Received Bone Marrow Research Articles

Effects of mesenchymal stem cells on pulmonary fibrosis

Objective To investigate the effects of mesenchymal stem cells (MSCs) on pulmonary fibrosis under different conditions (different types, different doses and different transplantation times in the same dose). Methods Seventy-two male SD rats were equally divided into 6 groups by random number table method: Group A received intratracheal instillation of NS (0.1 ml). Group B, C, D, E and F received intratracheal instillation of bleomycin (5 mg/kg in 0.1 ml NS). On the 1st day after bleomycin administration, Group A and B received NS (1.0 ml) via the tail vein, while Group C received bone marrow mesenchymal stem cells (BMSCs) 1×106, Group D received umbilical cord blood mesenchymal stem cells (UCB-MSCs) 1×106, Group E received BMSCs 5×105 and Group F received BMSCs 1×107 via the tail vein, respectively. On the 8th day after bleomycin administration, Group E received BMSCs 5×105 again. Half of rats in each group were sacrificed respectively on the 28th and 42nd day after bleomycin administration. The pathologic changes, transforming growth factor-β1 (TGF-β1), hydroxyproline (HYP), matrix metalloproteinase-2 (MMP-2) and tissue inhibitor of metalloproteinase-1 (TIMP-1) of the lung tissues were investigated. Results Alveolar structure was severely in disorder, and massive collagenous fiber was deposited in Group B. In Group B, C, D, E, F and A, the scores of pulmonary fibrosis were decreased in sequence [(3.00±0.00), (2.17±0.75), (1.60±0.89), (1.33±0.52), (1.00±0.00), (0.00±0.00) on the 28th day after bleomycin administration, and (3.00±0.00), (2.40±0.55), (1.75±0.96), (1.50±0.84), (-), (0.00±0.00) on the 42nd day after bleomycin administration], while the levels of TGF-β1, HYP and MMP-2/TIMP-1 were respectively decreased in sequence as above. The levels of TGF-β1, HYP and MMP-2/TIMP-1 were positively associated with pulmonary alveolitis and fibrosis scores respectively (P=0.000). Conclusion MSCs transplantation can suspend the progress of pulmonary fibrosis, and the mechanism may be related to reducing the level of TGF-β1 and improving the imbalance of MMP/TIMP. Choosing UCB-MSCs or multiple transplantation may improve the therapeutic effects of pulmonary fibrosis. Key words: Pulmonary fibrosis; Bleomycin; Bone marrow mesenchymal stem cells; Umbilical cord blood mesenchymal stem cells

515 - Comparison of T & NK Cell Donor Chimerism & Hematopoietic Engraftment Following Allogeneic Hematopoetic Stem Cell Transplantation in Pediatric Recipients Receiving Bone Marrow (BM) or Umbilical Cord Blood and/or Human Placental Derived Stem Cells (HPDSC)

515 - Comparison of T & NK Cell Donor Chimerism & Hematopoietic Engraftment Following Allogeneic Hematopoetic Stem Cell Transplantation in Pediatric Recipients Receiving Bone Marrow (BM) or Umbilical Cord Blood and/or Human Placental Derived Stem Cells (HPDSC)

Expressions of CD96 and CD123 and Their Relationship with Prognosis of Patients with Myelodysplastic Syndrome

To explore the relationship between expression of CD96 and CD123 and prognosis of patients with myelodysplastic syndrome(MDS). Eight-nine MDS patients(MDS group) and 20 persons without hematologic disease as controls(Control group) were enrolled. The patients were grouped by the risk. All participants received bone marrow biopsy. Mononuclear cells were extracted, CD34+CD38-CD123+ and CD34+CD38-CD96+ cells were counted by using flow cytometry. Expressions of 2 type cells in control group, MDS group and its subgroups were analyzed. The proportion of CD34+ cells and CD34+CD38- cells in mononuclear cells of patients in MDS group was higher than in control group (P<0.05). The proportions of CD34+CD38-CD123+ cells and CD34+CD38-CD96+ cells in CD34+CD38- cells were significantly higher than that in control group(P<0.05) and the proportion increased with the risk. In the low-and middle-risk group, the rates of complete remission(CR) and partial remission(PR) of patients with CD123- and CD96- were higher than those in patients with CD123+ and CD96+; in the middle-2 and high risk patients, the PR of patients with CD123- was higher than that in patients with CD123+(P<0.05). The CR rate of patients with CD96- was higher than that of patients with CD96+(P<0.05). The differentiation of CD34+ cells in bone marrow of MDS patients is abnormal, and the high expression of CD123 and CD96 cells existes. These findings may partially explain the cause of hematopoietic stem cell malignant clone in MDS patients.

Unrelated Donor Transplantation in Children with Thalassemia using Reduced-Intensity Conditioning: The URTH Trial

Allogeneic hematopoietic stem cell transplantation (HSCT) can cure transfusion-dependent thalassemia (TDT). In a multicenter trial we investigated the efficacy of reduced-intensity conditioning (RIC) before unrelated donor (URD) HSCT in children with TDT. Thirty-three children, ages 1 to 17 years, received bone marrow (BM) or umbilical cord blood (UCB) allografts. Median time to neutrophil engraftment was 13 days (range, 10 to 25) and 24 days (range, 18 to 49) and platelet engraftment 23 days (range, 12 to 46) and 50 days (range, 31 to 234) after BM and UCB allografts, respectively. With a median follow-up of 58 months (range, 7 to 79), overall and thalassemia-free survival was 82% (95% CI, .64% to .92%) and 79% (95% CI, .6% to .9%), respectively. The cumulative incidence of grades II to IV acute graft-versus-host disease (GVHD) after BM and UCB allografts was 24% and 44%; the 2-year cumulative incidence of chronic extensive GVHD was 29% and 21%, respectively; 71% of BM and 91% of UCB recipients discontinued systemic immunosuppression by 2 years. Six patients who had Pesaro risk class 2 (n = 5) and class 3 (n = 1) died of GVHD (n = 3), viral pneumonitis (n = 2) and pulmonary hemorrhage (n = 1). Outcomes after this RIC compared favorably with URD HSCT outcomes for TDT and supported engraftment in 32 of 33 patients. Efforts to reduce GVHD and infectious complications are being pursued further.

Role of Infiltrating Monocytes in the Development of Radiation-Induced Pulmonary Fibrosis.

Lung exposure to radiation induces an injury response that includes the release of cytokines and chemotactic mediators; these signals recruit immune cells to execute inflammatory and wound-healing processes. However, radiation alters the pulmonary microenvironment, dysregulating the immune responses and preventing a return to homeostasis. Importantly, dysregulation is observed as a chronic inflammation, which can progress into pneumonitis and promote pulmonary fibrosis; inflammatory monocytes, which are bone marrow derived and express CCR2, have been shown to migrate into the lung after radiation exposure. Although the extent to which recruited inflammatory monocytes contribute to radiation-induced pulmonary fibrosis has not been fully investigated, we hypothesize that its pathogenesis is reliant on this population. The CC chemokine ligand, CCL2, is a chemotactic mediator responsible for trafficking of CCR2+ inflammatory cells into the lung. Therefore, the contribution of this mediator to fibrosis development was analyzed. Interleukin (IL)-1β, a potent pro-inflammatory cytokine expressed during the radiation response, and its receptor, IL-1R1, were also evaluated. To this end, CCR2-/-, IL-1β-/- and IL-1R1-/- chimeric mice were generated and exposed to 12.5 Gy thoracic radiation, and their response was compared to wild-type (C57BL/6) syngeneic controls. Fibrotic foci were observed in the periphery of the lungs of C57 syngeneic mice and CCR2-/- recipient mice that received C57 bone marrow (C57 > CCR2-/-) by 16 and 12 weeks after irradiation, respectively. In contrast, in the mice that had received bone marrow lacking CCR2 (CCR2-/- > C57 and CCR2-/- syngeneic mice), no pulmonary fibrosis was observed at 22 weeks postirradiation. This observation correlated with decreased numbers of infiltrating and interstitial macrophages compared to controls, as well as reduced proportions of pro-inflammatory Ly6C+ macrophages observed at 12-18 weeks postirradiation, suggesting that CCR2+ macrophages contribute to radiation-induced pulmonary fibrosis. Interestingly, reduced proportions of CD206+ lung macrophages were also present at these time points in CCR2-/- chimeric mice, regardless of donor bone marrow type, suggesting that the phenotype of resident subsets may be influenced by CCR2. Furthermore, chimeras, in which either IL-1β was ablated from infiltrating cells or IL-1R1 from lung tissues, were also protected from fibrosis development, correlating with attenuated CCL2 production; these data suggest that IL-1β may influence chemotactic signaling after irradiation. Overall, our data suggest that CCR2+ infiltrating monocyte-derived macrophages may play a critical role in the development of radiation-induced pulmonary fibrosis.

Haploidentical T-Repleted Stem Cell Transplantation (SCT) Has Comparable Survival to 10/10 and 9/10 Unrelated SCT in Poor-Cytogenetics Risk Acute Myeloid Leukemia in First Complete Remission: A Study on Behalf of the Acute Leukemia Working Party (ALWP) of the European Society for Blood and Marrow Transplantation (EBMT)

Background: Allogeneic stem cell transplantation (SCT) is the most powerful therapy to prevent relapse in poor-cytogenetics risk acute myeloid leukemia (poorAML) patients (pts) in first complete remission (CR1). For pts who lack a matched related donor (MRD), SCT from an unrelated (UD) or alternative donor is indicated. Pts with poorAML and thus at high risk of relapse can theoretically benefit the most from SCT from haploidentical donors (HaploSCT), which is an attractive option as the time required to find a well-matched UD could be inacceptable. Several recent reports show comparable outcomes between HaploSCT and transplants from UD (Piemontese S, JHO 2017; Versluis J, Blood Advances 2017). Comparative studies able to include sufficient numbers of pts with poorAML in CR1 are limited; this prompted us to compare the outcomes of HaploSCT to those of 10/10 and 9/10 HLA-matched UD in this disease category.Methods: We retrospectively selected denovo poorAML pts in CR1 receiving T-repleted haplo (n=74), 10/10 UD (n=433) and 9/10 UD SCT (n=123) from 2007 to 2015 who were reported to the ALWP of EBMT Registry. PoorAML was defined as the presence of: complex karyotype (at least 3 structural abnormalities per clone); monosomal karyotype (1 autosomal monosomy plus 1 monosomy or structural abnormality); inv(3)/t(3;3); -5 or del(5q); -7 or abn(7q); t(v;11)(v;q23); abn(17p); t(6;9); t(9;22). Primary endpoints were leukemia-free survival (LFS) and overall survival (OS). Secondary endpoints were acute and chronic GVHD (aGVHD and cGVHD), relapse and nonrelapse mortality (NRM).Results: Main population characteristics are depicted in Table 1. Recipients of haplo-, 10/10 UD- and 9/10 UD-SCT were comparable concerning time from diagnosis to SCT and time from CR1 to SCT. HaploSCT more likely received bone marrow as stem cell source. In-vivo T cell depletion (TCD) with ATG was most likely adopted in UD-SCT, with a conversely increased use of high-dose post-transplant Cyclophosphamide as GvHD prophylaxis backbone in HaploSCT (65% Vs 3% for 10/10 UD and 2% for 9/10 UD, p<10¯³).LFS and OS at 2 years were not significantly different between haploSCT, 10/10 UD SCT and 9/10 UD SCT (53±12% and 59±12%, 43±5% and 50±6%, 44±9% and 50±9%, respectively, p=0.5 and p=0.5, respectively).In Haplo-SCT, the 100-day cumulative incidence (CI) of grade≥2 aGvHD was in line with the one reported for 10/10 and 9/10 UD (33±11% for haplo, 30±4% for 10/10 UD and 34±9% for 9/10 UD, p=0.6). Likewise, the 2-y CI of cGvHD (35±12%) of HaploSCT was similar to those of 10/10 UD (36±4%) and 9/10 UD (36±9%), p=0.8. The 2-y CI of NRM was 19±8% after haploSCT, 18±4% after 10/10 UD SCT and 18±6% after 9/10 UD SCT (p=0.9). Relapse incidence was not significantly affected by donor source, with a 2-y CI of 27±9% for haploSCT, 39±5% for 10/10 UD SCT and 37±9% for 9/10 UD SCT (p=0.3).After adjustment for centre effect, pts age, time from diagnosis to SCT, conditioning intensity, in-vivo TCD, donor/pts gender and CMV serostatus, the multivariable model showed that haploSCT recipients didn't experience worse outcomes compared to 10/10 and 9/10 UD. Indeed, compared to haploSCT (reference) the hazard ratio (HR) for LFS was 1.2 for 10/10 UD (p=0.3) and 1.2 for 9/10 UD (p=0.4). The hazards for OS in 10/10 and 9/10 UD did not differ from haplo-SCT (1.3, p 0.3 and 1.2, p 0.4, respectively). Moreover, compared to haplo, SCT from 10/10 and 9/10 UD was not associated with lower hazards for relapse (HR: 1.4, p=0.2 and 1.4, p=0.3, respectively), NRM (HR: 1, p=0.9 and 1, p=0.9, respectively), grade≥2 aGvHD (HR: 1.2, p=0.6 and 1.4, p=0.3, respectively) and cGvHD (HR: 1.2, p=0.5 and 1.3, p=0.4, respectively). The only factor associated with worse LFS and OS was pts age (for each 10-year interval: HR 1.1, p=0.02 and 1.2, p=0.001, respectively).Conclusions: In the present series of poorAML pts transplanted in CR1, haploSCT recipients experienced comparable outcomes with respect to 10/10 and 9/10 HLA-matched UDs. This suggests that the immunotherapeutic effect of allogeneic SCT is exerted similarly across these different donor sources in this peculiar population. Therefore, in the absence of a MRD, pts with poor risk cytogenetics who have a very high risk of relapse could be allocated to haploSCT in their first remission, especially in the context of the recent improvements, which fostered an abatement of GvHD and NRM rates, historically the main detrimental factors for Haploidentical transplants. [Display omitted] DisclosuresCiceri:GSK: Other: B-thalassemia gene therapy was developed by Fondazione Telethon and Ospedale San Raffaele and has been inlicenced by GSK that provides funding for the clinical trial, Research Funding. Mohty:Sanofi: Honoraria, Speakers Bureau.

Early Transplant Improves Outcomes in Patients with Chronic Granulomatous Disease: Results from United States Immunodeficiency Network (USIDNET)

Introduction: Chronic granulomatous disease (CGD) is a primary immunodeficiency due to defective nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, resulting in failure of killing intracellular pathogens. Clinical presentation and management of these patients remains heterogeneous. Current management involves non-transplant therapy with interferon-gamma and supportive care, or hematopoietic stem cell transplant (HSCT) in severe cases. There has been limited data on morbidity and outcomes with different treatment options using patient registries. There has been no registry-based study of CGD patients from the United States comparing different treatment options.Methods: We performed a multi-institutional analysis of data from the United States Immunodeficiency Network (USIDNET) registry, comparing morbidity, mortality, and different treatment options. Retrospective review of data from its inception in 1992 through 2016 was performed. We analyzed demographic, co-morbidity and treatment information (HSCT versus non-transplant therapy). Logistic regression was used to compare overall survival (OS), transplant related survival (TRS) and frequency of serious infections and co-morbidities among the treatment groups. Multivariate logistic regression was used to evaluate factors affecting survival.Results: Within the database, 507 CGD patients were identified, with 84% male. X-linked inheritance was most common (73%) with about 78% of patients having gp91 mutation. Of 507 patients, about 10% underwent HSCT (n=50). Overall mortality was 15% with relatively lower incidence in HSCT group (12%). In HSCT cohort, median age at transplant was 13.5 years (0.6-37.3 years), with 74% receiving reduced intensity conditioning (RIC). The most common graft source was peripheral blood stem cells (60%) from a matched-unrelated donor (72%). No mortality was reported in patients who received stem cells from matched siblings and in those who received bone marrow graft with RIC, at median follow-up of 24 months (range 3-402 months). Survival analysis included 302 patients. OS was not significantly different between HSCT and non-transplant groups (p=0.50; Figure 1 a, b). Among patients who underwent HSCT, there was significantly improved TRS in those transplanted before 14 years of age (p= 0.035, HR: 4.51; Figure 1 c, d). The proportion of patients who had >3 infections prior to transplant was significantly reduced by 2-years post-transplant (p < 0.0001, Figure 2). The rate of serious infections prior to transplant was significantly lower in patients transplanted at <14 years old compared to those transplanted at an older age (p< 0.001, 95% CI 19.2- 62.8). Stem cell source, graft type, conditioning regimen, and prior gamma-interferon treatment did not differ significantly between these two groups. Ten of 13 patients who had gastrointestinal granulomatous inflammation prior to transplant did not have documentation of disease at 1-35 months post-HSCT, with an additional subject experiencing relief of esophageal stricture post-transplant. In univariate analysis, patients with gp91 mutation had significantly worse survival compared to other mutations (p=0.008). In multivariate analysis for OS, gp91 mutation showed a trend towards significance within the entire cohort (p = 0.08, Table 1). Figure 3 shows comparison of mean quality of life (QoL) indicator for non-transplant (n=40) and post-transplant patients (n=12). The mean performance score was significantly higher in post-HSCT (93.33) patients compared to their non-transplant counterparts (84.73) for subjects 15 years or older (p=0.0021).Conclusion: This is the largest reported American CGD cohort. HSCT at <14 years of age is associated with improved survival, decreased burden of serious infections, and granulomatous inflammation. Post transplant CGD patients have significantly better QoL as compared to non-transplant ones. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Ten Years HIV Free: An Interview with “The Berlin Patient,” Timothy Ray Brown

It was just about 10 years ago that Timothy Ray Brown was cured of HIV infection by receiving bone marrow stem cell transplantations from an HLA matched donor who also was homozygous for the CCR5 delta 32 mutation that renders immune cells resistant to infection by most HIV viruses. During an October 2017 visit to Cleveland to help launch a number of clinical trials targeting the Cure, Timothy sat down with Pathogens and Immunity editor Michael Lederman and community activist Earl Pike to talk about his experiences.

Assessment of TREC, KREC and telomere length in long-term survivors after allogeneic HSCT: the role of GvHD and graft source and evidence for telomere homeostasis in young recipients.

Reconstitution of the adaptive immune system following allogeneic hematopoietic stem cell transplantation is crucial for beneficial outcome and is affected by several factors, such as GvHD and graft source. The impact of these factors on immune reconstitution has been thoroughly investigated during the early phase after transplantation. However, little is known about their long-term effect. Similarly, leukocyte telomere length (TL) shortening has been reported shortly after transplantation. Nevertheless, whether TL shortening continues in long-term aspect is still unsettled. Here, we assessed T-cell receptor excision circle (TREC), kappa deleting recombination excision circle (KREC) and leukocyte TL in recipients and donors several years post transplantation (median 17 years). Our analysis showed that, recipients who received bone marrow (BM) as the graft source have higher levels of both TREC and KREC. Also, chronic GvHD affected TREC levels and TL but not KREC levels. Finally, we show that recipient's TL was longer than respective donors in a group of young age recipients with high KREC levels. Our results suggest that BM can be beneficial for long-term adaptive immune recovery. We also present supporting evidence for recipient telomere homeostasis, especially in young age recipients, rather than telomere shortening.

Analysis of Clinical Characteristics and Prognosis of Mantle Cell Lymphoma With Gastrointestinal Involvement

Introduction: Mantle cell lymphoma (MCL) is an aggressive B-cell lymphoma characterized by rapid disease progression. Studies on gastrointestinal (GI) involvement of MCL are lacking. In this study, we investigated the clinical features and prognosis of MCL with GI involvement. Methods: We retrospectively analyzed 64 patients who diagnosed MCL from January 2009 to April 2017 consecutively. At the time of MCL diagnosis, patients who were identified to have GI involvement by endoscopy or radiologic examination were assigned to the GI-MCL group. And those who were confirmed to have no GI involvement were assigned to the non GI-MCL group. The clinical characteristics and the outcomes of both groups were compared. Results: The GI-MCL group included 29 patients (45.3%). There were no significant differences in age and gender between the GI and non GI-MCL group. The stage of GI-MCL group was higher compared to the non GI-MCL group and all of 29 patients were stage IV. The mean of International Prognostic Index (IPI) score was also significantly higher in GI-MCL (2.97 vs. 1.97, p=0.047). Bone marrow involvement in the GI-MCL group was much greater than non GI-MCL group (69.0% vs. 40.0%, p=0.021). R-CHOP (cyclophosphamide, doxorubicin, vincristine and prednisolone combined with rituximab) was the most abundant in 1st line chemotherapy (44/64, 69.8%). After 1st line chemotherapy, 56 of 64 patients showed complete remission (87.5%). Although there was no difference in complete remission rate between the two groups, recurrence rate was much higher in GI-MCL group (72.% vs. 41.2%, p=0.024). A total of 15 patients received bone marrow transplantation (15/64, 23.4%). The median follow up period was 37 months and 11 out of 64 patients died during this period (17.2%). There was no significant difference in mortality between the two groups. Conclusion: The patients with MCL invading GI tract tended to have advanced stage, higher IPI score, more bone marrow involvement and more recurrence than MCL not involving the GI tract. Endoscopic examination to rule out GI involvement in patients with MCL is warranted.

Diagnosis and treatment of adult acute myeloid leukemia with NPM-MLF1 fusion gene and literatures review

Objective To evaluate the clinical features, treatment and prognosis of acute myeloid leukemia (AML) with nucleophosmin (NPM)-myelodysplasia/myeloid leukemia factor (MLF)1 fusion gene. Methods On 16 February 2016, a case of AML patient with NPM-MLF1 fusion gene who was admitted to Changhai Hospital Affiliated to Second Military Medical University was included in this study. When the patient was admitted, bone marrow morphology, cytochemical staining and immunophenotype analysis of leukemia blasts, cytogenetics and molecular genetic analysis were conducted and diagnosis was made base on those test results. On 17 February 2016, the patient recieved two courses of daunorubicin+ arabinosylcytosin (DA) 3+ 7 induction therapy. Subsequently, the patient recieved one course of fludarabine, cytarabine and granulocyte colony-stimulating factor (FLAG) regimen and 5 courses of high-dose of cytarabine. In the meantime, bone marrow morphology and minimal residual disease (MRD) were conducted during the chemotherapy. On 6 April 2017, the patient received bone marrow morphology, immunophenotype analysis of leukemia blasts and molecular genetic analysis on her visit. The clinical features, diagnosis and treatment of this patient were analyzed retrospectively, and the related literatures were reviewed. Results ①When the patient was admitted, bone marrow morphology revealed hyper-cellular marrow with 48.0% leukemia blasts. Cytochemical staining showed that peroxidase (POX) staining in some leukemia blasts were positive, and naphthol AS-D chloroacetate esterase (CE) staining in a few leukemia blasts were positive, and non-specific estertase (NSE) staining in some leukemia blasts were positive. The results of immunophenotype analysis revealed the ratio of the blasts cell was about 39.9%, and the ratio of immature cell was about 13.6%. R banding karyotype analysis marked with 46, XX, t(3; 5)(q25; q34)[3]/46, XX[7]. Molecular genetic analysis showed NPM/MLF1 fusion gene was positive. According to above, the diagnosis of patient was AML-subtype M4 with NPM-MLF1 fusion gene. ②After the patient was treated with the first course of induction therapy, and the reexamined bone marrow morphology post therapy showed partial remission (PR). The patient received complete remission (CR) after the second course of induction therapy. After the patient received consolidation therapy, and bone marrow morphology showed the patient obtained sustained CR status. On the patient′s last visit, bone marrow morphology revealed a total of 25.0% leukemia blasts, consisting of 21.0% blasts and 4.0% immature cell, suggesting AML recurrenced. Immunophenotyping of leukemia blasts showed approximately 28.5% of the early myeloid cells in bone marrow which was consistent with AML recurrence. Molecular genetic analysis showed that the MPM-MLF1 fusion gene was positive (66.1%). During the treatment, bone marrow MRD decreased, then increased. The patient went back to local hospital for treatment after AML relapsed. Conclusions AML-subtype M4 patient with NPM-MLF1 fusion gene can achieve CR after chemotherapy, but prone to early recurrence, and NPM-MLF1 fusion gene may be a poor prognostic factor for AML. Key words: Leukemia, myeloid, acute; Drug therapy; NPM-MLF1; t (3;5)

Evaluation of the diagnostic performance of panfungal polymerase chain reaction assay in invasive fungal diseases.

Timely diagnosis of invasive fungal diseases (IFDs) is important, as delays in treatment initiation are associated with increased mortality rates. However, early diagnosis of IFDs in immunocompromised patients remains difficult. The conventional diagnostic methods currently used for IFDs are not sufficiently effective. Molecular tests, such as polymerase chain reaction (PCR)-based assays, have great potential to improve the early diagnosis of IFDs due to their sensitivity and specificity. In the present study, the diagnostic performance of panfungal PCR assays in IFD patients who received bone marrow transplantation was evaluated. The results suggested that panfungal PCR assay offered a quick and convenient guide for clinical decision-making by identifying higher numbers of fungal species in comparison with the conventional blood culture method. Furthermore, panfungal PCR assay exhibited a sensitivity of 93% and a specificity of 71% in the diagnosis of IFD patients based on the EORTC/MSG criteria. Thus, the present study concluded that the reported PCR-based method was effective and sensitive in early IFD diagnosis and should be integrated into clinical decision-making for the treatment of IFDs in the future.

Novel mutations of TCIRG1 cause a malignant and mild phenotype of autosomal recessive osteopetrosis (ARO) in four Chinese families

Human autosomal recessive osteopetrosis (ARO), also known as infantile malignant osteopetrosis, is a rare genetic bone disorder that often causes death. Mutations in T-cell immune regulator 1 (TCIRG1) are a frequent cause of human ARO. Six additional genes (TNFSF11, TNFRSF11A, CLCN7, OSTM1, SNX10, PLEKHM1) were also found to be associated with human ARO. In order to expand the mutation spectrum and clinical diversity for a better understanding of the ARO phenotype and to further investigate the clinical characteristics of benign subjects with ARO, we here report five individuals with ARO from four unrelated Chinese families. X-ray examination was conducted and bone turnover markers were assayed. The gene of T-cell immune regulator 1 (TCIRG1) was screened and analyzed. Monocyte-induced osteoclasts were prepared and their resorption ability was studied in vitro. We identified five novel mutations (c.66delC, c.1020+1_1020+5dup, c.2181C>A, c.2236+6T>G, c.692delA) in these patients. Four patients displayed a malignant phenotype, three of them died, and one who received bone marrow transplantation survived. The remaining one, a 24-year-old male from a consanguineous family, was diagnosed based on radiological findings but presented no neurological or hematological defects. He was homozygous for c.2236+6T>G in intron 18; this mutation influenced the splicing process. An in vitro functional study of this novel splicing defect showed no resorption pits on dentine slices. TCIRG1-dependent osteopetrosis with a mild clinical course was observed for the first time in Chinese population. The present findings add to the wide range of phenotypes of Chinese patients with TCIRG1-dependent ARO and enrich the database of TCIRG1 mutations.

Periodontitis in Chédiak-Higashi Syndrome: An Altered Immunoinflammatory Response.

Chédiak-Higashi syndrome (CHS), a rare autosomal recessive disorder caused by mutations in the lysosomal trafficking regulator gene (LYST), is associated with aggressive periodontitis. It is suggested that LYST mutations affect the toll-like receptor (TLR)-mediated immunoinflammatory response, leading to frequent infections. This study sought to determine the periodontal status of patients with classic (severe) and atypical (milder) forms of CHS and the immunoregulatory functions of gingival fibroblasts in CHS patients. In contrast to aged-matched healthy controls, atypical (n = 4) and classic (n = 3) CHS patients presented with mild chronic periodontitis with no evidence of gingival ulceration, severe tooth mobility, or premature exfoliation of teeth. As a standard of care, all classic CHS patients had undergone bone marrow transplantation (BMT). Primary gingival fibroblasts obtained from atypical and BMT classic CHS patients displayed higher protein expression of TLR-2 (1.81-fold and 1.56-fold, respectively) and decreased expression of TLR-4 (-2.5-fold and -3.85-fold, respectively) at baseline when compared with healthy control gingival fibroblasts. When challenged with whole bacterial extract of Fusobacterium nucleatum, both atypical and classic CHS gingival fibroblasts failed to up-regulate TLR-2 and TLR-4 expression when compared with their respective untreated groups and control cells. Cytokine multiplex analysis following F. nucleatum challenge showed that atypical CHS gingival fibroblasts featured significantly increased cytokine expression (interleukin [IL]-2, IL-4, IL-5, IL-6, IL-10, IL-12, interferon-γ, tumor necrosis factor-α), whereas classic CHS cells featured similar/decreased cytokine expression when compared with treated control cells. Collectively, these results suggest that LYST mutations in CHS patients affect TLR-2 and TLR-4 expression/function, leading to dysregulated immunoinflammatory response, which in turn may influence the periodontal phenotype noted in CHS patients. Furthermore, our results suggest that atypical CHS patients and classic CHS patients who undergo BMT early in life are less susceptible to aggressive periodontitis and that hematopoietic cells play a critical role in mitigating the risk of aggressive periodontitis in CHS. Knowledge Transfer Statement: Results from this study can be used to create awareness among clinicians and researchers that not all CHS patients exhibit historically reported aggressive periodontitis, especially if they have atypical CHS disease or have received bone marrow transplantation. LYST mutations in CHS patients may affect TLR-2 and TLR-4 expression/function leading to dysregulated immunoinflammatory response, which in turn may influence the periodontal phenotype noted in CHS patients.

ORAL MANIFESTATION OF MUCORMYCOSIS IN A BONE MARROW TRANSPLANTED PATIENT

Mucormycosis is an aggressive fungal infection that usually affects patients with immune disorders. The most important form is the rhino-orbito-cerebral mucormycosis that may demonstrate oral cavity involvement. We describe an original case of mucormycosis affecting a 61-year-old male patient that received bone marrow transplantation because of a B-cell acute lymphocytic leukemia. The patient was referred to our department with an extensive, painful, necrotic lesion in the left soft palate that contained a thick whitish membrane surrounded by black areas, consistent with necrosis. Biopsy was done and microscopic analysis demonstrated hyperplastic epithelium with the connective tissue revealing moderate inflammatory infiltrate, extensive tissue necrosis, and numerous irregular, broad, non-septate hyphae that branched in right angle frequently invading blood vessels. Grocott and PAS reactions were positive. These findings confirmed the diagnosis of mucormycosis. The lesion was surgically removed and the patient received antifungal, but died 2 months after diagnosis.

Mediator 1 Is Atherosclerosis Protective by Regulating Macrophage Polarization.

MED1 (mediator 1) interacts with transcription factors to regulate transcriptional machinery. The role of MED1 in macrophage biology and the relevant disease state remains to be investigated. To study the molecular mechanism by which MED1 regulates the M1/M2 phenotype switch of macrophage and the effect on atherosclerosis, we generated MED1/apolipoprotein E (ApoE) double-deficient (MED1ΔMac/ApoE-/-) mice and found that atherosclerosis was greater in MED1ΔMac/ApoE-/- mice than in MED1fl/fl/ApoE-/- littermates. The gene expression of M1 markers was increased and that of M2 markers decreased in both aortic wall and peritoneal macrophages from MED1ΔMac/ApoE-/- mice, whereas MED1 overexpression rectified the changes in M1/M2 expression. Moreover, LDLR (low-density lipoprotein receptor)-deficient mice received bone marrow from MED1ΔMac mice showed greater atherosclerosis. Mechanistically, MED1 ablation decreased the binding of PPARγ (peroxisome proliferator-activated receptor γ) and enrichment of H3K4me1 and H3K27ac to upstream region of M2 marker genes. Furthermore, interleukin 4 induction of PPARγ and MED1 increased the binding of PPARγ or MED1 to the PPAR response elements of M2 marker genes. Our data suggest that MED1 is required for the PPARγ-mediated M2 phenotype switch, with M2 marker genes induced but M1 marker genes suppressed. MED1 in macrophages has an antiatherosclerotic role via PPARγ-regulated transactivation.

Patients with TTC7A Deficiency Causing Multiple Intestinal Atresia with Combined Immunodeficiency (MIA-CID) Can Survive Well Beyond Infancy

Introduction: TTC7A gene mutations have recently been identified as a cause of Multiple Intestinal Atresia with Combined Immunodeficiency (MIA-CID), an extremely rare condition with few patients surviving beyond infancy. This case series reports the clinical characteristics of six surviving children. Methods: Records of all children with MIA-CID and TTC7A mutations confirmed on genetic testing at two tertiary intestinal rehabilitation centres were reviewed retrospectively to identify common phenotypic findings and current management strategies. Results: Six patients (three female) were identified. All patients had multiple intestinal atresias requiring early surgical intervention. All patients required additional operations for acquired strictures. Mean length of remaining small bowel in continuity was 8 cm (range 4 to 20 cm). Only one patient was able to tolerate any substantial enteral feed; all patients required lifelong parenteral nutrition, except one, who achieved full enteral autonomy after a multivisceral transplantation at five years of age. Four patients received bone marrow transplants with successful engraftment as treatment of their immunodeficiency, which was characterized primarily by severe T-cell lymphopenia. All patients developed intestinal failure associated liver disease. One child had a multivisceral transplantation, a second is listed and a third is undergoing evaluation for multivisceral transplantation. The other three have stable liver disease on non-soybean-based lipid formulations. There is variable occurrence of pulmonary, dermatological and cardiac disease in our cohort. Conclusions: Patients with TTC7A deficiency leading to MIA-CID can survive beyond infancy with intensive surgical and multidisciplinary medical management. Patients develop intestinal strictures requiring proximal stomas and are unable to establish enteral autonomy leading to parenteral nutrition dependence. Intestinal failure associated liver disease is seen in all patients. The optimal timing of bone marrow transplantation requires consideration, especially for patients considering multivisceral transplantation and for those who do not have a fully matched donor.

Outcomes of solid tumor malignancies in patients of Jehovah’s Witness faith.

e18176 Background: Jehovah's Witness (JW) patients with cancer present unique challenges due to their refusal of blood and blood products which limits supportive care options in the setting of cancer treatment. This study was aimed at determining their cancer specific outcomes. Methods: A retrospective analysis of case records of JW with solid tumors presented between 2005 and 2015 was conducted. Patient demographic data, diagnosis, treatment details and survival outcomes were collected and analysed. Results: A total of 63 JW were identified (39 females; 24 males); median age was 70 years (25-90). The most common cancer was breast (n=16) followed by colorectal (n=10). 27 JW had advanced disease (TNM stage 4), 48 patients had ECOG 0-1 and 22 JW had anaemia (Hb&lt;120g/L for women and &lt;130g/L for men) at the time of diagnosis. While surgery and radiation were uniformly accepted when recommended, 7 out of 42 refused chemotherapy. Median OS was 64 months in early stage (TNM stages 1-3) and 11 months in advanced stage. At presentation blood transfusion was considered unacceptable by all patients under any circumstances. 19 patients rejected transfusion even after symptomatic or life threatening anaemia. They had 1 year overall survival rate of 42% compared with 84% for those who did not require transfusion (HR 4.31, CI 1.96- 9.49, P=0.001). When adjusted for age and ECOG, these results were significant in early stage (P &lt;0.026) but not in late stage (P= 0.6). Treatment was suboptimal in 14 JW patients due to early treatment discontinuation or inferior chemotherapy regimen used to avoid pancytopenia. Median OS in this group was 15 months compared to 46 months who had optimal treatment (HR 2.5, CI 1.07- 6.27, P =0.034). 17 JW received bone marrow stimulating agents like erythropoietin or G-CSF. They helped with dyspnoea and maintaining chemotherapy dose intensity. Conclusions: Refusal of blood components may have negative impact on long term cancer outcome of JW due to compromised chemotherapy dose intensity. Strategies to deliver optimum therapy while being sensitive of religious perceptions will help in improving outcomes for this group.

Cervical Papanicolaou Smears in Hematopoietic Stem Cell Transplant Recipients: High Prevalence of Therapy-Related Atypia during the Acute Phase

Hematopoietic stem cell transplant (HSCT) recipients have a higher risk of cervical cancer. Papanicolaou (Pap) smear is the standard tool for screening cervical cancer, but there is limited research about the cervical cytology in HSCT recipients. Here, we retrospectively included adult female patients who underwent allogeneic or autologous HSCT at National Taiwan University Hospital during 2009 to 2015 and reviewed their Pap smears before and after HSCT. There were 248 allogeneic and 131 autologous HSCT recipients in our study. In allogeneic HSCT recipients, 38.7% (96 of 248) had pre-HSCT Pap smears and 17.1% (44 of 248) had post-HSCT Pap smears. In the autologous HSCT recipients, 35.1% (46 of 131) had pre-HSCT Pap smears and 13.7% (18 of 131) had post-HSCT Pap smears. Compared with allogeneic HSCT recipients without post-HSCT Pap smears, more recipients with post-HSCT Pap smears received bone marrow–derived stem cells (18.2% versus 4.9% respectively; P = .0077) and had longer overall survival (median overall survival, not reached versus 22.1 months; P < .0001). The abnormal rates of post-HSCT Pap smear were 13% (6 of 44) and 11% (2 of 18) in allogeneic and autologous recipients respectively, higher than in the general Taiwanese population (1.22%). Infections were rare in post-HSCT Pap smears. Of note, 11% (5 of 44) of post-HSCT Pap smears from allogeneic recipients showed therapy-related atypia, manifesting as enlarged hyperchromatic nuclei, vacuolated cytoplasm, and occasional tadpole-like cells. These atypical cytological features mimic precancerous lesions, but cervical biopsies and human papilloma virus tests were negative. The atypical cytological features resolved spontaneously in the subsequent follow-up Pap smears. On average, Pap smears with therapy-related atypia were sampled at day +77, significantly earlier than those without therapy-related atypia (P = .016). Therapy-related atypia was more frequent in post-HSCT Pap smears sampled within 100 days after HSCT (before day +100, 4 of 5, 80%, versus after day +100, 1 of 39, 2.56%; P = .0002). The strong temporal relationship suggests these atypical cytological changes resulted from conditioning regimen, most likely busulfan-containing chemotherapy. No therapy-related atypia were observed after total body irradiation or nonbusulfan-containing chemotherapy. In conclusion, therapy-related atypia was common in post-HSCT Pap smears sampled within 100 days after HSCT. Clinical information is critical for correct cytological diagnosis.

Vulvar and vaginal graft versus host disease: A healthcare clinic initiative

Objective:In patients receiving bone marrow transplantation (BMT), their mucosa becomes altered and sclerotic changes in the female external genital organs occur. Although a few studies have specifically addressed vulvar and vaginal graft versus host disease (VVGvHD) and its repercussions on the sexual health and quality of life of patients, VVGvHD can be overlooked by health practitioners. The objective of the study is to describe the initiation of a health care clinic specializing in VVGvHD in a general tertiary hospital.Methods:A VVGvHD clinic was founded as a part of BMT daycare in a joint initiative of the nursing staff and the medical director of the department and a gynecologist specializing in vulva and vaginal disease. Patients were assessed for vulvovaginal symptoms, such as dryness, burning, itching, pain to touch, pain during intercourse, and dysuria. These patients might be subsequently referred to the VVGvHD clinic according to their needs assessed by daycare nurses. Treatment guidelines were developed by the specialist gynecologist.Results:A total of 81 women aged 2–66 years (median age = 38 years) visited the clinic from 2009 to 2015. Of these women, 70 received an allogeneic transplant and 11 underwent autologous transplantation before consultation in our clinic. VVGvHD was detected in 54% of the patients.Conclusions:The VVGvHD clinic was developed to fulfill the specific needs of female patients who underwent BMT. The pioneer clinic was founded as a joint effort of the multidisciplinary team. Evidence supporting the optimum treatment for this condition is insufficient. This was the main reason for performing this study to explore the clinic that was newly based in Israel. VVGvHD may be a fluctuating condition with frequent deterioration and improvement. Therefore, regular clinical examinations are necessary.