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It is well known that Δ9 - tetrahydrocannabinol (THC), the main active component of marijuana, exerts mainly suppressive infl uence on the immune system. Here we present data indicating that THC induce immune enhancing effects. In this study we examined the effects of small doses of THC on the natural killer cell cytotoxicity (NKCC) (51Cr – release assay) and also on the number of NK cells (identifi ed as large granular lymphocytes, LGL) (Timonen method) in the peripheral blood in rats. Animals received acute or chronic (10 days) i.p. injections of THC at the doses of 0.2 and 0.5 mg/kg. It was found that acute injection of THC signifi cantly increased NKCC after the dose of 0.2 mg/kg (+ 131 Δ%; P<0.001 vs. control and + 52 Δ%; P<0.05 vs. 0.5 mg/kg dose). After chronic exposure to THC, a signifi cant NKCC increase was observed after the dose of 0.5 mg/kg (+ 209 Δ%; P<0.001 vs. control and + 97 Δ%; P<0.01 vs. 0.2 mg/kg dose). The number of LGL remained unchanged after both acute and chronic treatment with THC. It has been reported repeatedly that the doses of over a dozen milligrams of THC suppress NKCC. These results negate the common opinion about unidirectional suppressive effects of cannabinoids on the immune system. It seems that the pharmacodynamics rule of hormesis may also concern the infl uence of THC on the immune system.
We assesed the effectiveness of coping with stress during a 30 hours driving course by people with different levels of stress. Volunteers aged 18–30 took participated: 8 women and 7 men in the control group, and 9 women and 9 men preparing for the exam. Before the start of the course all participants took the Coping Inventory for Stressful Situations (CISS). Saliva samples to measure cortisol were collected from each of the participants: before the start of the course, before the 1st, 13th and 28th driving hour. Blood pressure was measured: before the course, during the 15th hour and the 30th hour of the course. Participants in the control group had one saliva sample taken and their blood pressure was measured once. The results suggest that the hour of the course is related to the level of the cortisol – the highest level is achieved before the first hour of the course and then it decreases in time (ANOVA: F3,64=14.9, P<0.001). Systolic pressure is related to the hour of the course (F2,48=11.3, P<0.001) and it reaches its peak before the state exam. There is a similar relation in case of diastolic pressure (F2,51=6.4, P=0.003). CISS test has shown that there are differences in copying with stress (between sexes as well as age-related).
Cocaine use has been associated with the increased risk for HIV and HCV infection in humans. The addicts have faster progression of HIV infection and increased incidence of AIDS suggesting that cocaine use results in a specifi c functional impairment of the immune system. Present study aimed to evaluate lymphocyte subsets number and distribution in peripheral blood after acute series of intravenous cocaine infusions. Adult male Wistar rats received 3 injections of cocaine hydrochloride (5 mg/kg/ml) in 30 min intervals. Blood samples were collected 30 min after the last infusion. Total leukocyte number, percentage numbers of leukocyte subpopulations and T, B, NK, T helper and T cytotoxic lymphocyte subsets were assessed. Intravenous cocaine caused signifi - cant decrease in total leukocyte and lymphocyte number and signifi cant fall in total numbers of T, B, NK and both T helper and T cytotoxic lymphocyte subsets. Furthermore, intravenous cocaine altered proportions between lymphocyte subsets by increasing T, mainly T helper and decreasing B, T cytotoxic and NK lymphocyte percentage portion. Immunosuppressive effect of intravenous cocaine infusions may thus be connected with alterations in lymphocyte subsets proportions. Such destabilization of lymphocyte subsets balance, especially T helper lymphocyte, could constitute basis of increased susceptibility for HIV and HCV infections.
Cannabidiol (CBD) is a major non-psychotropic compound of cannabis that has been reported to exert neuroprotective, antipsychotic and anxiolytic effects. CBD has promising anti-inflammatory properities, but despite therapeutic potential its mechanisms of action remain poorly understood. Our previous study revealed decrease in number of lymphocytes B and T in peripheral blood following repeated CBD administration. The present study aimed to assess effects of repeated administration of CBD on distribution of lymphocyte subsets in the spleen and the involvement of CB2 receptors. Adult male Wistar rats (n=35, 10 weeks of age at the start of study) received intraperitoneal injections of CBD at a dose of 5 mg/kg/day, or the vehicle, for 14 consecutive days. Total and relative numbers of lymphocyte T (T CD4+, and T CD8+), B, NK subsets were determined by flow cytometry. The selective CB2 receptor antagonist AM630 (1 mg/kg) was administered 15 min before CBD (or the vehicle) in order to block CB2 receptors. Repeated administration of CBD decreased total leukocyte number resulting from decreased numbers of lymphocytes B and T (both T CD8+ and T CD4+) in the spleen. Pretreatment with CB2 receptor antagonist partially inhibited CBD-induced decrease in lymphocyte number that was most pronounced in case of T CD8+ lymphocytes. AM630 itself produced slight decline in lymphocyte number that did not reach statistical significance. Observed effects were accompanied by a decrease in body weight gain, which was prevented by pretreatment with CB2 antagonist. The results indicate that CBD reduces lymphocyte number in the spleen, as it does in peripheral blood and that CBD has ability to affect the lymphocyte number via CB2 receptor.
BACKGROUND AND AIMS: Amphetamine, besides its well known psychological and behavioral effects, was found to influence the immune functions. However, the mechanism of amphetamine-induced changes in the immune system remains unknown. In search of a possible mechanism of immunomodulating effect of amphetamine, in the present study we tested the involvement of sympathetic nervous system in that effect. RESULTS: After pretreatment with 6-hydroxydopamine (3×75 mg/kg, ip), we evaluated the effect of acute amphetamine (1 mg/ kg, ip) administration on natural killer cell cytotoxicity (NKCC; Cr-51 release assay) and the number of NK (LGL) cells in the peripheral blood and spleen in male Wistar rats. Amphetamineinduced stimulation of blood and splenic NKCC was completely blocked by chemical sympathectomy. Blood NKCC in amphetamine-injected rats was 260% higher in comparison to a control group. Rats pretreated with 6-hydroxydopamine before amphetamine administration showed over 70% lower NKCC then rats which received amphetamine without chemical sympathectomy. Similarly to the peripheral blood, over 190% increase in NKCC in rats injected with amphetamine was observed in the spleen. Splenic NKCC in rats pretreated with 6-hydroxydopamine was about 60% lower after amphetamine in comparison with rats without chemical sympathectomy. The similar effects were observed in the case of LGL number. CONCLUSIONS: The data clearly show that AMPH-induced stimulation of NK cells numbers and function both in the peripheral blood and spleen are mediated by peripheral sympathetic nervous system.
It is well known that the main site of the action of amphetamine (AMPH) are the catecholaminergic neurons located both in central nervous system and in the sympathetic nervous system (SNS). To analyse the potential role of the SNS in the mechanism of AMPHinduced changes in natural killer cells cytotoxicity (NKCC) rats were sympathectomized by 6-hydroxydopamine (6-OHDA, 3 × 50 mg/kg, i.p.) prior to AMPH (1 mg/kg, i.p.) administration. In a separate experiment, rats were pretreated with a α-adrenergic receptor antagonist phentolamine (5 mg/kg, i.p.), β-adrenergic receptor antagonist propranolol (5 mg/kg, i.p.) or both. NKCC (51Cr-release assay) and the number of LGL (NK cells) were evaluated in the peripheral blood and spleen. In the peripheral blood AMPH-induced stimulation of NKCC was completely blocked by 6-OHDA. The increase in LGL number in the peripheral blood evoked by AMPH was partially inhibited by sympathectomy. In the spleen both effects of AMPH i.e. reduction of NKCC and decrease in LGL number were completely reversed by 6-OHDA. β-antagonist attenuated the AMPH-induced changes in NKCC and LGL number in the peripheral blood and spleen. In contrast, changes in NKCC and LGL number were not affected by α-blockade. These data clearly show that both AMPH-induced stimulation of the peripheral blood NKCC and suppression of spleen NKCC are mediated by the SNS. Furthermore, catecholamines elevated by AMPH modulate the NKCC via β-adrenergic, but not α-adrenergic mechanism.
High frequency stimulation (HFS) of the subthalamic nucleus (STN) is a surgical therapy for improving of motor symptoms in Parkinson’s disease. In this study, we assessed the HFS effects on cytometric analysis of the peripheral blood lymphocytes (T, B, NK, T helper, T cytotoxic) in freely moving hemiparkinsonian rats. Before unilateral lesion of the right substantia nigra pars compacta (6-hydroxydopamine), all rats were divided into two behavioral groups: high-HR and low-LR responders in novelty test. As compared to the sham controls, HFS of STN signifi cantly increases NK cell percentage number (25.86 ± 9.68% vs. 18.79 ± 5.63%, P<0.05). In contrast, signifi cantly (P<0.05) lower B lymphocytes level in stimulated (18.86 ± 3.91%) then sham group (22.80 ± 4.63%) was observed. These general changes in NK and B lymphocyte numbers were refl ected in HR stimulated animals (31.08 ± 7.19% vs. 20.44 ± 5.65% for NK cells number and 17.67 ± 2.57% vs. 23.23 ± 3.37% for B lymphocyte, stimulated vs controls, respectively, P<0.01). On the other hand, signifi - cantly lower T lymphocyte level in LR stimulated animals in comparison to LR controls was found (38.32 ± 4.18% vs. 45.91 ± 5.68%). Moreover, signifi cant differences between stimulated HRS and LRS in the NK and T cytotoxic percentages were found (31.08 ± 7.19% vs. 17.63 ± 6.74%, P<0.05; 8.46 ± 1.83% vs. 12.29 ± 1.76%, P<0.01). The results emphasize the importance of individual differences in reactivity to novelty on immune response to HFS of STN.
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Although addiction to amphetamine (AMPH) is a serious social and medical problem, the data concerning AMPH – immune interactions are still not numerous. To analyze the mechanism of AMPH-induced changes in the function of the immune system, rats were pretreated with ß-adrenergic receptor antagonist propranolol (PROP; 5 mg/kg, i.p.) prior to AMPH (1 mg/kg, i.p.) administration. Natural Killer cells cytotoxicity (NKCC) (51Cr-release assay), the number of LGLs (NK cells) (Timonen method), leukocytes, lymphocytes and monocytes, and plasma corticosterone level (CORT) (RIA) were evaluated in the peripheral blood and spleen. In the peripheral blood increases in NKCC (+331%), as well as in LGL (+33%) and monocyte (+65%) number observed after AMPH were partially inhibited by PROP (respectively by 30%, 19%, and 30%) in contrast to lymphopenia (-19%) and granulocytosis (+65%) which were not affected by ß-blockade. In the spleen AMPH-induced decreases in NKCC (-25%) and in all the leukocyte populations number (approximately -30%) were completely blocked by PROP. Plasma CORT level, highly elevated by AMPH (+337%), was attenuated nearly by 50% under ß-adrenergic blockade. These data indicate that AMPH-induced enhancement of cytotoxic activity of NK cell is related to ß-adrenergic mechanism.
To evaluate a possible mechanism of stress-induced lymphopenic effect we assessed the activity of lymphocyte lysosomal enzymes (LE) under immobilization. The effects of immobilization stress on LE (AP, acid phosphatase, cathepsin D and L, ß-N-acetyl-glucosamidase) activity in lymphocytes, number of lymphocytes and plasma cortisol (COR) level in the peripheral blood were examined in the cross-bred Pietrain pigs showing genotypic (presence or lack of RyR1 gene mutation) and phenotypic (reactivity to halothane) differences. It was found that immobilization stress evoked an increase in LE which was concomitant with lymphopenia and a rise of COR level. The most pronounced enhancement of LE, which may reflect a tendency to lymphocyte cytolysis, was found in the recessive homozygotes RyR1 (nn) phenotypically defined as stress/halothane susceptible as well as in the heterozygotes RyR1 (Nn) included in the group of stress/halothane resistant. Despite this individual variability the stress-induced increase in LE activity was present in all the animals. It seems that a possibility of destruction (lysis) of lymphocyte cells should not be excluded as one of the causes of stress lymphopenia.
The expression of dopamine, dopamine receptors and dopamine active transporters in peripheral lymphoid tissues and lymphocytes suggest that increased dopaminergic activity induced by cocaine may be involved in regulation of the distribution of lymphocyte subsets in the spleen. Adult male Wistar rats implanted with jugular vein catheter received cocaine hydrochloride (3 × 5 mg/kg/ml, i.v., at 30 min intervals) or saline infusions. Animals were pretreated with haloperidol (1 mg/kg/ml, i.v.) or vehicle 10 min prior to the first cocaine infusion. After each cocaine infusion the locomotor-activating effects of cocaine were measured. The spleens were collected 30 min after the third cocaine infusion and total splenocyte numbers and percentage numbers of leukocyte subpopulations were assessed using a morphological method. Three-color immunofluorescent antibody staining procedure (CD3-FITC/CD45RA-PC7/CD161A-APC and CD3-FITC/ CD4-PC7/CD8-APC) was used for determination of T, B, NK, T CD4+ and T CD8+ lymphocyte subsets. Plasma corticosterone and serum concentrations of IL-4, IFN-gamma and cocaine were assessed. In the spleen, administration of cocaine after pretreatment with haloperidol decreased numbers of splenocytes, lymphocytes and T CD4+ and B lymphocytes, significantly in comparison to rats treated with cocaine alone. The proportions between lymphocyte subsets and CD4/CD8 ratio in the spleen were not affected. Cocaine or/and haloperidol increased plasma corticosterone concentration. Serum cocaine concentration indicated the possibility of accumulation of cocaine in the applied schedule of administration. Serum concentrations of IL-4 and IFN-gamma were not altered. Haloperidol abolished locomotor response induced by cocaine infusions. In conclusion, dopaminergic activity following administration of cocaine is involved in retention of T CD4+ and B lymphocytes in the spleen of rats.
BACKGROUND AND AIMS: The subthalamic nucleus (STN) is the best target for deep brain stimulation (DBS) elevating motor symptoms in Parkinson’s disease (PD) patients. Although DBS is beneficial for patients with PD, it can also cause psychiatric and autonomic side effects of unknown etiology. Since the depression-like behavior is described after high frequency subthalamic stimulation (HFS-STN) in rats, it is important to examine impact of HFS-STN on hypothalamic-adrenal axis (HPA) activation and plasma proinflammatory cytokine levels. The aim of this study was to investigate the plasma corticosterone (CORT), TNF-α, IFN-γ and IL-6 levels following HFS-STN in hemiparkinsonian rats. METHODS: Unilateral, continuous HFS-STN (pulse width: 60 µs, frequency: 130 Hz, stimulation intensity: 30–115 µA, during 1 h. stimulation period) was provide in freely moving hemiparkinsonian rats. The model of PD was obtained by stereotactic microinjection of 6-hydroxydopamine into the right substantia nigra pars compacta. The blood samples were collected by heart puncture after the electrical or sham stimulation. The CORT level in plasma was measured by radioimmunoassay method, while concentrations of cytokines were quantified using ELISA method. RESULTS: We found that HFS-STN applied in hemiparkinsonian rats significantly increase plasma CORT level (t27=2.31, P≤0.05) in comparison to SHAM control group). We also observed an increase in IFN–γ (t27=2.89, P≤0.01) and TNF-α (t27=2.88, P≤0.01) concentrations, while the IL-6 level decrease (t27=3.38, P≤0.01) following HFS-STN. CONCLUSIONS: These data shows that HFS-STN influence endocrine and immune parameters in peripheral blood in hemiparkinsonian rats. One hour, continuous HFS-STN activated HPA axis (measured by plasma corticosterone level) and elevated concentrations of pro-inflammatory cytokines. Our results suggest that HFSSTN provokes peripheral immune and endocrine effects similar to observed in behavioral depression.
The mesolimbic dopamine system, that plays a pivotal role in emotionally motivated behaviours, originates in the ventral tegmental area (VTA) and sends major projections to the nucleus accumbens (Acb). In the present work, we assessed the effects of manipulation in the mesolimbic dopaminergic system identified with the “brain reward system” on the percentage of peripheral blood lymphocytes in freely moving rats differing in locomotor activity to novelty (high responders; HR or low responders; LR). T and B lymphocyte populations were determined by flow cytometry (CD3-FITC/CD45RA-PC7). Peripheral blood was collected from rats exposed to chronic 14 day VTA electrical stimulation to obtain feeding or exploration response (constant current 0.1 ms duration, cathodal pulses, 50 Hz, 30-min daily session) and with a lesion electrode aimed at Acb: 3 weeks after electrode implantation, after 2 weeks of chronic VTA stimulation, on the 2nd day after the Acb lesion (2mA for 15s) and on the 14th VTA stimulation day following the Acb lesion. As compared to the respective sham animals, chronic stimulation of the VTA caused significant (p<0.05) decrease in T cell percentage in both HR (33.32 ± 5.39% vs 48.04 ± 7.45%) and LR (44.99 ± 2.94% vs 51 ± 3.57%) animals. In addition, significantly (p<0.05) lower level of T cell percentage in HR animals (40.13±3.64% vs 50.94 ± 5.87%) rats were observed. On the 14th VTA stimulation day following the Acb lesion, significant (p<0.05) decrease in B cell percentage in both HR (12.47 ± 3.55% vs 19.45 ± 1.58%) and LR (7.09 ± 2.48% vs 16.5 ± 3.46%) animals was observed. On the other hand, chronic stimulation of the VTA (20.99 ± 3.38% vs 17.65 ± 0.49%) and Acb lesion (25.14 ± 2.48% vs 20.62 ± 0.85%) caused significant increase (p<0.05) in B cell percentage in HR animals. The results emphasize the importance of individual differences (HR vs LR) in the influence of the mesolimbic motivational system on blood lymphocyte distribution.
The medial septal nucleus (MS) is a forebrain limbic structure involved in learning and memory mechanisms. In previous study we found that electrolytic lesion of the medial septal nucleus caused depression of the peripheral blood natural killer cell cytotoxicity (NKCC) and the leukocyte number. In the sham operated group mere insertion of electrodes into the MS evoked transient NKCC enhancement, probably resulting from mechanical MS stimulation. To check this effect in the present study, we evaluated both spleen and blood NKCC (51Cr-release assay) and large granular lymphocytes (LGL) number (a morphological method) after chronic electrical stimulation (constant current 0.1 ms duration cathodal pulses delivered at a frequency of 50 Hz during 30-min daily session for 14 consecutive days) of MS in conscious, freely moving rats (n=12). Additionally, peripheral blood leukocyte, lymphocyte and neutrophile number was mesaured. Chronic MS stimulation caused signifi cant blood NKCC augmentation and LGL number (25.86 ± 9.31% vs. 15.75 ± 4.75%, P<0.01) in comparison to the sham group (n=13). No signifi cant effect was found in the spleen (27.14 ± 9.99% vs. 28.58 ± 8.04%). A week after termination of the stimulation procedure all measured parameters returned to the baseline. The results obtained indicate that such limbic structure as medial septum enhances antitumor and antiviral function and number of NK cells.
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