Cannabinols and the rosette forming properties of lymphocytes in vitro.

Preincubation of lymphocytes, for 60 minutes at 37°C with tetrahydrocannabinol (THC), obtained from controls produced reduction in early but not total T cell rosettes compared to aliquots treated with vehicle alone. Similar reductions in early rosettes were seen after preincubation with cannabinol, cannabidiol, and olivetol. The marijuana smokers' lymphocytes were less affected by preincubation with THC than those from controls. Presumably their lymphocytes may already have been affected by prior exposure to cannabinols. These data show that THC and other cannabinols can affect the rosetting properties of T cells $\text{in vitro}$ and may provide a model for the study of THC effects on these immunocompetent cells.     

Spontaneous and pokeweed mitogen-induced in vitro IgM production in children and adolescents with insulin-dependent diabetes mellitus

Serum IgA, IgG and IgM levels, spontaneous and pokeweed mitogen (PWM)-induced in vitro IgM production (determined by ELISA) and blastogenic responses of peripheral mononuclear cells to PWM were evaluated in 4 insulin-dependent (IDDM) children, at the onset and after 4, 8, 12 months of disease, and in 32 children and adolescents with IDDM of 1-14 years duration (mean 4.8 +/- 3.8 years). Fifteen age-matched healthy subjects served as controls. Serum immunoglobulin levels were normal in 31 (86%) patients. Spontaneous in vitro IgM production showed no significant difference between IDDM patients and controls. The PWM-stimulated lymphocytes from IDDM patients at onset or after 4 months of disease produced significantly lower concentrations of IgM compared to long-standing IDDM patients or to controls. No different blastogenic response to PWM was observed in IDDM patients compared to controls. No correlation was present between the immunological parameters evaluated and metabolic control. Our data suggest that a defect of antibody producing B lymphocytes or an alteration of T cell can occur during the early stages of diabetes.     

Expression of cell surface markers on T and B lymphocytes after long-term chemotherapy of acute leukemia

The aim of this study was to determine the distribution of peripheral blood T and B lymphocytes in children with ALL in remission, before and after cessation of 3 yr of immunosuppressive therapy. Immunofluorescence of viable cells and rosette formation were the two methods used to identify B and T cells, respectively. Though combination chemotherapy depresses the total lymphocyte population, B lymphocytes were more depressed than T lymphocytes. On the last day of therapy, the population of lymphocytes bearing IgG-M (B cells) was markedly reduced, but the percentage of RFL (T cells) was within normal values. After chemotherapy was stopped, and in the absence of extrinsic antigenic stimulation, there was an immunological rebound in the B cell compartment. During the first 3 months off therapy, there was an increase in intensity of fluorescence and in the proportion of IgG-M-bearing lymphocytes above the levels of normal controls. Assays of lymphocytes bearing IgG, IgM and IgA indicate that there was a rebound of IgG and IgM but not IgA lymphocytes. Changes in the proportion of RFL as a function of time off therapy were inversely proportional to those observed for IgG-M-bearing lymphocytes, that is, the percentage of RFL decreased during the first 3 months off therapy. When the absolute numbers of B and T cells were compared, it was found that B lymphocytes reached a plateau phase after 2–3 months off therapy, but T lymphocytes continued to rise beyond this period, reaching normal levels at 12+ months off therapy.These results provide evidence, at the single cell level, of the immunological rebound that occurs after cessation of therapy and suggest that the kinetics of recovery are different for T and B lymphocytes.     

Immunological features in multiple sclerosis.

A clinical and laboratory profile of the immunological system of patients with multiple sclerosis (MS) strongly suggested that many specific immune deficiencies exist in MS. The immunological history showed that patients with MS had had more tonsillectomies, appendicectomies, and childhood infections than matched controls, which suggested that there had been problems in controlling various types of childhood infections. The cell-mediated immune response and the circulating antibody titres were specifically impaired against a variety of antigens. Patients with MS had significantly lower serum antibody titres than controls against many naturally occurring antigens-namely, diptheria and tetanus toxoids, adenovirus, and mumps viruses. Raised serum antibody titres were found against measles and varicella zoster viruses while no difference was found towards other antigens. The delayed hypersensitivity reaction and the immunological memory of patients with MS were also greatly reduced against the mumps skin test antigens. There were normal amounts of circulating T and B lymphocytes, and the phytohaemagglutinin, concanavallin A, pokeweed mitogen, and encephalitogens lymphocyte transformation was not different from that in controls. These results indicated that patients with MS have more infectious problems than normal people and that both their T and B cell systems cannot mount a fully normal immunological response to some viral and bacterial antigens, while they give an increased response to others.     

Marihuana, tetrahydrocannabinol and T-cell function.

Conflicting reports exhist on the effect of marihuana smoking on thymus derived lymphocytes (T-cells). Marihuana smoking has been reported to both reduce the formation of T-rosettes and affect mitogenic stimulation of T-cells in man. This present study was conducted to evaluate the effects of marihuana smoking on T-cells during a 24-hour period after smoking. Chronic marihuana smokers, who had abstained from smoking for one month, smoked “street” marihuana, marihuana cigarettes with a known quantity of delta-9-tetrahydrocannabinol (THC), or placebo marihuana cigarettes. In two of three subjects who used “street” marihuana, T-cell rosette formation was reduced 24 hours after smoking. Response to phytohemagglutinin (PHA) stimulation was reduced in one of the above subjects. In a second group of subjects, rosette formation was decreased in five of six subjects 3 to 6 hours after smoking marihuana cigarettes containing 10 mg of THC. However, values in all but one individual returned to control levels within 24 hours. A 50% reduction in PHA stimulation was also observed in this subject 6 hours after smoking. PHA stimulation was not affected following placebo. In the sixth subject, a stimulatory effect on rosette formation was observed following both the use of the THC-containing and placebo marihuana cigarettes. The results of these studies indicate that while marihuana smoking affects certain $\text{in}$$\text{vitro}$ tests of T-cell function, the effects are variable, transitory, and may be associated with factors other than THC.     

Peripheral T-cell subsets in asymptomatic hepatitis B-virus carriers

To ascertain whether the abnormalities of circulating T-cell subsets in patients with hepatitis B virus (HBV)-related chronic liver diseases represent the primary immunological process or are secondary to liver disease process, peripheral T-cell subsets were analyzed by indirect immunofluorescence using monoclonal antibodies against total T cells (OKT3), T helper/inducer cells (OKT4), and T suppressor/cytotoxic cells (OKT8), in 30 asymptomatic HBV carriers without biochemical or histological evidence of liver disease, and the results were compared to 15 HBV-induced chronic active liver diseases. The results revealed that OKT4/OKT8 ratios were significantly reduced in 15 hepatitis B e antigen (HBeAg)-positive asymptomatic carriers as compared with controls, with decreased OKT4-positive cells and increased OKT8-positive cells, while T-cell subsets and ratios were normal in 15 hepatitis B e antibody (anti-HBe)-positive asymptomatic carriers. The changes of circulating T-cell subsets in 15 HBe-Ag-positive asymptomatic carriers showed no significant difference from those of 15 HBeAg-positive patients with chronic active liver diseases. These findings suggest that the deranged T-cell subsets in chronic HBV infection are not secondary to liver cell damage, but might represent the underlying immunological abnormalities which are closely related to HBeAg/anti-HBe status, and that the pathogenetic mechanism of liver cell damage in chronic HBV infection may not be simply related to circulating T-cell subsets.     

Cigarette smoking may reduce plasma leptin concentration via catecholamines

BACKGROUND: Leptin might influence body weight among smokers. DESIGN: (A) Screening of plasma leptin levels in 222 sedentary, smoking and non-smoking middle-aged men. (B) Double-blind, placebo-controlled smoking intervention on smokers (n=31). (C) Non-smokers (n=40) received chewing gum with nicotine (2mg nicotine, n=23) or without nicotine (n=19). (D) The effects of nicotine (0.05 and 0.5 microg/mL) were monitored on leptin secretion and mRNA levels in a human placental cell line (BeWo) expressing leptin, a murine adipocyte cell line (3T3-L1) and human adipose tissue explants. RESULTS: (A) Plasma leptin levels in smoking men (8.4+/-8.4 ng/mL, n=100) was lower as compared to non-smokers (10.3+/-7.3 ng/mL, n=122) (P<0.001), even when adjusted for differences in body mass index (BMI) (P<0.001). (B) A significant reduction (P=0.02) in plasma concentration of leptin was found already after smoking one cigarette. Concomitant with the 3-5 fold increase in plasma nicotine concentration after the first cigarette, we observed increased plasma adrenaline levels (P=0.005). (C) There was no effect of nicotine on plasma leptin levels in non-smokers receiving nicotine-containing chewing gum, and plasma concentrations of catecholamines were unaltered. (D) There was no effect of nicotine on leptin mRNA expression after incubation with cells or adipose tissue. CONCLUSION: Cigarette smoking reduced plasma leptin concentration in vivo, whereas nicotine had no direct effect on leptin expression in vitro. Nicotine might indirectly reduce leptin secretion via enhanced plasma catecholamine concentration.     

Detection of HLA-DRw (Ia-like) antigens on human T lymphocytes grown in tissue culture.

Human T lymphocytes that proliferate in the presence of conditioned medium from PHA-stimulated allogeneic peripheral blood cells were shown to express IPA antigens after the 8th culture day. Ia antigens as detected by xenogeneic antisera were present on 80 to 90% of the cultured cells which were also strongly reactive with xenogeneic antisera defining a human T cell antigen and formed E rosettes. Control cultures with PHA or no conditioned medium expressed T cell but not Ia antigens. These cultured T cells also express the same HLA-DRw determinants as the B cells of the donor they were derived from. Absorption of xenogeneic Ia, and HLA-DRw alloantisera with cultured T cells completely removed the reactivity of these sera for enriched peripheral blood B lymphocytes from normal donors.     

Products of 5-lipoxygenase and myeloperoxidase activities are increased in young male cigarette smokers

Abstract The significance of 5-lipoxygenase and myeloperoxidase activities has not been extensively studied among young male smokers. Leukotriene B(4), 20-hydroxy-leukotriene B(4), 20-carboxy-leukotriene B(4) and 3-chlorotyrosine were measured in plasma and urinary samples of young male smokers at 8 hours following cigarette abstinence and an hour after cigarette smoking. Leukotriene B(4) and 3-chlorotyrosine were determined in neutrophils isolated from these individuals. The levels of these markers were compared with those of age-matched controls. In vitro studies were performed to evaluate the production of leukotriene B(4) and 3-chlorotyrosine from human neutrophils following exposure to nicotine and cotinine. Thirty male smokers (mean age, 27.4 years) and 28 male non-smokers (mean age, 28.7 years) were studied. Plasma levels of leukotriene B(4), 20-carboxy-leukotriene B(4) and 3-chlorotyrosine were higher in smokers than in non-smokers; leukotriene B(4) and 20-carboxy-leukotriene B(4) levels increased further an hour after cigarette smoking. Peripheral neutrophils isolated from smokers showed greater expressions of myeloperoxidase and 5-lipoxygenase activities compared with non-smokers, while plasma leukotriene B(4) and 3-chlorotyrosine were correlated significantly with high-sensitivity C-reactive protein and plasma nicotine concentrations. Exposure of human neutrophils to nicotine and cotinine resulted in a higher production of leukotriene B(4) and 3-chlorotyrosine. To conclude, leukotriene B(4) and 3-chlorotyrosine levels are increased in young male cigarette smokers. These results suggest that cigarette smoking aggravates neutrophil-mediated inflammation by modulating the activities of myeloperoxidase and 5-lipoxygenase pathways.     

The CD3epsilon proline-rich sequence, and its interaction with Nck, is not required for T cell development and function

The CD3epsilon proline-rich sequence (PRS) binds to the cytosolic adaptor molecule Nck after TCR ligation. It has been proposed that this interaction is essential for immunological synapse formation and T cell activation. To assess the physiological importance of the CD3epsilon PRS, we have generated mice that lack this motif (CD3epsilon.PRS(M)). Pull-down experiments demonstrated the inability of Nck to bind to the CD3epsilon PRS in thymocytes from mutant mice after TCR ligation. Surprisingly, no differences were observed in the number and percentage of T cell subsets in the thymus and spleen, and there was no apparent defect in positive or negative selection. Furthermore, the proliferative response of CD3epsilon.PRS(M) T cells to staphylococcal enterotoxin B and anti-CD3 Ab was normal. TCR surface expression, constitutive internalization, and Ag-induced down-modulation were also normal. These data suggest that the interaction between the CD3epsilon PRS and Nck, or any other Src homology 3 domain-containing molecule, is not essential for T cell development and function.     

The effects of 9-ene-tetrahydrocannabinol on hormone release and immune function

We investigated effects of 9-ene-tetrahydrocannabinol (THC) on endocrine and immunological function. Seventeen male volunteers entered into a double blind, randomized study to receive oral THC (10 mg t.i.d. for 3 days and on the morning of the fourth day) or placebo, after at least 2 weeks of abstinence. Plasma prolactin, ACTH, cortisol, luteinizing hormone and testosterone were not altered during or after THC, compared with baseline concentrations. Tests of lymphocyte function showed no differences compared to baseline between THC and placebo groups. As the relatively low dosing regimen of THC (10 mg t.i.d.) resulted in no alterations, another group of 6 men were administered higher doses of THC by inhalation (18 mg/marijuana cigarette) following the same dosing regimen. No endocrine or immunological alterations were observed. When the subjects were grouped according to their history of THC use prior to admission, heavy THC users had lower prolactin concentrations than light users. No differences were observed in concentrations of other hormones or in tests of immune function.     

Response of rhesus monkey lymphocytes to short-term administration of THC

Four Rhesus monkeys were subjected to daily administration of 2.5 mg of tetrahydrocannabinol (THC)/kg body wt, after establishing the norms for complete blood count, T- and B-cell concentrations, and the dose response of thymidine incorporation after PHA stimulation. THC was administered daily for 3 weeks, the treatment was stopped, and then the animals were allowed to recover for 4 weeks. Cellular responses, incorporation studies and fibrinogen levels were determined during the treatment and recovery phases. Compared to 4 vehicle-treated animals, the THC-treated animals experienced significant augmentation of both their total white cell and their neutrophil counts during the recovery phase which returned to normal levels during the recovery phase. There was no alteration in total lymphocyte count or T- or B-cell concentrations. Fibrinogen levels of the THC-treated animals during the treatment phase were also elevated compared to controls, and the levels diminished to the same values as the vehicle-treated animals during recovery phase. Possible mechanisms for the response of Rhesus monkeys to short-term administration of THC are discussed.     

The effect of low exposure-rate gamma irradiation on T and B lymphocyte function in the mouse

The effect of chronic irradiation on T and B cell numbers and function was studied in mice. Cobalt 60 gamma radiation at 6 R/hour reduced the numbers of anti-SRBC PFC in the spleen, with minimal levels recorded after total exposures of 1000-2000 R. Recovery was incomplete after 1000 R, reaching only 40-50 per cent of normal in four months and remaining at that level for the animal's lifetime. The long-term deficiency in PFC formation was not due to a quantitative lack of T or B cells since normal cell numbers were observed in the spleen 60-144 days after 1000 R. Adoptive transfer studies with combinations of bone marrow and thymus cells, or of splenic T and B cells, from normal and irradiated mice, revealed functional defects in both cell compartments during the first two months. Normal and near normal function of T and B cells occurred 100 days postirradiation, a time when the splenic in vivo response was still only 50 per cent of the controls. The latter observation suggests that the microenvironment of the chronically irradiated spleen alters factors regulating T and B cell interactions in response to a T-dependent antigen.     

Antigen recognition by T lymphocytes. IV. Differences in antigen-binding characteristics of T- and B-RFC: a cause for variations in the evaluation of T-RFC

Some of the characteristics of antigen binding by macrophages, B and T cells, have been examined using the rosette technique. Rosette-forming macrophages were labeled with carbon and T-RFC were defined by their sensitivity to inhibition by anti-θ serum and azathioprine. T rosettes have been further characterized by the RBC number in the rosette since T rosettes appeared to bear statistically less RBC than B rosettes. Increasing the RBC/WBC ratio principally augments the number of B-RFC. T rosettes are formed more rapidly than B rosettes but are more sensitive to disruption by turbulent resuspension. These data are valid both for unimmunized and immunized spleen cells, although they are less clear-cut with immunized mice. These differences between antigen binding characteristics of B and T-RFC may explain variations in T-RFC evaluation. They suggest that T-RFC bear fewer receptors than B-RFC and that the avidity of their receptors is probably more homogeneous than that of B-RFC.     

Circadian and Ultradian Rhythms in Heart Rate and Motor Activity of Smokers,Abstinent Smokers,and Nonsmokers

In a field study, heart rate and motor activity were assessed continuously in 12 male smokers during 2 smoking and 2 abstinence days and in 12 male nonsmokers during 4 days. A circadian analysis revealed earlier activity acrophases in smokers than nonsmokers and earlier heart rate acrophases in abstinent than smoking smokers. Furthermore, heart rate acrophases of smoking smokers significantly anticipated activity acrophases;, whereas in abstinent smokers and nonsmokers the two parameters oscillated in phase. With the use, in smoking smokers, of the individual average smoking interval as a hypothetical ultradian period length, significant periodicities were found for heart rates in 16 and for activity in 15 of 24 observation days. These rhythms were nicotine independent and based on heart rate and activity increases prior to lighting up the cigarettes. Individual frequency spectra for the 16 h after getting up and the 7 h after going to bed did not reveal single dominant frequencies but rather complex frequency distributions. Power spectra of the daytime data revealed no group differences for activity and no heart rate differences between smoking smokers and nonsmokers. In abstinent smokers, however, a significant reduction of heart rate frequencies slower than 1 cycle/135 min and a significant increase of heart rate frequencies faster than 1 cycle/20 min were observed as compared with all other groups. This effect persisted over the 2 abstinence days, suggesting an activity-independent change in the frequency distribution of heart rates after quitting smoking.     

Circadian and Ultradian Rhythms in Heart Rate and Motor Activity of Smokers, Abstinent Smokers, and Nonsmokers

In a field study, heart rate and motor activity were assessed continuously in 12 male smokers during 2 smoking and 2 abstinence days and in 12 male nonsmokers during 4 days. A circadian analysis revealed earlier activity acrophases in smokers than nonsmokers and earlier heart rate acrophases in abstinent than smoking smokers. Furthermore, heart rate acrophases of smoking smokers significantly anticipated activity acrophases;, whereas in abstinent smokers and nonsmokers the two parameters oscillated in phase. With the use, in smoking smokers, of the individual average smoking interval as a hypothetical ultradian period length, significant periodicities were found for heart rates in 16 and for activity in 15 of 24 observation days. These rhythms were nicotine independent and based on heart rate and activity increases prior to lighting up the cigarettes. Individual frequency spectra for the 16 h after getting up and the 7 h after going to bed did not reveal single dominant frequencies but rather complex frequency distributions. Power spectra of the daytime data revealed no group differences for activity and no heart rate differences between smoking smokers and nonsmokers. In abstinent smokers, however, a significant reduction of heart rate frequencies slower than 1 cycle/135 min and a significant increase of heart rate frequencies faster than 1 cycle/20 min were observed as compared with all other groups. This effect persisted over the 2 abstinence days, suggesting an activity-independent change in the frequency distribution of heart rates after quitting smoking.     

A role for autophagic protein beclin 1 early in lymphocyte development

Autophagy is a highly regulated and evolutionarily conserved process of cellular self-digestion. Recent evidence suggests that this process plays an important role in regulating T cell homeostasis. In this study, we used Rag1(-/-) (recombination activating gene 1(-/-)) blastocyst complementation and in vitro embryonic stem cell differentiation to address the role of Beclin 1, one of the key autophagic proteins, in lymphocyte development. Beclin 1-deficient Rag1(-/-) chimeras displayed a dramatic reduction in thymic cellularity compared with control mice. Using embryonic stem cell differentiation in vitro, we found that the inability to maintain normal thymic cellularity is likely caused by impaired maintenance of thymocyte progenitors. Interestingly, despite drastically reduced thymocyte numbers, the peripheral T cell compartment of Beclin 1-deficient Rag1(-/-) chimeras is largely normal. Peripheral T cells displayed normal in vitro proliferation despite significantly reduced numbers of autophagosomes. In addition, these chimeras had greatly reduced numbers of early B cells in the bone marrow compared with controls. However, the peripheral B cell compartment was not dramatically impacted by Beclin 1 deficiency. Collectively, our results suggest that Beclin 1 is required for maintenance of undifferentiated/early lymphocyte progenitor populations. In contrast, Beclin 1 is largely dispensable for the initial generation and function of the peripheral T and B cell compartments. This indicates that normal lymphocyte development involves Beclin 1-dependent, early-stage and distinct, Beclin 1-independent, late-stage processes.     

Lymphocyte subpopulations in peripheral blood and bone marrow in patients with idiopathic myelofibrosis

In 8 patients with idiopathic myelofibrosis (IM) T and B cells were studied in view of the possibility that immunological dysfunctions are involved in initiating or contributing to the bone marrow fibrosis. In peripheral blood the absolute numbers of E-SRBC and OKT3+ lymphocytes were significantly reduced; in addition a significant decline was observed in the proportion and absolute numbers of OKT8+ cells, resulting in a reversed Leu-3a/OKT8 ratio. An impaired B cell function was observed in 4 of the 8 patients, characterized by a disturbed in vitro pokeweed mitogen stimulated immunoglobulin synthesis and low serum immunoglobulin levels. Immuno-histological studies of the bone marrow demonstrated a scarcity of T cells but normal numbers of B cells. However, no correlation was noted between the observed deviations of B and T cells and the degree of bone marrow fibrosis determined by means of bone marrow histology and serum procollagen-III levels. These data are not sufficient to support the hypothesis that immunological changes in IM are primarily involved in the process of bone marrow fibrosis.     

Cellular immune responses to the hepatitis B virus polymerase

CD4 T cells play an important role in hepatitis B virus (HBV) infection by secretion of Th1 cytokines that down-regulate HBV replication, and by promoting CD8 T cell and B cell responses. We have identified and characterized 10 CD4 T cell epitopes within polymerase and used them to analyze the immunological effects of long-term antiviral therapy as compared with spontaneous recovery from HBV infection. Candidate epitopes were tested for binding to 14 HLA-DR molecules and in IFN-gamma ELISPOT and cytotoxicity assays using peripheral blood lymphocytes from 66 HBV-infected patients and 16 uninfected controls. All 10 epitopes bound with high affinity to the most prevalent HLA-DR Ags, were conserved among HBV genomes, and induced IFN-gamma responses from HBV-specific CD4+ T cells. Several epitopes contained nested MHC class I motifs and stimulated HBV-specific IFN-gamma production and cytotoxicity of CD8+ T cells. HBV polymerase-specific responses were more frequent during acute, self-limited hepatitis and after recovery (12 of 18; 67%) than during chronic hepatitis (16 of 48 (33%); p=0.02). Antiviral therapy of chronic patients restored HBV polymerase and core-specific T cell responses during the first year of treatment, but thereafter, responses decreased and, after 3 years, were no more frequent than in untreated patients. Decreased T cell responsiveness during prolonged therapy was associated with increased prevalence of lamivudine-resistant HBV mutants and increased HBV titers. The data provide a rationale for the combination of antiviral and immunostimulatory therapy. These newly described HBV polymerase epitopes could be a valuable component of a therapeutic vaccine for a large and ethnically diverse patient population.     

Deficits in inhibitory control in smokers during a Go/NoGo task: an investigation using event-related brain potentials

Introduction

The role of inhibitory control in addictive behaviors is highlighted in several models of addictive behaviors. Although reduced inhibitory control has been observed in addictive behaviors, it is inconclusive whether this is evident in smokers. Furthermore, it has been proposed that drug abuse individuals with poor response inhibition may experience greater difficulties not consuming substances in the presence of drug cues. The major aim of the current study was to provide electrophysiological evidence for reduced inhibitory control in smokers and to investigate whether this is more pronounced during smoking cue exposure.

Methods

Participants (19 smokers and 20 non-smoking controls) performed a smoking Go/NoGo task. Behavioral accuracy and amplitudes of the N2 and P3 event-related potential (ERP), both reflecting aspects of response inhibition, were the main variables of interest.

Results

Reduced NoGo N2 amplitudes in smokers relative to controls were accompanied by decreased task performance, whereas no differences between groups were found in P3 amplitudes. This was found to represent a general lack of inhibition in smokers, and not dependent on the presence of smoking cues.

Conclusions

The current results suggest that smokers have difficulties with response inhibition, which is an important finding that eventually can be implemented in smoking cessation programs. More research is needed to clarify the exact role of cue exposure on response inhibition.