Prolonged circulation time in vivo of large unilamellar liposomes composed of distearoyl phosphatidylcholine and cholesterol containing amphipathic poly(ethylene glycol).

The effect of poly(ethylene glycol) (PEG) on the circulation time of liposomes in mice was examined by employing amphipathic PEGs (phosphatidylethanolamine (PE) derivatives of PEG) with average molecular weights of 1000, 2000, 5000 and 12,000. The activity of dioleoyl phosphatidylethanolamine-PEG (DOPE-PEG) in prolonging the circulation time of egg phosphatidylcholine/cholesterol large unilamellar liposomes (ePC/CH LUVs) (200 nm) was proportional to the molecular weight of PEG, i.e., 12000 = 5000 greater than 2000 greater than 1000. On the other hand, inclusion of distearoylphosphatidylethanolamine-PEG (DSPE-PEG) or dipalmitoyl-phosphatidylethanolamine-PEG (DPPE-PEG) of low molecular weight such as 1000 and 2000 in distearoylphosphatidylcholine (DSPC)/CH LUVs or dipalmitoyl phosphatidylcholine (DPPC)/CH LUVs effectively increased their blood circulation time. At least 3 mol% of amphipathic PEG in liposomes was required for activity. Addition of CH, which has a bilayer-tightening effect, to DSPC/CH/DSPE-PEG2000 LUVs further increased the blood residence time. A size of less than 300 nm was essential for prolonging the residence time of amphipathic PEG-containing liposomes in blood. DSPC/CH/DSPE-PEG2000 LUVs (1:1:0.13, m/m) containing 6 mol% of PEG and 200 nm in diameter remained in the circulation for over 24 h after injection and may be clinically useful for sustained release of an entrapped drug in the bloodstream and for drug accumulation in solid tumors.

Targetability of novel immunoliposomes modified with amphipathic poly(ethylene glycol)s conjugated at their distal terminals to monoclonal antibodies.

Distearoyl-N-(3-carboxypropionoyl poly(ethylene glycol) succinyl)phosphatidylethanolamine (DSPE-PEG-COOH) was newly synthesized and used to prepare novel immunoliposomes carrying monoclonal antibodies at the distal ends of the PEG chains (Type C). Liposomes were prepared from egg phosphatidylcholine (ePC) and cholesterol (CH) (2;1, m/m) containing 6 mol% of DSPE-PEG-COOH, and a monoclonal IgG antibody, 34A, which is highly specific to pulmonary endothelial cells, was conjugated to the carboxyl groups of DSPE-PEG-COOH to give various amounts of antibody molecules per liposome. Other immunoliposomes with PEG coating (Type B) or without PEG coating (an earlier type of immunoliposome, Type A) were prepared for comparison. The average molecular weight of PEG in Type B or C immunoliposomes was 2000. Type B and Type C liposomes without antibodies showed prolonged circulation time and reduced reticulo-endothelial system (RES) uptake owing to the presence of PEG. These three different types of 34A-immunoliposomes with 30-35 antibody molecules per vesicle were injected into mice to test the immunotargetability to the lung. The efficiency of lung binding of 34A-Type B was one-half of that of 34A-Type A, though a large amount of 34A-Type B remained in the blood circulation for a long time, suggesting that the steric hindrance of PEG chains reduced not only the immunospecific antibody-antigen binding, but also the RES uptake. The degree of lung binding of 34A-Type C was about 1.3-fold higher than that of 34A-Type A, indicating that recognition by the antibodies attached to the PEG terminal was not sterically hindered and that the free PEG (i.e., that not carrying antibody) was effective in increasing the blood concentration of immunoliposomes by enabling them to evade RES uptake. The latter phenomenon was confirmed by using nonspecific antibody-Type C immunoliposomes (14-Type C), which showed a high blood level for a long time. Our approach provides a simple means of conjugating antibodies directly to the distal end of PEG which is already bound to the liposome membrane, and should contribute to the development of superior targetable drug delivery vehicles for use in diagnostics and therapy.

Asymmetric intramyocardial lymphocytic accumulation during acute rejection in rat heart and heart-lung transplantation.

Lymphocytic accumulation in different areas of the myocardium during different phases of acute rejection were studied in rat heterotopic cardiac and cardiopulmonary allograft and isograft transplantation models. Indium 111-labeled syngeneic lymphocytes were injected 1 to 7 days after transplantation, and accumulation of the labeled cells in the graft was determined 24 hours later. The cardiac graft was divided into three segments: right ventricular (RV) free wall, epicardial portion of the left ventricle (LV) plus RV septum, and endocardial portion of the LV. In the subsequent heart-lung transplantation group, the LV epicardial portion plus RV septum segment was subdivided into LV epicardial portion and RV septum. Lymphocytic accumulation in each segment was compared between three groups according to the day the animal was killed after transplantation: days 2, 3 (group 1), days 4, 5 (group 2), and days 6, 7, 8 (group 3). Lymphocytic accumulation caused by rejection in group 1 and group 2 of both the cardiac and cardiopulmonary allografts showed significantly different intramyocardial distribution (p less than 0.01). Lymphocytic accumulation was consistently highest in RV free wall and lowest in LV endocardial portion. In group 3 specimens, lymphocytic accumulation was symmetrical in both the cardiac and cardiopulmonary allografts. In the cardiac allograft, the lymphocytic accumulation increased sharply on day 4 then declined to a nadir. This transient increase was not so pronounced in the cardiopulmonary allografts. The significantly decreased intramyocardial lymphocytic accumulation in group 2 of the cardiopulmonary allografts when compared with the cardiac allografts (p less than 0.01) is compatible with the clinical observation of a lower incidence of acute heart rejection in heart-lung transplant recipients. These data show an asymmetric accumulation of lymphocytes during acute heart rejection, which is due to increased lymphocytic affinity for the RV free wall.

Right atrial mural thrombus associated with pericarditis.

A 33-year-old man was hospitalized with right heart failure. He was diagnosed as having right atrial mural thrombus complicated with pericarditis on echocardiography, thoracic computed tomography, and cardiac catheterization. Pericardiectomy and thrombectomy with a partial resection of the right atrial wall were performed under extracorporeal circulation. It was suggested that the cause of right atrial thrombus was congestion, atrial fibrillation, and pericarditis. The cause of pericarditis could not be determined by pathologic examination and laboratory data. Surgical treatment should be performed as soon as possible to prevent pulmonary embolism.

Effects of intraaortic balloon position on renal artery blood flow.

Debate continues over what happens to renal blood flow when intraaortic balloons are adjacent to the renal arteries. Fourteen dogs were prepared by implanting instruments to measure heart rate; right atrial, pulmonary arterial, carotid arterial, and femoral arterial pressures; cardiac index; mixed venous oxygen saturation; urine output; and left and right renal blood flows. A 12-mL intraaortic balloon was inserted through the left (n = 9) or right (n = 5) femoral artery. The position of the balloon was randomized so that it was initially placed in either the control (thoracic) or renal position (at the level of the renal arteries). Intraaortic balloon pumping was performed for 4 hours in each position. In 8 dogs, at least one of the renal arteries had partial occlusion, 23% to 98% decrease in flow (mean decrease, 66%), while the intraaortic balloon was in the renal position. An intraaortic balloon in the renal position results in lower renal blood flow as well as a high risk (57%) of selective renal artery occlusion. Decreased renal blood flow is not apparent using conventional monitoring, as hemodynamics do not change.

A multipurpose grasping forceps for retrieval of an embolized catheter fragment.

Transjugular removal of an embolized catheter fragment from the right atrium of a 6-month-old infant was successfully performed using a multipurpose grasping forceps. All procedures for the retrieval were completed in only 10 minutes without complications or difficulty.

Pedicle transposition of the greater omentum for sternal osteomyelitis and mediastinitis after cardiac operation.

Three patients underwent pedicle transposition of the greater omentum for sternal osteomyelitis and mediastinitis after cardiac operation. These patients were previously operated on with coronary artery bypass grafting, aortic valve replacement and redo Fontan operation in each. This procedure was carried out on 4, 6, 8, and 14th postoperative day. The complications cured within about one month after the procedure in all cases. It is suggested that this maneuver is effective for obtaining healing osteomyelitis and mediastinitis following cardiac operation.

Leaf senescence under various gravity conditions: relevance to the dynamics of plant hormones.

Effects of simulated microgravity and hypergravity on the senescence of oat leaf segments excised from the primary leaves of 8-d-old green seedlings were studied using a 3-dimensional (D) clinostat as a simulator of weightlessness and a centrifuge, respectively. During the incubation with water under 1-g conditions at 25 degrees C in the dark, the loss of chlorophyll of the segments was found dramatically immediately after leaf excision, and leaf color completely turned to yellow after 3-d to 4-d incubation. In this case kinetin (10 micromolar) was effective in retarding senescence. The application of simulated microgravity conditions on a 3-D clinostat enhanced chlorophyll loss in the presence or absence of kinetin. The loss of chlorophyll was also enhanced by hypergravity conditions (ca. 8 to 16 g), but the effect was smaller than that of simulated microgravity conditions on the clinostat. Jasmonates (JAs) and abscisic acid (ABA) promoted senescence under simulated microgravity conditions on the clinostat as well as under 1-g conditions. After 2-d incubation with water or 5-d incubation with kinetin, the endogenous levels of JAs and ABA of the segments kept under simulated microgravity conditions on the clinostat remained higher than those kept under 1-g conditions. These findings suggest that physiological processes of leaf senescence and the dynamics of endogenous plant hormone levels are substantially affected by gravity.

Growth and development in higher plants under simulated microgravity conditions on a 3-dimensional clinostat.

Growth and development of etiolated pea (Pisum sativum L. cv. Alaska) and maize (Zea mays L. cv. Golden Cross Bantam) seedlings grown under simulated microgravity conditions were intensively studied using a 3-dimensional clinostat as a simulator of weightlessness. Epicotyls of etiolated pea seedlings grown on the clinostat were the most oriented toward the direction far from cotyledons. Mesocotyls of etiolated maize seedlings grew at random and coleoptiles curved slightly during clinostat rotation. Clinostat rotation promoted the emergence of the 3rd internodes in etiolated pea seedlings, while it significantly inhibited the growth of the 1st internodes. In maize seedlings, the growth of coleoptiles was little affected by clinostat rotation, but that of mesocotyls was suppressed, and therefore, the emergence of the leaf out of coleoptile was promoted. Clinostat rotation reduced the osmotic concentration in the 1st internodes of pea seedlings, although it has little effect on the 2nd and the 3rd internodes. Clinostat rotation also reduced the osmotic concentrations in both coleoptiles and mesocotyls of maize seedlings. Cell-wall extensibilities of the 1st and the 3rd internodes of pea seedlings grown on the clinostat were significantly lower and higher as compared with those on 1 g conditions, respectively. Cell-wall extensibility of mesocotyls in seedlings grown on the clinostat also decreased. Changes in cell wall properties seem to be well correlated to the growth of each organ in pea and maize seedlings. These results suggest that the growth and development of plants is controlled under gravity on earth, and that the growth responses of higher plants to microgravity conditions are regulated by both cell-wall mechanical properties and osmotic properties of stem cells.

[Acute ventricular septal perforation in a patient with autoimmune hemolytic anemia].

A 71-year-old woman with autoimmune hemolytic anemia underwent an emergency endocardial patch repair for ventricular septal perforation after acute myocardial infarction. Use of washed red blood cells was effective in averting hemolytic crisis throughout perioperative period. In spite of improvement of her hemodynamics, liver dysfunction which had been present preoperatively deteriorated after the operation. Finally she died of hepatic failure on the 21st postoperative day. Deterioration of liver function could not be associated with autoimmune hemolytic anemia. To date, little information is available concerning the influence of cardiopulmonary bypass on hemolysis in patients with autoimmune hemolytic anemia. Therefore, prudent management and use of washed red blood cells transfusion would prevent hemolytic aggravation even in open heart surgery.

STS-95 space experiment for plant growth and development, and auxin polar transport.

The principal objective of the space experiment, BRIC-AUX on STS-95, was the integrated analysis of the growth and development of etiolated pea and maize seedlings in space, and the effect of microgravity conditions in space on auxin polar transport in the segments. Microgravity conditions in space strongly affected the growth and development of etiolated pea and maize seedlings. Etiolated pea and maize seedlings were leaned and curved during space flight, respectively. Finally the growth inhibition of these seedlings was also observed. Roots of some pea seedlings grew toward the aerial space of Plant Growth Chamber. Extensibilities of cell walls of the third internode of etiolated pea epicotyls and the top region of etiolated maize coleoptiles which were germinated and grown under microgravity conditions in space were significantly low. Activities of auxin polar transport in the second internode segments of etiolated pea seedlings and coleoptile segments of etiolated maize seedlings were significantly inhibited and extremely promoted, respectively, under microgravity conditions in space. These results strongly suggest that auxin polar transport as well as the growth and development of plants is controlled under gravity on the earth.

[Total anomalous pulmonary venous connection associated with Klippel-Feil syndrome: a case report].

Klippel-Feil syndrome is characterized by congenital fusion of the cervical vertebrae. Anomalies in other organs sometimes combine. Many kinds of congenital cardiac anomalies, most frequently ventricular septal defect, considerably associate with this syndrome. Few cases of surgical corrections of cardiac anomalies with Klippel-Feil syndrome have been reported up to today. Therefore surgical problems with this syndrome have not been solved yet, completely. We have experienced a very rare case of a 18-year-old female of total anomalous pulmonary venous connection (Darling Ia type) associated with Klippel-Feil syndrome, that was repaired successfully. No difficulty in the surgical and anesthesiological management referring to the fusion of cervical vertebrae was encountered. The result of surgical correction of congenital cardiac anomalies in Klippel-Feil syndrome mainly depends on the severity of the heart disease and not on Klippel-Feil syndrome itself, as literature and the present case suggest.

[Nonsurgical retrieval of intravascular catheter fragments].

The incidence of catheter fragment induced by iatrogenic accident has been increasing because of frequent use of intravenous catheters and catheterization in daily practice. Migrated intravascular fragments from broken catheters were retrieved transcutaneously in four cases. All fragments were removed safely using biotome, multipurpose grasping forceps and basket catheter. But in a case which the fragment was lodged in the pulmonary artery, retrieval was not easy and it took many times. Fragments lodged in the right ventricle and in the pulmonary artery cause many complications including deaths. So broken fragments should be removed as soon as possible, before it migrated to there.

[Traumatic cardiac tamponade due to the nail stabbing].

A 45-year-old man was admitted to our hospital complaining of mild chest pain. The stab wound on the chest was very small and the nail had been completely buried under the skin. Emergency operation was mandatory for cardiac tamponade by the nail stabbing. Contused right ventricular wall was repaired using Teflon felt under cardiopulmonary bypass. It was important to have a suspicion as cardiac tamponade, when the patient showed uncertain hypotension.

[Noonan's syndrome associated with cardiovascular anomalies; report of three cases].

Noonan's syndrome has a phenotype similar to Turner's syndrome, but has no chromosome abnormalities. This syndrome is frequently associated with congenital heart diseases, particularly valvular pulmonary stenosis. Three patients of Noonan's syndrome with congenital heart disease were successfully operated upon. Diagnostic criteria, frequency of anomalies and the association of cardiac malformations have been discussed together with special surgical points of view.

[Simplified technique for closure of median sternotomy incision: the use of bone stapler].

A new technique with the bone stapler (Staplizer) was applied in 11 patients undergoing open-heart surgery for closure of a median sternotomy incision. The staplers were used for fixation of the sternum following partial trans-sternal fixation with two wires. The method was quick and easy to perform, and effective in establishing firm and permanent approximation of the sternum. There were no instances of dehiscence among all the patients with stapler closure. We therefore recommend this technique in median sternotomy closure except for patients with the osteoporotic or elastic sternum.

[Perforation and localized dissection of the left common iliac artery during the modified Bentall procedure: a case report].

We performed the modified Bentall procedure for annuloaortic ectasia in a 41-year-old man with Marfan's syndrome. Approximately 15 minutes after initiation of cardiopulmonary bypass with an arterial delivery via the left common femoral artery, perfusion volume gradually decreased and the abdomen of the patient was distended. Retrograde dissection was strongly suggested. Perforation and localized dissection of the left common iliac artery were recognized. Following repair of the lesion, the modified Bentall operation was carried out. Postoperative course was uneventful. The patient has been well for 22 months after the operation.

[Surgery for ventricular septal defect with aortic regurgitation].

Twenty-two patients (mean age of 12.9 years) with ventricular septal defect (VSD) associated with aortic regurgitation (AR) were treated surgically. Sixteen patients had subpulmonic VSD and six had infracristal VSD. Fourteen patients with subpulmonic VSD underwent VSD closure alone. VSD was closed by pulling up the inferior rim to the pulmonary valve thus support the prolapse aortic cusp. AR improved in two and has remained stable in eleven for 0.5 to 12.7 years (mean 4.4 years). Aortic valve replacement (AVR) was necessary in an adult patient later. Plication of the aortic valve was effective in a young patient but ineffective in an adult patient. In infracristal VSD, AVR was necessary in two adult patients. In a young patient, AR improved by plication. Direct VSD closure ceased progression of AR in three patients. The long-term follow-up shows that VSD closure with our technique has been sufficient to arrest progression of AR in subpulmonic VSD. Early closure of VSD should be emphasized to prevent aortic valve prolapse and regurgitation.

[Nonsurgical removal of the embolized catheter fragments from the heart and great vessels].

Embolized catheter fragments in the heart and great vessels were successfully removed in seven patients by nonsurgical techniques. All procedures for the retrieval were completed without complications or difficulty using instruments such as multipurpose grasping forceps, stone basket, loopsnare and myocardial biopsy catheters. Embolized catheter fragments should be removed promptly before fatal complications develop, and can be taken off safely and simply by nonsurgical technique.

[Myocardial protection with cold blood potassium cardioplegia: experimental and clinical studies].

One hundred and eighty-one patients (101 men and 80 women) underwent 61 congenital and 120 acquired heart operations using cold blood potassium cardioplegia as the method of myocardial protection at Kagoshima University from August, 1978 to August, 1981. Hospital deaths occurred in 18 patients (9.8%), and 4 cases were late death (2.2%). Multivariate analysis revealed no significant relationship between type of heart disease and operative mortality or the occurrence of post-operative low output syndrome. To evaluate the superiority of cold blood cardioplegia for myocardial protection, we carried out 20 orthotopic heart transplants in dog; 16 transplanted grafts could maintain the circulation of recipient without cardiopulmonary bypass. Cardiac contractility following 90-minute ischemic time showed 97 to 100% of the control values without any inotropic drugs. These clinical and experimental findings suggest that cold blood potassium cardioplegia provides excellent myocardial protection during the ischemia of the heart.