Interaction of prenatal exposure to cigarettes and MAOA genotype in pathways to youth antisocial behavior.

Genetic susceptibility to antisocial behavior may increase fetal sensitivity to prenatal exposure to cigarette smoke. Testing putative gene x exposure mechanisms requires precise measurement of exposure and outcomes. We tested whether a functional polymorphism in the gene encoding the enzyme monoamine oxidase A (MAOA) interacts with exposure to predict pathways to adolescent antisocial behavior. We assessed both clinical and information-processing outcomes. One hundred seventy-six adolescents and their mothers participated in a follow-up of a pregnancy cohort with well-characterized exposure. A sex-specific pattern of gene x exposure interaction was detected. Exposed boys with the low-activity MAOA 5' uVNTR (untranslated region variable number of tandem repeats) genotype were at increased risk for conduct disorder (CD) symptoms. In contrast, exposed girls with the high-activity MAOA uVNTR genotype were at increased risk for both CD symptoms and hostile attribution bias on a face-processing task. There was no evidence of a gene-environment correlation (rGE). Findings suggest that the MAOA uVNTR genotype, prenatal exposure to cigarettes and sex interact to predict antisocial behavior and related information-processing patterns. Future research to replicate and extend these findings should focus on elucidating how gene x exposure interactions may shape behavior through associated changes in brain function.

Renal localization of the membrane attack complex in systemic lupus erythematosus nephritis.

The membrane attack complex (MAC) of the complement system was localized in both glomeruli and peritubular regions of 22 kidneys manifesting systemic lupus erythematosus (SLE) nephritis. A similar distribution was observed for immune complex markers (IgG, Clq, and C3) and MAC in glomeruli, although the deposits of MAC were more discrete and showed lesser immunofluorescence staining intensity compared with immunoglobulins and complement components. In contrast, peritubular immune complexes were present in only 7 out of 22 kidneys, involved comparatively small clusters of tubules, exhibited weaker immunofluorescence staining than MAC, and failed to correlate with interstitial foci of inflammation. Granular or irregular, linear aggregates of the MAC were observed at the periphery of larger groups of tubules contiguous to areas of interstitial inflammation. Comparable amounts of IgG, Clq, C3, and MAC were present in blood vessel walls in areas of fibrinoid necrosis. These data suggest that the MAC is a direct mediator of tissue injury occurring in renal glomeruli, tubules, and blood vessels. The discordance between immune complexes and MAC localized in the peritubular region, but not in glomeruli or blood vessels, raises the possibility that both immune complexes and nonimmune agents, such as bacterial antigens, may activate the classical or alternative complement pathways and thereby play a role in the pathogenesis of tubulointerstitial lesions of SLE nephritis.

Localization of the membrane attack complex (MAC) in experimental immune complex glomerulonephritis.

The role of the membrane attack complex (MAC) as a mediator of renal tissue injury was evaluated in rats affected by bovine serum albumin (BSA)-induced immune complex glomerulonephritis. Immunofluorescence studies revealed concurrent deposits of IgG, BSA, C3, and the MAC along glomerular capillary walls, although the MAC manifested a more restricted distribution than that observed for immune complexes. Immunoelectron microscopic techniques were utilized to demonstrate immune complexes, C3, and the MAC within dense deposits in the subepithelial aspect of the basement membrane. Visceral epithelial foot processes were fused in areas overlying large dense deposits and exhibited intense staining for the MAC, lesser reactivity for C3 but IgG was absent from the foot process membranes. Smaller granular deposits of immune complexes, C3, and the MAC were observed in the subendothelial region of the lamina rara interna and the lamina densa. Immune complexes may activate the classical complement pathway causing diffuse injury to the glomerular basement membrane (GBM), allowing subepithelial accumulation of complexes. These observations implicate the MAC as a mediator of GBM and juxtaposed podocyte membrane injury, thereby contributing to disruption of the glomerular filtration barrier. IgG and C3 were demonstrated within tubulointerstitial regions on the surface of collagen fibers in close proximity to the tubular basement membrane (TBM) of proximal convoluted tubules. Within the TBM, C3 localization was prominent with diminished reactivity for the MAC, but IgG was not detectable. The demonstration of C3 and scant MAC deposits in the TBM of nonimmunized control rats without evidence of interstitial IgG and C3 deposits suggests that both nonimmune and immune processes play a role in the pathogenesis of extraglomerular lesions. Evidence derived from these morphologic studies indicates that the MAC is associated with injury to the GBM, foot process membranes of visceral epithelium, and the TBM. Further experiments designed to selectively enhance or inhibit the deposition of MAC and assess consequent renal dysfunction are required to substantiate hypotheses concerning the in vivo membranolytic potential of the MAC in experimental immune complex glomerulonephritis.

C5b-9 dimer: isolation from complement lysed cells and ultrastructural identification with complement-dependent membrane lesions.

The membrane attack complex (MAC) of complement was extracted from the membranes of cells lysed by human complement and its properties were compared with those of the fluid phase complex SC5b-9. Upon sodium dodecyl sulfate polyacrylamide gel electrophoresis and immunochemical analysis, the two isolated complexes had identical subunit compositions, except that the MAC lacked the S-protein. The sedimentation coefficient and molecular weight of the extracted and isolated MAC were, respectively, 33.5 S and 1.7 x 10(6) daltons, compared to 23 S and 1.0 x 10(6) dalton for SC5b-9. Because the molecular weight of the MAC is approximately two times greater than that of C5b-0 (800,000 daltons), the MAC is considered the dimer of C5b-9. Under specified conditions, the 33.5 S dimer could be converted to the 23 S monomer without dissociation of subunits. The MAC had the electron microscopic appearance and dimensions that are characteristic for the complement produced ultrastructural membrane lesions. SC5b-9 had a different ultrastructure that is dissimilar to the morphology of the lesions. The isolated MAC could be reincorporated into phospholipid bilayers and assumed on the surface of the resultant lipid vesicles the orientation and appearance of typical complement lesions.

Membrane attack complex of complement: a structural analysis of its assembly.

This study was conducted to gain insight into the process of assembly of the membrane attack complex (MAC) of complement through structural analysis. Four intermediate complexes and the MAC were examined by electron microscopy and by sucrose density-gradient ultracentrifugation. The C5b-6 complex has a sedimentation rate of 11S, an elongated, slightly curved shape and dimensions of 160 x 60 x 60 A. At protein concentrattions greater than 1 mg/ml, and physiologic ionic strength and pH, the complex forms paracrystals that have the appearance of parallel strands. Equimolar quantities of C5b-6 and C7 mixed in the absence of lipids or detergents give rise to C5b-7 protein micelles which are soluble in aqueous media and have a sedimentation rate of 36S, suggesting a tetrameric composition. Ultrastructurally, C5b-7 protein micelles consist of four half-rings, each measuring 200 x 50 A, which are connected to one another by short stalks extending from the convex side of the half-rings. C5b-7 bound to dioleoyl lecithin (DOL) vesicles has a similar ultrastructural appearance. After extraction with deoxycholate (DOC), C5b-7 has a sedimentation velocity of 36S which further suggests the occurrence of C5b-7 in the form of tetrameric protein micelles. Attachment of C8 to vesicle-bound C5b-7 results in dissociation of the protein micelles. An individual C5b-8 complex appears as a half-ring attached to the DOL-vesicle via a 100-A-long and 30-A-wide stalk. After extraction from the DOL-vesicles with DOC, C5b-8 has a sedimentation velocity of approximately 18S. Binding of C9 to DOL-vesicle bound C5b-8 induces the formation of the typical ultrastructural complement lesions. C5b-9 extracted from the vesicles with DOC has a sedimentation rate of 33S, which is characteristic of the C5b-9 dimer. It is concluded that dimerization is a function of C9. C5b-9 monomers are visualized when a single C5b-9 complex or an odd number of complexes were bound per DOL-vesicle. The C5b-9 monomer has an ultrastructural appearance that is theoretically expected of a half-dimer: a 200- x 50-A half-ring which is attached to the DOL-vesicle by a 100- x 80-A appendage. Extracted with DOC, the C5b-9 monomer has a sedimentation rate of 23S. At a higher multiplicity of MAC per DOL-vesicle, large structural defects in the lipid bilayer are seen which are attributed to direct physical destruction of membranes by the known lipid-binding capacity of the MAC. It is proposed that protein micelle formation at the C5b-7 stage of MAC assembly and dissociation of these micelles upon binding of C8 are events that facilitate dimerization of C5b-9 and thus MAC formation.

Identification of an Arg-Gly-Asp (RGD) cell adhesion site in human immunodeficiency virus type 1 transactivation protein, tat.

Tat, the transactivation factor of human immunodeficiency virus type 1 (HIV-1), contains the highly conserved tripeptide sequence Arg-Gly-Asp (RGD) that characterizes sites for integrin-mediated cell adhesion. The tat protein was assayed for cell attachment activity by measuring the adhesion of monocytic, T lymphocytic, and skeletal muscle-derived cell lines to tat-coated substratum. All cell lines tested bound to tat in a dose-dependent manner and the tat cell adhesion required the RGD sequence because tat mutants constructed to contain an RGE or KGE tripeptide sequence did not mediate efficient cell adhesion. The tat-mediated cell attachment also required divalent cations and an intact cytoskeleton. In addition, cell adhesion to tat was inhibited in the presence of an RGD-containing peptide GRGDSPK or an anti-tat mAb that recognizes the RGD epitope. These results strongly suggest that cells are bound to tat through an integrin. Interestingly, myoblast cells bound to tat remained round, whereas the same cells attached through an integrin for a matrix protein typically flatten and spread. The role of this RGD-dependent cellular adhesion of tat in HIV-1 infection remains to be determined.

Structure of complement poly-C9 determined in projection by cryo-electron microscopy and single particle analysis.

The ring-like complement 'lesions' found on membranes of complement lysed cells comprise a complex of components C5b through C9 that coalesce to form hollow cylinders which penetrate the membrane bilayer and create lytic pores. Walls of these C5b-9 membrane attack complex cylinders may consist primarily of the C9 component, since samples of purified, isolated C9 can polymerize into cylindrical structures which appear identical with the fully assembled C5b-9 complex. The structure of these poly-C9 molecules has been investigated using the techniques of cryo-electron microscopy and single particle analysis. Sets of single poly-C9 particles viewed as rings were selected from cryo-EM images, then particles were aligned and treated by correspondence analysis to identify the principle interparticle similarities and variations. The highest ranking variation found was the presence or absence of a dense inner ring of protein density. Other important variations were interpreted as different types of particle tilt. These results were used in selecting a subgroup of untilted particles for averaging and symmetry analysis. The rotational power spectrum of the initial average suggested 13-fold symmetry. The 13-fold symmetry was used to select and group particles for further analysis. Individual particles were 13-fold rotational averaged and those with enhanced peripheral features were placed into either a right-handed subgroup or into a left-handed subgroup based on orientation of the peripheral features. Particles within each group were aligned and averaged, and a poly-C9 structure was produced which shows important structural details and from which the C9 monomer structure can be deduced. The poly-C9 structure contains a dense inner ring of diameter between 113-181 A and which is modulated into 13 discrete peaks with peak-to-peak separation of approx. 35 A. The dense inner ring is surrounded by a less dense, concentric outer rim extending to 254 A diameter. The outer rim contains projections that are contiguous with the inner peaks but are skewed relative to the ring radius to produce the appearance of a pin-wheel. These projections correspond with the peripheral features picked up in the rotationally averaged individual particles; the left- or right-handed orientation of projections may result from the up/down orientation of individual particles in ice. The C9 monomer structure within the cylinder is suggested by the density distribution. The monomer would be a rod with diameter of 35 A, oriented parallel to the cylinder axis and would be roughly perpendicular to a membrane.(ABSTRACT TRUNCATED AT 400 WORDS)

Immunology of myasthenia gravis.

Anti-acetylcholine-receptor antibody is demonstrable in more than 90 per cent of patients with myasthenia gravis. Serum antibody titers do not show a direct correlation with disease severity, although in certain patients antibody levels increase in association with disease activity. Impairment of neuromuscular transmission results from the loss of junctional receptors, either as a result of receptor internalization or destruction of junctional folds containing the acetylcholine receptor. Myasthenia gravis manifests immunologic, genetic, and clinical similarities to rheumatic syndromes, suggesting a generic immune dysfunction common to these disorders.

Patient acceptance of local anesthesia for breast biopsy.

Three out of every four patients undergoing breast biopsy found the outpatient local-anesthesia method acceptable in this study. Patient acceptance was greater among those who experienced less pain and anxiety. This suggests that acceptance could be increased by more complete preoperative explanation to the patient, adequate premedication, properly administered local anesthesia and gentle technique. Acceptance would also probably be far greater in a private practice setting than in the instititutional and sometimes impersonal setting of a large federal hospital. Breast biopsy under local anesthesia has not compromised survival rates or increased local recurrence. When it is done on an outpatient basis, hospital costs have been reduced at least threefold. It is apparent, then, that patient objection is not a deterrent to the use of breast biopsy under local anesthesia on an outpatient basis. For patients with dominant breast masses, this modality can be added to any plan of management which seeks to reduce patient risk and inconvencience, diminish hospital costs and alleviate bed demand all without impairing diagnostic accuracy or long-term survival.

The complement SC5b-9 complex mediates cell adhesion through a vitronectin receptor.

Adhesion of cells to the terminal complement complex of C5b through C9 containing the serum S-protein (SC5b-9) was investigated using a microtiter plate attachment assay with L8 myoblast indicator cells. The skeletal muscle-derived L8 myoblasts bound and spread on substratum coated with SC5b-9, and with the vitronectin/S-protein component of SC5b-9. The myoblasts did not adhere to substratum coated with collagen, laminin, or fibronectin. The cell attachment was blocked by antibody to vitronectin/S-protein, whereas antibody to the other components C5, C6, C7, C8, or C9 had minimal effect. The cells were not bound to free vitronectin because attachment activity was removed by adsorption with an anti-C6 antibody column. The L8 cell attachment was dependent on divalent cations, was blocked by synthetic peptides containing the amino acid sequence Arg-Gly-Asp, and was inhibited by antivitronectin receptor antibody. These results indicate that cells adhere to the SC5b-9 complex through interaction of the vitronectin component with an integrin vitronectin receptor. Cell attachment to terminal C complexes could be used for leukocyte adherence and migration during inflammation, and also for attachment of tissue cells during regeneration after disease or traumatic injury.

Resistance to experimental autoimmune myasthenia gravis in genetically inbred rats. Association with decreased amounts of in situ acetylcholine receptor-antibody complexes.

Genetically related susceptibility for experimental autoimmune myasthenia gravis was investigated in nine inbred strains of rats immunized with heterologous acetylcholine (AChR) from Torpedo californica. Wistar Munich and Fischer strain animals consistently developed severe, fatal disease associated with impaired neuromuscular transmission and increased sensitivity to low doses of curare. A lower incidence of disease was induced in Wistar Kyoto, ACI, Brown Norway, Buffalo, and Lewis strain animals. In contrast, Wistar Furth and Copenhagen strain animals were resistant to experimental autoimmune myasthenia gravis, electrophysiologic responses were normal, and animals were insensitive to curare. All strains of animals manifested equivalent amounts of serum antibody to AChR and total muscle AChR was reduced to the same extent in both resistant and susceptible animals. In contrast, the amount of antibody-bound AChR was greater in susceptible Wistar Munich animals than the amount observed in resistant Wistar Furth animals. These data suggest that impaired neurotransmission is correlated with the extent of antibody binding to the AChR. The discordance in the amount of antibody bound to the AChR of resistant and susceptible animals may result from heritable differences in antibody properties. Cross-breeding experiments with Wistar Munich and Wistar Furth animals show that resistance for development of experimental autoimmune myasthenia gravis is recessive and indicate that disease susceptibility is linked to one or two genetic loci.

Inhibition of acute passive transfer experimental autoimmune myasthenia gravis with Fab antibody to complement C6.

The role of the lytic complement C5b-9 membrane attack complex (MAC) in acute passive transfer experimental autoimmune myasthenia gravis (EAMG) produced in rats was investigated by in vivo inhibition of MAC formation with anti-C6 Fab. Anti-C6 Fab totally inhibited in vitro serum hemolytic activity, but did not consume or inhibit early complement pathways. Injection of rats with 0.12 mg/ml anti-C6 Fab reduced serum C6 to 8% and inhibited the muscle weakness, electrophysiologic abnormalities and loss of acetylcholine receptor (AChR) associated with acute EAMG. This level of C6 inhibition reduced the total serum complement hemolytic activity to 29% of normal but did not reduce the serum levels of complement components C3, C5, or C7. Treatment of rats with lower amounts of anti-C6 Fab (0.08 mg/ml) also inhibited clinical and electrophysiologic signs of EAMG, however, the lower amount of anti-C6 did not prevent the loss of muscle AChR. Both the higher and the lower amount of anti-C6 Fab inhibited the accumulation of macrophages at muscle motor end-plates. The inhibition by anti-C6 indicates that muscle weakness and electrophysiologic abnormalities associated with EAMG are dependent on the complement MAC, and that muscle weakness results from tissue injury in addition to loss of muscle membrane and AChR.

The ninth component of human complement: purification and physicochemical characterization.

A procedure for the isolation of the human complement (C) protein C9 is described. The procedure allowin. The purified protein has the electrophoretic mobility of an alpha-globulin, and is a single polypeptide chain with a m.w. of 71,000. No impurities were detected either on gel electrophoretic or immunochemical examination. C9 is a glycoprotein containing 7.8% carbohydrate, and in terms of residues per mole, 3.0 glucosamine, 17.6 neutral hexose, and 7.4 sialic acid. Its amino acid composition is typical of a globular serum protein. Upon automated Edman degradation of reduced and alkylated C9, no amino acid residues were released, suggesting a blocked N-terminus. The concentration of C9 in normal human serum is 58 +/- 8 microgram/ml. A high titer rabbit antiserum was produced and employed to immunochemically deplete serum of C9. The CH50 of the C9-depleted serum was identical to that of whole human serum; however, membrane fragments of erythrocytes lysed by C9-depleted serum lacked the typical ultrastructural C lesions, which constitute the dimeric membrane attack complex.

Complement activation in muscle fiber necrosis: demonstration of the membrane attack complex of complement in necrotic fibers.

The membranolytic C5b-9 complement membrane attach complex (MAC) is assembled after activation of either the classic or the alternative complement pathway. The quaternary configuration of the MAC macromolecule presents neoantigenic determinants not present on precursor molecules. Consequently, antibodies specific for these neoantigen(s) do not detect nonspecifically bound native complement precursors of MAC. By means of antibodies rendered specific for MAC neoantigen(s), MAC was localized by the immunoperoxidase reaction in cryostat sections of human muscle. In 66 biopsy specimens containing necrotic muscle fibers (Duchenne dystrophy, 13; other dystrophies, 15; inflammatory myopathies, 31; miscellaneous myopathies, 7) all of the necrotic fibers reacted for MAC neoantigen(s). C3 and C9 were also consistently localized in necrotic fibers, but localization of C1q, C4, and IgG was variable and often did not exceed background staining. None of the nonnecrotic fibers reacted for immunoglobulin or complement. Detection of MAC neoantigen(s) in necrotic fibers in a wide variety of muscle disease unambiguously shows that (1) the lytic complement pathway is consistently activated and participates in muscles fiber necrosis in vivo, and (2) complement reaction products are generated than can stimulate cellular infiltration and phagocytosis of the necrotic fiber. The findings also suggest that cell necrosis in general may involve participation of complement.

Infancy predictors of emotional availability in middle childhood: the roles of attachment security and maternal depressive symptomatology.

This investigation examined the longitudinal prediction of emotional availability in mother-child interaction during middle childhood from two indicators of socioemotional functioning in infancy: security of infant-mother attachment; and maternal depressive symptoms. Forty-five children and their mothers were seen during infancy: security of attachment was assessed in the laboratory Strange Situation; and mothers completed a self-report of depressive symptoms. At age 7, children were observed with their mothers in a lab playroom. The dyad's emotional availability was assessed during reunion following an hour-long separation. Results demonstrated significant associations between infancy and middle childhood socioemotional organization, both for mothers and for children. Security of attachment in infancy was related to maternal sensitivity and structuring, and to child responsiveness and involvement at age 7. Maternal depressive symptoms in infancy were associated with maternal sensitivity and structuring at age 7. Greatest differentiation was found between infants with secure attachments and those with insecure-disorganized attachments.