bluuebunny/crossref_metadata_2025_split huggingface dataset:
CD26 Inhibition Enhances Allogeneic Donor Cell Homing and Engraftment after In Utero Bone Marrow Transplantation.
Authors: Peranteau William H. , Endo Masayuki , Adibe Obinna O. , Merchant Aziz , Zoltick Philip , Flake Alan W. 
Published: April 2020

Abstract
In utero bone marrow transplantation (IUBMT) induces donor-specific tolerance for postnatal cellular or organ transplantation. Consistent induction of tolerance requires a threshold of mixed hematopoietic chimerism (&gt;1%). CD26 is a peptidase whose inhibition has been demonstrated to influence granulocyte colony-stimulating factor induced mobilization of hematopoietic stem cells and increase homing and engraftment of BM cells in adult transplantation models. We hypothesized that CD26 inhibition would increase the frequency and levels of allogeneic hematopoietic chimerism after IUBMT.
Methods: B6 GFP BM was injected intravenously into E14 Balb/c fetal mice at a dose of 20e6 cells/fetus with or without CD26 inhibition with diprotin A. Early kinetic analysis was performed to assess donor cell homing to fetal liver (FL). Peripheral blood (PB) was analyzed up to 20 weeks after birth for donor cell chimerism and multilineage engraftment by flow cytometry. PB was also analyzed for donor cell chimerism at the same time points from Balb/c mice receiving 10e6 CD26 inhibited GFP BM cells coinjected with 10e6 noninhibited congenic B6Ly5.2 BM cells at E14 to assess for an in utero competitive advantage provided by CD26 inhibition.
Results: CD26 inhibition increased donor cell homing to the FL at 24, 48 and 96 hours after injection (69.27±8.19 vs 30.21±6.44, 57.20±14.63 vs 36.80±14.20, 46.14±15.79 vs 12.09±7.01; p&lt;0.05 at all time points). The frequency and levels of engraftment at 4 weeks of life were increased in those mice receiving CD26 inhibited BM compared to noninhibited BM (50.0% vs 22.5%; 20.48±14 vs 6.96±8.44, p&lt;0.05). Chimerism was multilineage and maintained at 20 weeks of age (figure 1a) supporting improvement of engraftment at the stem or early progenitor cell level (* p&lt;0.05 comparing chimerism levels between inhibitied and noninhibited cells). The coinjection of CD26 inhibited and noninhibited cells resulted in higher levels of engraftment of inhibited cells at all time points up to 16 weeks of age arguing for a competitive engraftment advantage of early progenitor cells provided by CD26 inhibition (figure 1b).
Conclusion: CD26 inhibition of donor BM prior to IUBMT results in an increased efficiency of donor engraftment and higher levels of chimerism. CD26 inhibition offers a potential mechanism to increase the level of engraftment and the rate of donor specific tolerance and may facilitate combined pre and postnatal strategies for cellular and organ transplantation.
Figure Figure
Immunoregulatory Effects of Allogeneic Mixed Chimerism Induced by T-Cell Depleted, Nonmyeloablative Bone Marrow Transplantation on Chronic Inflammatory Arthritis and Autoimmunity Developed in Interleukin-1 Receptor Antagonist-Deficient Mice.
Authors: Cho Seok-Goo , Park Min-Chung , Min So-Youn , Cho Young-Gyu , Lee Seok , Cho Mi-La , Chang Hong-Seok , Lee Jong-Wook , Min Woo-Sung , Kim Ho-Youn , Kim Chun-Choo 
Published: April 2020

Abstract
Objective: To investigate the immunoregulatory effects of allogeneic mixed chimerism induced by T-cell depleted, nonmyeloablative bone marrow transplantation (TCD-NMT) on chronic inflammatory arthritis and autoimmunity developed in interleukin-1 receptor antagonist-deficient (IL-1Ra−/ −) mice.
Methods: IL-1Ra−/ − mice (H-2kd) were treated with anti-asialoGM1 Ab, TBI 500 cGy, and TCD-NMT from C57BL/6 mice (H-2kb). Engraftment and chimerism were evaluated on peripheral blood (PB), lymph node, and spleen by multi-color flow cytometry. The severity of arthritis was evaluated by clinical score and histopathology. IgG1 and IgG2a subtype of anti-type II collagen (CII) were measured in PB samples. After T cells were stimulated with CII, ovalbumin, and phytohemagglutinin, T-cell proliferation response and cytokines production (INF-g, TNF-a, IL-10, and IL-17) in culture supernatant were assayed.
Results: All the transplanted IL-1Ra mice showed marked improvement of arthritis within 3 weeks after transplantation as well as successful induction of mixed chimerism. Mice in mixed chimerism showed higher level of anti-CII IgG1 and lower level of anti-CII IgG2a and weaker T cell proliferative response than in control groups, such as no-treatment and conditioning only without BM rescue. In mixed chimera, INF-g, TNF-a and IL-17 production from CII-stimulated T cells was significantly suppressed and IL-10 production was significantly increased as compared to the control groups.
Conclusion: These observations indicate that the introduction of allogeneic mixed chimerism has a strong immunoregulatroy potential to correct established chronic inflammatory arthritis and autoimmunity originating from dysregulated proinflammatory cytokine network.
Use of Engineered Autologous BOEC for Gene Therapy of Canine Hemophilia A.
Authors: Hebbel Robert P. , Milbauer Liming , Roney Mark , Lillicrap David , Voorberg Jan , Nichols Timothy C. 
Published: April 2020

Abstract
Hemophilia A is an attractive candidate disease for corrective gene therapy because relatively small amounts of the missing protein, FVIIII, have a significant biological effect. We previously described a novel approach: intravenous administration of ex vivo expanded BOEC (blood outgrowth endothelial cells) engineered to express FVIII. Robust therapeutic results were obtained using NOD/SCID mice and human BOEC stably transfected to express human FVIII (Blood99:457, 2002). We are now conducting scale-up studies in the canine hemophilia A model. Based on the previous mouse data, we estimated that 400 million cells would be required to achieve a therapeutic effect in the dog. First, two normal dogs were infused with autologous unmanipulated BOEC, which revealed removal from the circulation via single-pass kinetics. Then, BOEC cultures were obtained from three hemophilia A dogs, and an onco-retroviral vector (MIRG/cFVIII) was used to obtain transduced BOEC producing B-domainless canine FVIII. Then, for each treatment, autologous engineered BOEC were given IV to the original donor dogs, with dose divided over 3 daily infusions, and whole blood clotting time (WBCT, in minutes) was followed.
Dog Date # cells infused pre Rx WBCT effect on WBCT D28 12/04 274x106 &gt;60 nadir 36 at day 14, Still 39 at day 185 D28 6/05 386x106 39 nadir 24.5 at day 23, now at 30 at day 44, being followed E64 7/04 220x106 &gt;60 nadir 34.5 at day 3, &gt;60 min by day 38 E64 12/04 274x106 &gt;60 still 37.5 at day 189 E64 6/05 185x106 37.5 nadir 34.5 at day 2, at 36 on day 32, being followed H17 6/05 272x106 &gt;60 nadir 29 at day 44, being followed
WBCT (nl = 8-12 min) needs to be &lt;20 min to reflect &gt;0.1% FVIII activity; but the observed WBCT are clearly improved compared to baseline. One dog (E64) was treated for a mouth bleeds on d37 in cycle 2, and d36 in cycle 3. One animal (E64) developed hypotension and tachypnea during one of the infusions (probably because cells were not kept sufficiently in suspension) but recovered. Four of the infusions were associated with mild transient thrombocytopenia. Based on these initial scale-up experiments, it appears that use of engineered BOEC for gene therapy is an approach worthy of continued study, as cell doses can be increased and other improvements in the method are readily envisioned. It has a number of advantages, including use of autologous carrier cells with which any expression vector can be paired, ex vivo exposure to vector rather than in vivo, use of BOEC expansion in culture to simultaneously achieve vector expansion, and the ability to chose a clone of engineered cells having a single, known (and studied) insertion site.
High Dose Cyclophosphamide (CY) for Severe Aplastic Anemia (SAA): Safety and Long Term Follow-Up.
Authors: Brodsky Robert A. , Chen Allan , Dorr Donna M. , Brodsky Isadore , Jones Richard J. 
Published: April 2020

Abstract
Background: Acquired SAA is a potentially life-threatening bone marrow failure disorder that is usually immune-mediated. SAA can be effectively treated with bone marrow transplantation (BMT), immunosuppressive therapy (IST), or CY. BMT can cure the disease but ideally requires a matched sibling donor. IST has a high response rate, but up to 50% of patients relapse, become dependent on cyclosporine, or develop secondary clonal disease such as paroxysmal nocturnal hemoglobinuria (PNH) or myelodysplastic syndromes (MDS). We previously showed that CY induces durable remissions in the majority of SAA patients, but concern has been raised about the safety of this approach.
Methods: Since 1996, we treated 38 previously untreated, and 17 IST failed, SAA patients with CY (50 mg/kg/d x 4) followed by daily G-CSF (5 ug/kg/day) until the neutrophil count (ANC) reached 1000/dl. Response was defined as ANC &gt; 1000/dl and transfusion independence without growth factor support for &gt; 3 months. Relapse was defined as no longer meeting criteria for response. PNH was monitored by flow cytometry.
Results: The median age of the newly diagnosed patients was 40 (range 2–68) years. With a median follow-up of 41 (range, 6 – 111) months, 33/38 patients survive (actuarial survival of 86%, 95% CI 72–95%) with 28 (74%, 95% CI 58–85%) achieving remission, most being complete. Median time to ANC of 500, last platelet and red cell transfusion was 50, 99, and 181 days, respectively. Before treatment, 15 patients met criteria for very (v) SAA (ANC &lt; 200). Mortality within 6 months after CY occurred in 4 (10.5%) patients, all with vSAA; 1 additional patient died from bacterial sepsis 18 months after CY. 22/23 (96%) SAA patients survive (20 in remission) compared to 11/15 (73%) with vSAA (10 in remission). Eight patients had a severe infection at the time of beginning treatment and 5 survive in remission. PNH screening revealed a PNH population ranging from 0.5–40% of granulocytes in 12 patients, and all 12 achieved a durable remission (p = 0.039). No patient in this series has progressed to PNH or MDS, and the PNH clone is regressing in all 12 patients. Two patients have relapsed. One patient, whose first remission lasted 5 years, was retreated with CY into a persisting second complete remission 3 years ago; another patient recently relapsed 3 years after achieving remission. Ten of the 17 patients who failed IST (median age of 31, range 6–58) are alive and nine are in remission.
Conclusions: Cy is safe and highly effective therapy for both untreated and relapsed SAA. Relapses after CY are rare and progression to PNH or MDS in previously untreated patients has not been observed in this series with now 15 patients out beyond 5 years. The presence of a PNH population may be a favorable risk factor, perhaps by excluding non-immune mediated forms of SAA.
Figure Figure
Prevention of Acute Graft-Versus-Host Disease by CD4+CD25+ Regulatory T Cells Is Dependent on the CD30/CD153 Pathway.
Authors: Zeiser Robert , Nguyen Vu , Buess Martin , Karimi Mobin , Bjorck Pia , Baker Jeanette , Negrin Robert S. 
Published: April 2020

Abstract
CD4+CD25+ regulatory T (Treg) cells suppress acute graft versus host disease (aGVHD), prevent autoimmunity and delay allograft rejection. CD30 and other TNF-R family members have been demonstrated to be expressed by Treg and to function as alternative costimulatory pathways for T cell activation. In this study we assessed the significance of the CD30/CD153 pathway in Tregs suppression of aGVHD in a murine major MHC mismatch BMT model. Using bioluminescence imaging proliferation of donor derived luciferase-labeled CD4+ and CD8+ T cells was quantified at serial time points after transplantation. Treg suppressed the early expansion of alloreactive T-cells. Immunofluorescence microscopy revealed a predominant infiltration of donor derived Treg in CD153 positive regions of secondary lymphoid organs, namely parafollicular T cell zones of lymph nodes and the subepithelial dome regions of Peyers Patches. In vivo blockade of the CD30/CD153 pathway with anti CD153 Ab did not alter Treg migration to secondary lymphoid organs but reduced their suppressive effect. Proliferation of donor T cells as measured in photons/second/mouse was significantly higher in animals receiving Treg and CD153 blocking antibodies as compared to recipients of Treg only (p=0.0038). Gene expression profiling of Treg with DNA microarrays indicated a Treg signature that was consistently found in different mouse strains. This Treg signature was altered after CD153 blockade in vitro. Importantly, aGVHD lethality was significantly increased (p=0.021) when CD30-CD153 interaction was blocked during Treg transfer. This study provides direct evidence that the TNF-R family member CD30 is critical for Treg cell function in the regulation of pathological T cell responses that lead to aGVHD.
Inhibition of Graft-Versus-Host Disease (GVHD) by Histone Deacetylase Inhibitor (HDAC) SAHA Leads to Downregulation of STAT1 Activation.
Authors: Leng Corinna , Li Cuiling , Ziegler Judy , Lokshin Anna , Lentzsch Suzanne , Mapara Markus Y. 
Published: April 2020

Abstract
Histone deacetylase (HDAC) inhibitors have been shown to reduce development of graft versus host disease [GVHD] following allogeneic bone marrow transplantation [BMT]. Administration of the HDAC inhibitor suberonylanilide hydroxamic acid [SAHA] resulted in a significantly reduced GVHD-dependent mortality following fully MHC-mismatched allogeneic BMT. Median Survival Time (MST) for vehicle and SAHA-treated mice were 7.5 days and 38 days respectively. However, SAHA treatment did not affect T cell activation nor T cell expansion in vitro and in vivo as determined by MLR assays, phenotypic analysis of donor T cells with regard to expression of the CD25 activation antigen and calculation of donor CD4+ and CD8+ T cell numbers on days +3 and +6 post-BMT. Thus, SAHA treatment was not able to inhibit the strong upregulation of CD25 antigen on CD8+ T cells observed during induction of GVHD on days +3 and +6 post-BMT. We therefore focused on the effects of SAHA treatment on efferent immune effects including cytokine secretion and intracellular signaling events in vitro and in vivo following GVHD induction. SAHA treatment broadly inhibited lipopolysaccharide [LPS] and allo-antigen-induced cytokine/chemokine secretion in vitro like MIP-1-α, IP-10, IFN-γ, TNF-α and IL-6 and led also to a significant decrease in IFN-γ and TNF-α levels in vivo following induction of GVHD. Concomitantly, SAHA treatment inhibited phosphorylation of STAT1 and STAT3 in response to LPS and allo-activation in vitro. Furthermore, analysis of liver tissue and spleens from SAHA-treated animals with GVHD showed a significant decrease in phosphorylated STAT1. In contrast SAHA treatment had only moderate effects on p38 or ERK1,2 Mitogen-activated Protein Kinase (MAPK) pathway underscoring the relevance of the inhibition of the STAT1 pathway. In conclusion, GVHD is associated with a strong induction of phosphorylation of STAT1 in the liver and spleen and SAHA-dependent reduction of GVHD is associated with systemic and local inhibition of pSTAT1 and modulation of the inflammatory cytokine milieu during the efferent immune response.
Immunomodulatory Effects of the Triterpenoid CDDO after Allogeneic Bone Marrow Transplantation in Mice: Reduction of Acute Graft-Versus-Host Disease Lethality.
Authors: Murphy William J. , Frolova Olga , Konopleva Marina , Andreeff Michael , Ma Weihong , Wilkins Danice , Welniak Lisbeth A. , Sun Kai 
Published: April 2020

Abstract
The use of hematopoietic stem cell transplantation (HSCT) in cancer treatment is seriously hampered by the occurrence of graft-versus-host disease (GVHD) and cancer relapse. During acute GVHD, inflammatory cytokines play a pivotal role in the amplification of GVHD. Therefore, assessment of agents with known anti-neoplastic activity that also reduce cytokine production may be useful in both the prevention of GVHD and cancer relapse. The synthetic triterpenoid, CDDO (2-cyano-3, 12-dioxooleana-1, 9-dien-28-oic acid) is multifunctional molecule which has shown potent anti-cancer activities both in vitro and in vivo through the induction of apoptosis. We first examined the effects of CDDO on both human and murine T cell mitogen responses in vitro. CDDO significantly inhibited mitogen responsiveness of both human and murine T cells in vitro with evidence of cell cycle arrest of the human T cells. We then proceeded to examine the effects of CDDO on acute GVHD induction and progression. In these studies, lethally irradiated C57BL/6 mice received 10 million bone marrow cells (BMC) and 40 million spleen cells from fully MHC-mismatched BALB/c donors. All of the control mice succumbed rapidly due to acute GVHD. In contrast, the mice that received CDDO (120 ug/BID) given from days 0-3 following BMT exhibited significant improvement in survival (P &lt; 0.001). Body weights from the treated mice also were significantly increased compared to untreated controls. We found that the timing of CDDO administration was a critical factor for protection from GVHD as protection only occurred when CDDO was administered early after BMT. Importantly, donor myeloid reconstitution was not adversely affected by CDDO treatment as determined by peripheral blood cell count and donor chimerism assessment on day +14 post-transplant. No adverse toxicities or effects on reconstitution were observed in the mice receiving BMC alone with CDDO being administered continuously. Given the reported direct anti-tumor effects of CDDO, it will be of particular interest in examining the effects of CDDO and allogeneic BMT in tumor-bearing recipients. Our findings suggest that CDDO can enhance the efficacy of allogeneic BMT by decreasing acute GVHD in mice.
Protein-Based Identification of Inflammatory Signaling Pathways in Graft-Versus-Host-Disease (GVHD): Evidence for STAT1 Pathway as a Key Regulator in GVHD.
Authors: Li Cuiling , Ziegler Judy , Leng Corinna , Lokshin Anna , Lentzsch Suzanne , Mapara Markus Y. 
Published: April 2020

Abstract
The pathogenesis of GVHD is influenced by the immunogenetic disparities between donor and recipient, the presence of host-derived antigen-presenting cells, the inflammatory reaction in response to conditioning-induced tissue damage and cytokine secretion. Intestinal damage and subsequent translocation of LPS into the circulation have been shown to be central to the subsequent pathogenetic events occurring in development of GVHD. Interfering with this proinflammatory response by targeted therapy might be an attractive new approach for inhibiting the development of GVHD. We have therefore started to comprehensively analyze the inflammatory signaling pathways, which are activated during the induction phase of GVHD in order to identify key molecular regulators of GVHD, which might serve as targets for future therapy. For this purpose GVHD was in induced in the fully MHC-mismatched BALB/c to B6 strain combination following lethal irradiation with 9.5 Gy. Spleens as well as GVHD target organs liver, skin and small bowel and colon were harvested on days 1, 3, and + 6 post-BMT to study activation of JAK-STAT, MAPK, PI3-Kinase and NF-κB signaling transduction pathways. Studies were performed using western blot, multiplex assays for analysis of phoshotyrosine proteins and transcription factor binding activity and also standard gel shift assays. Furthermore, GVHD liver tissue was subjected to microarray gene expression profiling. Most strikingly, in contrast to syngeneic and allogeneic controls induction of GVHD was associated with a strong activation of JAK-STAT pathway as determined by Tyr701-specific STAT1 phosphorylation and also Tyr705-specific STAT3 phosphorylation in the liver and spleen as early as on day +1 post-BMT and remained elevated until day +6 post-BMT. In line with these results we observed a significant increase of STAT1 dependent gene expression (e.g. CXCL10 5.6-fold, IRF7 4.5-fold, CXCL9 4.2-fold) as studied by microarray gene expression profiling in the liver. Furthermore, multiplex DNA-binding and standard gel shift studies in the liver revealed that GVHD led to an increased DNA binding activity of most prominently NF-κB but also AP-2, EGR1 and NF1. In conclusion we provide evidence that induction of GVHD is associated with strong activation of the STAT1 pathway in the spleen and liver as early as on day +1 post-BMT in additon to activation of the NF-κB pathway. Therefore this study might help to identify candidate molecules to create more specific therapies targeting the inflammatory component of GVHD and thus reducing the development of GVHD without mitigating the GVL response.
“Virtual Flow Cytometry” of Immunostained Lymphocytes on Microscopic Tissue Slides.
Authors: Cualing Hernani D. , Moscinski Lynn C. 
Published: April 2020

Abstract
A method and approach were developed for fully automated measurements of immunostained lymphocytes in tissue sections by means of digital color microscopy and patent pending advanced cell analysis. The validation data for population statistic measurements of immunostained lymphocytes in tissue sections using tissue cytometry is presented. Segmentation of a 512 x 474 RGB image and display of statistical results table took 12 to 15 seconds using in-house developed algorithms. We used a panel of 7 antibodies for validation on 14 cases of mantle cell lymphoma giving percentage positive, total lymphocytes, density staining. An average of 2027 image frames with 810,800 cell objects were evaluated. Antibodies to CD3, CD4, CD8, Bcl-1, Ki-67, CD20, CD5 were subjected to virtual flow cytometry on tissue. The results of tissue cytometry were compared with manual counts of expert observers and with the results of flow cytometry immunophenotyping of the same specimen. The correlation coefficient and 95 % confidence interval by linear regression analysis yielded a high concordance between manual human results (M), flow cytometry results(FC), and tissue cytometry (TC) results per antibody, (r =0.9365 manual vs TC, r =0.9537 FC vs TC). We noted a lower CD8 tumor infiltrating lymphocytes in blastic form than in non-blastic form of mantle cell and a higher Ki67 in the latter subtype than in the former. These results suggest the technology of tissue cytometry could be a clinically valid surrogate for both manual and flow cytometry analysis when only tissue immunohistochemistry is available for diagnosis and prognosis. The application for cancer diagnosis, monitoring, and prognosis is for objective counting of immunostained cells in tissues with percentage results. These results could be also be used in standardization of cut off percentage used in diagnosis; for exemplary tool for leukemic blasts(CD34) &gt;20% for acute leukemia, for determining automated 30 % cut off for positivity for prognostic markers in diffuse large B-cell lymphoma for bcl-2, bcl-6, mum-1, and CD10.