Hepatitis C virus long-term persistence in peripheral blood mononuclear cells in patients with haemophilia. Detection of occult genotype 1.

Peripheral blood mononuclear cells (PBMC) from chronic hepatitis C virus-infected persons can harbour viral variants that are not detected in plasma samples. We explored the presence and persistence of HCV genotypes in plasma and PBMC cultures from 25 HCV-monoinfected and 25 HIV/HCV-coinfected patients with haemophilia. Cell cultures were performed at different time points between 1993 and 2010-2011, and the HCV genome was examined in culture supernatants. Sequential plasma samples were studied during the same time period. Analysing sequential plasma samples, 21% of patients had mixed-genotype infections, while 50% had mixed infections determined from PBMC culture supernatants. HIV coinfection was significantly associated with the presence of mixed infections (OR = 4.57, P = 0.02; 95% CI = 1.38-15.1). In our previous study, genotype 1 was found in 72% of 288 patients of this cohort. Similar results were obtained with the sequential plasma samples included in this study, 69% had genotype 1. However, when taking into account plasma samples and the results from PBMC supernatants, genotype 1 was identified in 94% of the population. The PBMC-associated variants persisted for 10 years in some subjects, emphasizing their role as long-term reservoirs. The presence of genotype 1 in PBMC may be associated with therapeutic failure and should not be disregarded when treating haemophilic patients who have been infected by contaminated factor concentrates. The clinical implications of persistent lymphotropic HCV variants deserve further examination among multiple exposed groups of HCV-infected patients.

Severe haemophilia A patients have reduced numbers of peripheral memory B cells.

The development of inhibitors is a complication of replacement treatment in Haemophilia. Loss of factor VIII-specific memory B cells in the spleen is associated with down regulation of antibodies in mice treated with high doses of FVIII, but changes in B cell memory have not been described in haemophilic patients. The aim of this study was to evaluate the phenotype of circulating lymphocytes in severe haemophilia A. Twenty patients with inhibitors (PI), 22 without inhibitors (P), nine patients during immune tolerance induction (ITI) treatment and 20 healthy donors (HD) were included. Peripheral blood lymphocytes were examined using flow cytometry. Anti-FVIII antibodies were measured using Bethesda and flow cytometry. Percentages of T subsets and B lymphocytes were similar in all groups. In contrast, memory B cells (CD27+) were decreased in PI and P compared with HD, but the level of significance was higher in PI (P = 0.001) than P (P = 0.01). PI with high level of anti-FVIII antibodies presented the lowest B memory values. CD70 expression was also lowest in PI. Non-switched CD27+ subpopulation (IgD+) was prevalent in PI, but did not show statistical significance. When ITI failed, the percentages of CD27+ B cells after 12 months of ITI were lowest. In a longitudinal study performed in four patients, an increased percentage of CD27+ and CD70+ B cells during ITI was found. This work suggests that different peripheral lymphocyte markers, such as CD27 and CD70 on B cells, may be helpful to evaluate anti-FVIII response and to monitor the success of ITI.

A flow cytometry evaluation of anti-FVIII antibodies: correlation with ELISA and Bethesda assay.

In this study, we describe a flow cytometry (FC) system for detecting antibodies to factor VIII (FVIII) and compare its results with those of enzyme-linked immunosorbent assay (ELISA) that detects both inhibitory (I-Ab) and non-inhibitory (NI-Ab) antibodies and the Nijmegen modification of the Bethesda method, detecting I-Ab. FC was set up in our laboratory. Recombinant FVIII (rFVIII) was coupled to microspheres (FVIII-m) and reacted with different plasma dilutions. Microspheres without rFVIII were used as control (control-m). Captured anti-FVIII antibodies were detected using anti-human IgG. Plasma samples from the following patients with severe haemophilia A (SHA) patients were evaluated: 17 P (patients without I-Ab, <0.5 BU mL(-1)); 13 PI (patients with I-Ab, 1.1-8200 BU mL(-1)). Of these 13, two PI were referred during immune tolerance induction (ITI), and plasmas from 12 healthy donors (HD) were evaluated. Semiquantitative results were given as an index (the highest mean fluorescence intensity ratio between FVIII-m and control-m multiplied by the inverse of the corresponding plasma dilution). Both plasma and serum were suitable for the test. FC agreed with the Bethesda method (r = 0.8; P = 0.0001). FC and ELISA had 80% of coincidence. Four of 17 patients (23.5%) had NI-Ab by FC, and two of them developed high levels of I-Ab later on. This test provides a useful alternative for measuring FVIII antibodies supplementing Bethesda assay. FC is fast and easy to perform. No more than 200 µL of plasma or serum is required especially making it useful for paediatric patients.

C1r, subunit of the first complement component: purification, properties, and assay based on its linking role.

A method to obtain C1r, a subunit of the first complement component, in a highly purified state has been described for the first time. The stepwise method starts with a neutral euglobulin precipitation, after diethylaminoethyl- and carboxymethyl-cellulose chromatography and a final preparative polyacrylamide electrophoresis step. Such C1r preparations are devoid of C1q and C1s activities and show only one protein band on analytic polyacrylamide electrophoresis. Rabbits injected with this preparation produced antisera showing only one precipitation band. The stability of C1r activity was determined under different conditions, and C1r was found to be labile at 37 degrees C, pH 7-8 and low ionic strength. The electrophoretic mobility of purified C1r is that of a beta-globulin on disc acrylamide electrophoresis and on agarose electrophoresis at pH 8.6. Its molecular weight as estimated by sephadex chromatography is 168,100.A sensitive hemolytic assay based on the property of C1r to link C1s to C1q and thereby to generate macromolecular C[unk]1 is described. The number of C[unk]1 molecules generated is stoichiometrically related to the concentration of C1r for a fixed C1q and C1s concentration provided that the titration is carried out below the plateau zone. Macromolecular C1 can be separated from free C1s as the former is cell bound. This method of purification and assay should allow the development of monospecific antisera and further chemical study of C1r.

IgG subclasses of FVIII inhibitors in an argentine cohort of severe hemophilia A patients: Analysis by flow cytometry.

INTRODUCTION: FVIII inhibitors consist of a polyclonal population of antibodies. Previous studies have demonstrated different distribution of IgG subclasses. IgG4 was associated to high level of FVIII inhibitors and failure of immune tolerance induction (ITI) treatment. This study monitored the relative distribution of IgG subclasses of anti-FVIII in patients with severe hemophilia A (SHA). METHODS: Anti-FVIII antibodies were measured employing an immunomethod, developed in our laboratory, that combines flow cytometry (FC) with microspheres coupled (FVIII-m) or not (Control-m) to FVIII. Seventy-five patients with SHA were studied, 17 without inhibitors (Group I); 58 with inhibitor history, 13 low responders: (LR: Group II), and 45 high responders (HR: Group III). Eight patients undergoing ITI were also included. RESULTS: We found anti-FVIII antibodies in 11 of 27 patients (40%) without inhibitors and in 45 of 48 with inhibitors at the moment of the study. IgG4 was predominant only in the Group III: P=0.02 in patients with low level of inhibitors and P=0.0001 with high titer of inhibitors. Longitudinal analysis performed on patients undergoing ITI showed a gradual decrease of IgG4 values that was associated to improvement of clinical parameters during treatment. CONCLUSION: We suggest the use of the FC method to supplement functional traditional assays and to help to improve the management of patients with SHA.

Biphenotypic acute leukemia with t(15;17).

Biphenotypic acute leukemias (BAL) represent 5% of all acute leukemias. The most frequent cytogenetic abnormalities described are Philadelphia chromosome and 11q23 involvement. Here we report a BAL case, with blasts showing lymphoblast morphology and positivity for myeloperoxidase (in 6% of the blast cells). Immunophenotype revealed the compromise of myeloid and B-lymphoid lineages. Cytogenetic analysis showed the t(15;17) and 8 trisomy. PML/RARa rearrangement was detected by fluorescent in situ hybridization (FISH) on interphase nuclei while PML/RARa fusion transcript was detected in the bone marrow and peripheral blood by molecular biology studies (RT-PCR). This report describes a BAL case with an unfrequent cytogenetic abnormality, and highlights the importance of correlating the results of multiple diagnostic methods in order to establish a correct diagnosis and treatment in BAL patients.

Tyrosine kinase regulatory action on ileal muscarinic effects of IFN-gamma.

Interferon-gamma (IFN-gamma) is a pleiotropic cytokine that has a large number of immunologic and nonimmunologic functions. We have described that IFN-gamma could activate muscarinic cholinergic receptors (mAchR) of rat intestine, stimulating ileal motility. We also observed that mAchR activation induced inhibition of cAMP levels and stimulation of cGMP formation. The objectives of our work were to clarify the signal transduction pathways involved in regulation of ileal motility through mAchR activation by IFN-gamma. Our results demonstrate that this cytokine produces an ileal cholinergic response through tyrosine kinase activity. The activation of tyrosine kinase mediates ileal contractility, phosphoinositide hydrolysis by phospholipase C, nitric oxide synthase via protein kinase C, and cGMP synthesis. The increment in ileal motility is probably due to hyperproduction of prostaglandin E2 (PGE2) by ileal tissue. This prostanoid is an important mediator because it stimulates ileal motility. We conclude that IFN-gamma not only immunomodulates the gut microenvironment but also exerts a local nonimmunologic regulation on intestinal motility.

Reservoirs of HIV replication after successful combined antiretroviral treatment.

Sustained reduction of viral replication can be achieved in HIV infected patients after treatment with combinations of drugs (HAART) that inhibit the viral reverse transcriptase, and protease enzymes. However, replication competent virus can still be recovered from latently infected resting memory CD4+ T-cell lymphocytes. Moreover, "covert" virus replication has been demonstrated in patients who experienced reductions in plasma viremia to levels below the limit of detection of the most sensitive PCR assays. In most studies, preferential attention has been given to latent resting CD4+ T-lymphocytes as a source of HIV persistence. However, insufficient suppression of HIV replication could also lead to viral re-emergence after HAART interruption. In addition to CD4+ T- lymphocytes, other host cells such as long-lived resident macrophages or recently infected blood monocytes could also contribute to maintain persistent HIV replication after HAART. Establishing the origin of re-emerging HIV in patients under HAART upon treatment interruption is important to design optimal treatment schemes. Therapeutic strategies aimed at reducing the number of latently infected cells involve immune activation with IL-2, or other stimulatory factors, in the presence of antiretroviral drugs. Elimination of replication-competent virus would require intensification of HAART, or the use of antiretroviral drugs achieving an effective concentration at the site of HIV replication. In this review the mechanisms of HIV persistence and the methods that can be used to distinguish latent from covert HIV replication in different cell types will be discussed.

Expression of alpha-adrenoceptors in a human transformed lymphoblastoid cell line.

The presence of alpha-adrenergic receptors (absent in normal lymphocytes) has been demonstrated in transformed human lymphocytes of the Raji cell line. Binding properties of beta-adrenergic receptors were similar to those reported for normal lymphocytes. A single population of alpha 2-adrenergic receptors was characterized in intact Raji lymphoblasts by binding and saturation assays with the alpha 2-adrenergic antagonist yohimbine. Competition curves with [3H]yohimbine indicate the presence of typical alpha 2-adrenoceptors. Reaction of Raji with the alpha 2-adrenergic agonist clonidine (10(-6) M) stimulated their growth rate. In contrast, the alpha 1-adrenergic agonist methoxamine (10(-6) M) had no effect. Previous work indicates that Raji can actively produce thromboxanes (TX) and that these decreased atrium contractility. In agreement with these results and with the binding studies, it is now shown that clonidine stimulation enhanced the negative inotropic effects of Raji on isolated rat atria. This reaction was prevented by incubation of Raji with yohimbine (10(-6) M) but not with the alpha 1-adrenergic antagonist prazosin (10(-6) M) or the beta-adrenergic antagonist propranolol (10(-7) M). The biologic effect of Raji on rat atria was probably due to production of cyclooxygenase metabolites of arachidonic acid, because it was blocked by preincubation of the cells with the cyclooxygenase inhibitors indomethacin (10(-6) M) and aspirin (10(-4) M) or the thromboxane synthetase inhibitors nictindol (10(-5) M) and imidazole (10(-4) M).(ABSTRACT TRUNCATED AT 250 WORDS)

Cholinergic response of isolated rat atria to recombinant rat interferon-gamma.

Addition of recombinant rat interferon-gamma (IFN-gamma) to beating rat atria decreased the contractile strength in a dose-dependent manner. The effect was specific of IFN-gamma since it was abrogated by monoclonal anti-rat IFN-gamma. It required the activation of the cholinergic system of the heart as inhibition of both nicotinic (10(-7) M hexametonium) and muscarinic cholinoceptors (10(-7) M atropine) prevented the reaction. Hemicholinium (2 x 10(-5) M) and tetrodotoxin (5 x 10(-7) M) also reduced the response. Likewise, IFN-gamma potentiated the action of the muscarinic agonist carbachol. IFN-gamma simulated the biological effect of cholinergic agonists because: (a) it increased cGMP formation; (b) it decreased cAMP formation; and (c) it reduced heart contractility at doses that can be considered physiologic. IFN-gamma also modified the muscarinic receptor by interfering with the binding of the radiolabelled antagonist quinuclidinyl benzilate [( 3H]QNB). It is suggested that IFN-gamma binding to IFN-gamma receptors in the heart may lead to a cholinergic response by interaction of both receptor systems on the surface of atrial cells.

Interleukin 2 stimulates heart contractility in the presence of exogenous arachidonate or the calcium ionophore A 23187.

An increase in the isometric developed tension (IDT) of isolated rat atria was observed shortly after the addition of human interleukin 2 (IL-2) to the organ preparation with subthreshold concentrations of either arachidonate (AA, 1.98 X 10(-6)M) or the calcium ionophore A 23187 (1.9 X 10(-6)M). Both natural purified IL-2 (nIL-2) and yeast recombinant IL-2 (rIL-2) were active in this experimental system. It was determined that this lymphokine was active at 2 X 10(-11)M, considering as a reference the specific activity of rIL-2. Anti-IL-2 monoclonal antibody (anti-IL-2 MAb) abolished this reaction. Inhibition of atrial phospholipase C activity by nitrocarboxyphenyl N,N-diphenylcarbamate (NCDC, 5 X 10(-6)M) prevented the development of the inotropic positive effect of IL-2 in the presence of either AA or A 23187. The synthetic diacylglyceride 1-oleoyl, 2-acetyl-glycerol (OAG) replaced the IL-2 as stimulatory signal but NCDC had no effect on the reaction. The results suggest that IL-2 can alter the physiologic behaviour of the heart and that its mechanism of action is probably similar to the one proposed for other IL-2 targets (IL-2 receptor-positive T lymphocytes, T cell lines).

Leukoagglutinins in patients with hemophilia.

Anti-leukocyte antibodies may occur in hemophilic patients as a consequence of replacement therapy with blood derivates. In this report we describe the presence of leukoagglutinins (LA) in serum of HIV-infected hemophiliacs (HIV + He) and their absence in HIV-negative patients (HIV-He). LA activity was recovered in IgG fractions from HIV + He, in the polyethylene glycol (PEG) precipitates from these sera, and in some cases in the supernatant fractions of PEG precipitates. Although the amount of PEG precipitates corresponding to circulating immune complexes (CIC) was higher in HIV + He than in HIV-He and normals, there was no direct relationship between CIC levels and LA. LA reacted both with autologous and with allogeneic polymorphonuclear leukocytes (PMN). In contrast to PMN isolated from HIV-He, HIV + He PMN had membrane associated IgG. In HIV + He, LA activity was more frequently observed in patients with more advanced stages of HIV infection than in asymptomatic individuals. Our results suggest that LA activity could be one of the manifestations of autoreactivity associated with progressing HIV infection in patients with hemophilia.

Effect of lymphokines on natural killer cytotoxicity in patients with high risk of developing the acquired immune deficiency syndrome.

Regulation of natural killer (NK) activity of peripheral blood mononuclear cells (PBMC) from patients with the acquired immune deficiency syndrome (AIDS) and from individuals at high risk of developing AIDS (R-AIDS) was studied. The response of untreated PBMC to the interferon inducer polyinosinic polycytidilic acid (Poly I:C) was lower in AIDS and R-AIDS than in normal controls and PBMC from R-AIDS were more susceptible to stimulation with lymphokine rich supernatants (Con A-SN, PHA-SN, lectin free IL-2) than AIDS and normal controls. To determine the role of the different T lymphocyte subsets in the regulation of NK activity, PBMC were selectively treated with monoclonal non-cytotoxic anti-Leu 2a and anti-Leu 3a antibodies and then stimulated with lymphokine rich supernatants. These results indicate that the effect of crude supernatants was the combination of opposite effects. Leu 2a-blocked R-AIDS-PBMC enhanced NK cytotoxicity when exposed to IL-2 rich supernatants whereas Leu 3a-blocked R-AIDS-PBMC suppressed the cytotoxic reaction.

Inhibition of normal natural killer cytotoxicity by sera from hemophilic patients.

In this study we searched for circulating antibodies or other serum factors that could account for the natural killer (NK) defect observed in hemophiliacs (He) infected with the human immunodeficiency virus (HIV). We analyzed the effect of negative or positive sera for HIV from He on normal NK activity. We showed that sera from He interfered with normal NK cytotoxicity. The inhibitory activity was higher in HIV+ sera and increased as the HIV disease progressed. HIV- sera also inhibited NK function, although to a lesser extent than HIV+, and it was probably due to isoimmunization through replacement treatment with plasma-derived concentrates. For each individual, no direct correlation was found between NK inhibition (NK-INH) of sera and the NK activity of He peripheral blood mononuclear cells (PBMC). Furthermore, He serum was poorly inhibitory on autologous PBMC. Preincubation of allogenic effector or target cells with He sera revealed that the inhibitory effect was the result of the reaction with these cells. A positive correlation was found by comparing NK-INH of whole He sera with the serum levels of circulating immune complexes. When the NK-INH assay was performed using the same concentration of DEAE-purified IgG from N, HIV- or HIV+, we found that HIV+ AIDS IgG was more inhibitory than the others.

Anti-leukocyte antibodies as a consequence of HIV infection in HIV+ individuals.

Antibodies (Ab) that induce antibody-dependent cell-mediated cytotoxicity (ADCC) against non-T lymphocytes, anti-HLA class II specific Ab and anti-PMN were tested in hemophilic (He) patients who were alloimmunized because they had received replacement treatment with blood derivates and become infected with HIV, as well as in those who remained seronegative. In addition, the serum reactivity of spouses of HIV+ individuals and their children was studied to determine the effect of HIV infection in the absence of concomitant alloimmunization. The results of this study indicate that ADCC Ab were already present in HIV- He, suggesting the influence of alloimmunization. Their titer increased after appearance of HIV disease. While low reactivity against class II antigens was observed in HIV- He, activity augmented sharply after HIV infection and increased further with disease progression. Anti-PMN reactivity followed a similar pattern. Anti-class II, ADCC Ab and anti-PMN were also detected in the asymptomatic HIV+ spouses of HIV+ patients in titers that were similar to those of asymptomatic HIV+ He. In children born to HIV+ mothers in whom HIV infection was confirmed, anti-class II, ADCC Ab and anti-PMN reactivity were also observed, and activity increased after the onset of disease. These results suggest that induction of anti-leukocyte Ab occurs in the absence of massive allostimulation after HIV infection. HIV infection may enhance preexisting class II and anti-leukocyte response in allostimulated individuals.

HIV infection and natural killer cytotoxicity in hemophilic patients.

In this study we analyzed the ability of peripheral blood mononuclear cells (PBMC) from hemophilic patients (He) with negative or positive serology for the human immunodeficiency virus (HIV), to increase natural killer (NK) cytotoxicity upon stimulation with physiological and non physiological agents. Purified interleukin-2 (IL-2), the interferon (IFN)-inducer polyinosinic polycytidylic acid (PIC), recombinant alpha- and gamma-IFN and the protein kinase activator phorbol myristate acetate (PMA) were used as stimulatory agents. The NK functional response was correlated with the presence of PBMC bearing phenotypic markers of activated cells (IL-2 receptor, IL-2R) and of different NK cell maturation stages. Our results demonstrate that NK effector cells with slight lytic activity (Leu 7+ CD16-) predominated in HIV+ He patients. On the other hand the occurrence of IL-2R positive cells was similarly high in both HIV+ and HIV- individuals and was probably more related to chronic replacement treatment with Factor VIII or Factor IX concentrates than to HIV infection. The ability to respond to physiological NK regulators such as IL-2 and IFNs, or to the IFN-inducer PIC was impaired in HIV+ He, especially in HIV+ LAS individuals, suggesting that the inability of these cells to increase NK cell activity after appropriate induction was due to an intrinsic defect. Since phosphoinositide turnover and subsequent protein kinase C activation are thought to be part of the physiological mechanism of NK cytotoxicity, we studied the effect of PMA on PBMC from each group of patients. The ability to respond to PMA was lost only in PBMC from HIV+ LAS patients, indicating that impairment of the NK lytic mechanism progresses as the disease gets worse.(ABSTRACT TRUNCATED AT 250 WORDS)

alpha-Adrenoceptor stimulated lymphocytes trigger the mechanical response of vas deferens: participation of arachidonic acid metabolites.

Normal human lymphocytes (L) (8 X 10(5) ml-1) incubated with methoxamine (Me) (1 X 10(-7) M) (Me-L) triggered the mechanical response of the isolated vas deferens of the rat. L or Me alone did not modify this contractile activity at the concentrations cited above. Me alone (10(-6) to 10(-3) M) increased the tension of the vas. In the presence of L (8 X 10(5) ml-1) the dose-response curve to Me shifted to the left and the efficacy of Me was enhanced. Inhibitors of alpha 1-adrenoceptors completely blocked the reaction between Me and L while drugs that block alpha 1 and alpha 2-adrenoceptors reduced the reaction between Me-L and the vas deferens. Direct contact of Me-L with the assay organ was not necessary. Cell-free supernatants of L exposed to Me (Me-L supernatants) elicited the reaction in the same way as Me-L. This effect required the continuous presence of Me since dialyzed Me-L supernatants were inactive. Inhibitors of lipoxygenase(s) completely blocked the positive inotropic effect of Me-L or of Me-L supernatants. Inhibitors of cyclo-oxygenase potentiated this effect. These results suggest that Me reacts with alpha 1-adrenoceptors of L. From this reaction, soluble factors are released that potentiate the alpha-adrenoceptor stimulatory effect of Me on the vas deferens as a consequence of the release of oxidative products of the lipoxygenase/s pathway of arachidonic acid.

Lymphocytes sensitize rat isolated atria to the inotropic and chronotropic effects of sodium arachidonate.

Normal human lymphocytes (4 X 10(5) ml-1) incubated with sodium arachidonate (8 X 10(-7)M) (NaA-L) induced a strong enhancement of the tension and frequency of spontaneously beating rat atria. Normal human lymphocytes (L) or NaA alone at 8 X 10(-7)M did not modify this contractile activity. Between 2 X 10(-6)M to 1 X 10(-5)M NaA alone increased the tension of the atria without effect on the rate. In the presence of L (4 X 10(5) ml-1) the dose-response curve to NaA shifted to the left, the potency and the efficiency of NaA were enhanced and the chronotropic action was triggered. Inhibitors of cyclo-oxygenase (indomethacin 1 X 10(-6)M or acetylsalicylic acid (ASA) 1.8 X 10(-4)M) completely blocked the positive inotropic effect induced by NaA alone. Inhibitors of lipoxygenase/s (nordihydroguaiaretic acid (NDGA) 1 X 10(-5)M or 5,8,11,14-eicosatetraynoic acid (ETYA) 1 X 10(-7)M did not modify this effect. Indomethacin and ASA did not block the positive inotropic and chronotropic effects of the lower concentration of NaA-L and significantly reduced the inotropic effect of the higher ones. NDGA and ETYA shifted to the right the inotropic and chronotropic dose-response curve to NaA-L. FPL-55712 (1 X 10(-7)M), the slow reacting substance of anaphylaxis (SRS-A) antagonist, significantly reduced the overall inotropic and chronotropic effect of NaA-L. Direct contact of NaA-L with the atria was not necessary. Cell-free supernatants of L exposed to NaA increased the tension and the frequency of beating rat atria. 7 The stimulatory effect of NaA-L supernatants did not occur if rat atria had been previously incubated with NDGA 1 x 10-5 M. On the other hand, the generation of stimulatory products from NaA-L was not prevented by preincubating L with 1 x 10-5 M NDGA. Hence SRS-A and/or other oxidative metabolites of arachidonic acid were produced by the atria. 8 These results suggest that NaA-L react in vitro with spontaneous beating rat atria, inducing inotropic and chronotropic effects. Moreover, the stimulatory action of NaA itself was potentiated by L. These reactions involved a balance between cyclo-oxygenase and lipoxygenase oxidative products with a central role for SRS-A.

Autoantibodies in HIV-infected patients that modulate the cholinergic activity of heart and gut tissue.

In human immune deficiency virus (HIV) disease, direct infection of heart tissue with HIV and repeated intestinal infections with opportunistic pathogens are thought to be the main cause of cardiac disease and diarrhoea respectively. A role for autoimmune phenomena may also be involved in the pathogeny of HIV disease. In this study, we demonstrate that immunoglobulins from the A and G classes from HIV positive patients are able to interfere with the function of the muscarinic cholinergic receptors from heart and gut. Both IgA and IgG HIV+ preparations decreased the tension of isolated atria and increased the tension of isolated ileum. The mechanical effect of carbachol was inhibited in both atria and ileum preparations, when they were preincubated with either IgA or IgG HIV+ fractions. An inhibitor of muscarinic cholinergic receptors (atropine) impaired the negative inotropic action of HIV+ immunoglobulins (Ig) on the heart and prevented the positive inotropic effect of HIV+ Igs on ileum. HIV+ IgA fraction was approximately ten fold more potent to interfere with the cholinergic function as compared to the IgG fraction. These results suggest that antibodies present in HIV+ serum may also modulate muscle's cholinergic activity in the heart and ileum from HIV+patients.