Peripheral Blood-derived Mononuclear Cells Research Articles

Editorial

Editorial

The human Pk histo-blood group antigen provides protection against HIV-1 infection

Several human histo-blood groups are glycosphingolipids, including P/P1/P(k). Glycosphingolipids are implicated in HIV-host-cell-fusion and some bind to HIV-gp120 in vitro. Based on our previous studies on Fabry disease, where P(k) accumulates and reduces infection, and a soluble P(k) analog that inhibits infection, we investigated cell surface-expressed P(k) in HIV infection. HIV-1 infection of peripheral blood-derived mononuclear cells (PBMCs) from otherwise healthy persons, with blood group P(1)(k), where P(k) is overexpressed, or blood group p, that completely lacks P(k), were compared with draw date-matched controls. Fluorescence-activated cell sorter analysis and/or thin layer chromatography were used to verify P(k) levels. P(1)(k) PBMCs were highly resistant to R5 and X4 HIV-1 infection. In contrast, p PBMCs showed 10- to 1000-fold increased susceptibility to HIV-1 infection. Surface and total cell expression of P(k), but not CD4 or chemokine coreceptor expression, correlated with infection. P(k) liposome-fused cells and CD4(+) HeLa cells manipulated to express high or low P(k) levels confirmed a protective effect of P(k). We conclude that P(k) expression strongly influences susceptibility to HIV-1 infection, which implicates P(k) as a new endogenous cell-surface factor that may provide protection against HIV-1 infection.

Recovery and functional activity of mononuclear bone marrow and peripheral blood cells after different cell isolation protocols used in clinical trials for cell therapy after acute myocardial infarction

Clinical trials showed contradictory results in functional recovery after intracoronary infusion of autologous mononuclear (bone marrow) cells in patients with acute myocardial infarction. A recent study suggests that this might be related to the isolation protocol used. In The Netherlands, a comparable randomised multicentre trial (HEBE) was designed. To validate the isolation method of bone marrow and peripheral blood-derived mononuclear cells, we compared our processing protocol with methods comparable to the ASTAMI (no beneficial effect) and the REPAIR-AMI study (beneficial effect). The effect of several factors (density gradient, washing buffer and centrifugation speed) has been studied on recovery and function (migration and clonogenic capacity) of mononuclear cells. Significantly lower cell recoveries were found at a centrifugation speed of 250 g, compared to 600 or 800 g, respectively. Furthermore, washing buffer without supplemented human serum albumin and heparin resulted in significantly lower cell recovery and functional impairment as measured by clonogenic capacity. The results of our study justify the cell-processing protocol as applied in the HEBE trial (600 g, human serum albumin supplemented washing buffer). This protocol results in viable and functional cells of which the quantity and quality is at least comparable to a successful study like the REPAIR-AMI.

Repair of large full‐thickness articular cartilage defects by transplantation of autologous uncultured bone‐marrow‐derived mononuclear cells

The objective of this study was to investigate the feasibility of autologous uncultured bone marrow-derived mononuclear cell transplantation in large full-thickness cartilage regeneration. After fixing with a hinged external fixator, the entire surface of the left tibial plateau was resected and large full-thickness cartilage defects were formed in 48 rabbits. Animals were divided into four groups: autologous uncultured bone marrow-derived mononuclear cells with fibrin gel (BMC), autologous uncultured peripheral blood-derived mononuclear cells with fibrin gel (PBC), fibrin gel alone (GEL), or nothing (CON) transplanted to the articular cavity 7 days after the operation. The rabbits were killed 8 or 12 weeks after the operation. The repair of defects was investigated histologically and scored using a histological and histochemical grading scale that was modified from the International Cartilage Repair Society Visual Histological Assessment Scale. To evaluate the regenerated cartilage, we also morphometrically measured the staining area positive for Safranin-O or type II collagen and calculated the percentages of the positive staining areas with respect to the regenerated soft tissue area. Histological findings showed that the BMC group had superior cartilage repair compared with the other groups, and that the PBC and CON group showed better cartilage repair than did the GEL group. Histological scores and morphometrical measurements also showed the same results quantitively. The transplantation of autologous uncultured bone marrow-derived mononuclear cells contributes to articular cartilage repair. The easy and safe method used in this study is potentially viable for clinical application.

Diverse regulatory activity of human heat shock proteins 60 and 70 on endotoxin-induced inflammation

Tissue injury is often associated with bacterial infection. Intracellular heat shock proteins (HSPs) are released from damaged tissue, come in contact with cells of the immune system, and might affect the immune response against bacteria. In the present study, we investigated the capacity of highly purified human HSP60 and HSP70 to modulate the response of human peripheral blood-derived mononuclear cells (PBMC) to lipopolysaccharide (LPS). HSP70 but not HSP60 decreased the LPS-induced secretion of TNF-α when added simultaneously with LPS. In contrast, HSP60 and HSP70 primed PBMC for enhanced secretion of TNF-α when added 24h prior to the stimulation with LPS. Neither HSP60 nor HSP70 alone induced the release of TNF-α. The capacity of LPS to bind to monocytes was not affected by HSPs, but HSP70 increased the expression of Toll-like receptor 4. Thus, HSP60 and HSP70 released upon tissue damage might play a role in the regulation of bacteria-induced inflammation.

ORIGIN OF IMMUNOMODULATION AFTER SOFT TISSUE TRAUMA

Severe injury may lead to immunosuppression, multiple organ failure, and death. The aim of the study was to investigate the direct impact of soft tissue destruction on the development of trauma-associated immunomodulation. Hip surgery was considered to represent an isolated soft tissue trauma that allowed for the examination of changes taking place locally at the site of trauma or systemically with regard to monocyte function and leukocyte redistribution. Peripheral blood and wound fluid collected from the drains of 21 patients after hip surgery were analyzed to determine the cellular composition and/or the responsiveness of mononuclear cells (MNCs) to lipopolysaccharide (LPS). Different factors present in the wound fluids were tested for their capacity to modulate the MNC of healthy individuals with regard to cytokine and chemokine secretion. We found that various factors, including heat-shock protein (HSP) 60 and HSP70, were locally released at the site of soft tissue trauma and could be detected in wound fluids. The wound fluid-derived MNC (but not the peripheral blood-derived MNC) showed an impaired capacity to release TNF-alpha after LPS stimulation. Cell-free wound fluid suppressed in healthy individuals the LPS-induced TNF-alpha secretion by MNC. After surgery, granulocytosis was found in peripheral blood and in wound fluids, but monocytopenia was restricted to wound fluids. In parallel, wound fluids induced in healthy individuals the release by MNC of distinct chemokines specific for granulocytes and monocytes. These wound fluid-mediated effects of TNF-alpha suppression and chemokine induction could be mimicked by recombinant human HSP70 and, in part, by HSP60. Thus, tissue-derived factors, such as HSP70 released after injury, suppress monocyte function and, therefore, might favor the development of immunosuppression after severe injury.

Genome-Wide Differential Gene Expression Profiling of Human Bone Marrow Stromal Cells

Bone marrow stromal cells (BMSCs) reside in bone marrow and provide a lifelong source of new cells for various connective tissues. Although human BMSCs are regarded as highly suitable for the development of cell therapeutics and regenerative medicine, the molecular factors and the networks of signaling pathways responsible for their biological properties are as yet unclear. To gain a comprehensive understanding of human BMSCs at the transcriptional level, we have performed DNA microarray-based, genome-wide differential gene expression analysis with the use of peripheral blood-derived mononuclear cells (MNCs) as a baseline. The resulting molecular profile of BMSCs was revealed to share no meaningful overlap with those of other human stem cell types, suggesting that the cells might express a unique set of genes for their stemness. By contrast, the distinct molecular signature, consisting of 92 different genes whose expression strengths are at least 50-fold higher in BMSCs compared with MNCs, was shown to encompass largely a gene subset of umbilical cord blood-derived adherent cells, suggesting that adherent cells derived from bone marrow and umbilical cord blood may be defined by a common set of genes, regardless of their origin. Intriguingly, a large number of these genes, particularly ones for extracellular matrix products, coincide with normal or tumor endothelium-specific markers. Taken together, our results here provide a BMSC-specific genetic catalog that may facilitate future studies on molecular mechanisms governing core properties of these cells.

Vitamin C Reversed Malfunction of Peripheral Blood-Derived Mononuclear Cells in Smokers Through Antioxidant Properties

Smoking impairs neovascularization, possibly, through the impaired function of peripheral blood-derived mononuclear cells (PB-MNCs). Thus, the mechanism of impaired function of PB-MNCs caused by chronic smoking was examined, and whether vitamin C reversed the malfunction of PB-MNCs in smokers was investigated. The cohort comprised 27 healthy male volunteers (16 smokers and 11 age-matched non-smokers). For evaluation of the colony-forming activity of PB-MNCs, the number of endothelial colony-forming units (e-CFUs) was counted in a culture assay. Migration activity of PB-MNCs was evaluated by the modified Boyden chamber method. In smokers, the number of e-CFUs was reduced to 56% and migratory activity of PB-MNCs to 40% compared with non-smokers (p<0.01). The urinary level of 8-isoprostane, an oxidative stress marker, was greater in smokers than in non-smokers (p<0.05). There was an inverse correlation between migratory activity of PB-MNCs but not between the number of e-CFUs and urinary level of 8-isoprostane. Furthermore, oral administration of vitamin C for 2 weeks ameliorated the impaired migratory activity of PB-MNCs in smokers. Chronic smoking impairs the function of PB-MNCs. Smoking-induced oxidative stress may be involved in impaired migratory activity of PB-MNCs.

Intracoronary Transplantation of Non-Expanded Peripheral Blood-Derived Mononuclear Cells Promotes Improvement of Cardiac Function in Patients With Acute Myocardial Infarction

Transplantation of non-expanded peripheral blood mononuclear cells (PBMNCs) enhances neovessel formation in ischemic myocardium and limbs by releasing angiogenic factors. This study was designed to examine whether intracoronary transplantation of PBMNCs improves cardiac function after acute myocardial infarction (AMI). After successful percutaneous coronary intervention (PCI) for a ST-elevation AMI with occlusion of proximal left anterior descending coronary artery within 24 h, patients received an intracoronary infusion of PBMNCs within 5 days after PCI (PBMNC group). PBMNCs were obtained from patients by COBE spectra-apheresis and concentrated to 10 ml, 3.3 ml of which was infused via over-the-wire catheter. The global left ventricular ejection fraction (LVEF) change from baseline to 6 months followup in th ePBMNC group that underwent standard PCI for similar AMI [corrected]. The primary endpoint was the global left ventricular ejection fraction (LVEF) change from baseline to 6 months' follow-up. The data showed that the absolute increase in LVEF was 7.4% in the control group and 13.4% (p=0.037 vs control) in the PBMNC group. Cell therapy resulted in a greater tendency of DeltaRegional ejection fraction (EF) or significant improvement in the wall motion score index and Tc-99m-tetrofosmin perfusion defect score associated with the infarct area, compared with controls. Moreover, intracoronary administration of PBMNCs did not exacerbate either left ventricular (LV) end-diastolic and end-systolic volume expansion or high-risk arrhythmia, without any adverse clinical events. Intracoronary infusion of non-expanded PBMNCs promotes improvement of LV systolic function. This less invasive and more feasible approach to collecting endothelial progenitor cells may provide a novel therapeutic option for improving cardiac function after AMI.

Benidipine, a Dihydropyridine-Ca2+ Channel Blocker, Increases the Endothelial Differentiation of Endothelial Progenitor Cells In Vitro

Benidipine is a dihydropyridine-Ca2+ channel blocker used in the treatment of hypertension and angina pectoris. In the present study, we examined the effects of benidipine on the endothelial differentiation of circulating endothelial progenitor cells (EPCs) using an in vitro culture method. Peripheral blood derived mononuclear cells (PBMCs) containing EPCs were isolated from C57BL/6 mice, and then the cells were cultured on vitronectin/gelatin-coated slide glasses. After 7 days of culture, endothelial cells differentiated from EPCs were identified as adherent cells with 1,1'-dioctadecyl-3,3,3',3'-tetramethyl-indocarbocyanine-labeled acetylated low density lipoprotein (Dil-Ac-LDL) uptake and lectin binding under a fluorescent microscope. Incubation of PBMCs for 7 days with benidipine (0.01-1 micromol/l) significantly increased the number of Dil-Ac-LDL+/fluorescein isothiocyanate-lectin (FITC-Lectin)+ cells. Wortmannin, a phosphoinositide-3 kinase (PI3K) inhibitor, selectively attenuated the effect of benidipine on the endothelial differentiation. In addition, benidipine treatment augmented the phosphorylation of Akt, indicating that the PI3K/Akt pathway contributed, at least in part, to the endothelial differentiation induced by benidipine. These results suggest that the treatment with benidipine may increase the endothelial differentiation of circulating EPCs and contribute to endothelial protection, prevention of cardiovascular disease, and/or an improvement of the prognosis after ischemic damage.

Novel Mechansims for Pk (Gb3) Inhibition of HIV Infection.

Many blood group antigens belong to a class of lipid-sugar conjugate molecules known as glycosphingolipids (GSLs), including ABH, Lewis and P blood group systems. GSLs are implicated in HIV-host cell fusion, and several bind to HIV-gp120 in vitro. In addition, inhibiting glycolipid biosynthesis has been shown to prevent HIV membrane fusion. The Pk blood group antigen, also known as globotriaosylceramide (Gb3), has been shown to augment HIV fusion in in vitro models; however its exact role in HIV infection is not known. We have previously developed a soluble analogue of Pk, a highly effective gp120 ligand, which inhibits HIV-1 and HIV-2 membrane fusion. We hypothesize that Pk influences susceptibility to HIV infection, inhibiting at the level of membrane fusion and entry. We have presently investigated the effects of soluble and expressed Pk on HIV-1 infection in vitro. Soluble Pk effects were considered by pre-treating monocyte (M)-tropic HIV-1Ba-L or T-cell (T)-tropic HIV-1IIIB with soluble Pk prior to infection of PHA or PHA/IL2 activated peripheral blood-derived mononuclear cells (PBMCs) or Jurkat T-cells. Productive M-tropic and/or T-tropic HIV-1 infection of PBMCs and Jurkat cells was inhibited by more than 90%. Inhibition of membrane fusion and entry was visualised by transmission electron microscopy. Soluble Pk had minimal effects on cell viability or HIV receptor (CD4) and co-receptor (CCR5 or CXCR4) expression as measured by flow cytometry. Exogenously introduced Pk in Jurkat T-cells was accomplished by liposome fusion, and susceptibility to T-tropic HIV-1IIIB infection was determined. Pk transfer in Jurkat T-cells also reduced productive HIV-1 infection by 50%, without affecting cell viability or HIV receptor and co-receptor expression. Effects of differential Pk expression on HIV susceptibility was investigated using PBMCs with Pk accumulation, from patients with Fabry disease and P1k blood group individuals, or which lack Pk, from p blood group individuals. PBMCs representative of these two groups were infected with M-tropic HIV-1Ba-L or T-tropic HIV-1IIIB. PBMCs with Pk accumulation from patients with Fabry disease were resistant to productive M-tropic, but not T-tropic HIV-1 infection. These PBMCs had significantly decreased expression levels of the M-tropic HIV-1 co-receptor CCR5. Pk accumulation in P1k blood group PBMCs shows resistance to T-tropic and M-tropic HIV-1 infection, unrelated to levels of chemokine receptors. PBMCs lacking Pk from p blood group individuals were hypersensitive to M-tropic HIV-1 infection, while T-tropic HIV-1 infection levels were variable. Interestingly, p blood group PBMCs showed significantly higher levels of the CCR5 co-receptor. Our findings suggest Pk is inhibitory to HIV infection, which may translate into natural resistance in individuals that express high levels of Pk. This may be linked to interactions with CCR5, through receptor trafficking to the cell surface, or associations within lipid rafts. Furthermore, soluble Pk or potential mechanisms to increase cellular Pk expression, provide novel strategies for prevention of HIV infection.

Lack of susceptibility of cells from patients with Fabry disease to productive infection with R5 human immunodeficiency virus

A lack of viral replication after HIV-1Ba-L (R5) but not HIV-1IIIB (X4) infection was found using in-vitro activated peripheral blood-derived mononuclear cells from patients with Fabry disease, who have a defect in the catabolism of globotriaosylceramide. CCR5, but not CD4 or CXCR4 expression levels, were lower and the surface expression of globotriaosylceramide was negligible on activated patients' cells. Our findings suggest a novel resistance mechanism to productive infection with R5 HIV-1 that potentially involves abnormal globotriaosylceramide catabolism.

Kinin B1 Receptor Expression on Multiple Sclerosis Mononuclear Cells

We have previously shown that the inducible kinin B(1) receptor is expressed on T lymphocytes during relapses and progression in multiple sclerosis. To evaluate the correlation between the expression of B1 receptor on peripheral blood mononuclear cells derived from patients who have multiple sclerosis with serial, clinical magnetic resonance imaging and immunological study-derived measures. Using frozen samples obtained from a high-frequency magnetic resonance imaging-immunological study, we analyzed B1 receptor messenger RNA (mRNA) expression in peripheral blood-derived mononuclear cells serially collected from 6 patients with multiple sclerosis and 8 healthy control subjects by semiquantitative radioactive duplex reverse transcriptase-polymerase chain reaction amplification. Time-course kinin B1-actin mRNA ratios were subsequently compared with corresponding clinical magnetic resonance imaging and immune parameters. The time-course kinin B1-actin mRNA ratio correlated positively with the Expanded Disability Status Scale index (P<.001), occurrence of clinical relapse (P = .02), volume of lesion on T2-weighted images (P<.003) and interleukin 2 receptor and major histocompatibility complex class II expression on CD4+ lymphocytes, but not with gadolinium-enhancing lesions. The time-course kinin B1-actin mRNA ratios were 5 to 25 times lower in samples derived from healthy controls. The correlation of kinin B1 receptor mRNA levels with dynamic clinical and magnetic resonance imaging measures suggests that expression of this receptor can serve as an index of disease activity in multiple sclerosis.

末梢動脈閉塞症に対する骨髄単核球移植療法と血管内皮前駆細胞の定量的評価

We performed therapeutic angiogenesis by cell transplantation (TACT) for patients with peripheral obstructive arterial disease using autologous bone marrow mononuclear cells. The patients' symptom has begun to improve within several days after therapy, as did laboratory data (i. e., anklebrachial blood pressure index, thermography, percutaneous measurement of oxygen pressure). We quantified the mRNA expression of endothelial progenitor cell (EPC) -specific molecules (e. g., Flk-1, CD133, VE-cadherin, etc.) in bone marrow- or peripheral blood-derived mononuclear cells obtained from patients with ischemic limbs, using real time reverse transcription-PCR. The mRNA expression level of EPC markers was significantly lower in the patients than in healthy controls. We furthermore revealed the different expression pattern of EPC markers in several sources, including bone marrow and peripheral blood obtained from healthy volunteers or recombinant G-CSF-treated donors. The finding that the expression of EPC-specific molecules was decreased in the patients' marrow and blood suggests that patients with peripheral obstructive arterial diseases may have lower angiogenic potential. However, TACT with autologous marrow mononuclear cells was effective and increased the number of circulating EPCs in the patients' blood.

Molecular evaluation of endothelial progenitor cells in patients with ischemic limbs: therapeutic effect by stem cell transplantation.

Although some patients with limb ischemia have recently undergone therapeutic angiogenesis by cell transplantation, their angiogenic potential has not been well characterized. It is also important to evaluate endothelial progenitor cell (EPC) contents in different stem cell sources to choose the best material for therapeutic angiogenesis. We quantitated the mRNA expression of EPC-specific molecules (eg, Flk-1, Flt-1, CD133, VE-cadherin, etc) in bone marrow-derived or peripheral blood-derived mononuclear cells obtained from patients with ischemic limbs, using real-time reverse-transcription polymerase chain reaction technique. The mRNA expression level of EPC markers was significantly lower in the patients than in healthy controls, which was consistent with results of flow cytometric analysis. However, the implantation of autologous bone marrow mononuclear cells increased the circulating EPCs in the peripheral blood of patients. We furthermore revealed the different expression pattern of EPC markers in possible sources for stem cell transplantation, including normal bone marrow, peripheral blood obtained from recombinant granulocyte colony-stimulating factor-treated donor, and umbilical cord blood. Patients with peripheral obstructive arterial diseases may have lower angiogenic potential because of decreased expression of EPC specific molecules in their marrow and blood. Therapeutic angiogenesis by transplantation of autologous marrow mononuclear cells increased circulating EPCs in the patients and improved ischemic symptoms.

The spontaneous remission of juvenile idiopathic arthritis is characterized by CD30+ T cells directed to human heat-shock protein 60 capable of producing the regulatory cytokine interleukin-10.

To test the hypothesis that T cell reactivity to self heat-shock protein 60 (Hsp60) in patients with remitting juvenile idiopathic arthritis (JIA) is part of an antiinflammatory, regulatory mechanism. Using peripheral blood-derived mononuclear cells (PBMCs) and synovial fluid-derived mononuclear cells (SFMCs) obtained from patients with JIA, we analyzed the expression of CD30 and the induction of regulatory cytokines in response to human and mycobacterial Hsp60. In oligoarticular JIA patients, in vitro activation of PBMCs and SFMCs with Hsp60 induced a high expression of CD30 on CD4+, activated (HLA-DR-positive), memory (CD45RO+) T cells. The expression of CD30 induced by human Hsp60 was much higher than that induced by mycobacterial Hsp60. In oligoarticular JIA patients with active disease, the expression of CD30 in response to human Hsp60 was paralleled by a high interleukin-10 (IL-10):interferon-gamma (IFNgamma) ratio. In addition, restimulated human Hsp60-specific T cell lines from oligoarticular JIA patients showed a high production of IL-10 and a low production of IFNgamma. In contrast, PBMCs and SFMCs from polyarticular JIA patients responded to human Hsp60 with virtually no expression of CD30 and a low IL-10:IFNgamma ratio. The results show that T cells responding to human Hsp60 in oligoarticular JIA patients express CD30, and during active phases of the disease, these T cells have a cytokine profile with a high IL-10:IFNgamma ratio. These findings suggest that in oligoarticular JIA patients, human Hsp60-specific CD4+ cells have a regulatory function and contribute to disease remission.

Phagocytosis of apoptotic inflammatory cells by microglia and its therapeutic implications: termination of CNS autoimmune inflammation and modulation by interferon-beta.

Apoptosis of autoaggressive T-cells in the CNS is an effective, noninflammatory mechanism for the resolution of T-cell infiltrates, contributing to clinical recovery in T-cell-mediated neuroinflammatory diseases. The clearance of apoptotic leukocytes by tissue-specific phagocytes is critical in the resolution of the inflammatory infiltrate and leads to a profound downregulation of phagocyte immune functions. Adult human microglia from surgically removed normal brain tissue was used in a standardized, light-microscopic in vitro phagocytosis assay of apoptotic autologous peripheral blood-derived mononuclear cells (MNCs). Microglia from five different patients had a high capacity for the uptake of apoptotic MNCs in contrast to nonapoptotic target cells with the phagocytosis rate for nonapoptotic MNCs amounting to only 61.6% of the apoptotic MNCs. A newly described phosphatidylserine receptor, critical in the phagocytosis of apoptotic cells by macrophages, is also expressed at similar levels on human microglia. The effects of the therapeutically used immunomodulatory agent interferon-beta (IFNbeta) were investigated using Lewis rat microglia and apoptotic, encephalitogenic, myelin basic protein-specific autologous T-cells. Also, rat microglia had a high capacity to phagocytose apoptotic T-cells specifically. IFNbeta increased the phagocytosis of apoptotic T-cells to 36.8% above the untreated controls. The enhanced phagocytic activity was selective for apoptotic T-cells and was not mediated by increased IL-10 secretion. Apoptotic inflammatory cells may be efficiently and rapidly removed by microglial cells in the autoimmune-inflamed human CNS. The in vitro increase of phagocytosis by IFNbeta merits further investigations whether this mechanism could also be therapeutically exploited.

Involvement of cytokines in the pathogenesis of systemic lupus erythematosus.

Systemic lupus erythematosus (SLE) is characterized by overt polyclonal B-cell activation and autoantibody (Ab) production. By contrast, cellular immune responses against allo- or recall antigens are significantly impaired. Many evidences indicate that IL-10 overproduction plays a pivotal role in the disease and the contribution of the IL-10/IL-12 imbalance to the pathophysiology of SLE will be extensively discussed. The authors will further summarize the available data about the involvement of IFN-γ, TNF-α, TGF-β and TALL-1. Other cytokines (IL-1, IL-2, IL-4, IL-6, IL-16, IL-17 and IL-18) will be briefly discussed. Numerous abnormalities of the cytokine network have been described in patients suffering from systemic lupus erythematosus as well as in murine lupus models. Some of them were shown to play a pivotal physiopathological role in certain T-cell, B-cell or antigen-presenting cell (APC) dysfunctions characteristic of the disease, while others are more likely to be innocent bystanders. It should be stressed, moreover, that not all the data discussed herein fit into a single picture. The heterogeneity of human SLE, the influence of disease activity and therapy on cytokine production and function, the relative shortage of human lupus peripheral blood-derived mononuclear cells (PBMC), together with the lack of a perfectly-matched animal model, further sophisticate the issue. For these very reasons, this review will focus on the cytokines that the authors wrongly or rightly consider as the major players in the game, namely IL-10, IL-12, TNF-α, IFN-γ and the recently described TALL-1/BLys/BAFF. Before doing so, we felt worthwhile to briefly glance through the major immune abnormalities observed in SLE.

Hepatitis A virus suppresses monocyte-to-macrophage maturation in vitro.

To analyze the pathogenetic mechanism of hematopoietic dysregulation associated with hepatitis A virus (HAV) infections, we studied the influence of HAV on monocyte (MO)-to-macrophage (MAC) maturation in vitro. Exposure of peripheral blood-derived mononuclear cells (MNC) to HAV led to diminished adherence of MO to plastic. Furthermore, HAV inhibited the ability of peripheral blood MO to differentiate toward MAC. Freshly isolated and 14-day-old MO cultures demonstrated reduced differentiation and decreased phagocytic capacity after challenge with HAV. Viral replication in MO/MAC cultures was confirmed by titration of infectious virus. We also determined the influence of HAV on the MO/MAC population in human long-term bone marrow cultures (LTBMCs). Inoculation of bone marrow MNC with HAV suppressed the establishment of an adherent stromal layer containing a reduced number of MAC. Furthermore, increased MO numbers in the nonadherent fraction of HAV-challenged LTBMCs are indicative of the disturbance of MO adherence. These findings suggest that HAV infection leads to a disorder of the mononuclear phagocytic system which may contribute to functional abnormalities of the bone marrow stroma.

Comparative Antiproliferative and Apoptotic Effects of Resveratrol, ϵ-viniferin and Vine-shots Derived Polyphenols (Vineatrols) on Chronic B Lymphocytic Leukemia Cells and Normal Human Lymphocytes

Trans-resveratrol, its dimer k -viniferin and two preparations of vineatrol (a grape-derived polyphenol fraction isolated from vine-shots extracts) were compared for their effects on the proliferation and survival of normal and leukemic human lymphocytes. The two different batches of vineatrol (vineatrol 10 and 25%) was obtained by HPLC fractionation and contained 10 and 25% trans-resveratrol, respectively. The different polyphenols were added to cultures of leukemic cells from chronic B cell malignancies (B-cell chronic lymphocytic leukemia, B-CLL or hairy cell leukemia, HCL) or normal peripheral blood-derived mononuclear cells (PBMC) as a control. The different polyphenols displayed anti-proliferative effect on the leukemic cells, as estimated by the observed inhibition of tritiated thymidine uptake and the reduction of cell recovery. Vineatrol 10% was the most potent whereas vineatrol 25% and resveratrol displayed comparable activity, k -viniferin only exhibiting slight effets. The same order of potency was observed for their capacity to induce apoptosis in leukemic B cells. In contrast, the survival of normal peripheral blood mononuclear cells (PBMC) was little affected in the presence of these polyphenolic compounds and higher concentrations were required in order to elicit cell death. Polyphenol-driven apoptosis in chronic leukemic B cells was shown to correlate with an activation of caspase 3, a drop in the mitochondrial transmembrane potential, a reduction in the expression of the anti-apoptotic protein bcl-2, as well as a reduction in the expression of the inducible nitric oxide synthase (iNOS). Our data therefore indicate that vine-shoots may be a convenient and natural source of material for the purification of resveratrol and other polyphenolic compounds of putative therapeutic interest.