High Level Secretion of HLA-G5 Might Inhibit the Incidence of Gvhd,

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 4068-4068
Author(s):  
Xiuli Wu ◽  
Yinkui Chen ◽  
Qifa Liu ◽  
Li Xuan ◽  
Yiwen Ling ◽  
...  

Abstract Abstract 4068 Human leucocyte antigen G (HLA-G) is a nonclassical HLA class I molecule, the tolerogenic role of HLA-G is highly supported in pregnancy immunization, tumor immune escape and organ transplant. HLA-G molecules also act as an inhibitor of the T-lymphocytes, NK cells and antigen presenting cells. HLA-G can be expressed as 7 different isoforms including 4 membrane-bound (HLA-G1 to -G4) and 3 soluble (HLA-G5 to-G7) proteins. The soluble HLA-G5 isoform encoded by intron-4 retaining spliced transcript has been previously detected in vivo in sera and grafts from solid transplanted patients who had significantly better graft acceptance. In the present study, we analyzed the expression levels of HLA-G in patients after hematopoietic stem cell transplantation (HSCT) and investigated the relationship between HLA-G and GVHD. The levels of soluble HLA-G (sHLA-G) in the plasma of peripheral blood from 90 patients underwent HSCT were determined by enzyme-linked immunosorbent assay (ELISA). Flow cytometry was used to detect the expression of membrane bound HLA-G (mHLA-G). The levels of mHLA-G of patients at pre-HSCT, +15d post-HSCT and +30d post-HSCT were similar with the normal individuals. High levels of sHLA-G in the plasma of recipients were found after allogeneic HSCT (allo-HSCT). The soluble HLA-G5 and HLA-G6 levels in the plasma of recipients at +15d post-HSCT and +30d post-HSCT were significantly higher than that at pre-HSCT, but the soluble HLA-G7 levels in the plasma of recipients at +15d post-HSCT or +30d post-HSCT were similar with that at pre-HSCT. In contrast, the levels of sHLA-G in the plasma of recipients had no significant change after autologous HSCT (auto-HSCT). High level secretion of HLA-G5 after allo-HSCT (the levels of sHLA-G increased more than 1.0 fold compared to the pre-HSCT group) significantly reduced the incidence of acute GVHD and chronic GVHD; High level secretion of HLA-G6 after allo-HSCT also significantly reduced the incidence of chronic GVHD, but had no influence to the incidence of acute GVHD; High level secretion of HLA-G7 after allo-HSCT had no influence to the incidence of acute GVHD or chronic GVHD. The increased levels of HLA-G5 after allo-HSCT which compared with the pre-HSCT group had a significant negative correlation with the severity of GVHD. In conclusion, the high level secretion of HLA-G5 might inhibit the incidence of acute and chronic GVHD. Detected the secretion level of HLA-G5 after HSCT might benefit to predict and evaluate the GVHD. Disclosures: Liu: National Natural Science Foundation of China (No.30971300), Science and Technology Planning Project of Guangdong Province of China (No.2009A030200007): Research Funding.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 5080-5080 ◽  
Author(s):  
Johannes Schetelig ◽  
Martin Bornhaeuser ◽  
Christian Thiede ◽  
Brigitte Mohr ◽  
Uta Oelschlaegel ◽  
...  

Abstract Recently we demonstrated that RIC with busulfan, fludarabine and ATG followed by allogeneic hematopoietic stem cell transplantation (HSCT) induced molecular remissions in patients (pts) with advanced CLL. However, this approach was hampered by severe GVHD. In an attempt to lower the rate of severe GVHD we replaced ATG by campath in a new study protocol. Patients and Methods: 20 pts with a median age of 54 years (range, 43 to 64) and advanced CLL were included. A median of 3 prior chemotherapy regimens had been given before HSCT, including fludarabine-containing regimens in all but two pts with autoimmune hemolysis. High risk cytogenetic features (17p−, 11q−, +12) were present in 9 pts. After conditioning with busulfan (8 mg/kg), fludarabine (150 mg/m2) and campath (75 mg) on days −9 to −5 peripheral blood stem cells from matched related (n=4) or unrelated donors (n=16) were transplanted. GVHD prophylaxis consisted of CSA monotherapy. Campath levels were analysed in frozen serum samples by BioAnaLab, Oxford, UK. Results: Two pts had no detectable campath level at the day of HSCT, while four pts had levels between 0.5 to 1.8 microgram/mL. Regeneration of neutrophils (>0.5/nl) and platelets (>20/nl) required a median of 17 (range, 14–25) and 10 (range, 0–27) days, respectively. Incomplete T-cell chimerism (<50%) was observed in 7 pts and subsequently 3 pts experienced secondary graft failure on days 134, 152 and 324. Six pts received donor lymphocyte infusions (DLI) for the conversion of incomplete T-cell chimerism (N=4) or progressive disease (N=2). Sponaneous acute GVHD II° to IV° occurred in 9/20 pts. After DLI four additional pts developed acute GVHD II° to IV°. Limited chronic GVHD occurred in 9 and extensive disease in 2 pts. In CMV seropositive pts the day 100 probability of CMV infection was 74% (95% CI, 44% to 100%). Severe encephalitis (HHV6, EBV and JC virus as suspected agents) was observed in 5 pts. Two pts recovered without sequelae, 2 pts are cognitively handicaped and one pt died. Hemorrhagic cystitis (CTC 2/3) occurred in 2 pts. After a median follow-up of 13 months (range, 6 – 26 months), 15 pts are alive. Four pts died from treatment related complications. Causes of death were pneumonia of unknown etiology (N=2), encephalitis (N=1) and GVHD grade IV (N=1). One pt died from severe acute GVHD subsequent to the treatment of relapse with DLI. One-year overall and progression-free survival was 75% (95% CI, 55% to 95%) and 50% (95% CI, 25% to 75%), respectively. The one-year probability of non-relapse mortality was 20% (95% CI, 2% to 38%). The number of binding sites for campath is highly variable in pts with progressive CLL resulting in interindividually highly variable pharmacokinetics. Differences in the extent of in vivo T-cell depletion might therefore explain the individually varying T-cell engraftment pattern. In addition, the high incidence of severe viral infections reflects impaired immunoreconstitution. Including pts after DLI we observed a substantial rate of severe GVHD. Based on these data we decided to skip the strategy of in vivo T-cell depletion with campath in patients with CLL.


Blood ◽  
1986 ◽  
Vol 67 (5) ◽  
pp. 1302-1308
Author(s):  
T de Witte ◽  
J Hoogenhout ◽  
B de Pauw ◽  
R Holdrinet ◽  
J Janssen ◽  
...  

Bone marrow from 22 histocompatible siblings was depleted of 98% of the lymphocytes using a combination of density flotation centrifugation followed by counterflow elutriation. Even with the marrow suppressive influence of methotrexate (MTX), the viability of the hematopoietic stem cells was not affected, as indicated by the normal repopulation after grafting in the evaluable patients. One patient (UPN 9) showed a primary graft failure, possibly resulting from persisting septicemia and long-term antibiotic therapy. Two patients have persistent host lymphocytes, one of whom was examined during relapse; the other remains in remission. Two patients did not receive immunosuppression after bone marrow transplantation (BMT), and acute graft-v-host disease (GVHD) developed in both. Nine patients received MTX as immunosuppression following BMT. GVHD did not develop in any of them, but fatal infections in the immediate posttransplant period developed in five patients. Eleven patients received cyclosporine (CsA) after transplantation. Beginning in week 5 after BMT, CsA was gradually replaced by MTX. Acute GVHD, substantial chronic GVHD, or fatal infections did not develop in any of these patients. Removal of 98% of the lymphocytes by counterflow centrifugation prevents development of acute GVHD, provided that immunosuppression is administered after BMT. Graft rejection was not observed, but the number of evaluable patients is limited at present.


JBMTCT ◽  
2020 ◽  
Vol 1 (1) ◽  
pp. 53-66
Author(s):  
Vaneuza A. M. Funke ◽  
Maria Claudia Rodrigues Moreira ◽  
Afonso Celso Vigorito

Graft versus host disease is one of the main complications of Hematopoietic stem cell, in­volving about 50% to 80% of the patients. Acute GVHD clinical manifestations and therapy is discussed, as well as new NIH criteria for the diagnosis and classification of chronic GVHD. Therapy for both refractory chronic and acute GVHD is an important field of discussion once there is no superiority for the majority of the agents after primary therapy has failed. Hence, this review is meant to be a useful tool of consultation for clinicians who are dealing with this complex complication.


2000 ◽  
Vol 44 (6) ◽  
pp. 1418-1427 ◽  
Author(s):  
L. E. Alksne ◽  
P. Burgio ◽  
W. Hu ◽  
B. Feld ◽  
M. P. Singh ◽  
...  

ABSTRACT Protein secretion is an essential process for bacterial growth, yet there are few if any antimicrobial agents which inhibit secretion. An in vivo, high-throughput screen to detect secretion inhibitors was developed based on the translational autoregulation of one of the central protein components, SecA. The assay makes use of a SecA-LacZ fusion reporter construct in Escherichia coli which is induced when secretion is perturbed. Several compounds, including two natural product extracts, which had the ability to induce the reporter fusion were identified and the MICs of these compounds forStaphylococcus aureus strain MN8 were found to be ≤128 μg/ml. Enzyme-linked immunosorbent assay, Western blotting, and immunoprecipitation techniques were used to analyze the affects of these compounds on protein secretion. Six representative compounds presented here appear to be bona fide secretion inhibitors but were found to have deleterious effects on membranes. It was concluded that, while the method described here for identifying inhibitors of secretion is valid, screens such as this, which are directed against the membrane-bound portion of a pathway, may preferentially identify compounds which affect membrane integrity.


Cells ◽  
2020 ◽  
Vol 9 (6) ◽  
pp. 1559 ◽  
Author(s):  
Lukas M. Braun ◽  
Robert Zeiser

Myeloproliferative diseases, including myeloproliferative neoplasms (MPN) and myelodysplastic syndromes (MDS), are driven by genetic abnormalities and increased inflammatory signaling and are at high risk to transform into acute myeloid leukemia (AML). Myeloid-derived suppressor cells were reported to enhance leukemia immune escape by suppressing an effective anti-tumor immune response. MPNs are a potentially immunogenic disease as shown by their response to interferon-α treatment and allogeneic hematopoietic stem-cell transplantation (allo-HSCT). Novel immunotherapeutic approaches such as immune checkpoint inhibition, tumor vaccination, or cellular therapies using target-specific lymphocytes have so far not shown strong therapeutic efficacy. Potential reasons could be the pro-inflammatory and immunosuppressive microenvironment in the bone marrow of patients with MPN, driving tumor immune escape. In this review, we discuss the biology of MPNs with respect to the pro-inflammatory milieu in the bone marrow (BM) and potential immunotherapeutic approaches.


2019 ◽  
Vol 48 (1) ◽  
pp. 197-209 ◽  
Author(s):  
Hongyao Xu ◽  
Xiangjie Zou ◽  
Pengcheng Xia ◽  
Mohammad Ahmad Kamal Aboudi ◽  
Ran Chen ◽  
...  

Background: Meniscal injury is very common, and injured meniscal tissue has a limited healing ability because of poor vascularity. Platelets contain both pro- and anti-angiogenic factors, which can be released by platelet selective activation. Hypothesis: Platelets release a high level of vascular endothelial growth factor (VEGF) when they are activated by protease-activated receptor 1 (PAR1), whereas the platelets release endostatin when they are activated by protease-activated receptor 4 (PAR4). The PAR1-treated platelets enhance the proliferation of meniscal cells in vitro and promote in vivo healing of wounded meniscal tissue. Study Design: Controlled laboratory study. Method: Platelets were isolated from human blood and activated with different reagents. The released growth factors from the activated platelets were determined by immunostaining and enzyme-linked immunosorbent assay. The effects of the platelets with different treatments on meniscal cells were tested by an in vitro model of cell culture and an in vivo model of wounded meniscal healing. Results: The results indicated that platelets contained both pro- and antiangiogenic factors including VEGF and endostatin. In unactivated platelets, VEGF and endostatin were contained inside of the platelets. Both VEGF and endostatin were released from the platelets when they were activated by thrombin. However, only VEGF was released from the platelets when they were activated by PAR1, and only endostatin was released from the platelets when they were activated by PAR4. The rat meniscal cells grew much faster in the medium that contained PAR1-activated platelets than in the medium that contained either PAR4-activated platelets or unactivated platelets. The wounds treated with PAR1-activated platelets healed faster than those treated with either PAR4-activated platelets or unactivated platelets. Many blood vessel–like structures were found in the wounded menisci treated with PAR1-activated platelets. Conclusion: The PAR1-activated platelets released high levels of VEGF, which increased the proliferation of rat meniscal cells in vitro, enhanced the vascularization of menisci in vivo, and promoted healing of wounded menisci. Clinical Relevance: Our results suggested that selective activated platelets can be used clinically to enhance healing of wounded meniscal tissue.


2020 ◽  
Vol 21 (22) ◽  
pp. 8448
Author(s):  
Chun-Hao Hung ◽  
Keh-Yang Wang ◽  
Yae-Huei Liou ◽  
Jing-Ping Wang ◽  
Anna Yu-Szu Huang ◽  
...  

Erythroid Krüppel-like factor (EKLF/KLF1) was identified initially as a critical erythroid-specific transcription factor and was later found to be also expressed in other types of hematopoietic cells, including megakaryocytes and several progenitors. In this study, we have examined the regulatory effects of EKLF on hematopoiesis by comparative analysis of E14.5 fetal livers from wild-type and Eklf gene knockout (KO) mouse embryos. Depletion of EKLF expression greatly changes the populations of different types of hematopoietic cells, including, unexpectedly, the long-term hematopoietic stem cells Flk2− CD34− Lin− Sca1+ c-Kit+ (LSK)-HSC. In an interesting correlation, Eklf is expressed at a relatively high level in multipotent progenitor (MPP). Furthermore, EKLF appears to repress the expression of the colony-stimulating factor 2 receptor β subunit (CSF2RB). As a result, Flk2− CD34− LSK-HSC gains increased differentiation capability upon depletion of EKLF, as demonstrated by the methylcellulose colony formation assay and by serial transplantation experiments in vivo. Together, these data demonstrate the regulation of hematopoiesis in vertebrates by EKLF through its negative regulatory effects on the differentiation of the hematopoietic stem and progenitor cells, including Flk2− CD34− LSK-HSCs.


Blood ◽  
2000 ◽  
Vol 96 (1) ◽  
pp. 347-354 ◽  
Author(s):  
Werner Krenger ◽  
Simona Rossi ◽  
Luca Piali ◽  
Georg A. Holländer

Abstract Reconstitution of the peripheral T-cell compartment is a critical aspect for the success of bone marrow transplantation and is also dependent on the reestablishment of normal thymic structure and function. Graft-versus-host disease (GVHD), however, exacerbates posttransplant immunodeficiency through a deleterious effect on thymic function. To investigate the mechanisms of GVHD-mediated thymic disease, 2 murine parent→F1transplantation models of acute and chronic GVHD, respectively, were studied. Acute GVHD was associated with changes in thymic architecture and a reduction in cellularity mainly because of the decrease in CD4+CD8+, or double-positive (DP) thymocytes, to less than 15% of values found in mice without GVHD. Simultaneously, mature donor-derived T cells expanded in the confines of the allogeneic thymic microenvironment, leading to local inflammation. Through analysis of in vivo cell proliferation, we demonstrated that the ensuing depletion of DP thymocytes was secondary to a decreased commitment of resident pro-T and pre-T cells to enter the cell cycle. Moreover, DP cells themselves showed altered proliferative capacities in the presence of acute GVHD. These findings suggested that thymic atrophy in acute GVHD is effected by impaired cellular proliferation of immature host thymocytes and that the failure of these cells to enter the cell cycle is dependent on an interferon (IFN)-γ–driven immune response. In contrast, interleukin-4–driven chronic GVHD was not accompanied by a sustained thymic infiltration of donor T cells. Consequently, there was a lack of apparent structural changes, a restricted in situ transcription of inflammatory cytokines, and a virtually unchanged cell cycle progression in vivo.


Blood ◽  
1999 ◽  
Vol 94 (7) ◽  
pp. 2271-2286 ◽  
Author(s):  
M. Rosenzweig ◽  
T.J. MacVittie ◽  
D. Harper ◽  
D. Hempel ◽  
R.L. Glickman ◽  
...  

Optimization of mobilization, harvest, and transduction of hematopoietic stem cells is critical to successful stem cell gene therapy. We evaluated the utility of a novel protocol involving Flt3-ligand (Flt3-L) and granulocyte colony-stimulating factor (G-CSF) mobilization of peripheral blood stem cells and retrovirus transduction using hematopoietic growth factors to introduce a reporter gene, murine CD24 (mCD24), into hematopoietic stem cells in nonhuman primates. Rhesus macaques were treated with Flt3-L (200 μg/kg) and G-CSF (20 μg/kg) for 7 days and autologous CD34+ peripheral blood stem cells harvested by leukapheresis. CD34+ cells were transduced with an MFGS-based retrovirus vector encoding mCD24 using 4 daily transductions with centrifugations in the presence of Flt3-L (100 ng/mL), human stem cell factor (50 ng/mL), and PIXY321 (50 ng/mL) in serum-free medium. An important and novel feature of this study is that enhanced in vivo engraftment of transduced stem cells was achieved by conditioning the animals with a low-morbidity regimen of sublethal irradiation (320 to 400 cGy) on the day of transplantation. Engraftment was monitored sequentially in the bone marrow and blood using both multiparameter flow cytometry and semi-quantitative DNA polymerase chain reaction (PCR). Our data show successful and persistent engraftment of transduced primitive progenitors capable of giving rise to marked cells of multiple hematopoietic lineages, including granulocytes, monocytes, and B and T lymphocytes. At 4 to 6 weeks posttransplantation, 47% ± 32% (n = 4) of granulocytes expressed mCD24 antigen at the cell surface. Peak in vivo levels of genetically modified peripheral blood lymphocytes approached 35% ± 22% (n = 4) as assessed both by flow cytometry and PCR 6 to 10 weeks posttransplantation. In addition, naı̈ve (CD45RA+and CD62L+) CD4+ and CD8+cells were the predominant phenotype of the marked CD3+ T cells detected at early time points. A high level of marking persisted at between 10% and 15% of peripheral blood leukocytes for 4 months and at lower levels past 6 months in some animals. A cytotoxic T-lymphocyte response against mCD24 was detected in only 1 animal. This degree of persistent long-lived, high-level gene marking of multiple hematopoietic lineages, including naı̈ve T cells, using a nonablative marrow conditioning regimen represents an important step toward the ultimate goal of high-level permanent transduced gene expression in stem cells.


Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 1818-1818
Author(s):  
Justin P. Kline ◽  
Rajiv Swamy ◽  
Dezheng Huo ◽  
Laura Michaelis ◽  
Richard A. Larson ◽  
...  

Abstract Conditioning regimens using in vivo alemtuzumab (Campath-1H, humanized anti-CD52) are characterized by low rates of acute and chronic GVHD, but may also result in delayed immune reconstitution. Optimization of such regimens will depend on an understanding of the relation between alemtuzumab exposure, immune reconstitution and GVHD. We have conducted a prospective study of fludarabine 30 mg/m2/d x 5 days, melphalan 140 mg/m2 x 1 day and alemtuzumab 20 mg/d x 5 days as conditioning for related and unrelated allografts. Using an enzyme-linked immunosorbent assay (ELISA), we determined serum free and total Campath levels in 46 patients with hematologic malignancies (45) or sickle cell disease (1) on day 0, day 7, day 14, day 28, day 50, day 75, day 100, day 150 and at one year after transplant (HSCT). 26 (57%) had a matched sibling donor, 16 (35%) a matched unrelated donor (MUD) and 4 (9%) a mismatched related or unrelated donor. 44 pts engrafted and are included in the analysis. Median follow up for survivors was 2.8 years. Grade II–IV aGVHD occurred in 8 pts after a median of 42 days (range 22 to 60). Eight pts developed cGVHD after a median of 107 days (range 89–140). 15/41 (37%) at risk pts developed CMV reactivation. The half-life of free alemtuzumab (fA) was 26 days after HSCT with wide interpatient variation in fA pharmacokinetics (e.g. Coefficient of variation was 138% on day 0). On day 0, 1 patient had an undetectable level of fA. By day 28, 50, and 100, there were 6 (14.6%), 12 (35.3%), and 13 (52%) pts with undetectable fA, respectively. Figure 1 shows the fitted means and 95% confidence intervals of fA over time. Using log-rank and Cox proportional hazard models, there was no association between fA on day 0, day 28, or the last available fA, and development of acute GVHD. However, pts with higher average free and total Campath levels in the first month had a lower risk of developing cGVHD (p=0.02). The median fA concentration in the first month for pts with cGVHD was 0.32 (inter-quarter range IQR: 0.22–0.41), as compared with 0.97 (IQR: 0.23–3.31) in those without cGVHD. No significant association between absolute lymphocyte count and fA concentration was found after adjusting for time (p=0.28). Finally, among pts at risk, a higher fA concentration on day 0 (p=0.002), and in the first month (p=0.003) was significantly associated with CMV viremia. In summary, the estimated half-life of serum fA is 26 days after HSCT, but with considerable interpatient variability. Higher concentrations of fA were associated with a decreased incidence of cGVHD, but an increased risk of CMV reactivation. In contrast to a previous preliminary analysis, no association existed between fA and lymphocyte reconstitution. Variation in alemtuzumab pharmacokinetics may predict important clinical outcomes, such as cGVHD and CMV reactivation. Future studies are warranted to determine an optimal alemtuzumab exposure that hastens immune reconstitution while minimizing chronic GVHD. Fig 1. Fitted Mean Free Campath and 95% CI Fig 1. Fitted Mean Free Campath and 95% CI


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