Cationic lipids enhance cellular uptake and activity of phosphorothioate antisense oligonucleotides.

We have investigated the use of a cationic lipid preparation to enhance antisense oligonucleotide activity in human umbilical vein endothelial cells. A liposomal preparation containing the cationic lipid N-[1-(2,3-dioleyloxy)propyl]-N,N,N-trimethylammonium chloride (DOTMA) was found to increase by at least 1000-fold the potency of an antisense oligonucleotide (ISIS 1570) that hybridizes to the AUG translation initiation codon of human intercellular adhesion molecule-1. In the presence of 8 microM DOTMA, 6-15-fold more 35S-ISIS 1570 associated with cells, at oligonucleotide concentrations from 0.01 to 5 microM, than did in the absence of DOTMA. Both 35S-ISIS 1570 association with cells and antisense activity were increased as a function of DOTMA concentration and with increasing time of incubation with the cationic lipid. Fluorescein-labeled ISIS 1570 was used to assess the intracellular distribution of the oligonucleotide in the presence and absence of DOTMA. In the absence of DOTMA, the oligonucleotide localized to discrete structures in the cytoplasm of the cell, resulting in a punctate fluorescence pattern. In the presence of DOTMA, cellular fluorescence markedly increased and the oligonucleotide localized within the nucleus, as well as to discrete structures in the cytoplasm. Accumulation of the oligonucleotide in the nucleus in the presence of DOTMA was time and temperature dependent. Nuclear accumulation was inhibited by preincubation of the cells with monensin but not chloroquine, NH4Cl, nocodazole, colcemid, or brefeldin A. These data demonstrate that cationic lipids increase antisense activity by increasing the amount of oligonucleotide associated with cells and altering intracellular distribution of the oligonucleotide.     

Ictal brain imaging in presurgical evaluation of patients with medically intractable complex partial seizures

At the Indiana University Medical Center, 99 patients with medically intractable complex partial seizures (MI-CPS) had presurgical evaluation with subsequent anterior temporal lobectomy. The majority of the patients had single photon emission tomography (SPECT) performed interictally as well as during an actual epileptic seizure (ictal scan). Decreased regional cerebral perfusion (rCP) was seen in 54/94 (57%) of the interictal scans corresponding to the eventual site of the surgery. However, ictal scans provided a higher yield; increased rCP in the temporal lobe during an actual seizure was observed in 60/82 (73%) concordant to the side of surgery. SPECT is a useful, noninvasive method of localizing the epilepti-form focus in patients with MI-CPS considered for resective surgery. Both interictal and ictal SPECT need to be performed; combined interictal hypo-perfusion and ictal hyperperfusion in the same focal area are unique to epileptogenic lesions. Ictal SPECT studies can be performed in the majority of patients during the period of continuous video/EEG monitoring with only a little additional effort. Combining the results of functional brain imaging (interictal and ictal SPECT, PET) with clinical semiology of seizures, surface and sphenoidal EEG, magnetic resonance imaging and other non-invasive tests, anterior temporal lobectomy can be recommended in approximately two-thirds of the patients without resorting to potentially dangerous intracranial EEG monitoring.     

An Unusual Form of Skin Tuberculosis Following B.C.G. Vaccination

A 25-year-old female has had brown to erythematous telangiectatic patches and grouped papules on her face, neck, arm, and trunk since childhood following B.C.G. vaccination. Histopathologically, the lesions consisted of hyperkeratosis, slight acanthosis, tuberculoid granulomas with some Langerhans type giant cells in the mid-dermis. Although various forms of cutaneous tuberculosis after B.C.G. vaccination have been reported, it was difficult for us to assign the patient's skin lesion to any specific disease entity. Remission of her cutaneous lesions occurred clinically and histopathologically after treatment with isoniazid and rifampin.     

Long-term results and complications of preoperative radiation in the treatment of rectal cancer

A retrospective study of 149 patients with rectal cancer diagnosed between 1972 and 1979 was undertaken to compare survival, disease-free survival, recurrence sites, and long-term complications of 40 patients who received 4000 to 4500 rads of preoperative adjuvant radiotherapy (radiation group) with those of 109 patients treated by resection alone (control group). After a mean follow-up of 84 months and 99 months, respectively, survival of the irradiated patients was significantly better than that of controls (68% versus 52%, p less than 0.05). Disease-free survival of those patients rendered free of disease by treatment was also superior for the irradiated group (84% versus 57%, p less than 0.005). Local recurrence without signs of distant metastases developed only one-third as often in irradiated patients (6% versus 18%). Distant metastases, alone or in combination with local recurrence, were also less common after radiation (12% versus 27%). Second primary tumors developed in 15% and 10% of the respective groups, a difference that was not statistically significant. When we consider the survival benefit of preoperative radiation therapy, long-term complications were relatively mild. Delayed healing of the perineum was noted in two irradiated patients. Persistent diarrhea was severe enough to warrant treatment in only one case, and one patient required a colostomy for intestinal obstruction from pelvic fibrosis.     

Reduction of flurazepam's antiseizure efficacy persists after stress.

Twenty-four hours after mice were forced to swim for up to 10 min in cold (6 degrees C) water, the ability of flurazepam to antagonize the electrical precipitation of seizures was reduced. This stress-induced reduction in flurazepam's antiseizure efficacy persisted for at least 72 h; but was absent 1 week after the single session of swim stress. The data may be relevant to stress-related psychiatric disorders and suggest that the therapeutic efficacy of benzodiazepines may be altered after a severe stress.     

Recognition and management of patients with high-risk vesicovaginal fistulas: Implications for teaching and research

Vesicovaginal fistulas (VVFs) occurring as a result of obstetric trauma are a vast problem in Nigeria and Ghana, where at least 20 000 women await repair, and fewer than 50 physicians have the necessary expertise. Through a series of conferences those VVFs that are at high risk and those at low-risk for repair failure, were identified. A clinic was established where repair of low-risk VVFs was done on an ongoing basis in a remote region of Ghana. A visiting surgical team was utilized to repair the difficult, or high-risk, VVFs, which included 4–6 cm VVFs (3), recurrent VVF (1), combined VVF and RVF (rectovaginal fistula), a large 5 cm juxtacervical VVF (1), and a vesicouterine fistula (1). Management of these patients and others with VVF repair complications is discussed.     

How accurate was GMENAC?--A retrospective review of supply projections for diagnostic radiologists.

In 1982, the Graduate Medical Education National Advisory Committee (GMENAC), a prominent national panel, predicted there would be 25,650 full-time equivalent (FTE) diagnostic radiologists, a 34% oversupply, by 1990. The radiologists involved in GMENAC, however, using models developed by the American College of Radiology, projected 19,800 FTE diagnostic radiologists in 1990, which was similar to the GMENAC estimate of need. The disagreement arose principally from different assumptions about residents entering the specialty. Recent data show there actually were approximately 21,900 FTE diagnostic radiologists in 1990. The radiologists' projection was 10% below this figure; the GMENAC projection was 17% above it. GMENAC erred principally in assuming diagnostic radiology residencies would not replace general radiology residencies, but rather be an addition to them. The radiologists erred principally in their assumption about the effects of the financial problems of hospitals on the number of residency positions. Accurate long-term projection of physician supply in individual specialties may well not be feasible.     

α-Bungarotoxin blocks the nicotinic receptor mediated increase in cell number in a neuroendocrine cell line

Exposure of H69 small cell lung carcinoma cells to nicotinic agonists resulted in a significant increase (up to 100%) in cell number after 6 to 12 days. The effect of nicotine (10⁻⁸ M to 10⁻⁴ M) was both dose and time dependent as was that of another nicotinic agonist cytisine (10⁻⁶ M to 10⁻⁴ M). Interstingly, both the nicotine and cytisine induced increases in H69 cell number were blocked by α-bungarotoxin, as well as d-tubocurarine a nicotinic blocker which appears to interact with most nicotinic receptors. These results suggest that the nicotine induced increase in cell number is mediated through an interaction at the nicotinic α-bungarotoxin receptor. This idea is further supported by experiments which show (1) that H69 cells possess high affinity α-bungarotoxin sites (K_d = 25 nM, B_max = 10.4 fmol/10⁶ cells) with the characteristics of a nicotinic α-bungarotoxin receptor and (2) that the potencies of nicotinic receptor ligands in the α-bungarotoxin binding assay were similar to those observed in the functional studies. Northern analysis showed that mRNA for α7, a putative nicotinic α-bungarotoxin binding subunit, and for α5 were present in H69 cells. The present data provide further evidence that nicotine increases cell number in small cell lung carcinoma and are the first to show that this effect is mediated through an interaction at the nicotinic α-bungarotoxin receptor population. These results suggest that the α-bungarotoxin site may be involved in modulating proliferative responses in neuroendocrine derived SCLC cells.     

Plasma Histamine After Methacholine, Allergen, and Aspirin Challenges

Plasma histamine levels were measured by radio-enzymatic technique in seven patients following 10 challenges: five methacholine challenge tests, four antigen inhalation challenge tests, and one oral aspirin challenge test. Baseline plasma histamine was the same in all patients except in the aspirin-challenged patient, who had a higher baseline histamine level. There was no statistical change in the level of histamine throughout the test in either the methacholine-challenged or the antigen-challenged patients, whereas there was a marked increase in histamine levels in the aspirin challenged patient. A possible explanation is that methacholine and antigen are inhaled and therefore have primarily local effects on the lung, whereas oral aspirin has a systemic effect with consequently systemic changes in histamine which are detectable as changes in plasma level.     

Electrophysiological properties of neurons in the rostral ventrolateral medulla of normotensive and spontaneously hypertensive rats

Single unit activities were recorded from the rostral ventrolateral medulla (RVL) of pentobarbital-anesthetized normotensive Wistar Kyoto rats (WKY) and spontaneously hypertensive rats (SHR). Throughout the recording period, arterial blood pressures of WKY (mean arterial pressure, MAP = 103.1 mm Hg) and SHR (MAP = 159.2 mm Hg) remained stable at the respective basal levels. The units recorded in this study were all spontaneously active and cardiac-locked. Two types of discharge patterns, namely single and double discharges, were identified. These single and double discharge units were found to distribute randomly in RVL. In WKY, 92.6% of RVL neurons exhibited single discharges whereas in SHR, the majority (57%) of RVL neurons exhibited double discharges. The mean firing rate of single discharge units in RVL of SHR was significantly higher than that of WKY, whereas the mean firing rate of double discharge units in WKY was similar to that of SHR. About half of the units studied were also tested for antidromic collision; all units tested could be antidromically activated from the intermediolateral column (IML) of the thoracic spinal cord and the lowest threshold sites were consistently localized within IML. In both groups of rats, the axonal conduction velocity of RVL neurons showed a bimodal distribution viz. the fast and slow conducting axons. The mean conduction velocities of each of these two groups of neurons in WKY and SHR were similar. Most of the double discharge units in WKY and SHR belonged to the fast conducting type.(ABSTRACT TRUNCATED AT 250 WORDS)