Aki Sato

Inhibition of PAI-1 induces neutrophil-driven neoangiogenesis and promotes tissue regeneration via production of angiocrine factors in mice

Plasminogen activator inhibitor-1 (PAI-1), an endogenous inhibitor of a major fibrinolytic factor, tissue-type plasminogen activator, can both promote and inhibit angiogenesis. However, the physiologic role and the precise mechanisms underlying the angiogenic effects of PAI-1 remain unclear. In the present study, we report that pharmacologic inhibition of PAI-1 promoted angiogenesis and prevented tissue necrosis in a mouse model of hind-limb ischemia. Improved tissue regeneration was due to an expansion of circulating and tissue-resident granulocyte-1 marker (Gr-1(+)) neutrophils and to increased release of the angiogenic factor VEGF-A, the hematopoietic growth factor kit ligand, and G-CSF. Immunohistochemical analysis indicated increased amounts of fibroblast growth factor-2 (FGF-2) in ischemic gastrocnemius muscle tissues of PAI-1 inhibitor-treated animals. Ab neutralization and genetic knockout studies indicated that both the improved tissue regeneration and the increase in circulating and ischemic tissue-resident Gr-1(+) neutrophils depended on the activation of tissue-type plasminogen activator and matrix metalloproteinase-9 and on VEGF-A and FGF-2. These results suggest that pharmacologic PAI-1 inhibition activates the proangiogenic FGF-2 and VEGF-A pathways, which orchestrates neutrophil-driven angiogenesis and induces cell-driven revascularization and is therefore a potential therapy for ischemic diseases.

Unrelated Cord Blood Transplantation after Myeloablative Conditioning in Adults with Acute Myelogenous Leukemia

We analyzed the disease-specific outcomes of adult acute myelogenous leukemia (AML) patients treated with unrelated cord blood transplantation (CBT) after myeloablative conditioning. Between August 1998 and February 2008, 77 adult patients with AML were treated with unrelated CBT. All patients received 4 fractionated 12 Gy total body irradiation (TBI) and chemotherapy as myeloablative conditioning. The median age was 45 years, the median weight was 55 kg, the median number of nucleated cells was 2.44 x 10(7)/kg, and the median number of CD34-positive cells was 1.00 x 10(5)/kg. All patients received a single and HLA mismatched cord blood unit. The cumulative incidence of neutrophil recovery at day 50 and platelet recovery at day 200 was 94.8% and 91.7%, respectively. A higher CD34-positive cell dose was associated with faster hematopoietic recovery. The cumulative incidence of grade III to IV acute graft-versus-host disease (aGVHD) and extensive-type chronic GVHD (cGVHD) was 25.1% and 28.6%, respectively. With a median follow-up of 78 months, the probability of event-free survival (EFS) at 5 years was 62.8%. The 5-year cumulative incidence of treatment related-mortality (TRM) and relapse was 9.7%, 25.8%, respectively. In multivariate analyses, the risk factor identified for event free survival (EFS) was disease status and cytogenetics. These results suggest that unrelated CBT after myeloablative conditioning could be safely and effectively used for adult patients with AML.

Allogeneic stem cell transplantation for hepatosplenic gammadelta T-cell lymphoma.

Hepatosplenic gammadelta T-cell lymphoma (HSTCL) was first described by Farcet et al. in 1990 [1]. Most cases of HSCTL occur in young men. Patients typically present with hepatosplenomegaly and bone marrow infiltration with resultant cytopenias but comparatively little lymphadenopathy. The prognosis of HSCTL is poor with reported median survivals of 8 – 16 months and few instances of longterm disease free survival with conventional chemotherapy [2,3]. Recently, a limited number of cases treated with allogeneic SCT for HSTCL have been reported [2 – 15]. We previously reported a patient of HSTCL successfully treated with allogeneic bone marrow transplantation (BMT) from an HLAidentical sibling [6]. In this report, we provide an update of this patient with 7-years follow-up and review the literature for allogeneic SCT for HSTCL. A 23-year-old Japanese male was admitted to our hospital in June 1999 with abdominal distension, malaise, and night sweats. As previously described, the patient was diagnosed with hepatosplenic gamma/delta T-cell lymphoma and treated with intensive B-NHL86 protocol chemotherapy. After the two courses of chemotherapy, he achieved complete remission (CR). Thereafter, he received allogeneic BMT in August 1999 from an HLAidentically matched younger sister. The conditioning regimen consisted of four fractionated 12-Gy total body irradiation (TBI) on Day 79 and Day 78 and high-dose etoposide (60 mg/kg), which was administered as 24-h continuous intravenous infusion on Day 74. Graft-versus-host disease (GVHD) prophylaxis consisted of intravenous cyclosporine and short-term methotrexate. He had evidence of grade II acute GVHD of the skin and gut, which required no steroid treatment. Chimerism evaluation assessed by fluorescent in situ hybridization (FISH) using a mixture of X and Y chromosome-specific probes revealed complete donor chimerism of bone marrow cells on Day þ28. Because of high risk of disease relapse after transplantation, cyclosporine was tapered rapidly and finished on Day þ70. Thereafter, mild chronic GVHD of the liver and skin developed at 4 months after BMT, requiring treatment with oral cyclosporine for 8 months. At 5 years post-BMT, bone marrow examination revealed complete donor chimerism by sex mismatched FISH. Seven years after BMT, the patient is alive and free of disease. Treatment modalities for HSTCL have considerable heterogeneity [3], including splenectomy, corticosteroids, purine analogue, anthracycline containing regimens such as CHOP (cyclophosphamide, hydroxydaunomycin, vincristine and prednisone) or CHOP-like regimen, second or third generation aggressive lymphoma regimen such as IEV (ifosphamide, epirubicin and etoposide) or modified MACOP-B (methotrexate, etoposide instead of adriamycin, cyclophosphamide, vincristine, predonisone and bleomycin), alemtuzumab and autologous and allogeneic stem cell transplantation. However, such treatments have limited efficacy and the vast majority of patients will die from progressive disease. Although allogeneic SCT, which is the only potentially curative therapy, has been attempted to treat

MT1-MMP plays a critical role in hematopoiesis by regulating HIF-mediated chemokine/cytokine gene transcription within niche cells

HSC fate decisions are regulated by cell-intrinsic and cell-extrinsic cues. The latter cues are derived from the BM niche. Membrane-type 1 matrix metalloproteinase (MT1-MMP), which is best known for its proteolytic role in pericellular matrix remodeling, is highly expressed in HSCs and stromal/niche cells. We found that, in MT1-MMP(-/-) mice, in addition to a stem cell defect, the transcription and release of kit ligand (KitL), stromal cell-derived factor-1 (SDF-1/CXCL12), erythropoietin (Epo), and IL-7 was impaired, resulting in a trilineage hematopoietic differentiation block, while addition of exogenous KitL and SDF-1 restored hematopoiesis. Further mechanistic studies revealed that MT1-MMP activates the hypoxia-inducible factor-1 (HIF-1) pathway via factor inhibiting HIF-1 (FIH-1) within niche cells, thereby inducing the transcription of HIF-responsive genes, which induce terminal hematopoietic differentiation. Thus, MT1-MMP in niche cells regulates postnatal hematopoiesis, by modulating hematopoietic HIF-dependent niche factors that are critical for terminal differentiation and migration.

Unrelated cord blood transplantation after myeloablative conditioning in adults with advanced myelodysplastic syndromes

We analyzed the disease-specific outcomes of adult patients with advanced myelodysplastic syndrome (MDS) treated with cord blood transplantation (CBT) after myeloablative conditioning. Between August 1998 and June 2009, 33 adult patients with advanced MDS were treated with unrelated CBT. The diagnoses at transplantation included refractory anemia with excess blasts (n=7) and MDS-related secondary AML (sAML) (n=26). All patients received four fractionated 12 Gy TBI and chemotherapy as myeloablative conditioning. The median age was 42 years, the median weight was 55 kg and the median number of cryopreserved nucleated cells was 2.51 × 107 cells per kg. The cumulative incidence of neutrophil recovery at day 50 was 91%. Neutrophil recovery was significantly faster in sAML patients (P=0.04). The cumulative incidence of plt recovery at day 200 was 88%. Plt recovery was significantly faster in CMV seronegative patients (P<0.001). The cumulative incidence of grade II–IV acute GVHD (aGVHD) and extensive-type chronic GVHD was 67 and 34%, respectively. Degree of HLA mismatch had a significant impact on the incidence of grade II–IV aGVHD (P=0.021). TRM and relapse at 5-years was 14 and 16%, respectively. The probability of EFS at 5 years was 70%. No factor was associated with TRM, relapse and EFS. These results suggest that adult advanced MDS patients without suitable related or unrelated BM donors should be considered as candidates for CBT.

Unrelated cord blood transplantation after myeloablative conditioning in adults with ALL.

We analyzed the disease-specific outcomes of adult ALL treated with cord blood transplantation (CBT) after myeloablative conditioning. Between October 2000 and November 2007, 27 adult patients with ALL were treated with unrelated CBT. All patients received four fractionated 12 Gy TBI and chemotherapy as myeloablative conditioning. The median age was 36 years, the median weight was 57 kg and the median number of nucleated cells was 2.47 × 107/kg. All patients received a single and HLA-mismatched cord blood unit. The cumulative incidence of neutrophil recovery at day 30 and platelet recovery at day 200 was 92.6 and 92.3%, respectively. With a median follow-up of 47 months, the probability of EFS at 5 years was 57.2%. The 5-year cumulative incidence of TRM and relapse was 3.7 and 27.4%, respectively. These results suggest that unrelated CBT after myeloablative conditioning could be safely and effectively used for adult patients with ALL.

Impact of ABO incompatibility on engraftment and transfusion requirement after unrelated cord blood transplantation: a single institute experience in Japan.

The impact of ABO incompatibility between donor and recipient on engraftment and transfusion requirement was studied in 95 adults who underwent unrelated cord blood transplantation (CBT). The patients included 27 ABO-identical, 29 minor, 21 major and 18 bidirectional ABO-incompatible recipients. Neutrophil engraftment did not differ between ABO-identical/minor ABO-incompatible and major/bidirectional ABO-incompatible recipients (hazard ratio (HR) 1.17, P=0.48). Cumulative incidence of platelet engraftment in ABO-identical/minor ABO-incompatible recipients was higher than in major/bidirectional ABO-incompatible recipients (HR 1.88, P=0.013). In addition, fewer platelet transfusions were required during the first 60 days after CBT in ABO-identical/minor ABO-incompatible recipients (HR 0.80, P=0.040). RBC engraftment did not differ between the two groups (HR 1.25, P=0.33). However, fewer RBC transfusions were required in ABO-identical/minor ABO-incompatible recipients than in major/bidirectional ABO-incompatible recipients (HR 0.74, P<0.005). No patients developed pure red-cell aplasia after CBT. These results indicate that ABO incompatibility affected platelet engraftment and transfusion requirement of RBC and platelet in CBT recipients. Further studies including larger patient numbers are required to elucidate the impact of ABO incompatibility on the clinical outcome of CBT.

Cardiovascular toxicity of cryopreserved cord blood cell infusion.

Although infusion of cryopreserved bone marrow or peripheral blood stem cell is associated with a variety of symptoms, there have been no reports detailing the data of infusion-related toxicities of cryopreserved cord blood (CB) units. We prospectively evaluated the incidence and significance of infusion-related toxicities in 34 adult patients undergoing unrelated CB transplantation. Cryopreserved CB units were thawed and immediately infused, unfiltered, through a central intravenous catheter without further manipulation. Heart rate, blood pressure, oxygen saturation and clinical symptoms were recorded during and after infusion. Twenty-four percent of patients experienced non-cardiovascular toxicities related to infusion. The incidence of systolic and diastolic hypertension and bradycardia was 58, 64 and 32%, respectively. Although three patients (9%) with severe systolic hypertension after the infusion required treatment with antihypertensive agents, no patients experienced life-threatening side effects or needed discontinuation of CB unit infusion. Patient or transplant characteristics had no effect on the hypertension and bradycardia related to the infusion of CB. These data suggest that infusion of cryopreserved CB without further manipulation after thawing is safe and well tolerated. However, cardiovascular toxicities including hypertension and bradycardia were frequently observed.

Inhibition of plasmin attenuates murine acute graft-versus-host disease mortality by suppressing the matrix metalloproteinase-9-dependent inflammatory cytokine storm and effector cell trafficking

The systemic inflammatory response observed during acute graft-versus-host disease (aGVHD) is driven by proinflammatory cytokines, a ‘cytokine storm’. The function of plasmin in regulating the inflammatory response is not fully understood, and its role in the development of aGVHD remains unresolved. Here we show that plasmin is activated during the early phase of aGVHD in mice, and its activation correlated with aGVHD severity in humans. Pharmacological plasmin inhibition protected against aGVHD-associated lethality in mice. Mechanistically, plasmin inhibition impaired the infiltration of inflammatory cells, the release of membrane-associated proinflammatory cytokines including tumor necrosis factor-α (TNF-α) and Fas-ligand directly, or indirectly via matrix metalloproteinases (MMPs) and alters monocyte chemoattractant protein-1 (MCP-1) signaling. We propose that plasmin and potentially MMP-9 inhibition offers a novel therapeutic strategy to control the deadly cytokine storm in patients with aGVHD, thereby preventing tissue destruction.

Donor cell-derived myelodysplastic syndrome after cord blood transplantation.

Donor cell-derived hematological malignancy is a rare complication after allogeneic SCT. Earlier studies reported that 0.12–5% of patients developed donor cell leukemia (DCL) after allogeneic SCT.1, 2 Recently, several reports have shown that donor cell-derived hematological malignancy occurred in patients after cord blood transplantation (CBT).3, 4, 5, 6, 7, 8, 9 Here, we report an adult case with ALL that subsequently developed donor cell-derived myelodysplastic syndrome (MDS) after CBT.