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PubMed2003

Absence of viral antigens on the surface of equine herpesvirus-1-infected peripheral blood mononuclear cells: a strategy to avoid complement-mediated lysis.

Equine herpesvirus-1 (EHV-1) may cause abortion in vaccination- and infection-immune horses. EHV-1-infected peripheral blood mononuclear cells (PBMCs) play an important role in virus immune evasion. The mechanisms by which infected PBMCs can avoid destruction by EHV-1-specific antibody and equine complement were examined. The majority of EHV-1-infected PBMCs (68.6 %) lacked surface expression of viral antigens and these cells were not susceptible to complement-mediated lysis. In infected PBMCs with surface expression of viral antigens, 63 % showed focal surface expression, whereas 37 % showed general surface expression. General surface expression rendered infected PBMCs susceptible to lysis by antibody and complement (from 5.4 to 31.2 % lysed cells depending on the concentration of antibody and complement). Infected PBMCs with focal surface expression showed significant lysis only in the presence of high concentrations of antibody and complement. Thus, the absence of surface expression protects infected PBMCs against complement-mediated lysis.

Animals↗

PubMed1997

In vitro immunologic and virologic effects of interleukin 15 on peripheral blood mononuclear cells from normal donors and human immunodeficiency virus type 1-infected patients.

Interleukin 15 (IL-15) is a cytokine that shares receptor subunits and functional activity, such as T-cell and B-cell stimulation, with IL-2. The effect of IL-2 on immune function and human immunodeficiency virus (HIV) viral load in HIV-infected patients is being actively studied. Thus, we examined how IL-15 compares with IL-2 in several in vitro immunologic and virologic assays in order to explore whether a rationale exists for pursuing initial clinical therapeutic trials with IL-15. The effects of IL-15 on induction of lymphokine-activated killer (LAK) cells, gamma interferon (IFN-gamma) production from HIV-positive peripheral blood mononuclear cells (PBMCs), and HIV production from PBMCs were studied. Induction of LAK cells by IL-15 was found in eight of eight HIV-positive donors. Incubation of PBMCs from some donors with IL-15 (1, 10, 50, and 100 ng/ml) induced production of IFN-gamma. The effect of IL-15 was compared with that of IL-2 on HIV replication in PBMCs from five HIV-positive patients and four HIV-negative donors whose PBMCs were infected in vitro with HIV. Levels of HIV p24 antigen were moderately lower in the presence of 10 ng of IL-15 per ml than with 10 ng of IL-2 per ml, but they were similar for 100 and 500 ng of each cytokine per ml. In summary, IL-15 can induce LAK cell activity in HIV-seropositive patients and can stimulate IFN-gamma production from PBMCs of some donors. IL-15 stimulates levels of HIV production from PBMCs which are similar to or moderately lower than those obtained with IL-2, depending on cytokine concentration.

Cells, Cultured↗

PubMed2002

Penicillium marneffei causes osteopontin-mediated production of interleukin-12 by peripheral blood mononuclear cells.

We investigated the role of osteopontin (OPN) in interleukin-12 (IL-12) production from peripheral blood mononuclear cells (PBMCs) stimulated with Penicillium marneffei. Kinetic studies showed that OPN synthesis preceded that of IL-12 at both mRNA and protein levels when PBMCs were cocultured with P. marneffei. Treatment with anti-OPN monoclonal antibodies (MAb) significantly suppressed IL-12 secretion. Furthermore, native OPN induced a profound level of synthesis of IL-12 from noninfected PBMCs. The major cellular source of OPN was monocytes, because depletion of CD14(+) cells resulted in the abrogation of such production. We also examined the regulatory role of granulocyte-macrophage colony-stimulating factor (GM-CSF) in OPN secretion from P. marneffei-stimulated PBMCs. Neutralizing anti-GM-CSF MAb significantly reduced OPN secretion, and treatment with this cytokine induced OPN production from both infected and noninfected PBMCs. Finally, antagonists against the mannose receptor but not the beta-glucan receptor almost completely abrogated the production of OPN. Our results demonstrated that OPN secreted from monocytes is involved in the production of IL-12 from PBMCs after stimulation with P. marneffei and that OPN production is regulated by GM-CSF. Our results also indicated the possible involvement of the mannose receptor as a signal-transducing receptor for triggering the secretion of OPN by P. marneffei-stimulated PBMCs.

AIDS-Related Opportunistic Infections↗

PubMed2005

Dysregulated innate immune responses in young children with autism spectrum disorders: their relationship to gastrointestinal symptoms and dietary intervention.

OBJECTIVE: Our previous study indicated an association between cellular immune reactivity to common dietary proteins (DPs) and excessive proinflammatory cytokine production with endotoxin (lipopolysaccharide, LPS), a major stimulant of innate immunity in the gut mucosa, in a subset of autism spectrum disorder (ASD) children. However, it is unclear whether such abnormal LPS responses are intrinsic in these ASD children or the results of chronic gastrointestinal (GI) inflammation secondary to immune reactivity to DPs. This study further explored possible dysregulated production of proinflammatory and counter-regulatory cytokines with LPS in ASD children and its relationship to GI symptoms and the effects of dietary intervention measures. METHODS: This study includes ASD children (median age 4.8 years) on the unrestricted (n = 100) or elimination (n = 77) diet appropriate with their immune reactivity. Controls include children with non-allergic food hypersensitivity (NFH; median age 2.9 years) on the unrestricted (n = 14) or elimination (n = 16) diet, and typically developing children (median age 4.5 years, n = 13). The innate immune responses were assessed by measuring production of proinflammatory (TNF-alpha, IL-1beta, IL-6, and IL-12) and counter-regulatory (IL-1ra, IL-10, and sTNFRII) cytokines by peripheral blood mononuclear cells (PBMCs) with LPS. The results were also compared to T-cell responses with common DPs and control T-cell mitogens assessed by measuring T-cell cytokine production. RESULTS: ASD and NFH PBMCs produced higher levels of TNF-alpha with LPS than controls regardless of dietary interventions. However, only in PBMCs from ASD children with positive gastrointestinal (GI(+)) symptoms, did we find a positive association between TNF-alpha levels produced with LPS and those with cow's milk protein (CMP) and its major components regardless of dietary interventions. In the unrestricted diet group, GI(+) ASD PBMCs produced higher IL-12 than controls and less IL-10 than GI(-) ASD PBMCs with LPS. GI(+) ASD but not GI(-) ASD or NFH PBMCs produced less counter-regulatory cytokines with LPS in the unrestricted diet group than in the elimination diet group. There was no significant difference among the study groups with regard to cytokine production in responses to T-cell mitogens and other recall antigens. CONCLUSION: Our results revealed that there are findings limited to GI(+) ASD PBMCs in both the unrestricted and elimination diet groups. Thus our findings indicate intrinsic defects of innate immune responses in GI(+) ASD children but not in NFH or GI(-) ASD children, suggesting a possible link between GI and behavioral symptoms mediated by innate immune abnormalities.

Autistic Disorder↗

PubMed2006

Telomerase activity is decreased in peripheral blood mononuclear cells of hemodialysis patients.

BACKGROUND: Telomerase preserves telomere length and structure, preventing cellular senescence, which is associated with alteration of the chromosomal ends. We hypothesized that telomerase activity is altered in peripheral blood mononuclear cells (PBMCs) of hemodialysis (HD) patients. To investigate this hypothesis as well as the relationship between telomerase and inflammation, we measured the activity of this reverse transcriptase as well as the level of several inflammatory markers in PBMCs and serum of an end-stage renal failure (ESRF) population and a non-renal-failure group of subjects. METHODS: In PBMCs isolated from 42 HD and 39 non-renal-failure subjects of the same age (51.0 +/- 12.4 and 51.4 +/- 12.1 years, respectively) telomerase activity was measured using PCR-ELISA; the method was based on the telomeric repeat amplification protocol. RESULTS: Telomerase activity in PBMCs was detected in 18 (42.9%) HD and 28 (71.8%) non-renal-failure subjects (p = 0.013). Among positive subjects, percent telomerase activity in PBMCs was significantly higher in non-renal- failure (117 +/- 112 %) than in HD (47.6 +/- 57.1 %) subjects (p = 0.008). Detectable telomerase activity was lower in long-term than in short-term HD patients (13.3 +/- 8.9 vs. 75.0 +/- 64.8%, respectively, p = 0.015). Although higher in HD group, inflammatory indexes (C-reactive protein, interleukin-6, IL-6, soluble IL-6 and soluble gp130) were not correlated to telomerase activity in PBMCs. CONCLUSION: Telomerase activity in PBMCs is reduced in HD patients. It seems that, at least in this type of cell in this population, defense from senescence, as assessed by telomerase activity, is altered and associated with the chronicity of uremia/HD procedure.

Adult↗

PubMed1996

Prolactin-induced immunoglobulin and autoantibody production by peripheral blood mononuclear cells from systemic lupus erythematosus and normal individuals.

Prolactin (PRL) has been shown to have immunoregulatory effects on a variety of immune responses. Its effect on B cell immune responses is suggested by in vitro data demonstrating a direct effect on B cell activation and differentiation, and also in vivo data demonstrating a biphasic stimulation of antibody production to sheep red blood cells. In addition, it has been shown both in animal models and patients with hyperprolactinemia that PRL may influence the presence of certain autoantibodies. The objective of this work was to study the effect of PRL on the induction of immunoglobulins, and anti-DNA and rheumatoid factor (RF) autoantibody production from peripheral blood mononuclear cells (PBMCs) from normal individuals and systemic lupus erythematosus (SLE) patients. Six female SLE patients and 10 normal individuals (5 females and 5 males) were studied. Ficoll-Hypaque-isolated PBMCs (1x10(6) cells/ml) with high concentrations of PRL (10(-4)-10(-8)M) and pokeweed mitogen (PWM) diluted to 1:400. An ELISA assay was used for immunoglobulins, RF and anti-dsDNA antibodies. PRL stimulated IgG and IgM production in a biphasic manner in normal PBMCs. Enhanced synthesis was observed at 10(-6) M, and a stimulatory effect was again observed at higher doses of PRL (10(-4))M. In contrast, only a mild stimulatory effect was observed in IgG synthesis by SLE PBMCs. These changes in Ig synthesis, however, did not reach statistical significance. PRL also induced IgG and IgM anti-dsDNA antibodies by both normal and SLE lymphocytes, but no differences were observed when compared to PWM stimulation. PRL induced IgM RF synthesis by normal lymphocytes but had no effect on SLE PBMCs. This study demonstrates that PRL induced immunoglobulin synthesis by normal and to a lesser degree by SLE lymphocytes, and also induced anti-dsDNA antibody by normal and SLE PBMCs, and IgM RF by normal PBMCs. However, the exact mechanism(s) of PRL action on the immune response awaits elucidation.

Adult↗

PubMed1998

Corticosteroid-resistant bronchial asthma is associated with increased c-fos expression in monocytes and T lymphocytes.

Unstimulated peripheral blood mononuclear cells (PBMCs) from corticosteroid-resistant (CR) but not corticosteroid-sensitive (CS) asthmatics demonstrate increased activating peptide-1 (AP-1)- and decreased glucocorticoid receptor (GR)-DNA binding. We test whether these abnormalities are associated with excessive generation of c-fos, the inducible component of AP-1. The c-fos transcription rate, mRNA and protein levels, and GR-DNA binding were quantitated in PBMCs, T cells, and monocytes from CS, CR, and nonasthmatic subjects. There was a 1.7-, 4.2-, and 2.3-fold greater increase in the baseline c-fos transcription rate, mRNA expression, and protein levels, respectively, in PBMCs derived from CR compared with CS patients. At optimal stimulation with PMA, there was a 5.7-, 3.4-, and 2-fold greater increase in the c-fos transcription rate, mRNA accumulation, and protein levels, respectively, in CR compared with CS PBMCs. These abnormalities were detected in both the T cell and monocyte subpopulations. PMA stimulation converted PBMCs from a CS to a CR phenotype and was associated with direct interaction between c-Fos and the GR. Pretreatment of PBMCs from CR patients with c-fos antisense oligonucleotides enhanced GR-DNA binding activity in CR PBMCs stimulated with dexamethasone. We suggest that increased c-fos synthesis provides a major mechanism for the increased AP-1- and decreased GR- DNA binding seen in CR asthma.

Adrenal Cortex Hormones↗

PubMed2004

MDR1 mRNA expressions in peripheral blood mononuclear cells of patients with ulcerative colitis in relation to glucocorticoid administration.

Overexpression of multidrug resistance (MDR) protein, P-glycoprotein (P-gp), on lymphocytes has been suggested to be implicated in the failure of glucocorticoid (GC) therapy in patients with ulcerative colitis (UC). However, whether the overexpression of P-gp in a class of patients with inflammatory bowel disease (IBD) is intrinsic or related to the administration of GC is unknown. Relative amounts of MDR1 mRNA expressed in peripheral blood mononuclear cells (PBMCs) were measured using the reverse-transcriptase polymerase chain reaction (RT-PCR) technique in 25 UC patients having no history of GC administration, 25 UC patients having experienced GC therapy, 19 patients with Crohn's disease (CD) with no history of GC therapy, and 27 healthy subjects. Relative amounts of MDR1 mRNA expressed in PBMCs were compared among the groups. The relationship between the amounts of MDR1 mRNA expressed, as well as the total dose of GC administered or the period of GC therapy in UC patients, was examined. The relative amounts of MDR1 mRNA expressed in PBMCs were not significantly different between the healthy subjects and CD patients or UC patients having no history of GC therapy. However, the mean MDR1 mRNA amount in PBMCs of UC patients having experienced GC therapy was significantly greater than that in PBMCs of UC patients with no history of GC administration (p = 0.0375). The amounts of MDR1 mRNA in PBMCs of UC patients having experienced GC therapy significantly correlated with the total dose of GCs administered (p = 0.0175). Overexpression of MDR1 mRNA in PBMCs of IBD patients is not intrinsic. However, high-dose administration of GCs for the treatment of UC may result in an increased expression of MDR1 mRNA, which may impair successful GC therapy in these patients.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMed2005

Impaired interferon-gamma production as a consequence of STAT4 deficiency after autologous hematopoietic stem cell transplantation for lymphoma.

Production of interferon gamma (IFN-gamma) is critical for optimal antitumor immunotherapy in several preclinical animal models. Interleukin-12 (IL-12)-induced IFN-gamma production is markedly defective after autologous stem cell transplantation. Quantitative deficiency in CD4 T cells, relative increase in CD25+CD4+ T cells, and bias toward T helper 2 (Th2) differentiation are not the primary mechanisms of defective IFN-gamma production. IL-12 receptor beta1 (IL-12Rbeta1) and IL-12Rbeta2 are expressed at equivalent or higher levels on posttransplantation patient peripheral blood mononuclear cells (PBMCs) as compared with control PBMCs. IL-12-induced tyrosine phosphorylation of signal transducer and activator of transcription 4 (STAT4) was undetectable or barely detectable in posttransplantation patient PBMCs, whereas IL-4-induced tyrosine phosphorylation of STAT6 did not differ in posttransplantation patient and control PBMCs. Levels of STAT4 protein were decreased by 97% in posttransplantation patient PBMCs. Levels of STAT4 mRNA were also significantly decreased in posttransplantation patient PBMCs. Incubation with IL-12 and IL-18 in combination partially reversed the defective IFN-gamma production by posttransplantation patient PBMCs. IFN-gamma production in response to IL-12 plus IL-18 did not require increased expression of STAT4 but was dependent on the activity of p38 mitogen-activated protein kinase (MAPK). These results indicate that defective IFN-gamma production is due to an intrinsic deficiency in STAT4 expression by posttransplantation patient lymphocytes and suggest strategies for circumventing this deficiency in cancer immunotherapy.

Case-Control Studies↗

PubMed2005

Durable engraftment of AMD3100-mobilized autologous and allogeneic peripheral-blood mononuclear cells in a canine transplantation model.

Peripheral-blood mononuclear cells (PBMCs) mobilized with AMD3100, a CXCR4 antagonist, combined with granulocyte colony-stimulating factor (G-CSF) have reconstituted autologous hematopoiesis in cancer patients following myeloablative conditioning. The engraftment potential of PBMCs mobilized with AMD3100 alone, however, has remained unproven. We therefore studied AMD3100-mobilized PBMCs in a canine model. Four dogs received 920 cGy total body irradiation (TBI) before infusion of autologous AMD3100-mobilized PBMCs (median CD34 cell count, 3.9 x 10(6)/kg). Neutrophil (> 0.5 x 10(9)/L [500/microL]) and platelet (> 20 x 10(9)/L [> 20 000/microL]) recoveries occurred at medians of 9 (range, 7-10) days and 25 (range, 23-38) days, respectively, after TBI, and all dogs had normal marrow function at 1 year after transplantation. To evaluate the long-term engraftment potential of AMD3100-mobilized PBMCs, 5 dogs were given 920 cGy TBI followed by infusion of AMD3100-mobilized PBMCs (median CD34 cell dose, 4.7 x 10(6)/kg) from their dog leukocyte antigen (DLA)-identical littermates. Neutrophil and platelet recoveries occurred at medians of 8 (range, 8-10) days and 26 (range, 26-37) days, respectively, after TBI. With a median follow-up of 53 (range, 33-61) weeks, recipients' marrow function was normal, and blood-donor chimerism levels were 97% to 100%. In summary, both autologous and allogeneic AMD3100-mobilized PBMCs led to prompt and durable engraftment in dogs after 920 cGy TBI.

Animals↗

PubMed2003

Comparison of suppressive potency between azathioprine and 6-mercaptopurine against mitogen-induced blastogenesis of human peripheral blood mononuclear cells in-vitro.

Azathioprine (AZ) is a prodrug of 6-mercaptopurine (6-MP), but little is known about the relative suppressive efficacy of these agents against blastogenesis of human peripheral blood mononuclear cells (PBMCs) in-vitro. We compared the pharmacological efficacy of AZ and 6-MP against T cell mitogen-induced blastogenesis of PBMCs in-vitro. PBMCs were obtained from seven healthy subjects. The potency of AZ and 6-MP to suppress PBMC-blastogenesis in-vitro was compared using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay procedures. Production of 6-MP from AZ in PBMC culture was examined by high-performance liquid chromatography. Both AZ and 6-MP dose-dependently suppressed PBMC blastogenesis. Mean +/- s.d. IC50 (concentration of drug that gave 50% inhibition) values for AZ and 6-MP were 230.4 +/- 231.3 and 149.5 +/- 124.9 nM, respectively. Thus, the potencies of AZ and 6-MP to suppress PBMC blastogenesis were not significantly different. A significant correlation was observed between the IC50 values of AZ and 6-MP (P < 0.01, n = 6). After incubation of PBMCs with 100 microM AZ for 4 days, 35.3-92.5 microM 6-MP was liberated into the culture medium. The ratio of 6-MP liberation from AZ was influenced by the presence of PBMCs, but not by the medium or fetal bovine serum. The results suggest that the suppressive potency of the prodrug AZ and its converted form 6-MP against blastogenesis of human PBMCs in-vitro is similar, although PBMCs appeared to convert AZ to 6-MP. AZ is suggested to be effective after conversion to 6-MP to express immunosuppressive efficacy in-vitro.

Adult↗

PubMed2003

Metabolism of tenofovir and didanosine in quiescent or stimulated human peripheral blood mononuclear cells.

OBJECTIVE: As tenofovir disoproxil fumarate substantially increases plasma concentrations of didanosine in patients with human immunodeficiency virus-1 infection, we sought to determine whether tenofovir and didanosine showed a similar intracellular interaction in human peripheral blood mononuclear cells (PBMCs). DESIGN: Comparative in vitro incubation of two antiretrovirals in lymphocytes. SETTING: Clinical research laboratory. MATERIAL: Radiolabeled tenofovir and didanosine in human PBMCs. MEASUREMENTS AND MAIN RESULTS: Phosphorylation of 2 and 20 microM didanosine to dideoxyadenosine triphosphate (ddATP) was determined in quiescent and stimulated PBMCs in the presence or absence of 5 microM tenofovir. Similarly, phosphorylation of 5 microM tenofovir to tenofovir diphosphate (TFVpp) was examined in the presence or absence of 2 and 20 microM didanosine. Intracellular amounts of ddATP and TFVpp were determined by incubating PBMCs with radiolabeled tenofovir or didanosine alone and together for up to 16 hours and then separating the anabolites by high-performance liquid chromatography for quantitation. The presence of tenofovir did not affect the amount of ddATP in quiescent or stimulated PBMCs with 2 or 20 microM didanosine. In addition, didanosine did not alter the amount of TFVpp that formed. The amount of ddATP was modestly (1.5-3-fold) but consistently higher in stimulated than in quiescent PBMCs, but the amount of TFVpp did not differ. CONCLUSION: There is no significant interaction between tenofovir and didanosine in human PBMCs as determined by the extent of formation of the phosphorylated anabolites. This suggests that adjusting didanosine dosage, when given with tenofovir, to achieve similar didanosine plasma concentrations, may be sufficient to accommodate the systemic drug interaction.

Adenine↗

PubMed1995

Chronic & infrequent opioid exposure suppresses IL-2R expression on rhesus monkey peripheral blood mononuclear cells following stimulation with pokeweed mitogen.

In the present investigation, infrequent and chronic (daily) exposure of rhesus monkeys to morphine was investigated for their effect on cytokine receptor expression, interleukin (IL)-2-production, and the nuclear factor kappa B (NF kappa B) levels following stimulation with PWM. In a time-dependent manner, peripheral blood mononuclear cells (PBMCs) from both infrequent- and daily-morphine treated monkeys displayed significantly less (40 +/- 7%) IL-2 receptor compared to PBMCs from saline-treated controls. However, PBMCs from chronic opioid- and infrequent opioid-treated monkeys displayed similar levels of IL-4 and IL-7 receptors compared to saline-treated animals. Following stimulation with PWM, PBMCs from chronic opioid-treated monkeys produced elevated levels of IL-2 (870 +/- 94 pg/ml) compared to IL-2 levels (463 +/- 88 pg/ml) of PBMCs from saline-treated monkey. Likewise, PBMCs from chronic-morphine exposed monkeys had elevated levels (43%) of NF kappa B compared to PBMCs from saline-treated (control) monkeys following 72 hours in culture with PWM. Collectively, these observations suggest morphine regulates selective molecular events that manifest in biologically altered immune function including T cell activation and IL-2 production.

Animals↗

PubMed1999

[Interleukin 10(IL-10) levels in serums and in culture supernatants of peripheral blood mononuclear cells from patients with chronic hepatitis B].

OBJECTIVE: To understand the impacts of interleukin 10(IL-10) in serums and in culture supernatants of peripheral blood mononuclear cells (PBMCs) from patients with chronic hepatitis B (CHB) on the persistent infection of hepatitis B virus (HBV). METHODS: The levels of IL-10 in serums and in culture supernatants of PBMCs from 15 patients with CHB and 6 normal controls were measured by ELISA method. PBMCs were cultured with or without Staphylococcus aureus enterotoxin B(SEB; 0.2 microgram/ml) or recombinant HBcAg(rHBcAg; 1.0 microgram/ml) for 48 hours in vitro. RESULTS: The levels of IL-10 in serums was greater in patients with CHB (123.11 +/- 13.89 ng/L) than in controls (95.97 +/- 11.68 ng/L, P < 0.01). Spontaneous, SEB-induced and rHBcAg-induced IL-10 production by PBMCs after culturing in vitro for 48 hours was also higher in patients with CHB (280.82 +/- 50.56 ng/L, 321.69 +/- 37.04 ng/L and 369.58 +/- 30.52 ng/L) than in controls (100.2 +/- 8.54 ng/L, 203.41 +/- 12.02 ng/L and 202.38 +/- 15.79 ng/L; P < 0.01). In addition, IL-10 production by rHBcAg-stimulated PBMCs was significantly greater than by SEB-stimulated PBMCs in patients with CHB. Patients with seropositive for HBV DNA had higher levels of IL-10 in serums and in culture supernatants of PBMCs than patients with seronegative for HBV DNA. But the levels of IL-10 were lower in patients with chronic heavy and severe hepatitis B than in patients with chronic mild and moderate hepatitis B. CONCLUSION: IL-10 may have an important role in persistent infection of HBV in patient with CHB.

Adolescent↗

PubMed2000

[Identification and visualization of virus-like particles in peripheral blood mononuclear cells].

OBJECTIVE: PBMCs from patients with chronic hepatitis C were examined by electron microscopy (EM), immune EM (IEM) combined with immunohistochemistry to trace the infection and morphology of HCV in the PBMCs. METHODS: The PBMCs from 28 patients with chronic hepatitis C were analyzed firstly for HCV RNA and HCV antigens by RT-PCR and immunohistochemistry respectively. The PBMCs with positive HCV RNA and HCV antigens were then observed by electron microscopy. RESULTS: The positive rate of HCV RNA and HCV antigens were 77.27% (17/22) and 75.00% (21/28), respectively. Two types of polymorphic HCV-like particles with diameter of approximately 65 nm and 110 nm were observed in cytoplasmic vesicles of the PBMCs from HCV Ag positive patients. 10 patients with HCV Ag over expression were studied by electron microscopy and immune election microscopy. The budding phenomenon of the particles could be also visualized in the cytoplasmic vesicles of the PBMCs. Some ultrastructural changes showed an increased ratio of cytoplasm to nucleus and pyknosis of nucleus was also suggestive of the virus infection. CONCLUSIONS: The EM study demonstrated the infection and replication of HCV in the PBMCs of chronic hepatitis C patients by morphology and morphogenesis.

Hepacivirus↗

PubMed2001

Effects of sex hormones on apoptosis in peripheral blood mononuclear cells from patients with systemic lupus erythematosus.

OBJECTIVE: To study the effects of dehydroepiandrosterone (DHEA) and/or estradiol (E2) on apoptosis in peripheral blood mononuclear cells (PBMCs) from patients with systemic lupus erythematosus (SLE). METHODS: The percentage of apoptosis of PBMCs from SLE patients and healthy blood donors were examined by means of AO staining 48 h after culture with DHEA and/or E2 at physiologic or pathologic concentrations. RESULTS: The percentage apoptosis of PBMCs from SLE patients is higher than that of healthy blood donors (P < 0.01). E2, whether at physiological or at pathological concentrations, had no effects no apoptosis of PBMCs from both SLE patients and healthy donors (P > 0.05). Both DHEA and DHEA plus E2 at physiologic concentrations, had no effect on apoptosis of PBMCs from healthy donors (P > 0.05), but significantly inhibited that of SLE patients (P < 0.05); at pathologic concentrations, they promoted apoptosis of PBMCs from SLE patients as well as healthy blood donors (P < 0.05). There were no significant differences between the effects of DHEA and that of DHEA plus E2 (P > 0.05). CONCLUSION: DHEA plays an important role in the apoptosis of PBMCs from SLE patients; low serum levels of DHEA may cause accelerated apoptosis.

Adult↗

PubMed2001

[The effects of adenosine, interleukin-1 and theophylline on the expression of A2 adenosine receptor mRNAs in peripheral blood mononuclear cells from asthmatic patients].

OBJECTIVE: To explore the roles of A2a and A2b adenosine receptors(A2aAR and A2bAR) in the pathogenesis of bronchial asthma. METHODS: Eleven asthmatics and 11 healthy subjects were included in this investigation. Peripheral blood mononuclear cells(PBMCs) were obtained after density centrifugation on Ficoll-Hypaque. The expression of A2aAR and A2bAR mRNAs in PBMCs was examined by use of the reverse transcription-polymerase chain reaction (RT-PCR) and image pattern analysis semiquantitation method. RESULTS: The expression of A2aAR mRNA in PBMCs from asthmatics was not enhanced, compared with that of healthy subjects(P > 0.05). Adenosine or IL-1 or theophylline was not observed to have effect on A2aAR mRNA (P > 0.05). A2bAR gene expression in PBMCs was more markedly in patients with asthma than in normal individuals (P < 0.05). Adenosine and IL-1 significantly increased A2bAR mRNAs in asthmatics. Theophylline inhibited A2bAR gene expressions. The expression of A2bAR mRNA in PBMCs was positively correlated to serum total immunoglobulin E (TIgE) level in asthmatics (r = 0.72, P < 0.01). The expressions of A2bAR mRNA elevated by adenosine or IL-1 were correlated positively with serum TIgE(r = 0.78 or r = 0.61; P < 0.05) and negatively with the forced expiratory volume in first second to predicted value ratio (FEV1%) (r = -0.62 or r = -0.81; P < 0.05) in asthmatics. CONCLUSION: These results indicated that the expression of A2bAR mRNAs in PBMCs was more remarkable in asthmatics than in healthy subjects; that adenosine or IL-1 potentiated the mRNA expression of A2bAR in PBMCs in asthmatic patients, which was correlated with allergy state and the degree of airway obstruction; and that theophylline might antagonize adenosine by inhibiting A2bAR mRNA.

Adenosine↗

PubMedDate not supplied

Production of TNF-alpha by peripheral blood mononuclear cells through activation of nuclear factor kappa B by specific allergen stimulation in patients with atopic asthma.

Stimulation by specific allergens induces inflammatory cytokine production from peripheral blood mononuclear cells (PBMCs) in patients with atopic asthma, but the mechanism remains unknown. PBMCs were collected from six patients with atopic asthma with an immunoglobulin E-radioallergosorbent test score to Dermatophagoides farinae of > or = 4 and six nonatopic healthy subjects (score = 0) using a dish adhesion method after density gradient centrifugation. CD23 expression in PBMCs was analyzed by the fluorescence-activated cell sorting method. PBMCs were incubated with D. farinae or lipopolysaccharide, and production of tumor necrosis factor (TNF) alpha into the supernatant was measured by enzyme-linked immunosorbent assay. After incubation, immunostaining nostaining of the PBMCs with anti-nuclear factor kappa B (NF-kappa B) antibody (anti-p65 antibody against p65 as the subunit of NF-kappa B) was performed, and NF-kappa B activation in extracted nuclear protein was examined by electrophoretic mobility shift assay. CD23 expression was significantly higher in PBMCs from patients with asthma than in the controls (p < 0.01). There was no significant difference in TNF-alpha production by lipopolysaccharide stimulation between the two groups, but D. farinae-specific TNF-alpha production was significantly higher in subjects with asthma than in the controls (p < 0.05). A significant translocation of NF-kappa B to nuclei by D. farinae stimulation was observed in cells from subjects with asthma (p < 0.01). Our results indicated that TNF-alpha production was induced by D. farinae in PBMCs of patients with atopic asthma by the activation of NF-kappa B via CD23. In patients with atopic asthma, CD23-mediated signals may cause proinflammatory cytokine production, which may lead to airway inflammation.

Adult↗