QuantiFERON®-CMV for Immune-Monitoring in Patients Undergoing Allogeneic Hematopoietic Stem Cell Transplantation

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 4369-4369
Author(s):  
Enrico Morello ◽  
Elisabetta Pagani ◽  
Vladia Monsurrò ◽  
Irene Cavattoni ◽  
Silvia Coin ◽  
...  
Keyword(s):  
At Risk ◽  
Whole Blood ◽  
Cmv Infection ◽  
Mortality And Morbidity ◽  
Viral Therapy ◽  
Ifn Γ ◽  
Cmv Disease ◽  
Anti Viral Therapy ◽  
Related Mortality

Abstract Background: CMV infection represents a major complication of allo-SCT affecting transplant related mortality and morbidity. Anti-viral therapy is toxic and prolonged treatment could affect graft function. Monitoring specific immune response against CMV could optimize the timing for anti-viral therapy administration in order to avoid related toxicity and to reduce CMV related mortality and morbidity. A recent ELISA based test (QuantiFERON®-CMV) could measure specific (anti-HCMV) and aspecific production of IFN-γ in whole blood. Aims: to test the reliability of QuantiFERON®-CMV in a cross sectional study in order to identify patients at risk of CMV disease after alloHSCT. Methods: QuantiFERON®-CMV is an in vitro diagnostic test using a peptide cocktail simulating human cytomegalovirus proteins (CMV) to stimulate cells in heparinised whole blood. Detection of interferon-γ (IFN-γ) by ELISA is used to identify in vitro responses to these peptide antigens that are associated with CMV infection. The IFN-γ response in the CMV Ag tube is considered positive if > 0.2 UI/mL as defined by the manufacturer. The mitogen-stimulated plasma sample was used as an IFN-γ positive control (PC) for each specimen tested. CMV reactivation and disease were defined according EBMT recommendations. Results: Among 92 tests no correlation between pp65 antigenemia and IFN-γ production was proved (p=0,346). However, among the 41 tests showing lower levels of anti-HCMV IFN-γ production (<0.2 IU/mL) 8 tests belonging 4 patients were associated with CMV disease, whereas among the 51 tests showing higher levels of anti-HCMV IFN-γ production (>=0.2 IU/mL) none were associated with CMV disease (p=0.001), RR 2.5 (CI95% 1.951–3.321). Conclusions: QuantiFERON®-CMV doesn’t seem to represent a significant reliable test for risk of viremia after alloHSCT, but patients with prolonged lower levels of anti-HCMV IFN-γ production (<0.2 IU/mL) are at risk of CMV disease. Prospective studies are required in order to identify the correlation between viremia and the need for treatment.

scholarly journals Comparison of Three Cellular Assays to Predict the Course of CMV Infection in Liver Transplant Recipients

Vaccines ◽  
2021 ◽  
Vol 9 (2) ◽  
pp. 88
Author(s):  
Smaranda Gliga ◽  
Melanie Fiedler ◽  
Theresa Dornieden ◽  
Anne Achterfeld ◽  
Andreas Paul ◽  
...  
Keyword(s):  
Liver Transplant ◽  
Solid Organ Transplantation ◽  
Solid Organ ◽  
Transplant Recipients ◽  
Cmv Infection ◽  
Cellular Assays ◽  
Risk Patients ◽  
Ifn Γ ◽  
Liver Transplant Recipients

To estimate protection from cytomegalovirus (CMV) replication after solid organ transplantation, CMV serology has been considered insufficient and thus CMV immunity is increasingly assessed by cellular in vitro methods. We compared two commercially available IFN-γ ELISpot assays (T-Track CMV and T-SPOT.CMV) and an IFN-γ ELISA (QuantiFERON-CMV). Currently, there is no study comparing these three assays. The assays were performed in 56 liver transplant recipients at the end of antiviral prophylaxis and one month thereafter. In CMV high- or intermediate-risk patients the two ELISpot assays showed significant correlation (p < 0.0001, r > 0.6) but the correlation of the ELISpot assays with QuantiFERON-CMV was weaker. Results of both ELISpot assays were similarly predictive of protection from CMV-DNAemia ≥500 copies/mL [CMV pp65 T-SPOT.CMV at the end of prophylaxis: area under curve (AUC) = 0.744, cut-off 142 spot forming units (SFU), sensitivity set to 100%, specificity 46%; CMV IE-1 T-Track CMV at month 1: AUC = 0.762, cut-off 3.5 SFU, sensitivity set to 100%, specificity 59%]. The QuantiFERON-CMV assay was inferior, reaching a specificity of 23% when setting the sensitivity to 100%. In conclusion, both CMV-specific ELISpot assays appear suitable to assess protection from CMV infection/reactivation in liver transplant recipients.

Utilisation of hepatocellular carcinoma screening in Australians at risk of hepatitis B virus‐related carcinoma and prescribed anti‐viral therapy

2018 ◽  
Vol 27 (13-14) ◽  
pp. 2673-2683
Author(s):  
Suzanne Sheppard‐Law ◽  
Iryna Zablotska‐Manos ◽  
Melissa Kermeen ◽  
Susan Holdaway ◽  
Alice Lee ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
At Risk ◽  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Viral Therapy ◽  
B Virus ◽  
Hepatocellular Carcinoma Screening ◽  
Anti Viral Therapy

Cytomegalovirus (CMV)-Specific CD8+ T Cells Are Associated with Control of Viral Reactivation Post-Hemopoietic Stem Cell Transplantation (HSCT) Only If Shown to Be Functionally Active.

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 2213-2213
Author(s):  
Stephanie Verfuerth ◽  
Karl S Peggs ◽  
Arnold Pizzey ◽  
Noha Chowdhry ◽  
Stephen Mackinnon
Keyword(s):  
T Cells ◽  
Cd8 T Cells ◽  
The Other ◽  
Viral Reactivation ◽  
Cd8 Cells ◽  
Ifn Γ ◽  
Antiviral Chemotherapy ◽  
Cmv Disease ◽  
Cmv Reactivation

Abstract Targeted antiviral chemotherapy has greatly reduced the incidence of CMV disease post-HSCT, but adverse side effects still make CMV seropositivity a negative prognostic factor for survival. Immune protection from CMV disease depends on the adaptive immune response, and various strategies for adoptive cellular therapy for CMV have been evaluated in clinical trials to reduce the dependency on antiviral drugs. Characterizing the development of post-transplantation CMV-specific T cell responses may help identify minimum requirements for successful immunologic control of CMV reactivation. We investigated CMV-specific T cell immune reconstitution in 15 patients who all received ACT for CMV 4 weeks post-HSCT. The CMV seropositive donor-derived ACT product consisted of 1x105 CD4+ and CD8+ T cells/kg, generated through 2-week in vitro co-culture with CMV antigen-pulsed autologous dendritic cells. No graft versus host disease &gt;grade II occurred in the study group. CMV-specific T cells were characterized at various time points before, during, and after CMV reactivation events by IFN-γ secretion assay, and/or by phenotyping with HLA CMV pp65 and/or IE-1 peptide pentamers. Three patients who remained CMV PCR negative throughout had no detectable levels of CMV-specific T cells, whereas high levels of CMV-specific T cells developed in the other 12 patients who all experienced episodes of viral reactivation. 11/12 of the patients with CMV reactivation had pre-emptive antiviral chemotherapy, 1/12 controlled CMV reactivation unaided. 3/12 patients experienced a second episode of viral reactivation, which they controlled unaided. At the time of first CMV DNA detection, 1 week before the start of pre-emptive chemotherapy, pentamer+ cells were already detectable in 3/5 patients tested, with absolute numbers ranging from 1150-4900/ml blood. Only after a further 1 to 4 more weeks did T cells secreting IFN-γ in response to in vitro restimulation with CMV peptides first appear in 5 patients tested. By the time viral DNA had become undetectable post-antiviral chemotherapy or post-immunological control of viral replication, pentamer+ cells were present in 8/8 patients, with absolute numbers of pentamer+ T cells &gt;10000/ml in 7/8 patients, a level often quoted as ‘protective’ (median: 37530 cells/ml, range: 3200–139300/ml). CMV-specific CD8 T cells were maintained at high levels long after resolution of CMV viraemia (median: 24200/ml, range: 5100–75900). Absolute numbers of IFN-γ + T cells also reached high levels shortly before to just after the end of a CMV reactivation episode in 5/5 patients tested (CD8+IFN-γ + median: 48490/ml, range: 2150–138290), but subsequently temporarily dropped again to undetectable levels in some patients. Four patients had CMV reactivation episodes that were immunologically controlled unaided. Just prior these four reactivation events there was a total lack of CD8+ T cells functionally responsive to CMV. This was in spite of the presence CMV-specific CD8+ cells by phenotype in 3/3 patients tested, at &gt;10000/ml in one patient. 0% of these pentamer+ cells secreted IFN-γ in response to CMV peptide in vitro. Soon after, by the time of the first CMV PCR+ result CD8+ IFN-γ + T cells were already detected in all patients tested (median: 2180 cells/ml, range: 550–48480, n=3), with up to 64% of pent+ cells secreting IFN-γ. Pent+ cells were above the 10000 threshold in 2/3 patients tested at this time point. These data show that characterizing T cells by phenotype alone does not permit conclusions about their functionality, because some patients experienced CMV reactivation events in the presence of high levels of pentamer+ cells. Viral reactivation episodes however appeared to be associated with a prior lack of CMV-specific CD8+ cells capable of IFN-γ secretion. Immunological control of reactivation, on the other hand, may be associated with the ability to make rapid CMV-induced IFN-γ responses, because only patients with self-resolving CMV infection developed CD8+IFN-γ + responses immediately at the onset of viraemia. However it is not possible to know how many of the patients who received antiviral chemotherapy would have been able to control the infection unaided.

scholarly journals Incubation of Whole Blood at 39°C Augments Gamma Interferon (IFN-γ)-Induced Protein 10 and IFN-γ Responses to Mycobacterium tuberculosis Antigens

2011 ◽  
Vol 18 (7) ◽  
pp. 1150-1156 ◽  
Author(s):  
Martine G. Aabye ◽  
Pernille Ravn ◽  
Isik S. Johansen ◽  
Jesper Eugen-Olsen ◽  
Morten Ruhwald
Keyword(s):  
Mycobacterium Tuberculosis ◽  
T Cell ◽  
Whole Blood ◽  
Incubation Temperature ◽  
Gamma Interferon ◽  
Content Type ◽  
Interleukin 7 ◽  
Ifn Γ

ABSTRACTA rarely challenged dogma in cell-mediated immune (CMI) assays is the incubation temperature, 37°C. Fever augments proinflammatory immune responsesin vivo, and the aim of this study was to explore whether incubation at fever-range temperature could increase antigen-specific biomarker responses. We compared CMI responses following incubation of whole blood at 37°C and 39°C. Whole blood was obtained from (i) 34 healthy subjects whose blood was incubated with TB10.4 antigen, present in theMycobacterium bovisbacillus Calmette-Guérin vaccine and many environmental mycobacteria; (ii) 8 TB patients and 8 controls incubated withMycobacterium tuberculosis-specific antigens in the QuantiFERON-TB Gold test (QFT-IT); and (iii) from both groups incubated with a T cell mitogen. T cell responses (gamma interferon [IFN-γ]) and responses from antigen-presenting cells (IFN-γ-induced protein 10 [IP-10]) were determined. We further evaluated the effect of adding interleukin-7 (IL-7) and blocking IL-10 during incubation. In TB patients, IFN-γ and IP-10 levels were increased 4.1- and 3.4-fold, respectively, at 39°C incubation (P< 0.001). Similar results were seen after mitogen stimulation. In subjects responding to TB10.4, the effects were less pronounced and significant only for IP-10. Incubation at 39°C increased IP-10 and IFN-γ responsiveness to both antigens and mitogen in persons with baseline or initial low responses. Adding IL-7 and blocking IL-10 augmented the effects in synergy with fever-range temperature. Incubation at fever-range temperature vividly increases CMI responsiveness to antigen stimulationin vitroin tuberculosis patients and may increase the sensitivity of CMI assays.

scholarly journals Cytomegalovirus in pregnancy and the neonate

F1000Research ◽  
2017 ◽  
Vol 6 ◽  
pp. 138 ◽  
Author(s):  
Vincent C. Emery ◽  
Tiziana Lazzarotto
Keyword(s):  
Vaccine Development ◽  
Cmv Infection ◽  
Chain Reaction ◽  
Congenital Cytomegalovirus ◽  
Symptomatic Disease ◽  
Diagnosis And Prognosis ◽  
Viral Therapy ◽  
Avidity Test ◽  
Polymerase Chain ◽  
Anti Viral Therapy

Congenital cytomegalovirus (CMV) remains a leading cause of disability in children. Understanding the pathogenesis of infection from the mother via the placenta to the neonate is crucial if we are to produce new interventions and provide supportive mechanisms to improve the outcome of congenitally infected children. In recent years, some major goals have been achieved, including the diagnosis of primary maternal CMV infection in pregnant women by using the anti-CMV IgG avidity test and the diagnosis and prognosis of foetal CMV infection by using polymerase chain reaction real-time tests to detect and quantify the virus in amniotic fluid. This review summarises recent advances in our understanding and highlights where challenges remain, especially in vaccine development and anti-viral therapy of the pregnant woman and the neonate. Currently, no therapeutic options during pregnancy are available except those undergoing clinical trials, whereas valganciclovir treatment is recommended for congenitally infected neonates with moderately to severely symptomatic disease.

scholarly journals Successful Treatment of Refractory CMV Chorioretinitis and Meningoencephalitis with Adoptive Transfer of Third Party CMVpp65 Specific T-Cell Lines

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 3157-3157
Author(s):  
Susan Prockop ◽  
Aisha Hasan ◽  
Ekaterina Doubrovina ◽  
H.R. Castro-Malaspina ◽  
Juliet N. Barker ◽  
...  
Keyword(s):  
T Cells ◽  
Stem Cell ◽  
Research Funding ◽  
Response Rate ◽  
Third Party ◽  
Hematopoietic Stem ◽  
Viral Therapy ◽  
Primary Donor ◽  
Cmv Disease ◽  
Anti Viral Therapy

Abstract Adoptive immunotherapy with transplant donor-derived virus specific T cells is an effective strategy for the treatment of CMV viremia and disease arising after an allogeneic hematopoietic stem cell (HSCT). At our center this approach has become a standard part of the armamentarium of CMV directed therapy. However, donor-derived CMVpp65-specific cytotoxic T cells (CMV-CTLs) are not available for patients transplanted from a seronegative or cord blood donor. In addition, in the HLA disparate transplant setting the CMV-CTL line derived from non-identical donors may be restricted by non-shared HLA alleles. For these patients as we have previously reported, treatment with in vitro expanded CMV-CTLs derived from an HLA partially matched third party donors is an option. One particularly difficult clinical situation is the treatment of patients who develop CMV disease involving the sanctuary sites of the central nervous system and retina. We have treated 12 patients with primary donor derived (n=1) or third party (n=11) CMV CTLs for retinitis (7) or meningitis/encephalitis (4) or both (1). Recipients of hematopoietic stem cell transplant (HSCT) had undergone T cell depleted (5) conventional (1) or cord blood (4) transplantation. One patient was treated after solid organ transplantation and one for HIV related CMV retinitis. Third party CMV CTLS were selected from a bank of 132 lines generated under GMP conditions from normal HSCT donors specifically consented for use of their T cells in patients other than their designated transplant recipient. Third party CMV-CTLs were selected on the basis of HLA matching at a minimum of 2/8 recipient alleles and HLA restriction of the T cells by one or more HLA alleles present in the patient. A total of 10 distinct third party CMV-CTL lines and one (1) donor derived line were used. Patients received infusions of CMV-CTLs after failing a median of 146 (43-419) days of prior therapy with a median of 4 anti-viral agents. Patients received 3 weekly infusions of CMV-CTLs at doses of 1 x 10e6 T cells/kg (n=10) 2 x 10e6 T cells/kg (n=1) and 0.5 x 10e6 T-cells/kg (n=1) and were eligible to receive additional cycles of cells if they had no toxicity five weeks after the start of cellular therapy. One patient was evaluated only for toxicity as efficacy was confounded by anti-viral therapy required for treatment of varicella zoster. Of the 11 evaluable patients, 7 achieved CR, 1 PR (clinical improvement in disease and a 3 log decrease in viral load) and 1 SD. All but one of these responses were durable. One patient who had achieved a CR had recurrence of low grade viremia which was not retreated due to his overall medical deterioration. The two patients with progression of disease ultimately died of CMV. Patients were monitored for expansion of CMV-CTLs by CMV-specific interferon gamma and where appropriate tetramer analysis. Responding patients consistently had detectable expansion of CMV-specific CTL populations. In addition, in one patient, undergoing serial lumbar punctures, we were able to detect third party CMV-CTLs in the CSF. There were limited toxicities associated with CMV-CTL infusions. No patient developed de novo GvHD or a flare of prior GvHD. This study demonstrates a high response rate among patients with otherwise refractory CMV chorioretinitis or meningoencephalitis following adoptive therapy with primary donor or third party CMV-CTLs; thus demonstrating the capacity of adoptively transferred CMV-CTLs to treat disease in these sanctuary sites without toxicity. When selected based on restriction through shared alleles, third party CMV-CTLs are effective despite significant HLA disparity. The bank of CMV specific T cells can provide an immediate source of HLA partially matched appropriately restricted T cells for adoptive immunotherapy to treat CMV disease affecting the CNS. This enables treatment early in the course of disease with CMV-CTL lines previously prepared and characterized in terms of HLA restriction. This approach is anticipated to maximize the response rate as well as minimize toxicity from anti-viral therapy. Disclosures Prockop: Atara Biotherapeutics: Other: I have no financial disclosures, but Atara Biotherapeutics has exercised a licensing agreement with Memorial Sloan Kettering Cancer Center and MSKCC and some investigators at MSKCC have a financial interest in Atara.. Hasan:Atara Biotherapeutics: Research Funding. O'Reilly:Atara Biotherapeutics: Research Funding.

Mitogen-Induced Interferon Gamma Production in Human Whole Blood: The Effect of Heat and Cations

2019 ◽  
Vol 20 (7) ◽  
pp. 562-572 ◽  
Author(s):  
Ji-Hyun Nam ◽  
Bomi Cha ◽  
Jun-Young Park ◽  
Fukushi Abekura ◽  
Cheorl-Ho Kim ◽  
...  
Keyword(s):  
Interferon Gamma ◽  
Whole Blood ◽  
Calcium Supplementation ◽  
Positive Control ◽  
Pokeweed Mitogen ◽  
Con A ◽  
Human Whole Blood ◽  
Heat Treated ◽  
Ifn Γ

Background: Interferon-gamma release assays (IGRAs) are blood tests used to measure the amount of interferon-γ (IFN-γ) released by T lymphocytes after stimulation by antigens specific for the diagnosis of latent tuberculosis infection. A mitogen serves as a positive control to assess the immune function in IGRAs. Methods: This in vitro study was conducted to evaluate IFN-γ production by human whole blood stimulated with heat-treated and/or cation-supplemented phytohemagglutinin (PHA), concanavalin A (Con A) and pokeweed mitogen (PWM), using QuantiFERON-TB Gold Kit ELISA tests. Results: The optimal concentrations of PWM, Con A and PHA for IGRAs were 2 µg/mL, 5 µg/mL and 10 µg/mL, respectively. The results showed that IFN-γ production in response to PWM was the highest and PHA was the lowest amount. The median values of three mitogens were in the following order: PWM≥Con A≥ positive control>PHA-P>negative control. PWM and PHA were heat stable, while Con A was heat sensitive. The mitogen response of lymphocytes to untreated or heat-treated PWM and heat-treated Con A was increased in 1 mM Ca2+-supplemented groups, whereas the response to heat-treated PHA was decreased. Exposure to 1 mM Mg2+ had no effect on untreated or heat-treated PWM, and a concentration of 1 mM Zn2+ inhibited the stimulation of un-treated PWM. We found that calcium supplementation improved the PWM-induced production of IFN-γ. Conclusion: Therefore, PWM is an appropriate mitogen for use as a positive control in IGRAs. It is a potential indicator of cytokine production in the diagnostic as well as research settings, and calcium supplementation improved stimulation.

Comprehensive Analysis of CD8 T Cell Immune Response Specific for Two Novel HLA-A*0201 Restriced CMV pp65 Peptides.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 3928-3928
Author(s):  
Laura Bracci ◽  
David F. Stroncek ◽  
Stefanie Slezak ◽  
Giulio C. Spagnoli ◽  
Maurizio Provenzano
Keyword(s):  
Gene Expression ◽  
Hla Class I ◽  
Cd8 T Cell ◽  
Comprehensive Analysis ◽  
Protein Release ◽  
Cmv Infection ◽  
Qrt Pcr ◽  
Ifn Γ ◽  
In Vitro Induction

Abstract In immune compromised subjects such as patients undergoing bone marrow, organ transplantation or immunosuppressive therapies, Cytomegalovirus (CMV) infection is associated with significant morbidity until the individual’s immune system is completely reconstituted. One method of preventing CMV infection during immune suppression in transplant patients is represented by adoptive administration of CMV peptide-specific cytotoxic T lymphocytes (CTLs) from HLA-matched donors. Despite the strong immunogenicity demonstrated by specific HLA class I peptides, the peptide’s responsiveness varies among individuals. Therefore, it is important to identify additional epitopes for each HLA determinant of interest. In this study we report a comprehensive analysis of CD8 T cell-specific activity against two novel CMV pp65 HLA-A*0201 associated peptides in CMV-experienced HLA-A*0201 subjects. PBMCs from CMV seropositive HLA-A*0201 restricted healthy subjects were peptide-stimulated ex vivo and IFN-γ gene expression was analyzed by qrt-PCR. An IFN-γ ELISPOT assay was carried out on peptide-specific elicited CD8 T lymphocytes to confirm the qrt-PCR results. Tetrameric HLA/epitope complexes (tHLA) were used to track the levels of peptide-specific CD8 T cells responsiveness to cognate epitopes. In addition, in vitro peptide-specific expanded populations of CTLs were used in 51Cr release assay. Based on qrt-PCR results, four out of eight HLA-A*0201 peptides identified by computer algorithms were selected. Two are preaviously published peptides: pp65495–503 (NLVPMVATV) and pp65347–355 (ALFFFDIDL), while two are novel: pp65340–348 (RQYDPVAAL) and pp65310–318 (LMNGQQIFL). In spite the four peptides induced comparable mRNA IFN-γ transcript production, peptides pp65495–503, pp65340–348 and pp65310–318 induced a consistent and sustained IFN-γ protein release and specific killing, while the pp65347–355 failed to induce IFN-γ protein secretion and killing activity (if we exclude a positive IFN-γ protein release after 2-week in vitro induction in one of the donors tested). Comparative assays carried on the functional activity of the three peptides pp65495–503, pp65340–348 and pp65310–318 revealed no intrinsic differences in term of IFN-γ protein release and cytotoxic activity save for the CTL affinity to the HLA-A*0201/epitope complexes (pp65495–503 ~2.6% in nearly 100% of donors vs pp65340–348 ~0.67% or pp65310–318 ~0.77% in nearly one third of the donors after 2-week in vitro induction). The tHLA binding results could be possibly ascribed to differencies in the peptide avidity and stability for the HLA class I. Taken together, these results lead to the conclusion that different peptides can induce a variable levels of immune responses ranging between mere cytokine gene expression and effective cytotoxic activity. In addition, the two novel peptides selected here broaden the panel of potential reagents useful for adoptive immune therapy. Thus, in anticipation of a specific epitope-targeted immune intervention, the three HLA-A*0201 peptides described could be used in combination for adoptive transfer of epitope-specific T cells or epitope-specific vaccination.

Human monocytes express functional receptors for granulocyte colony– stimulating factor that mediate suppression of monokines and interferon-γ

Blood ◽  
2000 ◽  
Vol 95 (1) ◽  
pp. 270-276 ◽  
Author(s):  
Eva-Maria Boneberg ◽  
Lars Hareng ◽  
Florian Gantner ◽  
Albrecht Wendel ◽  
Thomas Hartung
Keyword(s):  
Flow Cytometry ◽  
Whole Blood ◽  
Receptor Expression ◽  
Human Neutrophils ◽  
Enzyme Linked Immunosorbent Assay ◽  
Granulocyte Colony Stimulating Factor ◽  
Colony Stimulating Factor ◽  
Ifn Γ ◽  
Stimulating Factor

In a double-blind, placebo-controlled, randomized study, 10 healthy men received either a single dose of 480 μg granulocyte colony-stimulating factor (G-CSF) or saline. Blood taken from the volunteers was stimulated with 10 μg/mL endotoxin and released cytokines were measured by enzyme-linked immunosorbent assay. Expression of G-CSF receptors on leukocytes was examined by flow cytometry and reverse transcriptase-polymerase chain reaction. Functional activity of these receptors was tested by challenging isolated leukocyte populations to release cytokines with endotoxin in the presence of G-CSF. The G-CSF treatment attenuated the release of the proinflammatory cytokines tumor necrosis factor (TNF)-, interleukin (IL)-12, IL-1β, and interferon (IFN)-γ in ex vivo lipopolysaccharide (LPS)-stimulated whole blood. In blood from untreated volunteers the presence of G-CSF in vitro also attenuated the LPS-stimulated release of these cytokines. G-CSF in vitro also attenuated TNF- release from elutriation-purified monocytes. In the presence of 10 ng/mL recombinant TNF-, the attenuation of LPS-inducible IFN-γ release by G-CSF was blunted in whole blood. However, G-CSF had no such effect on IFN-γ release from isolated lymphocytes stimulated with anti-CD3 or a combination of TNF- and IL-12. G-CSF receptor expression was detected in human neutrophils and monocytes but not in lymphocytes by means of RT-PCR as well as flow cytometry. These results indicate that G-CSF receptors expressed on monocytes are functional in modulating monokine release. We conclude that the attenuation of IFN-γ release from lymphocytes is not a direct effect of G-CSF on these cells but is rather due to the inhibition of monocytic IL-12 and TNF- release by G-CSF. (Blood. 2000;95:270-276)

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