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PubMed1994

Proliferative responses towards native, heat-denatured and pepsin-treated ovalbumin by peripheral blood mononuclear cells from patients with hen's egg-sensitive atopic dermatitis.

In order to clarify the mechanism of food-antigen recognition, the proliferative responses of peripheral blood mononuclear cells (PBMCs) to native, heat-denatured or pepsin-treated ovalbumin (OA) were investigated in 16 hen's egg-sensitive patients with atopic dermatitis (AD). Seven of them had hypersensitivity to boiled hen's egg and others had not. The responses of PBMCs to heat-denatured OA were lower than those to native OA in the patients without hypersensitivity to boiled hen's egg. However, there were no differences of the responses of PBMCs between heat-denatured OA and native OA in the patients with hypersensitivity to boiled hen's egg. Moreover, the reduction of the responses of PBMCs to pepsin-treated OA was recognized in six out of seven patients. The primary structure of OA did not change by heating or pepsin treatment according to sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). These results suggested that the secondary structure of OA changed in connection with the reduction of the responses of PBMCs to denatured OA. In addition, we demonstrated the suppressive effect of anti-HLA-DR and anti-HLA-DQ monoclonal antibodies on the proliferative response of PBMCs to OA. The results suggested that the proliferative responses of PBMCs to OA were restricted by HLA-DR or HLA-DQ in hen's egg-sensitive patients with AD.

Antibodies, Monoclonal↗

PubMed2002

Induction of apoptosis of peripheral blood mononuclear cells by antithymocyte globulin (ATG) in aplastic anemia: an in vivo and in vitro study.

Antithymocyte globulin (ATG) is the standard therapy for aplastic anemia (AA) patients who are not eligible for allogeneic bone marrow transplantation (BMT). The exact mechanism of action of ATG is still not known. Profound lymphopenia is observed throughout the treatment period with ATG and appears to contribute to its immunomodulatory effect. One of the possible mechanisms, which could account for ATG-mediated lymphopenia, is by induction of apoptosis of peripheral blood mononuclear cells (PBMCs). However, there is no conclusive evidence to support this mechanism. We investigated whether ATG could induce an in vivo apoptosis in PBMCs of 12 AA patients undergoing ATG therapy by terminal deoxynucleotidyltransferase (TdT)-mediated dUTP nick-end labeling (TUNEL) assay using flow cytometry. A significant increase in the percentage of apoptosis was observed in six patients. The median percentage prior to therapy was 3 percent (range: 1-10 percent), which increased to a peak median value of 27 percent (range 17-66 percent) with therapy. ATG also induced an in vitro apoptosis of normal PBMCs as demonstrated by Annexin V and TUNEL staining using flow cytometry. Induction of apoptosis was dose dependent: 52 percent of the PBMCs exhibited Annexin V positivity after incubation with ATG at a dose of 500 microg/ml for 6 h, and 37 percent of the PBMCs exhibited DNA fragmentation after incubation with ATG at a dose of 1000 microg/ml for 24 h as demonstrated by TUNEL assay. Thus, ATG induces in vivo apoptosis in PBMCs of AA patients undergoing therapy as well as an in vitro apoptosis in normal PBMCs, and apoptosis may be an important mechanism for ATG-induced lymphopenia.

Anemia, Aplastic↗

PubMed1998

Analysis of peripheral blood mononuclear cell stimulated with pyruvate dehydrogenase complex, T-cell receptors from patients with primary biliary cirrhosis.

Progressive destruction of the intrahepatic bile ducts in patients with primary biliary cirrhosis (PBC) is thought to be mediated by cytotoxic T cells which recognize certain epitopes, such as the pyruvate dehydrogenase complex (PDC). To clarify the T-cell repertoire in PBC, we analyzed T-cell receptor (TCR) Vbeta-chain messages expressed in peripheral blood mononuclear cells (PBMCs) stimulated with PDC and in liver biopsy specimens. PBMCs from 12 PBC patients and 6 healthy controls were examined. The TCR Vbeta repertoires of unstimulated PBMCs and PBMCs stimulated with PDC purified from bovine heart were analyzed, using the reverse transcriptase-polymerase chain reaction (RT-PCR) and single-strand conformation polymorphism (SSCP). Liver biopsy specimens from 5 PBC patients were also analyzed. In the PBC patients, several different T-cell clones, some of which showed the same mobility, were evident in both the PDC-stimulated and unstimulated PBMCs, as demonstrated by SSCP analysis. In addition, TCR clonality of infiltrating lymphocytes in the liver was also observed in PBC patients, showing common clonal T-cell accumulation with that seen in PBMCs stimulated with PDC. These data indicate that common clonal T-cell accumulation specific for PDC may be present in both peripheral PBMCs and the liver of patients with PBC.

Case-Control Studies↗

PubMed2006

Anti-apoptotic activity of caffeic acid, ellagic acid and ferulic acid in normal human peripheral blood mononuclear cells: a Bcl-2 independent mechanism.

Polyphenols have been shown to induce apoptosis in a variety of tumor cells including leukemia both in vitro and in vivo. However, their action on normal human peripheral blood mononuclear cells (PBMCs) during oxidative stress remains to be explored. In this study, we have evaluated the anti-apoptotic and radical scavenging activities of dietary phenolics, namely caffeic acid (CA), ellagic acid (EA) and ferulic acid (FA). H2O2-induced apoptosis in normal human PBMCs was assayed by phosphotidylserine externalization, nucleosomal damage and DNA fragmentation. Incubation of PBMCs with 5 mM H2O2 led to increased Annexin-V binding to externalized phosphatidyl serine (PS), an event of pre-apoptotic stage of the cell. Peripheral blood mononuclear cells pretreated with phenolics could resist H2O2-induced apoptotic damage. Caffeic acid (60 and 120 microM) and EA (100 and 200 microM) caused no change in externalization of PS, whereas FA (100 and 200 microM) increased externalization of PS in PBMCs treated with H2O2. The effects of phenolics were abolished to a large extent by culturing the PBMCs for 24 h after washing the phenolics from the medium. Inhibitory activities of these phenolics on lipid peroxidation were in the order of EA<CA<FA. DPPH-scavenging activities of EA, CA and FA were found to be 31.2+/-1.36, 50+/-1.86 and 73.0+/-1.58 microM respectively. Although, the phenolics significantly inhibited DNA damage and lipid peroxidation, they could not alter the Bcl-2 expression in PBMCs. In conclusion, the anti-apoptotic effect of EA, CA and FA in PBMCs seems to be through the Bcl-2 independent mechanism.

Antioxidants↗

PubMed2006

Lithium regulates glycogen synthase kinase-3beta in human peripheral blood mononuclear cells: implication in the treatment of bipolar disorder.

BACKGROUND: Bipolar disorder has been linked to alterations in the multifunctional enzyme glycogen synthase kinase-3beta (GSK3beta). The mood stabilizer lithium inhibits GSK3beta in vitro and in mouse brain, and this is currently the strongest known potential therapeutic target of lithium. We tested whether lithium modified GSK3beta in vivo or in vitro in peripheral blood mononuclear cells (PBMCs) from healthy control and bipolar disorder subjects. METHODS: The PBMCs were obtained from 23 healthy control subjects, 9 bipolar subjects currently treated with lithium, and 13 lithium-free bipolar subjects. Immunoblot analyses were used to measure the inhibited, serine9-phosphorylated GSK3beta. RESULTS: The level of phospho-Ser9-GSK3beta in PBMCs was regulated by agents that modified kinases and phosphatases acting on GSK3beta and was increased by in vitro lithium treatment. More important, phospho-Ser9-GSK3beta levels were eightfold higher in PBMCs from lithium-treated bipolar than healthy control subjects. CONCLUSIONS: Signaling pathways regulating serine9-phosphorylation of GSK3beta can be studied in human PBMCs. Both in vitro and in vivo therapeutic lithium treatment is associated with a large increase in phospho-Ser9-GSK3beta in PBMCs. Therefore, the inhibitory serine9-phosphorylation of GSK3beta in human PBMCs may provide a biochemical marker to evaluate the association between GSK3beta inhibition and therapeutic responses to lithium treatment.

Adult↗

PubMed2006

Increased endothelial monocyte adhesiveness is related to clinical outcomes in chronic heart failure.

BACKGROUND: Vascular inflammation and endothelial dysfunction are evident in patients with chronic heart failure (CHF). We hypothesized that circulating peripheral blood mononuclear cells (PBMCs) may be activated and the resultant increased endothelial monocyte adhesion may be functionally and pathophysiologically relevant in CHF. In the present study, we investigated the clinical significance of the activity of PBMCs in patients with CHF. METHODS: PBMCs were isolated from 34 CHF patients, from 10 healthy volunteers (normal control group) and from 17 patients admitted for investigation of suspected coronary artery disease (disease control group). In each patient, the adhesiveness of PBMCs to cultured human aortic endothelial cells (HAECs) with or without tumor necrosis factor-alpha (TNF-alpha) stimulation was determined. Major adverse cardiac events (death, heart transplantation or hospitalization with worsening heart failure) were determined in the 34 CHF patients during a median follow-up period of 182 days. RESULTS: Compared with those from both control groups and from mild CHF patients, PBMCs isolated from severe CHF patients adhered more to the HAECs. The endothelial adhesiveness of PBMCs correlated positively with the circulating levels of CAMs and can supply prognostic information in CHF patients. The difference between event-free curves based on the median levels of endothelial-PBMC adhesion was significant (log rank test, p=0.0139). CONCLUSIONS: Endothelial adhesiveness of PBMCs is increased and correlated to clinical outcomes, and may be pathophysiologically relevant to the progression of CHF.

Aged↗

PubMed2006

TNF-alpha decreases hsp 27 in human blood mononuclear cells: involvement of protein kinase c.

Treatment of PBMCs with TNF-alpha decreased the levels of heat shock protein (HSP) 27, but had little effect on the level of HSP70. Parallel to the decrease of HSP27, TNF-alpha increased the level of HSP27 in the incubation medium of the cells. The decrease of HSP27 induced by TNF-alpha was suppressed by the pretreatment of PBMCs with the specific protein kinase C (PKC) inhibitor, GF109203X. Furthermore, phorbol myristate acetate (PMA), a PKC stimulant, but not dibutyryl cyclic AMP, a protein kinase A stimulant, decreased the levels of HSP27. To investigate the effect of TNF-alpha on the oligomerization state of HSP27 in PBMCs, we performed sucrose density gradient centrifugation with subsequent fractionation and immunoassay. Extract of vehicle-treated PBMCs contained mainly dissociated forms of HSP27. The amounts of dissociated forms of HSP27 in PBMCs was decreased by TNF-alpha, while the amounts of aggregated form of HSP27 was little changed. In intact PBMCs, HSP27 is constitutively phosphorylated at Ser78, but not at Ser15 or at Ser82. The amount of phosphorylated HSP27 at Ser78 was decreased by TNF-alpha. These results indicate that TNF-alpha reduces HSP27 in PBMCs through PKC activation. This decrease may be due to efflux of dissociated form of HSP27, phosphorylated HSP27 at Ser78, from the cells.

Carcinogens↗

PubMed2005

CD4-veCD8-ve CD30+ve T cells are detectable in human lung transplant patients and their proportion of the lymphocyte population after in vitro stimulation with donor spleen cells correlates with preservation of lung physiology.

INTRODUCTION: Survival following lung transplantation is less than 50% at 5 years, mainly due to immune-mediated chronic rejection. Recently a novel subset of T cells, CD4-veCD8-ve CD30+ve, so-called double negative (DN) CD30+ve T cells, has been described and shown to be responsible for tolerance in an animal model of skin transplantation. METHODS: We investigated 18 lung transplant recipients for the presence of DN CD30+ve T cells in resting peripheral blood and also following in vitro stimulation of recipient peripheral blood mononuclear cells (PBMCs) with donor spleen cells. RESULTS: Small percentages (0.2% to 6%) of DN T cells are detectable in resting PBMCs of human transplant patients (n = 18), but these did not correlate with allograft function, acute rejection episodes, HLA mismatch, or CMV status. On repeated stimulation of recipient PBMCs (two exposures) in vitro by donor spleen cells (2:1 ratio stimulators to responders) the percentage of DN CD30+ve T cells within the lymphocyte pool correlated with preservation of allograft lung function (both for FEV(1), P = .009, and FEF(25-75), P = .036) and was inversely correlated with grade of chronic rejection. On repeated exposure of recipient PBMCs to donor spleen cells with a 1:1 ratio the percentage of DN CD30+ve T cells correlated with the number of acute rejection episodes of grade 2 or greater. The total number of HLA mismatches correlated with the percentage DN CD30+ve T cells present after primary stimulation of recipient PBMCs with donor spleen cells (1:1 ratio). The number of mismatches at the B locus inversely correlated with the percentage of DN CD30+ve T cells after primary stimulation of recipient PBMCs with donor spleen cells (1:1 ratio; P = .031, n = 18). CONCLUSION: Percentages of DN CD30+ve T cells present following repeated stimulation of recipient PBMCs by donor spleen cells correlated with preservation of graft function following lung transplantation.

Antigens, CD↗

PubMed2005

Johne's disease in cattle is associated with enhanced expression of genes encoding IL-5, GATA-3, tissue inhibitors of matrix metalloproteinases 1 and 2, and factors promoting apoptosis in peripheral blood mononuclear cells.

Infection of ruminants with Mycobacterium avium subspecies paratuberculosis (M. paratuberculosis) leads to a chronic and often fatal granulomatous enteritis known as Johne's disease. Most infections with M. paratuberculosis occur during the first 6 months of life, and there is some evidence for transmission in utero. Once established, infections typically exist in a subclinical state for several years. Recent gene-expression profiling studies suggested the hypothesis that inherent gene-expression profiles in peripheral blood mononuclear cells (PBMCs) from M. paratuberculosis-infected cattle may be different than expression profiles in PBMCs from uninfected controls. If true, this would suggest that it is possible to identify an M. paratuberculosis infection "signature" through transcriptional profiling of peripheral immune cells. In addition, identification of groups or classes of genes showing inherently different expression in PBMCs from M. paratuberculosis-infected cattle relative to PBMCs from uninfected controls might highlight important interactions between this pathogen and the host immune system. In this report, we describe studies aimed at testing this hypothesis. Our novel results indicate that, indeed expression profiles of at least 42 genes are inherently different in freshly isolated PBMCs from M. paratuberculosis-infected cattle when compared to similar cells from uninfected controls. Gene-expression differences observed following microarray analysis were verified and expanded upon by quantitative real-time PCR (Q-RT-PCR). Our results indicate that T cells within PBMCs from M. paratuberculosis-infected cows have adopted a predominant Th 2-like phenotype (enhanced expression of IL-5, GATA 3, and possibly IL-4 mRNA), that cells within infected cow PBMCs may exhibit tissue remodeling deficiencies through higher expression of tissue inhibitor of matrix metalloproteinase (TIMP) 1 and TIMP2 RNA and lower expression of matrix metalloproteinase (MMP) 14 RNA than similar cells from healthy controls, and that cells within the PBMC population of M. paratuberculosis-infected cows are likely poised for rapid apoptosis (upregulation of CIDE-A, Bad, TNFRI, and Fas).

Animals↗

PubMed1998

Dermal microvascular injury in the human peripheral blood lymphocyte reconstituted-severe combined immunodeficient (HuPBL-SCID) mouse/skin allograft model is T cell mediated and inhibited by a combination of cyclosporine and rapamycin.

We have analyzed the mechanism of human endothelial injury in a human peripheral blood lymphocyte-severe combined immunodeficient (huPBL-SCID) mouse/human skin graft model of allograft injury and examined the effect of immunosuppressive drugs on this process. In this model, split-thickness human skin containing the superficial dermal microvessels was grafted onto immunodeficient C.B-17 SCID or SCID/beige mice and allowed to heal. Human peripheral blood mononuclear cells (PBMCs) allogeneic to the skin, when subsequently introduced by intraperitoneal injection, caused destruction of the human dermal microvasculature by day 16, evident as endothelial cell sloughing and thrombosis. In the same specimens, mouse microvessels that invaded the human skin graft were uninjured. Human microvascular cell injury was accompanied by a mononuclear cell infiltrate consisting of approximately equal numbers of human CD4+ and CD8+ T cells, some of which contained perforin-positive granules. We found no evidence of human natural killer cells and noted occasional human, but not mouse, macrophages at a frequency indistinguishable from that resident in skin on animals not receiving human PBMCs. These human T cell infiltrates did not extend into adjacent mouse skin. Human immunoglobulin G antibody was detected in the blood and was diffusely present throughout mouse and human tissues in SCID mice receiving PBMCs. Mouse C3 was detected on human dermal vessels in both unreconstituted control animals and those that received PBMCs. Blood and tissues from mice injected with PBMCs depleted of B cells showed no human immunoglobulin, but circulating CD3+ cells were detected by flow cytometry at levels comparable with those of animals receiving whole PBMCs. Significantly, skin graft infiltration by human T cells and human dermal microvascular injury were equivalent in the B cell-depleted and whole-PBMC-reconstituted mice. Mice inoculated with PBMCs depleted of CD8+ T cells developed microvascular injury and infiltrates containing perforin-expressing CD4+ T cells. These data suggested a cytolytic T cell-dependent mechanism of microvessel injury. We then tested the ability of T cell immunosuppressants, cyclosporine and rapamycin, to attenuate vessel damage. Neither cyclosporine nor rapamycin alone effectively reduced either mononuclear cell infiltration or vascular injury. However, a combination of the two agents reduced both parameters. We conclude that the huPBL-SCID/skin allograft model may be used both to study cytolytic T cell-mediated rejection and to test the effect of immunosuppressive drug strategies in vivo in a small-animal model of human immune responses.

Animals↗

PubMed2002

Optimisation of herpes simplex virus-based vectors for delivery to human peripheral blood mononuclear cells.

Peripheral blood mononuclear cells (PBMCs) represent a significant target for gene delivery both for therapeutic and experimental purposes. Thus far however, it has proved difficult to develop vectors capable of high efficient gene delivery to unstimulated PBMCs. We have tested a range of different vectors derived from herpes simplex virus (HSV) which differ in their degree of disablement in terms of their gene delivery efficiency to unstimulated human PBMCs and ability to deliver a reporter gene. None of the viruses had any significant toxic effect in PBMCs. However, optimal gene delivery to unstimulated PBMCs was obtained with a semidisabled virus lacking functional genes encoding ICP34.5 and Vmw65 which was more efficient than either nondisabled or more extremely disabled viruses. Expression of green fluorescent protein (GFP) with this virus was observed in up to 50% of PBMCs 1 day after infection, and reporter gene expression was detectable by Western blotting and immunofluorescence at undiminished levels at the longest time points tested, up to 5 days after infection. This optimised HSV vector may thus represent an effective tool for gene delivery to unstimulated PBMCs in culture.

Animals↗

PubMed1992

Decreased interferon gamma and increased interleukin-4 production in atopic dermatitis promotes IgE synthesis.

The mechanism(s) responsible for increased IgE synthesis in atopic dermatitis (AD) are unknown, but they may be related to either decreased interferon gamma (IFN-gamma) and/or increased interleukin (IL)-4 production. In this study we examined peripheral blood mononuclear cells (PBMCs) from 21 patients with AD, six patients with psoriasis, and 22 nonatopic healthy controls for IFN-gamma and IL-4 production after stimulation with concanavalin A (Con A). The Con A-induced proliferative response of AD PBMCs was similar to the response of healthy controls (p = 0.9). After mitogen stimulation, however, AD culture supernatants contained significantly less IFN-gamma (p = 0.001) but increased IL-4 (p = 0.001) compared with supernatants from nonatopic controls. In contrast, PBMCs from patients with psoriasis produced normal levels of IFN-gamma and IL-4 in vitro. Since IL-4 is known to decrease IFN-gamma synthesis, we examined the effect of neutralizing anti-IL-4 on IFN-gamma production. Anti-IL-4 significantly increased IFN-gamma production in patients with AD (p = 0.008) and nonatopic controls (p = 0.02) but did not normalize IFN-gamma production by AD PBMCs. Supernatants from AD PBMCs, but not supernatants from nonatopic PBMCs, induced IgE synthesis in PBMCs from nonatopic donors (p = 0.02). When an anti-IFN-gamma receptor antibody, which blocks cellular binding of IFN-gamma, was added to supernatants from nonatopic controls their capacity to induce IgE synthesis was significantly greater (p = 0.03). These results demonstrate an imbalance of IL-4 and IFN-gamma production, which may contribute to increased IgE synthesis in AD.

Dermatitis, Atopic↗

PubMed1998

Unique profile of IL-4 and IFN-gamma production by peripheral blood mononuclear cells in infants with atopic dermatitis.

Although the level of house dust mite (HDM)-specific lymphocyte proliferation is high in infants with atopic dermatitis (AD), the level of HDM-specific IgE antibody (HDM-IgE-RAST) is usually very low or negative. To elucidate the cause of the deficient HDM-specific IgE antibody formation in infants with AD, we examined the profile of IL-4 and IFN-gamma production by HDM-stimulated PBMCs. The amount of IL-4 production was higher in infants with AD and in children with AD (3 to 15 years) than in the nonatopic control subjects. Although the amount of IFN-gamma production in children with AD was lower than that found in nonatopic children, it was higher in infants with AD than in nonatopic infants. This result suggests that HDM-specific helper T lymphocytes in infants with AD have not yet differentiated into TH2 but rather stayed at the stage of TH0. The level of IgE-RAST for egg white (EW) is already elevated in infants with AD. The amount of IL-4 produced by EW-stimulated PBMCs was comparable to that produced by HDM-stimulated PBMCs in infants with AD. However, the amount of IFN-gamma produced by EW-stimulated PBMCs was distinctly lower than that produced by HDM-stimulated PBMCs in infants with AD. Although there was no correlation between the amount of IL-4 production by HDM-stimulated PBMCs and the level of HDM-IgE-RAST in infants with AD, the amount of IL-4 production by EW-stimulated PBMCs was closely correlated with the level of EW-IgE-RAST. These results suggest that it is not the lack of IL-4 but rather a relative increase in IFN-gamma production by HDM-specific helper T lymphocytes that causes the deficiency of HDM-specific IgE-antibody synthesis in infants with AD.

Adolescent↗

PubMed2002

Catecholamine production and tyrosine hydroxylase expression in peripheral blood mononuclear cells from multiple sclerosis patients: effect of cell stimulation and possible relevance for activation-induced apoptosis.

Sympathoadrenergic mechanisms may play a role in multiple sclerosis (MS). We examined catecholamine (CA) levels and production and tyrosine hydroxylase (TH) expression in peripheral blood mononuclear cells (PBMCs) from MS patients, and the correlation between CA production and apoptosis in PBMCs. PBMCs from MS patients had increased norepinephrine (NE) levels. However, phytohaemagglutinin (PHA)-stimulated PBMCs from MS patients with active disease synthesized less dopamine (DA) than cells from both healthy controls and patients with inactive disease. PBMCs from patients with inactive disease showed lower expression of TH. Pharmacological inhibition of TH in cultured PBMCs stimulated with PHA reduced the percentage of apoptotic cells. Since a failure of activation-induced apoptosis in immune cells may be involved in MS, it is suggested that altered CA production by PBMCs may be implicated in such dysregulation.

Adult↗

PubMed2001

Vascular endothelial growth factor mRNA synthesis by peripheral blood mononuclear cells in patients with acute myocardial infarction.

We studied vascular endothelial growth factor (VEGF) mRNA synthesis by peripheral blood mononuclear cells (PBMCs) in 30 patients with acute myocardial infarction (AMI) and 20 healthy individuals. PBMCs were isolated from all patients on days 3 and 14 after the onset of aMI, and from all of control individuals. To prepare samples containing identical amounts of GAPDH cDNA, competitive PCR was performed by co-amplifying serial dilutions of GAPDH mutant templates. Next, to measure VEGF cDNA quantitatively in the samples containing identical amounts of GAPDH, we also used competitive PCR by co-amplifying mutant templates of VEGF. The serum VEGF concentrations on day 14 in patients with aMI were measured by an ELISA method. Higher levels of VEGF mRNA in PBMCs were present on day 14 than either on day 3 or in the control group. Serum VEGF concentrations correlated with the VEGF mRNA levels of PBMCs on day 14. Peak serum CK levels correlated well with VEGF mRNA levels of PBMCs on day 14. The present findings suggest that PBMCs may be one of the candidates responsible for elevated circulatory VEGF protein following aMI. In addition, VEGF mRNA may be overexpressed in PBMCs in response to cardiac muscle damage.

Aged↗

PubMedDate not supplied

Quantification of hepatitis C virus RNA in peripheral blood mononuclear cells: a comparison between patients chronically infected by HCV and patients coinfected by HIV.

In patients chronically infected by hepatitis C virus (HCV), peripheral blood mononuclear cells (PBMCs) were shown to be targets for virus replication and in those coinfected with HIV, HCV viraemia was considerably increased. The purpose of this study was to quantify HCV RNA in PBMCs from 25 patients infected by HCV and from 25 patients coinfected by HCV and HIV. We used the branched DNA assay after extraction of total RNA on 5 x 10(6) cells to quantify HCV RNA, and the Inno LiPA assay to determine the HCV genotype. HCV RNA in PBMCs could be quantified in 8/25 patients in each group, but the HCV RNA concentration was very low in comparison with viraemia, since the highest result was 8.1 x 10(4) Eq genome/10(6) cells. In 10 ml of total blood, there was approximately 100 to 5,000 times less HCV RNA in PBMCs than in the plasma. It is therefore likely that PBMCs play only a minor part in the viral load present in the plasma. There was no preferential genotype associated with quantifiable HCV RNA in the PBMCs. In the case of HIV coinfection, there was no increase in the HCV-RNA concentration in PBMCs that could explain the increased viraemia observed in these patients. On the contrary, HCV RNA could not even be detected by RT-PCR in some of our coinfected patients.

Adult↗

PubMed2003

Configuration and replication competence of hepatitis B virus DNA in peripheral blood mononuclear cells from chronic hepatitis B patients and patients who have recovered from acute self-limited hepatitis.

Several studies have reported the presence of hepatitis B virus (HBV) DNA in peripheral blood mononuclear cells (PBMCs). However, the mode of existence, infectivity and replication competence of HBV DNA in PBMCs remain unclear. To clarify these questions, we assayed for covalently closed circular DNA (cccDNA) and integrated HBV DNA in PBMCs from ten patients who had recovered from acute self-limited hepatitis B (AHB) and 14 chronic hepatitis B (CHB) patients. HBV DNA was detected in all six CHB patients who were positive for hepatitis B e antigen (eAg), and cccDNA was detected in five. HBV DNA was detected in seven of the eight CHB patients who were negative for eAg, and cccDNA was detected in three. HBV DNA and cccDNA was also detected in five and two of the ten AHB patients, respectively. Integrated HBV was not detected in PBMCs of any patients. HBV DNA presence and replication competence in PBMCs are associated with eAg-positive phenotype. Sequence analysis revealed that the source of HBV DNA differed between PBMCs and serum. Even in patients who have recovered from AHB, replicable HBV may persist in PBMCs.

Journal Article↗

PubMed2002

Expression of type I interferon receptor in liver and peripheral blood mononuclear cells in chronic hepatitis C patients.

To assess whether peripheral blood mononuclear cells (PBMCs) can be substituted for liver tissue in monitoring the hepatic expression of interferon receptors (IFNAR1 and/or IFNAR2) in chronic hepatitis C, we quantified the mRNA of both subunits, using the competitive polymerase chain reaction, in the liver specimens from 37 chronic hepatitis C patients and in PBMCs from 29 of 37 patients. PBMCs from 10 healthy subjects were also quantified for both subunits. Hepatic expression of IFNAR1 and IFNAR2 were more frequent than PBMCs: IFNAR1, 23/27 (85%) vs 7/19 (37%), P = 0.0021, IFNAR2, 34/37 (92%) vs 19/29 (66%), P = 0.0182. Neither subunits was detected in PBMCs from control subjects. The expression level of IFNAR2 in the liver was related to that in PBMCs (r = 0.613, P = 0.0052). These results suggest that hepatitis C virus infection may up-regulate the expression of the type I interferon receptor and that the measurement of IFNAR2 expression in PBMCs may be useful for monitoring its expression in liver.

Adult↗