Serum lipase: a better test to diagnose acute alcoholic pancreatitis.

To determine whether serum lipase is a better test than serum amylase to diagnose acute alcoholic pancreatitis. Two hundred two asymptomatic chronic alcoholics (Group A) and 29 patients with image-proven pancreatitis (Group P). Serum lipase was measured using the Kodak Ektachem clinical chemistry slide. Serum amylase was estimated using the Kodak Ektachem clinical chemistry slide or the Beckman Astra amylase chemistry module. The level of serum amylase in Group A ranged from 17 to 347 U/L (mean 71, SD +/- 36 U/L) and in Group P from 180 to 2,985 U/L (mean 722, SD +/- 663 U/L). Thirteen of 29 patients (45%) with image-proven pancreatitis had levels that overlapped those found in asymptomatic alcoholics. The serum lipase levels in Group A ranged from 34 to 600 U/L (mean 186, SD +/- 111 U/L), while in Group P, the corresponding figures were 1,011 to 25,706 U/L (mean 5,822, SD +/- 5,664 U/L). None of the 29 patients with image-proven pancreatitis had levels that overlapped those found in asymptomatic alcoholics. Serum lipase is a better test that serum amylase to diagnose acute alcoholic pancreatitis.


Dual role of tumor necrosis factor-alpha in angiogenesis.

The role of tumor necrosis factor-alpha (TNF; cachectin) in angiogenesis has been controversial. In vitro TNF inhibits proliferation of endothelial cells (EC) whereas in the cornea it appears to stimulate vessel growth. The authors tested TNF in their recently developed disc angiogenesis system (DAS), designed to measure the proliferation of microvessels. The DAS, implanted subcutaneously in mice, was either fitted with a central pellet containing mouse recombinant TNF (mrTNF), or exposed to mrTNF delivered subcutaneously by an osmotic minipump. Low doses of mrTNF (0.01-1 ng) induced angiogenesis, which was maximum at 0.1 ng, whereas high doses (1, and 5 micrograms) inhibited it. Subcutaneous mrTNF delivered at the (high) rate of 15-60 ng/hr for 14 days inhibited angiogenesis. These results indicate bimodal, dose-dependent opposing effects and explain some of the in vitro versus in vivo paradoxical results. TNF (native or exogenous) may have opposing effects on microvessels of neoplasms and inflammatory reactions, depending on its local tissue concentrations.


Intermediate filaments as differentiation markers of normal pancreas and pancreas cancer.

Expression of intermediate filaments (IF) is regulated during development and differentiation. The authors have studied the expression of vimentin and cytokeratins (CK) 4, 7, 8, 13, 18, 19 in normal pancreas, chronic pancreatitis, and pancreas cancer using monoclonal antibodies. Immunohistochemical assays were performed on fresh frozen tissue sections and on cultured pancreas cancer cells using the streptavidin-peroxidase method. In normal pancreas, acinar cells expressed CK 8 and 18, whereas ductal cells expressed CK 7, 8, 18, and 19. CK 4 was expressed by 5-10% of pancreas duct cells in all specimens of normal pancreas. CK 13 was not detected in any epithelial cells of normal pancreas or pancreatitis. CK 7, 8, 18, and 19 were homogeneously expressed in all pancreas cancers, whereas CK 4 was expressed only in 5-50% of cells in 10/16 tumors. Foci of squamous metaplasia expressed CK 13 but showed partial loss of expression of CK 7, 8, 18, and 19. Thirteen pancreas cancer cell lines examined showed homogeneous expression of CK 7, 8, 18, and 19; 2/11 lines expressed CK 4 weakly, and 6/11 expressed vimentin. CK 13 was not detected in any of the lines. These results indicate that pancreas cancer cells consistently express cytokeratin polypeptides characteristic of ductal epithelial cells and that this phenotype is retained in pancreas cancer cell lines. In addition, squamous metaplasia is associated with a coordinate change in the expression of CK polypeptides.


Basal-cell keratins in cervical reserve cells and a comparison to their expression in cervical intraepithelial neoplasia.

Expression of keratins 5, 14 and 17 in endocervical subcolumnar reserve cells was detected by means of immunohistochemical studies using polypeptide specific monoclonal antibodies. These particular keratins that were found among others in basal cells could also be detected to a variable extent in metaplastic and dysplastic cervical lesions. In some cases of immature squamous metaplasia all three keratin subtypes were expressed throughout the full thickness of the epithelium. In contrast, in mature squamous metaplasia a compartmentalization of these keratins was observed. Mature squamous metaplastic epithelium showed a keratin distribution pattern comparable to ectocervical squamous epithelium, with the exception of keratin 17, which was only sporadically found in the basal layer of ectocervical epithelium and was always present in the basal cells of mature squamous metaplastic epithelium. During progression of cervical intraepithelial neoplasia a clear increase in the expression of keratin 17 was observed. However, also keratins 5 and 14 were expressed. Our results demonstrate that a considerable number of premalignant lesions of the uterine cervix express the same keratins as found in the progenitor reserve cells. Lesions that lack expression of keratin 17 may form a distinct group, which are regressive in nature and do not progress into cervical cancer.


Immunohistochemical evidence of oxidative [corrected] stress in Alzheimer's disease.

Membrane and cytoskeletal structures are known targets of oxidative injury. Brains from patients with Alzheimer's disease have cytoskeletal abnormalities and platelet and possible neuronal membrane lesions. The authors have recently demonstrated that superoxide anion is a powerful inducer of heat-shock protein synthesis, and have also shown that in response to oxidative stress or hyperthermia, intracellular levels of antioxidant enzymes increase to several folds. Whether the aforementioned mechanisms play a role in Alzheimer's disease has been suggested but is not totally established. While exploring this possibility, tissue sections from five brains with Alzheimer's disease and five neuropathologically normal age-matched controls were immunostained with polyclonal antibodies against superoxide dismutase (CuZn- and Mn- forms) and catalase. A standard avidin-biotin-peroxidase method was used for antigen detection. A subgroup of neurofibrillary tangles (15-25%) and senile plaques (50%) showed immunoreactivity for both enzymes with a staining pattern similar (but not identical) to that usually observed with antibodies against ubiquitin. Senile plaques displayed a granular pattern of immunostaining. Amyloid cores in mature classical plaques remained unstained. In addition, occasional elements with features consistent with reactive glial cells were strongly immunostained. Tangle-free neurons in both diseased and control brains showed weak to absent intracytoplasmic immunoreactivity. The immunoreactivity was totally abolished by preincubation of the primary antibodies with the corresponding purified antigens. These findings support the hypothesis that oxidative stress may be involved in the pathogenesis of Alzheimer's disease.


Expression of platelet-derived growth factor and its receptors in proliferative disorders of fibroblastic origin.

Platelet-derived growth factor (PDGF) is known to stimulate the proliferation of connective tissue-derived cells in vitro. Less is known about its functions in vivo, and the role of PDGF in the development of human tumors has not been clarified. The authors have investigated the occurrence of PDGF and PDGF receptors in a series of proliferative disorders of fibroblastic origin using immunohistochemical and in situ hybridization techniques. High expression of PDGF beta-receptor mRNA and protein was found in the malignant tumors, and also in some benign lesions, such as dermatofibroma. In all these cases, benign as well as malignant, the PDGF B-chain mRNA, and less clearly, the PDGF A-chain mRNA, were coexpressed with the beta-receptor. In contrast, high expression of PDGF alpha-receptor mRNA was only found in fully malignant lesions, i.e., malignant fibrous histiocytoma. These data indicate that an autocrine growth stimulation via the PDGF beta-receptor could occur in an early phase of tumorigenesis, and may be a necessary but insufficient event for the progression into fully malignant human connective tissue lesions.


Biologic and immunohistochemical analysis of interleukin-6 expression in vivo. Constitutive and induced expression in murine polymorphonuclear and mononuclear phagocytes.

Interleukin-6 (IL-6) is considered an important multifunctional cytokine involved in the regulation of a variety of cellular responses, including the induction of acute-phase protein synthesis, lymphocyte activation, and hematopoiesis. In vitro studies have identified many cells that can produce IL-6, but the cellular sources under physiologic conditions have yet to be identified. Using immunoaffinity purified goat anti-murine IL-6, the authors performed immunohistochemical studies to localize cells expressing IL-6 in selected organs of normal and endotoxin challenged NIH-Swiss outbred mice. In the blood, findings were correlated with cell-associated bioactivity using the standard B9 cell proliferation assay. In normal mice, constitutive expression was seen in granulocytes, monocytes and their precursors as well as in bone marrow and splenic stromal macrophages. Hepatic macrophages were negative, as were lymphocytes, megakaryocytes, erythroid precursors, and endothelial cells. In the absence of significant serum levels of IL-6, cell-associated IL-6 bioactivity was detected in circulating polymorphonuclear leukocytes (PMNs), but not lymphocytes. After endotoxin challenge, there was a threefold increase in PMN IL-6 content from 1 to 3 hours followed by almost complete depletion at 6 hours. This correlated well with a threefold increase of IL-6 mRNA in the bone marrow followed by a decrease at 6 hours. This pattern also correlated with serum levels of IL-6, which peaked at 3 hours and dropped significantly by 6 hours. By 24 hours, cell-associated IL-6 showed recovery with no increase in serum levels. In vivo findings showing IL-6 expression in bone marrow macrophages support in vitro studies suggesting a role for IL-6 in hematopoiesis. Furthermore, PMNs as well as macrophages are likely important sources of IL-6 during inflammatory and septic states.


Expression of intercellular adhesion molecule-1 in atherosclerotic plaques.

Immunohistochemistry of human atherosclerotic arteries demonstrates expression of the intercellular adhesion molecule-1 (ICAM-1) on endothelial cells, macrophages, and smooth muscle cells of the plaques. Normal arterial endothelial cells and intimal smooth muscle outside plaques give weaker or negative reactions; these differ from the strong endothelial expression in small vessels. Quantitative color-image analysis of the endothelial layer shows increased expression of ICAM-1 in all subtypes of atherosclerotic lesions, except fibrous plaques. Endothelial expression of ICAM-1 may be involved in the recruitment of monocytes to the lesion, as suggested by its role in the entry of leukocytes, including monocytes, into foci of inflammation. Collaboration with other mechanisms, particularly chemoattractant factors, may be important for this effect. ICAM-1 enhanced monocyte recruitment is a potential mechanism for the growth of an atherosclerotic plaque.


Vitronectin and its relationship to other extracellular matrix components in bronchoalveolar lavage fluid in sarcoidosis.

There is a need to find markers that can be used as indicators of early fibrotic changes in the lung in patients with sarcoidosis. The fibrotic reaction is accompanied by an increase in the connective tissue components, and the extracellular matrix molecules are characterized by an ability to interact with each other. We found increased concentrations of three components of the extracellular matrix, vitronectin (VN), fibronectin (FN), and hyaluronan (HA), in bronchoalveolar lavage (BAL) fluid from 56 patients with sarcoidosis compared with 38 healthy control subjects (p less than 0.001 for all). Using an enzyme-linked immunosorbent assay, the median value for VN in BAL fluid from sarcoid patients was 74 micrograms/L (interquartile range, 47 to 138) compared with 38 micrograms/L (IQR, 22 to 55) in control subjects. The median VN concentration in serum was 0.25 g/L in both groups. VN consists of various functional domains, and it may, together with FN and HA, contribute to repair or exaggeration of the interstitial changes that occur when sarcoidosis affects the lungs. VN correlated to the concentration of albumin in the BAL fluid (p less than 0.01) but even closer to the concentrations of FN and HA (p less than 0.001 for both). The extracellular matrix components did not show any correlation to the disease activity, roentgenographic stage, or functional signs of developed fibrosis. In conclusion, the increased concentrations of VN, FN, and HA may predict only an ongoing inflammation and not necessarily a fibrotic process.


c-myc protein distribution. Neoplastic tissues of the human colon.

There is an extensive literature documenting the increased or deregulated expression of the c-myc oncogene in human malignancies. The authors have recently devised a sensitive immunocytochemical method for studying the tissue localization of c-myc protein in tissue sections of human colon. We have compared nuclear c-myc staining using a polyclonal rabbit anti-c-myc antibody and a mouse monoclonal myc antibody NCM II 274. Microscopic observation of the tissue specific pattern of c-myc protein distribution shows that nuclear staining intensity varies in normal and neoplastic crypt cell nuclei in parallel with morphologic criteria of neoplasia. These studies yield further information on the usefulness of c-myc protein as a prognostic indicator.


Administration of alpha 1-proteinase inhibitor ameliorates bleomycin-induced pulmonary fibrosis in hamsters.

The effect of alpha 1-proteinase inhibitor (alpha 1Pi) administration on the acute lung injury and subsequent fibrosis induced by bleomycin (BLM) was examined in hamsters. Pulmonary lesions were quantitatively reduced in alpha 1Pi-administered BLM-treated (BLM-alpha 1Pi) animals compared with animals treated by BLM alone (BLM-control) at both 7 days (acute stage) and 30 days (fibrotic stage) after BLM treatment. Analysis of intraalveolar cells from bronchoalveolar lavage (BAL) fluid revealed that neutrophils and lymphocytes were significantly decreased in the BLM-alpha 1Pi animals at 7 days after BLM treatment and that 30 days after BLM treatment macrophages as well as neutrophils and lymphocytes were remarkably decreased in the BLM-alpha 1Pi animals. The elastase activity in supernatants of BAL fluid during 7 days following BLM treatment was detected, but there was no difference between the two groups. In vitro studies on neutrophil responsiveness to stimulation of BAL fluid at 3 days after BLM treatment revealed noticeable chemotaxis and generation of superoxide anion of isolated neutrophils, but alpha 1Pi did not show any inhibitory effects on neutrophil responsiveness. We suggest that alpha 1Pi administration ameliorates pulmonary fibrosis preceded by acute lung injury induced by BLM treatment in hamsters and that the inhibitory effects of alpha 1Pi on lung injury may not be brought about by altered elastase activity, chemotaxis, or superoxide generation in neutrophils. Alternative mechanisms are discussed.


Isolation and characterization of the complement-inhibiting protein CD59 antigen from platelet membranes.

Several groups have recently described the isolation of a 20 kDa membrane-attack-complex (MAC)-inhibiting protein, termed 'CD59 antigen', from human erythrocyte membranes. Antibodies raised against erythrocyte CD59 antigen detect antigen on the surface of many other cell types, and in some of these cells the antigen has been shown to have a molecular mass similar to that of the erythrocyte protein and to confer resistance to lysis by the MAC. A platelet-membrane form of CD59 antigen has been described and reported to be much larger than the erythrocyte protein. Here I report the isolation of CD59 antigen from platelet membranes and its molecular and functional characterization. The platelet protein is not significantly larger than the erythrocyte form and possesses similar MAC-inhibiting activity. Platelet CD59 antigen is anchored to the membrane via a glycosyl-phosphatidylinositol link, and consequently it is suggested that deficiency of this protein might be responsible for the increased thrombotic tendency observed in paroxysmal nocturnal haemoglobinuria.


Association of tetrapyrrole intermediates in the bacteriochlorophyll a biosynthetic pathway with the major outer-membrane porin protein of Rhodobacter capsulatus.

Rhodobacter capsulatus regulates synthesis of bacteriochlorophyll a in response to changes in oxygen partial pressure and light intensity. One early model proposed that this regulation involved a carrier polypeptide that functions to tether tetrapyrrole intermediates to the membrane. In the present study we isolated tetrapyrrole intermediates accumulated in three strains of R. capsulatus that contain mutations which block bacteriochlorophyll a biosynthesis at different steps of the magnesium branch of the pathway. Each of the tetrapyrrole intermediates was shown to be associated with the same 32 kDa polypeptide, as indicated by similar electrophoretic mobility and antigenic cross-reactivity with polyclonal antisera. The 32 kDa pigment-associated protein was further found to have an electrophoretic mobility, antigenic cross-reactivity and N-terminal sequence identical with those of the previously characterized major outer-membrane porin protein of R. capsulatus.


Epitope map of two polyclonal antibodies that recognize amyloid lesions in patients with Alzheimer's disease.

Two synthetic peptides with sequences identical with those of fragments of the extracellular domain of the Alzheimer's-disease amyloid precursor protein (APP) were used to raise antibodies. SP28 comprises positions 597-624 of the APP695 isoform, whereas SP41 extends towards the N-terminus (amino acids 584-624) and contains the entire SP28 peptide. Using e.l.i.s.a. and inhibition experiments we identified the two beta-turn-containing segments 602-607 and 617-624 as the epitopes recognized by anti-SP41 and anti-SP28 respectively. Both antibodies immunolabelled amyloid lesions in brains from Alzheimer's-disease patients and patients with related disorders, whereas they were unreactive in control brains. However, when probed on immunoblots, anti-SP28 failed to detect full-length APP from baculovirus-infected Sf9 cells, and anti-SP41 reacted weakly compared with other anti-APP antisera. The data suggest that these antibodies are directed to conformational epitopes not existent in the native molecules but present after alternative APP processing.


Natural polyamines stimulate G-proteins.

The natural polyamines spermine and spermidine, the biosynthetic precursor putrescine and their analogues cadaverine and tyramine stimulate the GTPase activity of purified GTP-binding proteins (Go/Gi) from calf brain reconstituted into phospholipid vesicles. The order of potency was spermine greater than spermidine greater than putrescine = cadaverine greater than tyramine. The physiological relevance of this observation was assessed, showing the same order of potency of polyamines in the stimulation of peritoneal and tracheal rat mast cells. The activation of rat mast cells by polyamines was inhibited by benzalkonium chloride or by a 2 h pretreatment of the cells with pertussis toxin. The increase in inositol phosphates evoked by polyamines was also inhibited by pertussis toxin. Therefore we propose that intracellular polyamines might control the basal level of second messengers and modulate extracellular signals transduced through G-protein-coupled receptors.


Molecular cloning and nucleotide sequence of a cDNA encoding hydroxyindole O-methyltransferase from chicken pineal gland.

Hydroxyindole O-methyltransferase (EC 2.1.1.4) is the enzyme that catalyses the synthesis of melatonin in the pineal gland and in the retina. Polyadenylated RNA from chicken pineal glands was used to prepare a cDNA library in lambda gt11. The library was screened with an antiserum directed against chicken hydroxyindole O-methyltransferase, and one cDNA clone was isolated. The fusion protein expressed by phage lysogens was identified on Western blots as a 165 kDA immunoreactive protein (beta-galactosidase, 110 kDa; hydroxyindole O-methyltransferase, 38 kDa). The fusion protein exhibited hydroxyindole O-methyltransferase activity. Its Km values for N-acetyl-5-hydroxytryptamine and S-adenosylmethionine were 5 times those of the natural enzyme. The intrinsic activity of the fusion protein was approx. 0.25% that of the natural enzyme. The cDNA consisted of 1436 nucleotides, including a 1038-nucleotide sequence encoding a full-length 346-amino-acid hydroxyindole O-methyltransferase. Comparison with bovine hydroxyindole O-methyltransferase [Ishida, Obinata & Deguchi (1987) J. Biol. Chem. 262, 2895-2899] revealed 52% identity in nucleotide sequences and 44% identity in peptide sequences. Northern-blot analysis revealed the presence of hydroxyindole O-methyltransferase mRNA transcripts in chicken pineal gland and retina, but not in the telencephalon.


Characterization of cDNA and genomic sequences encoding rabbit ELAM-1: conservation of structure and functional interactions with leukocytes.

Endothelial leukocyte adhesion molecule-1 (ELAM-1) is a cytokine-inducible endothelial cell surface glycoprotein involved in the adherence of neutrophils. ELAM-1 belongs to the selectin family of cell-surface molecules characterized by the general structure of an amino-terminal lectin domain followed by an epidermal growth factor domain, a variable number of complement regulatory elements, a single transmembrane sequence, and a short cytoplasmic tail. To study the in vivo regulation and expression of ELAM-1, we have isolated a complementary DNA (cDNA) clone encoding the rabbit homolog of human ELAM-1. The nucleotide sequence of the rabbit cDNA as well as its deduced amino acid sequence display extensive conservation compared to the human sequences. Rabbit ELAM-1 contains the characteristic protein domain organization of the selectin gene family and shares 74% amino acid identity with its human counterpart. However, rabbit ELAM-1 contains five complement regulatory elements whereas the human protein has six of these elements. Characterization of the genomic sequence encoding rabbit ELAM-1 indicated that individual extracellular protein domains are encoded by distinct exons. The genomic organization of rabbit ELAM-1 parallels that found for the human ELAM-1 gene and is similar to the pattern observed for other selectin family members (GMP-140, Lam-1), consistent with the hypothesis that the selectins evolved by duplication and rearrangement of individual exons. COS cells transiently expressing the rabbit ELAM-1 cDNA mediate the adhesion of rabbit and human polymorphonuclear leukocytes and are recognized by antibodies prepared against the human protein. Our results suggest that the specificity of molecular interaction between ELAM-1 and its ligand is highly conserved.


[The biological aftereffects of preoperative and palliative percutaneous biliary drainage].

Biological repercussions in 78 patients with malignant obstructive jaundice in whom percutaneous biliary drainage was performed, are reported. In 37 cases drainage was done during operation while 41 were palliative. Biochemistry, proteinogram, hematological studies, renal function and immunology were assessed 15.7 +/- 3.4 days postoperatively and 25.2 +/- 4.7 days in palliative drainage. Results show a significant improvement of all parameters, more important in preoperative drainages especially in those combining percutaneous and internal drainage techniques.


Intergeniculate leaflet and suprachiasmatic nucleus organization and connections in the golden hamster.

The intergeniculate leaflet (IGL) is a distinct subdivision of the lateral geniculate complex which receives retinal input and projects upon a circadian pacemaker, the suprachiasmatic nucleus (SCN). In the present study, we have analyzed the organization of the IGL and its connections in the hamster, a species commonly used in circadian rhythm studies. The location of the IGL is defined by the presence of retinal afferents demonstrated by anterograde transport of cholera toxin-HRP, neuropeptide Y-containing neurons and axons, cells retrogradely labeled from the regions of the SCN and contralateral IGL, and substance P-containing axons. It is a long nucleus extending the entire rostrocaudal axis of the geniculate. The most rostral IGL lies between the lateral dorsal thalamus, ventrolateral part, and the horizontal cerebral fissure. It then enlarges ventral to the rostral dorsal lateral geniculate, medial to the optic tract. The mid-portion of the leaflet is a thin lamina intercalated between the dorsal and ventral geniculate nuclei. The extended caudal portion of the nucleus lies lateral and ventral to the medial geniculate and is contiguous with the zona incerta and the lateral terminal nucleus. The IGL contains populations of neuropeptide Y (NPY+) and enkephalin (ENK+) neurons which project to the retinorecipient portion of the SCN. In addition to the immunoreactive perikarya, the IGL contains plexuses of NPY+, ENK+, substance P-, serotonin-, and glutamic acid decarboxylase-immunoreactive axons. Retrograde transport studies demonstrate that, in addition to the NPY+ neurons, there is a population of non-NPY+ neurons projecting upon the SCN.(ABSTRACT TRUNCATED AT 250 WORDS)


Organization and development of horizontal cells in the goldfish retina, II: Use of monoclonal antibody MH1.

We have produced and characterized a monoclonal antibody, MH1, which selectively labels rod horizontal cells and Müller cells in the goldfish retina. Biochemical and tissue distribution studies indicate that MH1 may recognize four out of five classes of intermediate filament proteins in goldfish: vimentin, desmin, glial fibrillary acidic protein (GFAP), and keratin, but not neurofilament. The intermediate filament which is labeled strongest in the retina is vimentin. In the goldfish retina, the only type of horizontal cells recognized by MH1 appear to be rod horizontal cells. This result suggests that the rod horizontal cell, an interneuron, and Müller (glial) cells share a common antigen: vimentin, which is usually only expressed in mesenchymal origin cells. The development of rod horizontal cells in the goldfish retina was also studied using MH1. The cells were not labeled by MH1 until 4-6 weeks posthatching, a stage in which the animals are already visually active. MH1 also did not label any horizontal cell in the region close to the ora terminalis in the goldfish retina. These results suggest that either the emergence and maturation of rod horizontal cells occur late during goldfish retinal development or the expression of vimentin itself occurs late in the development of rod horizontal cells.


The laminar distribution of macaque tectobulbar and tectospinal neurons.

The superior colliculus exerts its most direct influence over orienting movements, and saccades in particular, via its descending projections to the brain stem and spinal cord. However, while there is detailed physiological data concerning the generation of saccade-related activity in the primate superior colliculus, there is relatively little data on the detailed connectivity of this structure in primates. Consequently, retrograde transport techniques were utilized to determine the locations of the cells of origin of these descending pathways in macaque monkeys. Tectal cells that projected to the ipsilateral pontine reticular formation were mainly found in the deep gray layer and occasionally in the intermediate gray layer. Tectal cells that projected to the contralateral pontine reticular formation were predominantly located in the intermediate gray layer. The contralaterally projecting population could be subdivided into two groups. The cells in upper sublamina of the intermediate gray layer project primarily to the saccade-related regions of the paramedian reticular formation. Cells in the lower sublamina project primarily to more lateral regions of the pontine reticular formation and to the spinal cord. We conclude that the primate colliculus is provided with at least three descending output channels, which are likely to differ in their connections and functions. Specifically, it seems likely that the lower portion of the intermediate gray layer may be specialized to subserve combined head and eye orienting movements, while the upper sublamina subserves saccades.


Maternal serum unconjugated oestriol and human chorionic gonadotrophin levels in pregnancies with insulin-dependent diabetes: implications for screening for Down's syndrome.

To investigate maternal serum unconjugated oestriol (uE3) and human chorionic gonadotrophin (hCG) levels in pregnant women with insulin-dependent diabetes mellitus and to consider the implications of the results for antenatal screening for Down's syndrome. Descriptive study using stored antenatal serum samples. Stored serum samples collected from women receiving routine antenatal care in Oxford. 126 singleton pregnancies in 92 women with insulin-dependent diabetes mellitus and for each pregnancy, two pregnancies without diabetes matched for gestational age and duration of storage of the serum sample. None of the pregnancies was associated with fetal neural tube defect or Down's syndrome. Maternal serum uE3 and hCG levels at 15-22 weeks gestation. Alpha-fetoprotein (AFP) levels were also measured for comparison. The median uE3 level in the diabetic pregnancies was 0.92 multiples of the median (MoM) for pregnancies without diabetes at the same gestational age (P less than 0.05); and the hCG level was 0.95 MoM (P = 0.48). The median AFP level was also reduced to 0.77 MoM (P less than 0.001). The reduction in uE3 and AFP levels in insulin-dependent diabetic pregnancies is sufficiently great to be taken into account in maternal serum screening programmes for Down's syndrome. Dividing the uE3 and AFP levels in such pregnancies by the corresponding median for insulin-dependent diabetic pregnancies will yield a similar false-positive rate in pregnancies with and without insulin-dependent diabetes mellitus.


Effect of various blood-derived and semisynthetic nutrient media on in vitro uptake of 125I-albumin across the intact porcine aortic endothelial surface in an organ-support system.

The effects on porcine arterial structure and permeability of a 4-hour in vitro incubation at 37 degrees C in eight different blood-derived and synthetic nutrient media were examined. Changes in arterial permeability were inferred from the normalized, 1-hour, pressurized, transendothelial uptake (M/c0 cm) of porcine 125I-albumin in 60 porcine aortic tissue preparations using an organ-support system. The organ-support system provided experimental control of ambient gas composition, temperature, transmural pressure, flow (stirring), and nutrient media at a number of sites along the vessel. Light and electron microscopic (scanning and transmission) structural correlations with the observed permeability changes were examined. The M/c0 from the autogenous serum (AS) medium was used as the "control" measurement in each vessel preparation. (Grand mean M/c0 for AS from all studies was 0.312 +/- 0.011 [x10(-3)] [mean +/- SEM] cm, n = 60.) For brevity, M/c0 values from the other media are expressed below as a percentage of the corresponding paired M/c0 from the AS. Uptake from heparinized autogenous blood was 113 +/- 9% of that from AS (p = 0.119); from heparinized autogenous plasma was 135 +/- 10% (p = 0.048); from AS+heparin was 97 +/- 5% (p = 0.498); from pooled porcine serum was 113 +/- 9% (p = 0.037); from a synthetic medium was 131 +/- 8% (p = 0.004); and from a physiological hetastarch solution was 532 +/- 8% (p = 0.0002). Associated light microscopic structural changes and ultrastructural changes were not found. We conclude that 1) incubation with AS and heparinized blood (both of which are autogenous blood substances containing platelet products or platelets) provided the best support for the endothelial barrier function, whereas heparin plasma, pooled serum, a synthetic medium, and particularly hetastarch provided poorer support; 2) arterial permeability can increase significantly without discernible endothelial ultrastructural changes; and 3) AS and to a lesser extent heparin blood should provide a suitable nutrient medium for short-term (less than 4-hour) metabolic support of the endothelial surface and subjacent tissues.


Concerted modulation by myelin basic protein and sulfatide of the activity of phospholipase A2 against phospholipid monolayers.

The effect of myelin basic protein (MBP) on the activity of phospholipase A2 (PLA2, EC 3.1.1.4) against monolayers of dilauroylphosphatidylcholine (dlPC) or dilauroylphosphatidic acid (dlPA) containing different proportions of sulfatide (Sulf) and galactocerebroside (GalCer) was investigated. MBP was introduced into the interface by direct spreading as an initial constitutive component of the lipid-protein film or by adsorption and penetration from the subphase into the preformed lipid monolayers. The effect of MBP on PLA2 activity depends on the type of phospholipid and on the proportion of MBP at the interface. At a low mole fraction of MBP, homogeneously mixed lipid-protein monolayers are formed, and the PLA2 activity against dlPC is only slightly modified while the degradation of dlPA is markedly inhibited. This is probably due to favorable charge-charge interactions between dlPA and MBP that interfere with the enzyme action. The PLA2 activity against either phospholipid is increased when the mole fraction of MBP exceeds the proportion at which immiscible surface domains are formed. GalCer has little effect on the modulation by MBP of the phospholipase activity. The effect of Sulf depends on its proportions in relation to MBP. The individual effects of both components balance each other, and a finely tuned modulation is regulated by the interactions of MBP with Sulf or with the phospholipid.


Reconstitution of vacuolar ion channels into planar lipid bilayers.

Vacuolar ion channels were characterized after reconstitution into planar lipid bilayers. (1) Channel activity was observed after incorporation of tonoplast-enriched microsomal membranes, purified tonoplast membranes or of solubilized tonoplast proteins. (2) Channels of varying single-channel conductances were detected after reconstitution. In symmetrical 100 mmol l-1 KCl, conductances between 1 and 110 pS were frequently measured; the largest number of independent reconstitution events was seen for single-channel conductances of 16-25 pS (28 experiments), 30-42 pS (26), 49-56 pS (15) and 64-81 pS (15). Channel current usually increased linearly with voltage. (3) In asymmetrical solutions, cation-, non-selective and, for the first time for the tonoplast, anion-selective channels were detected. Ca(2+)-dependent regulation of channel opening was not observed in our reconstitution system. (4) Permeability was also observed for Cl-, NO3-, SO4(2-) and phosphate. (5) After fractionation of tonoplast proteins by size exclusion chromatography, ion channel activity was recovered in specific fractions. (6) Some of these fractions catalyzed sulfate transport after reconstitution into liposomes. The results suggest that different channels are active at the tonoplast membrane at a larger number than has been concluded from previous work.


Effect of fatty acyl domain of phospholipids on the membrane-channel formation of Staphylococcus aureus alpha-toxin in liposome membrane.

By use of carboxyfluorescein-loaded multilamellar liposomes prepared from synthetic phosphatidylcholine (PC) or sphingomyelin and cholesterol in a molar ratio of 1:1, we studied whether or not fatty acyl domain of the phospholipids affects the membrane-damaging action (or channel formation) of Staphylococcus aureus alpha-toxin on the phospholipid-cholesterol membranes. Our data indicated: (1) that toxin-induced carboxyfluorescein-leakage from the liposomes composed of saturated fatty acyl residue-carrying PC and cholesterol was decreased with increasing chain length of the acyl residues between 12 and 18 carbon atoms, although toxin-binding to the liposomes was not significantly affected by the length of fatty acyl residue; (2) that unsaturated fatty acyl residue in PC or sphingomyelin molecule conferred higher sensitivity to alpha-toxin on the phospholipid-cholesterol liposomes, compared with saturated fatty acyl residues; and (3) that hexamerization of alpha-toxin, estimated by SDS-polyacrylamide gel electrophoresis, occurred more efficiently on the liposomes composed of PC with shorter fatty acyl chain or unsaturated fatty acyl chain. Thus, hydrophobic domain of the phospholipids influences membrane-channel formation of alpha-toxin in the phospholipid-cholesterol membrane, perhaps by modulating packing of phospholipid, cholesterol and the toxin in membrane.


Thermal stability of hepatitis B surface antigen S proteins.

Thermal stability of hepatitis B surface antigen (HBsAg) has been studied by analyzing alterations in the native secondary structure and the antigenic activity. After heating for 19 h, circular dichrosim showed a cooperative transition with a midpoint at 49 degrees C. The conformational changes induced by temperature reduced the helical content of HBsAg S proteins from 49% at 23 degrees C to 26% at 60 degrees C and abolished the antigenic activity, as measured by binding to polyclonal antibodies. Furthermore, the six different antigenic determinants recognized by our panel of monoclonal antibodies were also shown to be dependent on the native structure of HBsAg proteins. Hence, it can be inferred that these epitopes are conformation-dependent. Binding of monoclonal antibodies to HBsAg protected the native structure of the corresponding antigenic determinant from thermal denaturation. In fact, binding of one of the monoclonals tested resulted not only in protection of the corresponding epitope, but also in a consistent increase of antibody binding with increasing temperature. Such an increase in antibody binding occurred simultaneously with an increase in the fluidity of surface lipid regions, as monitored by fluorescence depolarization of 1-(trimethylammoniophenyl)-6-phenyl-1,3,5-hexatriene. This correlation, along with the observation that lipids play an important role in maintaining the structure and antigenic activity of HBsAg (Gavilanes et al. (1990) Biochem. J. 265, 857-864), allow to speculate the certain epitopes of HBsAg which are close to the lipid-protein interface, are dependent on the fluidity of the surface lipid regions. Thus, any change in the physical state of the lipids could confer a different degree of exposure to the antigenic determinants.


Noise analysis and single-channel observations of 4 pS chloride channels in human airway epithelia.

Apical membranes of human airway epithelial cells have significant chloride permeability, which is reduced in cystic fibrosis (CF), causing abnormal electrochemistry and impaired mucociliary clearance. At least four types of chloride channels have been identified in these cells, but their relative roles in total permeability and CF are unclear. Noise analysis was used to measure the conductance of chloride channels in human nasal epithelial cells. The data indicate that channels with a mean conductance of 4.5 pS carry most of the chloride current, and that the mean number of such channels per cell is approximately 4,000. Chloride channels in this conductance range were also seen in single-channel recordings.


Defective glycosylphosphatidylinositol anchor synthesis in paroxysmal nocturnal hemoglobinuria granulocytes.

To investigate the biosynthesis of the glycosylphosphatidylinositol (GPI) anchor in the granulocytes of paroxysmal nocturnal hemoglobinuria (PNH), the glycolipids of granulocytes from PNH patients and normal volunteers were biosynthetically labeled with [3H]mannose in the presence of tunicamycin. Extracted glycolipids were examined by thin-layer chromatography and compared with known biosynthetic intermediates. Normal granulocytes consistently showed [3H]mannose incorporation into the complete GPI core, several GPI biosynthetic intermediates, and dolichol phosphate mannose (DPM). The granulocytes of 10 patients with PNH that had no expression of CD55 and CD59 on greater than 95% of the cells were able to incorporate [3H]mannose into DPM, but were not able to incorporate detectable amounts into the complete GPI core. THus, PNH granulocytes do not synthesize detectable amounts of the complete GPI core and this defect likely accounts for the absence of GPI-linked membrane proteins on hematopoietic cells in this syndrome.


Sustained human hematopoiesis in sheep transplanted in utero during early gestation with fractionated adult human bone marrow cells.

Sheep were transplanted in utero during early gestation with subpopulations of adult human bone marrow (BM) cells enriched for human progenitor and hematopoietic stem cells (HSC). Chimerism was documented in three of seven transplanted fetuses using monoclonal antibodies against human-specific hematopoietic cell lineages and/or cytogenetic analysis of BM and peripheral blood cells of recipients. Only chimeric sheep BM cells expressing CD45 (6.0% of total BM cells) formed human hematopoietic colonies in response to human recombinant cytokines as determined by cytogenetic analysis. Sorted CD45+ BM cells developed human T-cell colonies containing CD3+, CD4+, and CD8+ cells. DNA from chimeric BM cells obtained 3 months after birth displayed a finger printing pattern identical to that of DNA from the human donor of the HSC graft. These studies indicate that first trimester sheep fetuses are tolerant of adult human HSC grafts, thus permitting the creation of xenogeneic chimera expressing human myeloid and lymphoid lineages. The present findings also suggest that HSC grafts from immunologically competent, HLA-mismatched adult donors may be useful for correcting human genetic diseases in utero during early gestation.


Cytokine regulation of colony-stimulating factor (CSF) production in cultured human synovial fibroblasts. II. Similarities and differences in the control of interleukin-1 induction of granulocyte-macrophage CSF and granulocyte-CSF production.

Synovial fibroblasts are likely to be a significant source of granulocyte-macrophage colony-stimulating factor (GM-CSF) and granulocyte-CSF (G-CSF), which could be crucial to the pathogenesis of rheumatoid arthritis. Using specific enzyme-linked immunosorbent assays (ELISAs) and Northern analysis, GM-CSF and G-CSF expression were followed in human synovial fibroblast-like cells in response to a number of agents, either alone or in the presence of an optimal stimulatory concentration of interleukin-1 (IL-1). For both CSFs, interferon-gamma (100 U/mL) did not increase their levels but dramatically suppressed the stimulatory action of IL-1, while basic fibroblast growth factor (10(-8) mol/L), although nonstimulatory by itself, potentiated IL-1 action. The glucocorticoid, dexamethasone (10(-7) mol/L), inhibited IL-1-stimulated CSF production. However, evidence was obtained for noncoordinated CSF regulation. Cyclooxygenase inhibitors potentiated the action of IL-1 on GM-CSF synthesis but suppressed G-CSF synthesis, suggesting that endogenous cyclooxygenase products can have opposite effects in modulating the levels of each CSF. Also, the lymphokine, IL-4 (250 pmol/L), slightly inhibited GM-CSF formation in the presence of IL-1 but elevated the G-CSF levels in these cultures without having an effect by itself. Transforming growth factor beta (less than or equal to 20 ng/mL) did not modulate levels of either CSF. Mesenchymal cell production of both GM-CSF and G-CSF is generally viewed as being under coordinate control; our findings suggest that their synthesis in IL-1-stimulated human synoviocytes can be modulated by a number of agents, in some cases with divergent actions depending on which CSF is examined.


Effects of in vivo administration of interleukin-2 (IL-2) and IL-4, alone and in combination, on ex vivo human basophil histamine release.

Human basophils possess receptors for interleukin-2 (IL-2) and IL-4. The effect of 3 days of intravenous administration of IL-2 and/or IL-4 on basophil histamine release was examined in three groups of patients receiving IL-2, IL-4, or the combination of agents as part of a protocol to treat malignant melanoma or renal cell carcinoma. Because all patients received ranitidine for control of side effects, a control group of patients receiving ranitidine for Zollinger-Ellison's syndrome was also studied. IL-4 significantly inhibited IgE-mediated histamine release, while there was a trend for enhancement of IgE-mediated histamine release by IL-2. Administration of the combination of IL-2 and IL-4 did not alter IgE-mediated basophil histamine release. Both IL-2 and IL-4, alone and in combination, enhanced basophil histamine release induced by histamine releasing factors in human nasal washings. The effect of IL-2 alone was significantly greater than that of IL-4 alone or the combination of IL-2 plus IL-4. Taken together, the data suggest that when coadministered, IL-4 may inhibit the effects of IL-2 on basophils. Neither cytokine exerted any effect on basophil histamine release induced by the calcium ionophore A23187, nor did ranitidine cause any effects on histamine release induced by any of the stimulants. Thus, human basophil reactivity can be affected by IL-2 and by IL-4. The role that these two cytokines play in basophil function in vivo is likely to be complex.


Internalization and intracellular fate of anti-CD5 monoclonal antibody and anti-CD5 ricin A-chain immunotoxin in human leukemic T cells.

We have investigated the entry and subsequent intracellular fate of T101 monoclonal antibody (MoAb) and T101-ricin A-chain (RTA) immunotoxin (IT) directed against the CD5 antigen (Ag) expressed on human leukemic CEM cells. We provide direct evidence for the internalization of T101 MoAb and the corresponding IT. Both the MoAb and IT were internalized at a relatively low rate. This slow internalization process could be related to the partial recycling of the MoAb/Ag or IT/Ag complexes. Analysis of the internalized molecules showed that their molecular weight was only partially altered after internalization and that no free A-chain could be found inside the cells, indicating that lysosomal degradation and cleavage of disulfide-linked conjugates is a quantitatively minor phenomenon compared with the localization of internalized anti-CD5 ITs in an endosomo-Golgi compartment, followed by their recycling to the cell surface. We believe that this is the major factor explaining the low efficacy of anti-CD5 IT when assayed in the absence of potentiating substances. Finally, we showed that the presence of ammonium chloride and monensin, which both dramatically enhance the kinetics of IT activity, did not affect the rate of internalization or the intracellular localization of the conjugate, suggesting that these activators could act at the postendocytotic level on a limited number of IT molecules.


Isolation of the JMH antigen on a novel phosphatidylinositol-linked human membrane protein.

JMH is a high-frequency human erythrocyte blood group antigen. Previous work has shown that JMH is absent from complement-sensitive erythrocytes of patients with paroxysmal nocturnal hemoglobinuria (PNH); such cells have a broad defect in expression of phosphatidylinositol (PI)-linked proteins. Using both human JMH antisera and a JMH-like murine monoclonal antibody, we have identified a 76-Kd membrane protein present in JMH-positive but not JMH-negative erythrocytes. A similar 76-Kd JMH protein was also identified on a human lymphoid T-cell line, HSB-2. Using PI-specific phospholipase C, a small amount of JMH antigen could be cleaved from intact erythrocytes and immunoprecipitated from the supernate of treated erythrocytes, thus confirming that the protein bearing the JMH antigen is anchored by a PI-linkage to the erythrocyte membrane. This protein was further shown not to be identical to decay accelerating factor (70 Kd), a previously identified PI-anchored protein of somewhat similar molecular weight.


The use of colony stimulating factors in combination.

More gene products that influence hematopoiesis continue to become available. As a result, is now possible to carry out both in vivo and in vitro studies with purified erythropoietin, various colony stimulating factors and 11 interleukins. The identification and availability of the ligand for the c-kit gene product has had a profound influence in the past year.


Polymorphism of a long-chain cycloparaffin (CH2)120.

The polymorphism of a long-chain cycloparaffin (CH2)120 and chain packing in its crystals were discussed on the basis of some results obtained mainly by transmission electron microscopy. Monoclinic and orthorhombic single crystals of (CH2)120 were isothermally grown together from a dilute solution in p-xylene. Lozenge-shaped orthorhombic single crystals were more frequently observed than lath-shaped monoclinic ones. The basal surfaces of orthorhombic and monoclinic single crystal platelets were decorated with vapor-deposited polyethylene [PE]. Orthorhombic single crystals of (CH2)120 with the (110) twin boundary and monoclinic ones with the (100) twin boundary were also observed. Rod-like edge-on crystals of (CH2)120 were grown from a dilute p-xylene solution onto the (001) surface of alkali halides. The crystal system of the (CH2)120 edge-on crystals depended on the kind of substrate. The monoclinic crystal was grown on NaCl, the orthorhombic one on KBr and KCl. The monoclinic form of (CH2)120 edge-on crystal was transformed to the orthorhombic one by annealing on NaCl. In both monoclinic and orthorhombic edge-on crystals, the molecular plane determined by two zigzag stems in a molecule of (CH2)120 was parallel to the substrate surface and the molecular axis (crystallographic c-axis) was perpendicular to the longer side of the rod-like edge-on crystals. The sub-cell dimensions of the stem chains in both forms of (CH2)120 crystals were very similar to those of monoclinic and orthorhombic PE crystals, respectively.


Extraosseous Ewing's sarcoma: report of a case and review of literature.

Extraosseous Ewing's sarcoma is a primitive small round cell neoplasm found in children and young adults. Extraosseous Ewing's sarcoma has been recognized as being histologically indistinguishable from Ewing's sarcoma of the bone and other round cell tumors. Our study reports the clinical course and histopathologic features of this rare variant, occurring in a 13 year-old boy, who presented with a subcutaneous tumor of the right thigh without osseous involvement. Wedge resection of the tumor was done followed by chemotherapy with Actinomycin-D, Oncovin and Endoxan.


Free alpha-subunit, free beta-subunit of human chorionic gonadotropin (hCG), and intact hCG in sera of healthy individuals and testicular cancer patients.

To determine the serum concentrations of human chorionic gonadotropin (hCG), its free beta-subunit (hCG beta), and the free alpha-subunit (free alpha) common to all human glycoprotein hormones under physiological and pathological conditions, we developed monoclonal antibody-based immunoenzymometric assays. Free alpha-subunit was detected in the sera of all healthy individuals of both sexes; hCG was measurable in sera of 54% of the men, and 46% were positive for free hCG beta; in nonpregnant women, 69.5% were positive for hCG, 68.4% for the free beta-subunit. Pathological conditions, i.e., hCG-producing tumors, were studied in vitro and in vivo. In vitro, the concentrations of hCG, free hCG beta, and free alpha in tissue-culture supernates of a choriocarcinoma cell-line ("JAR") showed a parallel pattern during time-course analysis. In vivo, in long-term follow-up studies of 13 patients with testicular cancer, serum concentrations of the three analytes paralleled each other, whether the disease was in remission or not. Because of a selective increase of free hCG beta and free alpha in 27% of seminomatous tumor patients and in 13% of the nonseminomatous patients, the percentage of tumor-marker-positive sera was increased from 15% to 42% and 57% to 70%, respectively, by the additional measurement of free hCG beta and free alpha. Thus hCG, free hCG beta, and free alpha are physiologically present in a high percentage of the sera from healthy men, and the determination of free hCG beta and free alpha, although not of prognostic value, improves the diagnostic possibilities in patients with testicular cancer.


Impact of CF summer camp.

In two consecutive years, patients with cystic fibrosis were studied at the beginning and end of a nine-day summer camp program to assess the program's effects on weight gain and pulmonary function. The camp experience includes daily exercise and a high-protein and high-fat diet. There were a total of 58 children between 6 and 12 years of age (42 different patients) and 10 adult counselors from 19 to 30 years of age (eight different patients). On the first and eighth days patients were weighed, sputum cultures were collected, and spirometry was performed. In year 2, peak expiratory flow rate was monitored daily. Also in year 2, campers and counselors with CF were prescreened by sputum culture and excluded from camp if they had Pseudomonas cepacia in their sputum. Only one candidate screened was positive before camp. In year 1, no significant group changes in pulmonary function were identified. In year 2, significant increases on post-camp testing were found for FEF 25%-75% and PEF. Mean body weight for all patients increased significantly, by 0.4 kg in year 1 and 0.9 kg in year 2 (p less than .05). In year 1, a total of nine patients acquired a new organism in their follow-up sputum culture, including five who acquired a new Pseudomonas species. There was no intra-cabin pattern of spread. Four patients were positive for P. cepacia on day 1 culture. No new subjects acquired this organism on follow-up examination. In year 2, only one subject had P. cepacia on the first camp collection; he alone was positive on day 9.(ABSTRACT TRUNCATED AT 250 WORDS)


Bleomycin-induced lung injury in rats selectively abolishes hypoxic pulmonary vasoconstriction: evidence against a role for platelet-activating factor.

1. The role of platelet-activating factor in the attenuated hypoxic pulmonary vasoconstriction associated with lung injury was evaluated using specific platelet-activating factor antagonists and an isolated perfused lung preparation. 2. Intratracheal bleomycin was administered to rats to produce acute lung injury. Animals received intratracheal saline (control), intratracheal bleomycin or the platelet-activating factor antagonists BN 52021, WEB 2170 or WEB 2086 before and after bleomycin treatment. Forty-eight hours after intratracheal administration of bleomycin or saline the animals were killed. 3. The increases in pulmonary artery pressure during two periods of hypoxic ventilation and in response to 0.2 microgram of angiotensin II were measured. Acetylcholine-induced vasodilatation after pre-constriction with prostaglandin F2 alpha was also measured. To quantify lung injury, the wet/dry ratio of lung weight was determined. 4. Bleomycin treatment attenuated the first and second hypoxic pressor responses by 93% and 77%, respectively, but not the pressor response to angiotensin II nor the vasodilator response to acetylcholine. BN 52021 plus bleomycin augmented the first hypoxic pressor response compared with bleomycin treatment alone, but the structurally unrelated platelet-activating factor antagonists WEB 2170 and WEB 2086 had no significant effect on the bleomycin-induced attenuation of hypoxic pulmonary vasoconstriction. None of the platelet-activating factor antagonists blocked the increase in the wet/dry lung weight ratio induced by bleomycin. 5. Bleomycin-induced lung injury selectively attenuates hypoxic pulmonary vasoconstriction, an effect that does not appear to be mediated by platelet-activating factor. The mechanism remains to be elucidated, but may involve destruction of the hypoxic 'sensor' within the respiratory tract.


Ferrous ions detected in cerebrospinal fluid by using bleomycin and DNA damage.

1. During pathological states of iron-overload or oxidant stress, low-molecular-mass iron can become available within extracellular fluids. 2. This iron would be converted to the ferrous state were it not for the protective anti-oxidant protein caeruloplasmin. 3. The ferrous-ion-oxidizing activity of caeruloplasmin rapidly converts ferrous ions back to the less reactive ferric state so that they can bind to available binding sites on transferrin. 4. Cerebrospinal fluids, however, often appear to contain low-molecular-mass iron, high levels of ascorbate and low levels of ferroxidase activity with little or no iron-binding capacity. 5. When iron ions are present in cerebrospinal fluid they are therefore likely to be in the ferrous state. 6. The development and application of an assay to speciate and measure ferrous ions in simple aqueous solution and their redox cycling activity in biological fluids is described.


Systemic profiles of antigen-specific lymphocytes in animals chronically exposed to staphylococcus antigen in the mammary region.

Systemic profiles of lymphocytes were assessed in goats exposed chronically with Staphylococcus antigens in the supramammary region. Animals were inoculated three times subcutaneously in the right supramammary region with heat-killed Staphylococcus aureus antigen (HKS) at 1 month intervals. Prior to immunization and 1 week following each injection, 3 and 6 day cultures of peripheral blood mononuclear cells were made to determine proliferative responses of lymphocytes to HKS and the polyclonal T cell mitogen phytohemagglutinin (PHA). Peripheral blood lymphocytes responded significantly to both HKS and PHA in 3 day cultures after the second injection and showed peak responses after the final immunization, suggesting that repeated local injection of S. aureus antigen at the supramammary region, can induce an anamnestic response to the antigen in the peripheral blood of these animals with a concomitant increase in the responsiveness to the polyclonal mitogen, PHA. In contrast, initial antigen challenge induced little, if any, increase in responses to the specific antigen or mitogen when compared to pre-injection states. These data may also suggest that non-reactivity of peripheral blood lymphocytes to the HKS antigen immediately after the primary injection of antigen may be the result of local retention of antigen-reactive cells at the sites of infection.


Fixation of mammalian cells for flow cytometric evaluation of DNA content and nuclear immunofluorescence.

Mammalian tissue culture cells were fixed with 3 different alcoholic fixatives--acetone:methanol, EtOH, and MeOH. The quality of the resulting DNA histograms was evaluated by comparison of CV, G1/G2 ratio, G1 mode, cell aggregation, and debris formation; 81-90% MeOH (final concentration) was determined to be the optimal fixative by these criteria. A procedure was then examined using a prefix with paraformaldehyde followed by MeOH (PF/MeOH). This procedure produced cell preparations with reduced debris and aggregation, equivalent mode and ratio, but increased CV when compared with MeOH fixation. Both MeOH and PF/MeOH fixation procedures were then compared for their utility in dual staining for DNA and intracellular immunofluorescence for a nuclear protein, SV40 T antigen (Tag). Since alcohols are known to affect immunofluorescence staining of some antigens, fixation with paraformaldehyde followed by Triton X-100 permeabilization (PF/TX) was also included in this comparison to generalize the study by providing an alternative to MeOH permeabilization. The three procedures were evaluated for the quality of the sample by measuring the same descriptors of the DNA parameter as in the alcohol study. PF/TX consistently produced samples with decreased DNA CV and less debris and aggregation compared to MeOH methods. Two criteria were used to evaluate immunofluorescence--the amount of Tag measured and reproducibility. All MeOH methods were equivalently reproducible with CV's less than 3%. PF/TX was slightly less so with a CV of less than 6%. In contrast, different levels of Tag were measured for each procedure. For mouse 3T3 cells infected with a recombinant retroviral vector encoding T antigen, the level of T antigen measured after PF/MeOH was 21% greater than in MeOH fixed cells, and the level in PF/TX fixed cells was 37% less. The fraction of fluorescence specific to T antigen for these cells was 79-83% for all procedures. The lower levels measured after fixation by PF/TX were shown to be due to epitope masking. Why higher levels are measured with PF/MeOH procedures is unknown at present but may be due to antigen retention. Therefore, each of these fixation methods may be used with confidence in reliability but they are not equivalent with respect to the molecular architecture of the nucleus. It is postulated that PF/TX permeabilizes cells but cells retain native supramolecular structure, whereas MeOH based fixatives disrupt this structure and randomize availability of epitope to antibody. If so, the two procedures could be used as complementary procedures to study gene expression and function.


An autopsy case of late infantile and juvenile neuroaxonal dystrophy with diffuse Lewy bodies and neurofibrillary tangles.

The clinical and pathological features of a sporadic case of juvenile neuroaxonal dystrophy beginning at the age of 10 and leading to death at the age of 26 are described. Clinical manifestation began with cerebellar symptoms. The subject subsequently developed dementia, pes cavus (Friedreich's feet), epilepsy, myoclonus, and Parkinsonian syndrome, but demonstrated neither tremor nor choreoathetoid movement. Pathological examination showed typical generalized axonal dystrophy throughout the central nervous system (Seitelberger's disease). Iron-positive pigmentation was seen in the pallidonigral system, diffuse Lewy bodies (brainstem type and cerebral type) were demonstrated in the brainstem nuclei and cerebral cortex, and neurofibrillary tangles were observed.


Improved staining method for the simultaneous flow cytofluorometric analysis of DNA content, S-phase fraction, and surface phenotype using single laser instrumentation.

We have developed an improved technique for triple staining that permits the simultaneous flow cytofluorometric analysis of cell surface antigens, bromodeoxyuridine incorporation into DNA, and DNA quantification using 7-amino-actinomycin D. PHA-activated human peripheral blood lymphocytes were incubated with bromodeoxyuridine and stained for cell surface phenotype with phycoerythrin-labeled monoclonal antibodies. Stained cells were fixed serially with 1% paraformaldehyde and 45% ethanol. Fixed cells were sequentially stained with an anti-BrdUrd monoclonal antibody followed by a FITC-conjugated goat anti-mouse antibody and incubated with 7-amino-actinomycin D. Hypotonic buffer was employed for all procedures after fixation. Stained-fixed cells were analyzed by flow cytofluorometry for simultaneous green (525 nm), orange (570 nm), and red (greater than 650 nm) fluorescence. Utilizing this staining technique, we were able to analyze simultaneously cell phenotype, DNA synthesis, and total cellular DNA content with single laser excitation.


Flow cytometric analysis of whole blood lysis, three anticoagulants, and five cell preparations.

We studied the effects of anticoagulants and cell preparation methods on lymphocyte forward-angle scatter (FSC), autofluorescence, and immunofluorescent staining for CD45, CD14, and CD13. Blood samples collected in ethylenediaminetetracetic acid (EDTA), heparin, and acid citrate dextrose (ACD) were processed by using conventional Hypaque-Ficoll (HF) separation and four whole blood (WB) lysis techniques: Immuno-lyse, Q-Prep, FACS Lyse, and Gen Trak Lysis. Lymphocytes prepared by using three of the four whole blood methods gave FCS values comparable to those isolated by HF, while one method (FACS Lyse) gave consistently lower values. Autofluorescence values were comparable by all methods except Immuno-lyse, which showed consistently higher values in blood stored for 24 h with any anticoagulant. Immunofluorescent values for CD45-stained cells were quite consistent across all methods, and among the whole blood methods, FACS Lyse and Q-Prep uniformly gave the highest purity of CD45-positive cells in the lymphocyte light scatter gates. Additionally, propidium iodide (PI) analyses of CD45-stained whole blood, and analyzed without lysis, confirmed that ACD and heparin were superior to EDTA for maintaining viable leucocytes overnight. Future studies should focus on other commonly used reagents, a wide variety of abnormal samples, and cell viability.


Detection of granulocyte reactive oxygen species formation in whole blood using flow cytometry.

We have developed a technique for analysis of granulocyte reactive oxygen species formation in whole blood using flow cytometry and two color immunofluorescence. This technique relies upon the use of specific fluorescent dye (LDS-751) to stain nucleated cells, eliminating erythrocytes from analysis. Using LDS-751, forward angle light scatter, and 90 degrees side scatter, a granulocyte gate, monocyte gate, and lymphocyte gate were identified. Analysis with multiple FITC conjugated monoclonal antibodies demonstrated greater than 95% purity of a flow cytometrically identified granulocyte population in whole blood without physical manipulation of the blood. Utilizing 2'7' dichlorofluorescein diacetate (DCFH-DA), we were able to measure granulocyte intracellular reactive oxygen species production. Dose response curves were obtained for the effect of granulocyte agonists phorbol myristate acetate, FMLP, and heat fixed Staphylococcus aureus on reactive oxygen species production. The techniques described in this paper should be useful for measuring granulocyte activation in vivo with flow cytometry.


Multiparametric flow cytometric analysis of radiation-induced micronuclei in mammalian cell cultures.

A new flow cytometric method is presented that quantifies the frequency of radiation-induced micronuclei in mammalian cell cultures with high precision. After preparing a suspension of main nuclei and micronuclei stained with ethidium bromide and Hoechst 33258, both types of particles are measured simultaneously in a flow cytometer using forward light scatter and three fluorescence emission intensities excited by UV, 488 nm, and by energy transfer from Hoechst 33258 to ethidium bromide. Nonspecific debris overlapping the micronucleus distribution especially in the low fluorescence intensity region was discriminated from micronuclei by calculating ratios of the different fluorescences. The frequencies of radiation-induced micronuclei measured with this new technique agreed well with results obtained by conventional microscopy. The lower limit of the DNA content of micronuclei identified by this technique was found to be about 0.5%-0.75% of the DNA content of G1-phase nuclei. Dose effect curves and the time-dependent induction of micronuclei were measured for two different mouse cell lines.


Expression of proliferation associated antigens in the cell cycle of synchronized mammalian cells.

Flow cytometric bivariate analysis was used to investigate the expression of PCNA, p120 and p145 during the cell cycle of a mammalian cell line (CHO-K1). Initially, aliquots of cells in exponential and plateau (G0) phase were analyzed for proliferation associated antigen expression. Expression of PCNA and p145 during G0 was markedly depressed (less than 12% positive) while 54% of the G0 cells stained positive for p120. The fluorescent intensity (mean channel fluorescence) of these G0 positive p120 cells, however, was only slightly above the mean channel fluorescence (MCF) of cells stained with a negative isotype control. In asynchronous cultures, all three antigens were expressed in greater than 70% of the cells, with PCNA staining being greater than 95%. Cells were then synchronized using mitotic selection (mitotic index of 97%) and antigen levels were measured as cells progressed synchronously through the cell cycle. From DNA analysis histograms, it appeared that the degree of synchrony was approximately 90% throughout the remainder of the cell cycle. The bivariate DNA/PCNA, DNA/p120, and DNA/p145 histograms for mitotic cells indicated that both p120 and p145 expression were elevated (percent positive and MCF) while PCNA levels were near controls (MCF). In early G1, all three markers were depressed (less than 12% positive); however PCNA levels rose precipitously in mid-G1 (greater than 50% positive). In late G1 to early S, p145 levels increased concomitantly with increases in p120. All three antigens were elevated throughout S phase and began to decline as cells moved from G2/M to G1 of the next cell cycle with p145 expression decreasing first. This report indicates that all three proliferation associated antigens studied are differentially expressed in the cell cycle and therefore may be useful in detecting and assessing the proliferation state.


Comparative evaluation of several DNA binding dyes in the detection of apoptosis-associated chromatin degradation by flow cytometry.

Mouse thymocytes readily undergo apoptosis-associated DNA degradation upon exposure to glucocorticoids or ionizing radiation. It has been previously shown that flow cytometric cell cycle analysis of propidium iodide-stained apoptotic thymocytes results in the appearance of a distinct cell cycle region (the A0 region) below the G0/G1 region. Cells in this region were shown to be undergoing apoptosis, and determination of apoptosis by flow cytometric analysis was proposed as a superior method for evaluating thymocyte apoptosis. In this study, a variety of DNA binding dyes with diverse primary binding mechanisms were evaluated for their ability to detect glucocorticoid and ionizing radiation-induced apoptosis in mouse thymocytes. Apoptotic thymocytes stained with DNA binding dyes from the phenanthridinium, acridine, actinomycin, chromomycinone, anthracycline, and bisbenzimidazole groups all demonstrated clearly defined A0 regions with percentages comparable to those obtained for propidium iodide. These results indicate that the appearance of the A0 region is not dependent on a particular dye binding characteristic and may be the consequence of extensive changes in chromatin structure resulting in a significant degree of dye exclusion.


Flow cytometric DNA content of fresh tumor specimens using keratin-antibody as second stain for two-parameter analysis.

Studies concerning flow cytometric assessed DNA content reveal problems in interpretating DNA histograms of tumor specimens. The main problems are histograms with a broad coefficient of variation in the G0/G1 fraction; a high G2M fraction and samples with a low percentage of tumor cells. Therefore, in the present study, 382 fresh tumor specimens of carcinomas were analysed routinely, double labeled with, on the one hand, propidium-iodide for assessing DNA content and, on the other, a monoclonal keratin-antibody for marking epithelial and tumor cells. Of the 311 tumor samples, using single parameter analysis 165 (54%) were classified as DNA aneuploid and 146 (46%) as DNA "euploid." By double parameter analysis, 224 (72%) samples were keratin positive and 87 (27%) keratin negative and, of the 224 keratin positive tumors, 175 (78%) were DNA aneuploid and 49 (22%) DNA euploid. The DNA histograms of single and double parameter analysis were compared and it was concluded that in 24 cases (11%) keratin labeling was necessary to recognize DNA aneuploidy. In another 23 (10%) cases, keratin labeling was helpful in assessing DNA aneuploidy. Finally when the results of the 311 samples were combined, 215 (68%) were scored as DNA aneuploid and 99 (32%) DNA euploid. Thus the overall gain in assessing DNA aneuploidy using the double labeling technique is 14%. In conclusion, it is shown that keratin labeling on fresh tumor cell suspensions of epithelial tumors is of additional value in establishing DNA content. Because single parameter DNA assessment is adequate in approximately 60% of the tested samples, the double labeling technique can be performed routinely, or after initial single parameter DNA assessment. Histograms having a broad CV and/or a high G2M are good candidates for the double labeling technique. Using this technique, DNA-content assessment becomes more reliable.


Flow cytometric two-color staining technique for simultaneous determination of human erythrocyte membrane antigen and intracellular malarial DNA.

A novel fixative and permeabilization method is described which allows simultaneous flow cytometric detection of red blood cell membrane antigen and intracellular malaria parasites. To illustrate the method, red blood cells from patients with paroxysmal nocturnal hemoglobinuria were infected with Plasmodium falciparum and maintained in synchronous red blood cell culture. The infected red blood cells were immunolabeled with antibodies directed to the complement regulatory protein decay-accelerating factor (DAF) followed by subsequent fixations in paraformaldehyde and then glutaraldehyde in phosphate-buffered saline. Finally, DNA of the intraerythrocytic parasites was stained with propidium iodide. Using this technique, cellular morphology was well preserved, no cell aggregation was observed, and high-quality indirect immunofluorescence and parasite DNA staining were obtained with negligible nonspecific labelling. Simultaneous measurement of parasite DNA and red blood cell membrane determinants makes possible the investigation of alterations of red cell membrane proteins in association with development of intracellular malaria parasites.


Drug treatment of allergic conjunctivitis. A review of the evidence.

Allergic conjunctivitis, unlike several other ocular diseases, is seldom followed by permanent visual impairment; nevertheless, it is important because of both its frequency and its severity. Two major forms, seasonal and perennial, are considered in this review. To recognise the hallmarks of allergic conjunctivitis, clinicians have need of a thorough knowledge of its pathophysiological aspects and clinical features, enabling them to choose the best and most suitable therapy among the alternatives. The aims of treatment vary according to the symptoms, severity and characteristics of the allergic reactions; in general, treatment is based mainly on environmental control, pharmacotherapy and (sometimes) specific immunotherapy. Topical vasoconstrictors, decongestant compounds, standard antihistamines or combinations of these drugs have been used for a number of years to treat the acute and/or persistent symptomatology, and in order to prevent the side effects of a prolonged treatment with topical glucocorticosteroids. Nevertheless, the latter represent the most powerful anti-inflammatory drugs, and are particularly recommended in short term treatment (5 to 7 days) in severe acute symptomatology. Orally administered 'classic' antihistamines, i.e. histamine H1-receptor antagonists, are effective and very convenient in either short or long term treatment, largely because the new compounds also act on the inflammatory process secondary to the allergic events. Recently, other topical compounds such as sodium cromoglycate (cromolyn sodium), nedocromil and nonsteroidal anti-inflammatory drugs (NSAIDs) [i.e. piroxicam, aspirin] have become available. Sodium cromoglycate and nedocromil act as prophylactic compounds, able to prevent the allergic reaction; NSAIDs represent a valid and effective alternative to glucocorticosteroids in several situations.


An overview of the role of calcium antagonists in the treatment of achalasia and diffuse oesophageal spasm.

Early studies confirmed the beneficial effects of calcium channel blockers on the normal oesophagus, which included a decrease in lower oesophageal sphincter tone in achalasia and a decrease in oesophageal contractions and amplitude in diffuse oesophageal spasm. This resulted in the enthusiastic use of the drugs in both disorders. With further experience, and with increased recognition of side effects, the role of these drugs in the 2 disorders has been better clarified. Clinical trials in general have not reflected the improvement observed in the manometric parameters. Only a minority of patients appear to derive sustained symptomatic benefit. Calcium channel blockers may be the initial choice for high or moderate risk patients with achalasia prior to proceeding with pneumatic dilatation or surgical myotomy. In diffuse oesophageal spasm, they are a reasonable first choice for all risk categories.


Wolff-Parkinson-White syndrome. Identification and management.

Patients with Wolff-Parkinson-White (WPW) pattern of ventricular pre-excitation may develop paroxysmal re-entrant tachyarrhythmias through the Kent bundle and, less commonly, atrial fibrillation. WPW patients are at risk of sudden death when a rapid ventricular response occurs during atrial fibrillation due to conduction through the accessory pathway. Conduction properties of the accessory pathway and atrial vulnerability, which is the propensity to develop atrial fibrillation, are important parameters for evaluation in these patients. The former can be assessed by means of noninvasive tests, such as stress and pharmacological tests, and with electrophysiological study; the latter only by electrophysiological study. There is no indication for treatment of asymptomatic patients. Antiarrhythmic prophylaxis is required in patients with previous episodes of atrial fibrillation with rapid ventricular response, in patients with paroxysmal re-entrant tachycardias and rapid conduction through the accessory pathway, and in patients with frequent episodes of re-entrant tachycardias of long duration. Vaughan-Williams class IC anti-arrhythmic drugs (propafenone, flecainide) are the first choice for drugs in patients with rapid anterograde conduction through the accessory pathway due to their high efficacy and low incidence of adverse effects, while beta-blockers (atenolol, nadolol) are indicated for patients with re-entrant tachycardias and low conduction capacity through the bypass tract. When pharmacological therapy is ineffective, surgical or catheter ablation of the accessory pathway may be considered.


Menorrhagia. Current drug treatment concepts.

Since menorrhagia occurs in 9 to 14% of populations of healthy women, many general practitioners will encounter menorrhagia-related problems. Menorrhagia is difficult to objectify and the choice of treatment between the available drugs is not always an easy one. In this survey, the available knowledge on menorrhagia diagnosis, underlying pathophysiology and treatment, especially medicinal treatment, are discussed. Overall, a practical approach is emphasised. The desire for contraception as well as the underlying cause of menorrhagia determine the drug of choice in the treatment of menorrhagia. If contraception is desired, oral combination contraceptives and continuously dosed progestogens, orally or as a medicated intrauterine device (IUD), are the first choice drugs for essential menorrhagia, and for fibroid- and bleeding disorder-associated menorrhagia. If no contraception is desired, the first choice treatments are drugs that need to be administered only during menstruation, such as prostaglandin synthesis inhibitors or antifibrinolytics. Of these, antifibrinolytics reduce menstrual blood loss to the greatest extent, whereas prostaglandin synthesis inhibitors have the lowest incidence of side effects. Prostaglandin synthesis inhibitors also have the extra advantage of diminishing dysmenorrhoea. There is no place for ergometrine in the treatment of menorrhagia. No studies are available as yet on the combination of various drug treatment modalities, although such an evaluation would be desirable.


Current concepts in the pharmacological treatment of obsessive-compulsive disorder.

Obsessive-compulsive disorder (OCD) is a chronic and often disabling disease. OCD is characterised by intrusive, unwanted and persistently recurring mental events (obsessions) that usually evoke discomfort or anxiety, and/or repetitive ritualistic behaviours (compulsions) that are aimed at reducing discomfort and anxiety. However, the compulsions succeed only in achieving transient relief, followed by a growing sense of pressure. 10 years ago, OCD was considered a rare and treatment-refractory disorder. Recent well designed studies document a lifetime prevalence rate for OCD of more than 2% in the general population. The outlook for patients with OCD has changed in the last decade, with many well controlled studies showing that OCD patients respond to specific behavioural and pharmacological treatments. The specific form of behavioural therapy is in vivo exposure coupled with response prevention. Only serotonin reuptake inhibitors, such as clomipramine, fluoxetine and fluvoxamine, are effective in the treatment of both depressed and not depressed OCD patients. Fluoxetine and fluvoxamine lack the anticholinergic side effects of clomipramine and, thus, provide an alternative treatment for patients who cannot tolerate clomipramine. Other nonserotonergic antidepressants (tricyclics and monoamine oxidase inhibitors) and anxiolytic agents have not been found to be consistently effective in this disorder. Insufficient data on the efficacy of neuroleptics and their potentially irreversible side effects limit their use in OCD patients. Behavioural and the pharmacological treatment are complementary, and a combination of the 2 therapies is apparently more effective than either modality alone.


Optimum management of the discharging ear.

Discharge from the ear can be the result of many disease processes. The ear may discharge blood, pus, cerebrospinal fluid (CSF) or wax. Keratosis obturans, stenosis of the external meatus and benign tumours of the external meatus all lead to wax build-up, which may cause recurrent attacks of otitis externa. Malignant tumours, such as basal cell carcinoma, squamous cell carcinoma and tumours of ceruminous gland origin may also present with discharge. Tumours should be excluded by submitting all material removed from the external canal for histological examination. Single or multiple abscesses (known as furuncles) may occur in the hair follicles in the skin of the external acoustic meatus (EAM). Compulsive scratching, hearing aids and foreign bodies placed in the ear predispose to otitis externa, which is also often associated with infection by Pseudomonas aeruginosa, Staphylococcus aureus and faecal organisms. Management may be with aluminium acetate 14%, topical antibiotic/steroid drops, a gauze wick soaked with icthammol 10% in glycerin or polymyxin B sulphate--neomycin sulphate--hydrocortisone acetate cream placed into the EAM and replaced every 24 to 48 hours, or systemic antibiotics according to severity. Malignant (necrotising) otitis externa causes progressive destruction of the temporal bone, and cranial nerve palsies (usually facial first). Treatment is limited debridement of infected bone, accompanied by intravenous aminoglycosides, and local antibiotic treatment and aural cleanout or oral ciprofloxacin. Middle ear conditions causing discharge include acute otitis media, infected grommets, traumatic perforations and chronic suppurative otitis media, as well as tumours of the ear canal skin and middle ear, radiation-induced otitis externa and osteoradionecrosis of the temporal bone, tuberculosis, Langerhans cell histiocytosis, spontaneous or post-traumatic CSF leaks, Wegeners granulomatosis and immune deficiency states. Topical application of aminoglycoside antibiotics to the middle ear of laboratory animals such as rats, guinea pigs and chinchillas causes sensorineural hearing loss, an effect rarely seen clinically in humans. If the external acoustic meatus and tympanic membrane are obscured by discharge cotton buds, microsuction equipment or syringing are used to remove it. It is often useful to initiate treatment (usually with topical drops, wicks or an oral antibiotic) with a provisional diagnosis. A full examination and adequate visualisation of the tympanic membrane must eventually be performed, if necessary under anaesthesia, or else serious progressive conditions may be neglected. The most useful initial investigation is a swab sent for bacteriological assessment; other investigations are usually indicated by clinical findings and the provisional diagnosis.


Halofantrine. A review of its antimalarial activity, pharmacokinetic properties and therapeutic potential.

Halofantrine is an orally administered blood schizontocide which is active against both chloroquine-sensitive and chloroquine-resistant plasmodia. Dose-finding and noncomparative clinical trials have confirmed the efficacy of halofantrine in the treatment of falciparum malaria in areas of chloroquine- and sulfonamide/pyrimethamine-resistant malaria and vivax malaria. However, poor results obtained in patients who failed mefloquine prophylaxis suggest that the efficacy of halofantrine may not extend to mefloquine-resistant P. falciparum, although more studies are needed to confirm this. Data concerning halofantrine in the treatment of P. ovale and P. malariae infections are still limited. One comparative study indicates that halofantrine has an efficacy equivalent to that of mefloquine and may be better tolerated. Halofantrine is generally well tolerated in both adults and children, the most common drug-associated effects being abdominal pain, pruritus, vomiting, diarrhoea, headache and rash, although it is difficult to distinguish between disease- and treatment-related events. The development of parasite resistance to halofantrine, like other blood schizontocides, is inevitable. Poor absorption resulting in variable peak plasma halofantrine concentrations, and possible cross-resistance with mefloquine, may accelerate the emergence of resistance to halofantrine. Thus, it is of primary importance that halofantrine is used only in areas where chloroquine- and sulfonamide/pyrimethamine-resistance are established in order to preserve and sustain its efficacy. If used with care, halofantrine will provide an important treatment option for falciparum malaria, a widespread parasitic disease associated with considerable morbidity against which the number of effective drugs available is being increasingly compromised by the spread of resistance.


Terbinafine. A review of its pharmacodynamic and pharmacokinetic properties, and therapeutic potential in superficial mycoses.

Terbinafine is an orally and topically active allylamine antifungal agent with a primarily fungicidal action in vitro. Its spectrum of in vitro activity includes a broad range of dermatophyte, filamentous, dimorphic and dematiaceous fungi, and some yeast species. In clinical trials, mycological and overall efficacy rates of around 90 and 80%, respectively, have been achieved in cutaneous dermatophyte infections (tinea corporis/cruris and tinea pedis) with terbinafine, administered either orally (250 or 500 mg/day) or topically (a 1% cream applied twice daily). Similar rates of cure have been obtained with oral terbinafine in dermatophyte nail infections after relatively short treatment periods ranging from 3 to 12 months. Topical terbinafine has been effective in approximately 80% of patients with cutaneous candidiasis or pityriasis versicolor. Few comparative data have been published, but generally oral terbinafine appeared to be at least as effective as oral griseofulvin or ketoconazole in tinea corporis/cruris and more effective than griseofulvin in tinea pedis. Both oral and topical terbinafine have been very well tolerated in clinical trials to date, with only minor adverse effects reported. Although further research is required to establish the efficacy of terbinafine in comparison with other available therapies, as well as to fully clarify its tolerability profile, the early results obtained with terbinafine in superficial fungal infections are very encouraging. Terbinafine appears likely to become a first-line therapy for dermatophyte infections, particularly those affecting the nails.


Steady-state visual evoked responses in high and low alpha subjects.

Reactivity to photic stimulation was studied in 16 subjects who were divided into 2 groups based on the relative amounts of spontaneous EEG alpha produced during a 4 min eyes-closed baseline condition. During the experimental session the subjects were presented with thirteen 1 min periods of sinusoidally modulated light stimulation at frequencies ranging from 2 Hz below their spontaneous peak alpha frequency (SPAF) to 2 Hz above in 0.33 Hz intervals. Using FFTs, steady-state visual evoked responses (VERs) were extracted from each subject's EEG for each condition. VER magnitude for high alpha subjects varied with the proximity of the stimulus frequency to the SPAF, the VER for low alpha subjects did not. Conversely, low alpha subjects showed a similar effect in side-band alpha activity (once the VER had been extracted) whereas high alpha subjects did not. The results are explained in terms of a possible difference in the coupling strength between thalamic and cortical alpha sources.


Physiological functions of the ascending spinal tracts in HTLV-I-associated myelopathy (HAM).

Physiological functions of the spinothalamic tract (STT) and the posterior column (PC) were studied in 19 Japanese patients with HTLV-I-associated myelopathy (HAM). The former was evaluated by pain-related somatosensory evoked potentials (SEPs) following CO2 laser stimulation, and the latter by conventional SEPs following electrical stimulation. Conduction velocity of STT and PC was significantly decreased in 9 and 14 patients, respectively, and more than half of them showed no clinical impairment of sensations (subclinical abnormality).


Quantified EEG and cortical evoked responses in patients with chronic traumatic frontal lesions.

Eighteen frontal trauma patients and 17 age-matched control subjects had quantified EEGs and measurements of sensory (SEP) and auditory evoked potentials (P300) using a Biologic Brain Atlas III system. The findings were compared to the conventional paper EEG, and to the frontal lesion volumes, severity of head injury, and outcome variables. The quantified EEG confirmed the pathological findings detected by visual inspection, but some regional abnormalities were more easily detected by topographic mapping. The regional distribution of pathological slowing corresponded well with the morphological lesions in most patients. The modal frequency of EEG correlated both with lesion volume and injury severity and with the outcome variables. There were no pathological findings in the SEPs, and all but one patient had clearly distinguishable P300 responses. There was a significant reduction in P300 amplitude in the frontal patients at the anterior, but not at the posterior electrodes. The topographical distribution of the P300 changes corresponded well with the morphological lesions. Our findings indicate that the P300 potential is, in part, dependent upon the prefrontal cortical areas. The present study thus supports P300 investigations which have shown amplitude reduction in other disorders (e.g., schizophrenia) with a presumed prefrontal dysfunction.


Effects of temporal-parietal lesions on the somatosensory P3 to lower limb stimulation.

Temporal-parietal junction lesions reduce the auditory and upper limb somatosensory P3 event-related potential (ERP) to target and novel stimuli. The current study examined the somatosensory P3 to target and novel stimuli delivered to the sole of the foot in patients with unilateral temporal-parietal lesions (n = 6). Age-matched controls (n = 10) generated a parietal maximal target P3 and a frontal-central maximal novelty P3 ERP to foot stimulation. Unilateral temporal-parietal lesions abolished target and novelty P3 responses over all scalp sites for stimuli delivered contralateral to the lesion. The P3 was also reduced to ipsilateral stimuli at electrodes over the lesioned hemisphere with partial P3 preservation observed at electrode sites over the non-lesioned hemisphere. These results parallel the findings for upper limb stimulation and support the critical role of temporal-parietal cortex in P3 generation.


AEPs at the onset and offset of repetitive sound modulation, due to mismatch with the contents of an auditory sensory store.

Long latency auditory evoked potentials (AEPs), chiefly consisting of a negative peak at about 150 msec and a positivity at 250 msec, were recorded at the beginning and end of periods during which the interaural time difference of binaural noise was switched between 0.0 and 0.8 msec at a fast rate (ISI = 50 or 25 msec) or the frequency of continuous binaural clicks was switched between 167 and 200 Hz every 80, 50 or 25 msec. In the latter case the offset responses occurred later than onset by a mean of 89, 47 and 27 msec respectively, suggesting they were probably generated at the moment the next switch was expected but failed to occur. The offset responses must be non-specific with respect to the interaural delay or the frequency of clicks, since neurones which respond to particular delays or frequencies and are made refractory by a rapid rate of stimulation should not suddenly become less so at the last in a series of identical stimuli, or be activated by the absence of a further event. It is proposed that the potentials are due to a higher order of neurone which automatically responds to the occurrence of a "mismatch" between the immediate sound and an image of that which was previously present, encoded in a short-term sensory store. In addition to frequency content and interaural delay, the image must contain information about the temporal modulation pattern of the sound over the previous few seconds.


Effects of noise bandwidth on the late-potential masking level difference.

The masking level difference (MLD) is a psychoacoustic phenomenon derived from the subtraction of S pi No thresholds (signals pi radians out of phase and noise in phase at the two ears) from SoNo thresholds (signals and noise in phase at the two ears). The purpose of this study was to determine if the MLD derived from the late components (P1, N1, P2, N2) of the auditory evoked potentials was a physiological correlate of the behavioral MLD. Subjects were 15 young adults with normal hearing. Comparisons were made between behavioral and late potential thresholds to 500 Hz stimuli in So and S pi conditions in quiet, and to 500 Hz stimuli in SoNo and S pi No conditions in narrow band (50 Hz) and wide band (600 Hz) noise. No significant differences were seen for behavioral or late-potential thresholds to So and S pi conditions. The S pi No thresholds was significantly lower than the SoNo threshold, yielding an MLD for both the behavioral and physiological responses. The magnitudes of both the behavioral and late-potential MLD were larger with the narrow band noise than with the wide band noise. Evidence, therefore, is provided that the late-potential MLD reflects similar processes as are responsible for the behavioral MLD.


Human hand and lip sensorimotor cortex as studied on electrocorticography.

We investigated functional topography of human hand and lip sensorimotor cortex using somatosensory evoked potentials (SEPs) from chronically indwelling subdural grid electrodes (ECoG) in 3 epilepsy patients during stimulation of median nerve, ulnar nerve, and lower lip. We used dipole modeling to determine the cortical location of each peripheral sensory field. The cortical locations were in the postcentral gyrus and showed a clear somatotopic organization from medial superior to lateral inferior in the order: ulnar nerve, median nerve, and lip. The source localizations agreed with the results of cortical stimulations and anatomical features on intraoperative photographs. The cortical regions of median and ulnar nerve each could be modeled by sequential tangential and radial dipoles. The cortical region of lip was different and could be explained mostly by tangential dipoles. These findings suggest a difference in the cortical organization of human lip and hand sensory cortex and are consistent with a larger representation of lip in the posterior bank of central fissure in area 3b than on the gyral surface in area 1, similar to findings in macaque. Further studies in a larger population of patients with ECoG or normal subjects with scalp-EEG and MEG are warranted to test this hypothesis.


Midlatency auditory evoked responses: P1 abnormalities in adult autistic subjects.

MLR recordings from a group of 11 high-functioning adult autistic subjects were compared with those from a control group of 11 normal subjects. Components selected for analysis were "Pa", the maximum positivity in the 25-40 msec latency range following stimulus onset, "P1", the maximum positivity within the 50-65 msec latency range, and "Nb," the maximum negative deflection in the 40-50 msec latency range. Statistical analyses of amplitude and latency data were conducted using repeated measures analysis of variance and t test group comparisons. The Pa component showed no significant difference between autistic and control groups. However, 2 types of abnormality were noted in the P1 component: (1) the P1 component was significantly smaller in the autistic subjects at slow rates of stimulation, and (2) the autistic P1 did not change as rates of click stimulation increased from 0.5 to 10/sec, in contrast to the normally produced P1 decrement. Data from the P1 model in the cat, and complementary data from the human, closely link the generator substrate of the P1 potential to cholinergic components of the ascending reticular activating system (RAS) and their thalamic target cells. This is the first report of abnormal P1 responses in autism and strongly suggests that the RAS and/or its post-synaptic thalamic targets may be dysfunctional in this syndrome.