggested by experiments with a phosphodiesterase inhibitor. The results support an hypothesis where the initial stimulus at either alpha-adrenergic or muscarinic cholinergic receptors causes an immediate permeability change such that Ca2+ enters the cells resulting in K+ release. The loss of K+ is quickly countered by the ouabain-sensitive (Na+ + K+) ATPase which would be activated by the lowered intracellular K+ levels.


Polyol permeability of the human red cell. Interpretation of glucose transport in terms of a pore.

The kinetic equations describing transport through a pore that has a binding site and that undergoes a conformational change are identical to those of a carrier model. Therefore, in order to distinguish between the two models it is necessary to test specific predictions based on detailed mechanistic models. A pore model is described in which the substrate (glucose) is able to reach the single binding site only from the outside when the pore is in conformation I and only from the inside when it is conformation II. On the basis of this model it is predicted that solutes which do not have any specific affinity for the binding site should still have a finite permeability via the glucose transport system if they are the same size or smaller than glucose. This permeability should be proportional to the volume of distribution of the solute in the pore and should therefore decrease with increasing molecular size. A geometric pore volume can be estimated from this size dependence. In order to test these predictions, the glucose-dependent permeability of a series of 4-carbon (erythritol), 5-carbon (D-arabitol, L-arabitol and xylitol) and 6-carbon (D-mannitol, D-sorbitol and myo-inositol) polyols was measured. The permeability of all the polyols is decreased by the presence of glucose and the KI of this "inhibitable" component is similar to that D-sorbose, suggesting that this component is associated with the glucose transport system. Since these observations could be explained entirely in terms of a specific affinity for a carrier binding site, they do not exclude a carrier mechanism. However, as predicted for the pore model, this "inhibitable" permeability decreased with increasing molecular size and the calculated geometric pore volume was of a size that would be expected for a cell membrane pore.


Molecular orientation in the stacked dimer form of 5"AMP in aqueous solution. A study by the NMR-desert method.

Molecular orientation of 5"-AMP in its stacked dimeric form in neutral aqueous solutions has been investigated at room temperature through the NMR-DESERT method proposed earlier by Akasaka et al. (J. Magn. Resonance, (1975) 18, 328-343). The effect of deuterium substitution of H8 of the adenine ring on the relaxation rate of H2 has become appreciable with increasing concentration of 5"-AMP, which should be attributed to the intermolecular H2-H8 interaction between adjacent adenine rings in the stacked dimer of 5"-AMP. The reciprocal sixth-power-averaged distance between H8 and H2 of the adjacent adenine rings in the stacked dimeric form obtained from the differential relaxation rate for H2 has been found to be almost constant (3.6 +/- 0.2 A S.D.) in the whole concentration range studied (0.1--1.0 M). The result has presented a direct proof of the existence of trans-stacking with a relatively large proportion (more than 60%) in the stacked dimeric form of 5"-AMP.


Identification of human DNA sequences complementary to chromosomal RNA with an improved RNA/DNA gradient hybridization technique.

Native DNA components in the human genome were identified with an improved RNA/DNA gradient hybridization technique. 3H-Labelled complementary RNA transcribed from total human DNA was hybridized to homologous DNA which had been mildly sheared to generate single-stranded ends. Unhybridized RNA was eliminated from the reaction mixture by column chromatography before the hybrids were subjected to CsCl gradient centrifugation. Nine radioactive peaks representing RNA/DNA hybrids were reproducibly found at buoyant densities of 1.687, 1.693, 1.696, 1.699, 1.702, 1.705, 1.708, 1.711 and 1.715 g/cm3. The technique was also utilized to hybridize I125-labelled human chromosomal RNA to DNA. Three distinct radioactive peaks were found at the heavy buoyant densities of 1.702, 1.705, and 1.708 g/cm3, suggesting a differential enrichment of DNA components with sequences complementary to chromosomal RNA.


RNA synthesis in isolated nuclei of Xenopus laevis emproyos.

Nuclei were isolated from tailbud embryos of Xenopus laevis and their RNA synthetic activity was studied. The activity was sensitive to actinomycin D, and required the presence of all four ribonucleotide triphosphates and a divalent cation (Mg2+ or Mn2+). The effect of the divalent cation varied depending on the presence of 0.4 M KCl: in its absence more RNA was synthesized with Mg2+ than with Mn2+, but in its presence the situation was reversed. About 50% of the RNA synthesized was sensitive to the presence of alpha-amanitin. Polyacrylamide gel electrophoresis of the labeled RNA revealed that the RNA synthesized had a very heterogeneous size distribution ranging from 40 to 4 S, with a broad peak at around 28--18 S. The RNA which was distributed at around 28--18 S and 4 S was resistant to the presence of a low dose of alpha-amanitin, suggesting its ribosomal and soluble RNA nature. The RNA which was sensitive to this drug and hence may be assumed to be messenger RNA was distributed mainly between 18 and 4 S. From these results it appears that the isolated embryonic nuclei retain their activity to synthesize all of the major classes of RNA.


Isolation and characterization of 32P-labeled mitochondrial and cytosol ribosomal RNA from germinating wheat embryos.

With the aim of preparing highly labeled material, the incorporation of [32P]-orthophosphate into mitochondral and cytosol ribosomal RNA was examined in germinating wheat embryos. Nucleic acids were extracted from mitochondria and from post-mitochondrial supernatant (cytosol) prepared from homogenates of viable embryos (8g) imbibed for 24h in a medium containing [32P]orthophosphate (100mCi). High-molecular-weight ribosomal RNA was selectively precipitated in the presence of 3M NaCl and was further resolved on sucrose density gradients and polyacrylamide gels. Both the mitochondrial and cytosol NaCl-insoluble RNA fractions were found to contain two major radioactive components, corresponding to the large (26S) and small (18S)rRNA species. On non-denaturing gels, these species had apparent molecular weights of 1.3 and 0.67 million daltons (cytosol) and 1.3 and 0.75 million daltons (mitochondrial). The individual, purified [32P]rRNA components (isolated from sucrose gradients) had specific activities of 2--3-10(6) cpm/A260 unit, and were suitable for analysis of nucleotide composition and sequence. By hydrolyzing the individual [32P]rRNA specimens with purified snake venom phosphodiesterase and resolving the products by two-dimensional paper chromatography, it was possible to determine the specific activities (cpm/micronmol) of the four major 5'-nucleotide constituents. The results indicated that there had been no differential 32P-labeling of the nuclear and mitochondrial pools of ribonucleoside 5'-triphosphates (rNTP) during the 24 h imbibition period; however, as previously observed in this system (Lau, R.Y., Kennedy, T.D. and Lane, B.G. (1974) Can. J. Biochem. 52, 1110--1123), there had been unequal 32P-labeling of the individual rNTPs in both the mitochondria and nucleus. The relative specific activities of the 5'-nucleotide constituents of the mitochondrial and cytosol rRNA species were essentially the same, and in the order pA congruent to pU greater than pG greater than pC. By making suitable corrections for these differences in specific activity, the nucleotide composition of each of the [32P]rRNA specimens could be calculated...


The action of N-hydroxy-2-acetylaminofluorene on the synthesis of ribosomal and poly (A)-RNA in normal and regenerating liver.

Polyribosomal RNA prepared from normal and regenerating liver was chromatographed on poly(U)-Sepharose to obtain rRNA and poly(A)-RNA. Under the conditions employed for labeling the RNA with [5-3H]orotic acid, poly (A)-RNA from regenerating liver represented 1--3% of the total RNA content, and 14--18% of the total radioactivity in polyribosomal RNA at varying times after partial hepatectomy. The specific activity of poly (A)-RNA was elevated by 50% and 30% only at 2 h and 6 h after partial hepatectomy, respectively. Ribosomal RNA synthesis increased 2.5--4 fold at 2--20 h after operation. Treatment of normal or partially hepatectomized rats with N-hydroxy-2-acetylaminofluorene showed a differential dose-dependent effect on rRNA and poly (A)-RNA synthesis with rRNA synthesis being more sensitive to the inhibitory effect of the carcinogen. No change in the poly(A) content was apparent as a result of N-hydroxy-2-acetylaminofluorene treatment.


Genetic effects of photoadducts and photocross-links in the DNA of phage lambda exposed to 360 nm light and tri-methylpsoralen or khellin.

The furocoumarin, 4,5',8-trimethylpsoralen, sensitizes cells and viruses to 360 nm light, producing cross-links and monoadducts in their DNA. The furanochromone khellin is a less effective sensitizing agent than psoralen, but has been found to induce cross-links and adducts in DNA also. The number of cross-links increases as the square of the time of exposure to light. We found that greater fluences were required for khellin than for psoralen, possibly because of the less favorable angle of the distal unsaturated bonds for corss-linking pyrimidines in adjacent base pairs. By adjusting the time of exposure to 360 nm light, lambda phages were damaged with [3H]psoralen and [3H]khellin so as to produce equal numbers of cross-links. These exposures were found to produce 8-times more [3H]khellin than [3H]psoralen adducts in the DNA of the phages. Similar exposures were made with nonradioactive photosensitizers to determine the effectiveness of lambda phages carrying cross-links and monoadducts in producing genetic recombinants. Lambda phage-prophage genetic corsses were performed with psoralen and khellin-damaged phages under repressed conditions in which replication of the damaged DNA was blocked. It was estimated from the results that cross-links were about 20-times more effective than monoadducts for inducing recombination under repressed conditions. In tests on the survival of plaque forming ability on wild type bacteria, it was estimated that cross-links were about 15-times more effective than the adducts. The results support the conclusion that, in homoimmune crosses with psoralen-damaged lambda phages infecting wild type lysogens, more than three-quarters of the induced recombination can be attributed to cross-links rather than to monoadducts.


Mitochondrial DNA polymerase from rat liver.

A DNA-dependent DNA polymerase from rat liver mitochondria was partially purified and characterized. Mitochondrial DNA polymerase has been found to be quite different from other DNA-dependent DNA polymerases alpha and beta present in the rat liver in the following points: elution patterns in a DEAE-cellulose column chromatography, sedimentation coefficients determined by the glycerol gradient centrifugation in the presence of high salt, and sensitivities to N-ethylmaleimide, ethidium bromide and KCl.